DOI

Received: May 2022 – Accepted: July 2022

¹ Semmelweis University, Faculty of Health Sciences, Department of Dietetics and Nutritional Sciences

Keywords

polyphenolic compounds, diabetes, medicinal food, estimated glomerular filtration rate (eGFR), DPPH, ABTS, ORAC, FRAP, CUPRAC, Folin-Ciocalteu methods, gallic acid equivalent, catechin equivalent

2. Introduction

Bees have been around for 125 million years, and their evolutionary success has allowed them to become a perennial species that can utilize virtually every habitat on Earth. This ability to survive is largely due to the chemical composition and application of the special products they produce (honey, beeswax, venom, propolis, pollen and royal jelly). Propolis, the bees’ remedy against pathogenic microorganisms, has been used by mankind as a medicine since ancient times [1].

Propolis, known in Hungarian as bee glue, is a sticky, resinous substance, produced by bees (Apis mellifera L.) from beeswax, saliva and sap from the bark, buds and leaves of trees [2, 3]. They collect mainly from poplar, but also from birch, willow, horse chestnut, pine, oak, elm and alder [4]. The composition of propolis is mostly made up of plant resins, waxes, essential oils and pollen. In addition to these, it also contains smaller amounts of other substances, such as compounds partially produced by bees [2].

Bees use propolis in the hive many ways, including for disinfection, construction and maintenance of the hive, and for protection [2, 4, 5, 6], as well as to keep the humidity and temperature in the hive stable throughout the year, to seal holes and cracks and the inner wall of the hive. Propolis is also an important element of the so-called social immune system of honeybees, which, thanks to its antipathogenic (antimicrobial) properties, provides a certain general protection to the entire bee family against infections and parasites [8, 9].

3. Characteristics of propolis

The physical and chemical components of propolis, its quality and the possibilities of using it for physiological and medicinal purposes depend on the origin of the propolis, i.e., the climate, the botanical source and the species of the bees [4, 10]. The color of the product also depends on the origin, it is usually brown, but at the same time all the shades from yellow to black appear in it, in many cases with a reddish or greenish hue. The smell of propolis is aromatic, with notes of honey, resin, wax and vanilla mixed in it. Its taste is very characteristic [4].

Raw propolis typically consists of 50% plant resin, 30% wax, 10% essential and aromatic oils, 5% pollen, and 5% of other organic matter. More than 300 components have been identified in propolis, which differ depending on the source [9].

The compounds found in propolis include polyphenolic compounds (phenolic acids, flavonoids and their esters, e.g., caffeic acid phenyl ester), diterpenes, sesquiterpenes, lignans, aromatic aldehydes, alcohols, amino acids, fatty acids, organic acids, hydrocarbons, vitamins and minerals [9, 11].

The main bioactive components of propolis are flavonoids, which greatly contribute to the pharmacological effects of propolis. The amount of flavonoids is used as a criterion when evaluating the quality of temperate climate propolis. Flavonoids have a wide spectrum of biological properties, such as antibacterial, antiviral and anti-inflammatory effects [9].

Although volatile substances make up only 10% of the components of propolis, they are responsible for the characteristic resinous smell and contribute of the beneficial effects of propolis on health. Volatile substances are dominated by terpenoids, which play an important role in distinguishing good quality propolis from poor quality or counterfeit propolis, and also exhibit antioxidant, antimicrobial and other biological effects [9].

Although different bee species prefer different plants, even the chemical profile of propolis produced by the same species is not always the same. The composition of propolis varies by bee colony, location and season, and this makes it difficult to study it and make consistent health claims [12]. The protective properties of the bioactive substances found in propolis can also provide significant benefits in maintaining human health [5].

In recent years, several studies have confirmed that different propolis samples can be completely different in terms of chemical composition and biological activity [1, 7].

4. Propolis-containing products

A significant number of propolis-containing products are available on the market: medical and over-the-counter preparations, foods and drinks that help maintain health [7].

Propolis tincture is an extract of raw propolis made with a solvent (most often a mixture of water and ethanol). According to our knowledge, there are practical and application questions related to propolis tincture that should be answered and uniform regulations should be applied:

• Various preparation recipes are known;
• Soaking raw propolis for different lengths of time results in different tinctures;
• Differences in the extraction solvent (different amount and ethanol concentration) affect the composition of the preparations;
• The relationship between raw propolis and the composition of the tincture is not known.

In addition to tinctures, other propolis-containing foods are also available, such as lozenges, propolis honey, capsules filled with propolis extract [3].

In some countries, standardized propolis products with a constant bioactive substance concentration are already available [13].

5. Dose and safety

Clinical studies on mice and humans report that propolis and its constituents are generally well tolerated and non-toxic, except when used in very large amounts [5].

Determining the exact dose of propolis, on the basis of the studied population, the dosage regimen, compliance (accurate taking of the substance) and the purity of the product, faces significant difficulties, since the phenolic compounds found in propolis vary according to geographical origin, the bioactivity can also differ significantly, which makes it difficult to determine the correct dosage [14]. According to a particular study, based on previous animal experiments and applying a margin of safety, the safe dose of propolis for healthy humans is 70 mg/day [14].

Egy tanulmány szerint a korábbi állatkísérletek alapján és egy biztonsági tartalékot alkalmazva az egészséges emberek számára a propolisz biztonságos dózisa 70 mg/nap [15].

6. Physiological and therapeutic effects of propolis

Propolis has received increasing attention in recent years due to its beneficial effects on the human body. It is increasingly accepted as a preventive and therapeutic agent. However, the bioavailability of the useful substances found in propolis varies, which is also influenced by individual physiological conditions. According to a study, as a result of the consumption of propolis, its active ingredients can also be detected in the blood plasma [16].

6.1. Fighting infections, the immune system

Propolis can be considered as a potential medicinal food (“nutraceuticals”). Propolis extracts have anti-bacterial, antiviral and antifungal effects [3]. The immunoprotective and antioxidant properties of propolis are explained by its bioactive phytochemical components, regardless of its origin. A 2019 review cited immune system support as a health benefit of propolis [5].

The effect of propolis supplementation has also been studied among patients infected with the COVID-19 virus. In a recent, high quality (double-blind, placebo-controlled) study conducted in 2020, the effect of propolis on clinical symptoms was investigated. Infection was confirmed with a PCR test in participants aged 18 to 75. Members of the intervention group (n=40) received tablets containing 300 mg of Iranian green propolis extract three times a day for 2 weeks, while the control group (n=40) received no such treatment. The main result of the study was that the clinical symptoms of the disease improved faster in the group receiving propolis in terms of the duration and severity of the initial symptoms [17].

6.2. Cancerous diseases

Propolis has an antioxidant effect, which can be beneficial for the body in terms of neutralizing free radicals formed in excess [3], thus it can contribute to the regulation and control of inflammatory processes, tumor formation and aging processes. Its anti-inflammatory properties have been demonstrated in connection with propolis samples of Brazilian, Chinese and Malay origin. Its antitumor effect has been proven not only in in vitro, but also in vivo experiments taking place in living organisms) [3].

According to the results of another research, Brazilian red propolis had antioxidant properties and significantly reduced the percentage of survival of human tumor cells under laboratory conditions [10]. Alcoholic extracts of Turkish propolis also exhibited an inhibitory effect on the growth of tumor cells against human tumor cells (liver, colon, breast, cervix, prostate) [18].

The study, the aim of which was to find out whether propolis and the polyphenolic/flavonoid compounds contained in it can have an inhibitory effect on the growth of human bladder tumors in a cell culture, ended with promising results. Based on this, propolis may be suitable for auxiliary treatment of the disease in addition to surgery, to reduce or prevent the chance of tumor recurrence [19].

6.3. Diabetes

In relation to the effect of propolis on the human body, the reduction of blood sugar levels has also been studied [2]. According to the reliable, aggregated, comprehensive analysis of the results of several studies with similar objectives, the use of propolis reduced the fasting blood sugar level by 0.8 mmol/l compared to the subjects who did not receive treatment. In addition, taking propolis also reduced the value of HbA1c (A subunit of hemoglobin. The Ed.), which indicates the evolution of the blood sugar level of the examined person in retrospect over a period of 1 to 3 months. It is interesting to mention that the treatment did not affect the insulin level, so it follows that the drop in blood sugar level was not due to the effect of insulin. Almost 400 diabetic patients took part in the study, who were treated with 226 to 1,500 mg of propolis per day for 56 to 180 days. According to the authors, despite the positive results, further research is still needed regarding the type (composition) and dosage of propolis. This is so because the dose ranges were wide and the places of origin of the propolis samples used were varied [2]. In the studies related to propolis, it was stated that it is important to know the geographical and botanical origin, because they can affect the biological activity of the propolis, its effect and the composition of its organic components [3].

The objective of another study was to investigate the effect of Brazilian green propolis on type 2 diabetes patients through changes in blood test data. 80 people participated in the study, of which 39 received a placebo. The 41 people in the other group received 226.8 mg Brazilian green propolis per day during the 8-week period. The results indicate that Brazilian green propolis used in the aforementioned quantity and frequency can reduce then deterioration of uric acid levels and eGFR (estimated Glomerular Filtration Rate) values, which indicate kidney complications, in patients with type 2 diabetes [20].

In connection with the healing of diabetic leg ulcers, a favorable effect was reported in the study of Australian propolis samples. A favorable wound-healing role was also mentioned in connection with Chinese propolis extracts [3].

6.4. Cardiovascular diseases

In a human study published in 2017, changes in blood lipid levels were investigated as a result of oral application of propolis solutions. In the double-blind, placebo-controlled clinical trial, 35 of the 67 subjects received propolis, while 32 were given a placebo supplement (without propolis). In the propolis group, a significant increase in HDL (High Density Lipoprotein) was observed after 90 days. This effect may contribute to the reduction of the risk of cardiovascular diseases [21].

A 2019 review paper mentions lowering blood pressure as one of the health benefits of propolis. In this literature review, a total of 63 publications were reviewed, the majority of which were reports on animal experiments, but some key human studies were also included. According to the results, propolis can be an effective antioxidant and anti-inflammatory agent. Based on this, it is presumably effective against various chronic diseases, e.g., in preserving the health of the cardiovascular system, reducing atherosclerosis and reducing high blood pressure [5].

6.5. The skin and the nervous system

The components of propolis can be widely used to heal wounds and the human skin itself, and can also contribute to reducing the symptoms of some nervous system diseases (Alzheimer’s disease, Parkinson’s disease) [5].

In addition to research related to nervous system diseases, the protective effects of propolis on retinal cells have also been reported [22]. Propolis can also be used to prevent various eye diseases, such as macular degeneration in the aging population and myopia in the younger generation, but further studies are needed to prove this [5].

The range of commercially available propolis-containing skin care products is expanding, with creams and body lotions predominating. According to the advertisements, the majority of skin care products have a „soothing, moisture-rich, anti-aging” effect, and are also effective against eczema [23].

6.6. Alimentary canal

When examining the beneficial properties of various propolises, in the case of Brazilian green propolis, the stimulation of the functioning of the intestinal system was mentioned, as well as its beneficial effect in the treatment of gastric ulcers, while the liver protective function of propolis was proven in animal experiments [3].

The polyphenols in propolis can support the development and maintenance of a healthy intestinal flora by limiting the growth of pathogenic bacteria and, in addition, prevent their adhesion to human intestinal cells [24]. The possible therapeutic effect of propolis on inflammatory bowel diseases is still being investigated today, but many experiments still need to be performed before clinical application can begin [5].

6.7. Allergizing effect

In addition to its many beneficial physiological effects, propolis can also trigger allergic reactions (swelling, dermatitis, hives) in susceptible individuals. This is most common among beekeepers, but it also depends on individual sensitivity [3]. Therefore, it is recommended that the therapeutic use of propolis products is always carried out under medical supervision [5].

6.8. Summary of physiological and therapeutic effects

Several studies have proven that the observed beneficial physiological effects are not the result of a single prominent compound, but rather the combined effect of the complex components of propolis [9].

Overall, it can be stated that as a natural substance with good medicinal properties, propolis and its components can be used in a wide range of ways, including wound and skin healing, and in the treatment of some neurological diseases and atherosclerosis. Interest in the health effects of propolis and the number of publications have been continuously increasing in the last 30 years. However, even more human clinical studies are needed to confirm the beneficial effect of propolis for specific population groups. Preclinical studies support the antioxidant and anti-inflammatory effect of propolis, which can prevent or slow the progression of various chronic diseases, including heart disease, diabetes, high blood pressure, tumors and neurodegenerative diseases (e.g., Alzheimer’s disease) [5].

7. New areas of application of propolis

One of the areas of use of propolis can be to improve the growth performance and productivity of farm animals. Based on our knowledge so far, it can be said that propolis has a beneficial effect on the normal laboratory values, growth and productivity of the animals included in the studies. In addition, it is considered as a possible alternative to antibiotics in the production of animal feed, because it has the advantage that it does not induce resistance in microorganisms [25].

Another area of intensive research in the last few years have been the application of propolis in food preservation. Food preservatives primarily include antimicrobial and antioxidant agents. Antimicrobial agents added to foods serve two purposes: to stop the natural spoilage of food and to avoid/control contamination by microorganisms, including pathogenic microorganisms. Antioxidants are used to extend shelf life and prevent spoilage. Propolis favorably combines antioxidant and antimicrobial properties. However, its large-scale use as a food preservative has not yet been realized, as this would require proper standardization of the product [7].

8. Antioxidant properties of propolis

The antioxidant properties of propolis are manly determined by the bioactive components found in it, primarily the phenolic compounds, depending on the botanical and geographical origin. The phenolic compound profile of propolis is slightly different from that of honey. While in the former, the profile is mainly determined by the botanical origin, and the dominant flavonoids are quercetin, myricetin, chrysin, apigenin, luteolin, pinocembrin and pinobanksin, and of phenolic acids, p-hydroxybenzoic acid, p-coumaric acid, cinnamic acid, gallic acid, ferulic acid and caffeic acid, in propolis, which typically comes from poplar and birch in Central Europe, chrysin, kaempferol, apigenin, pinocembrin and pinobanksin are the most characteristic and, in addition to phenolic acids, their esters (e.g., caffeic acid and ferulic acid esters) also occur. Among the latter, the phenylethyl ester of caffeic acid is outstanding in terms of tumor prevention properties (although its effect also depends on the synergistic effect of other accompanying phenolic compounds). The polyphenols in propolis have been proven to inhibit the formation of amino, oxide and peroxide type free radical, as well as the formation of complexes between free radicals and transition metals, and also lipid peroxidation [26].

In addition to the differences depending on the origin of propolis, the literature is not uniform regarding the extraction method of the antioxidant compounds, and the differences can significantly influence the extraction results.

Based on the available data, extraction was mainly carried out with different mixtures of ethanol and water in the experiments, but extraction with methanol and other solvent also occurs. Regarding the methods for determining the antioxidant properties, only the results of experiments based on in vitro spectrophotometry have been reported, including the determination of radical scavenging properties (DPPH – 2,2-diphenyl-1-picrylhydrazyl, ABTS – 2,2’-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), ORAC (Oxygen Radical Absorbance Capacity), as well as total polyphenol content (Folin-Ciocalteu method) and total flavonoid content. Although in the case of propolis extracts from different locations, polyphenol contents of the same order of magnitude as in the case of honey were measured (typically in the 18-500 mg gallic acid equivalent/ml range), in the case of a Turkish sample there was also a value above 19,000 mg gallic acid equivalent/ml, and values above 1,000 mg gallic acid equivalent/ml were also measured for Brazilian samples. The situation is similar with regard to the total flavonoid content, where the majority of the samples stayed in the range typical of honey (1-25 mg catechin equivalent/ml), however, there were also extremely high values: nearly 5,000 mg catechin equivalent/ml in the case of an Algerian propolis, and a value over 29,000 mg catechin equivalent/ml in the case of a Turkish propolis. As for the radical scavenging ability, once again, values in the range of honey are reported by researchers (e.g., 50-80% inhibition in the case of DPPH radicals), but extreme values are also typical here (e.g., 90.7–99.34% inhibition in the case of a Malay honey). Similar to honey, numerous studies have confirmed in the case of propolis its effectiveness in the case of tests carried out on various animal and human bodily fluids and cell cultures, in terms of antioxidation properties.

9. Synergistic interaction of propolis and honey

Propolises of different origins can show a synergistic interaction not only with each other, but also when mixed with honey. Following the mixing of propolis extracts from Iraq, it was possible to prove a synergistic effect against various pathogens (E. coli, S. aureus, C. albicans) in microbiological tests. Similarly, in animal experiments, the extent of the wound healing effect (reepithelization) was increased in the case of a propolis mixture [27].

Due to the deterioration of sensory characteristics, propolis is typically mixed with honey in a proportion of no more than 1%. Even at this concentration, a four- to fivefold increase in the amount of phenolic compounds, phenolic acids and flavonoids was measured, and the anthocyanin and carotenoid content of the mixture also increased several times. Of the flavonoids, especially the amount of galangin, chrysin, pinocembrin and pinobanksin, while of phenolic acids, the amount of ferulic acid, caffeic acid and p-coumaric acid increased. The radical scavenging (ABTS, DPPH) and metal ion reducing capacity (FRAP – Ferric Reducing Antioxidant Power, CUPRAC – Cupric Reducing Antioxidant Capacity), measured by various in vitro methods, also showed multiple increases [26].

The synergistic interaction of propolis and honey was also confirmed in antimicrobial tests. In the research, in the case of antibiotic-resistant strains of E. coli, S. aureus and C. albicans, honey strengthened the effect of propolis both in cultures of individual strains and their mixtures [29].

10. Acknowledgment

Az anyag összeállításához Bencsik Boglárka demonstrátor hallgató is hozzájárult.

11. References

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[16] Yesiltas, B.; Capanoglu, E.; Firatligil-Durmus, E.; Sunay, A.E.; Samanci, T.; Boyacioglu, D. (2014): Investigating the in-vitro bioaccessibility of propolis and pollen using a simulated gastrointestinal digestion System. J. Apic. Res. 2014, 53, pp. 101–108.

[17] Miryan, M., Soleimani, D., Dehghani, L., Sohrabi, K., Khorvash, F., Bagherniya, M., Sayedi, S. & Askari, G. (2020): The effect of propolis supplementation on clinical symptoms in patients with coronavirus (COVID-19): A structured summary of a study protocol for a randomised controlled trial. Trials, 21.

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[20] Fukuda, T., Fukui, M., Tanaka, M., Senmaru, T., Iwase, H., Yamazaki, M., Aoi, W., Inui, T., Nakamura, N. & Marunaka, Y. (2015): Effect of Brazilian green propolis in patients with type 2 diabetes: A double-blind randomized placebo-controlled study. Biomedical Reports, 3(3), pp. 355-360.

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DOI

Received: May 2022 – Accepted: July 2022

¹ University of Debrecen, Institute of Food Science

Keywords

spices, bread, fortification, macroelement, nutrient reference value (NRV)

2. Introduction

Conscious food consumers have recognized and accepted that the consumption of “healthier” foods can prevent certain diseases. (The term “healthier” food can be misleading, because according to EU laws, “unhealthy” food cannot be placed on the market. In the present case, I accept that this term represents a comparative. The Ed.) In addition to researchers, industry also strives to develop and produce „healthier” foods [1, 2]. Bread and bakery products play an important role in the human diet. Wheat bread is generally an efficient source of energy and contains irreplaceable nutrients. The fortification of these products with functional components is widespread in order to improve health protection [3]. Examples of such components are spices and herbs [1, 2], as well as byproducts of cereals, pseudo-cereals, vegetable or fruit products [3].

Several publications have reported on the fortification of breads with various substances, which was also detailed by Varga-Kántor et al. [4].

Fortified breads are more valuable than plain bread from a nutritional physiology point of view, as they contain ingredients that have a beneficial effect on health. Spices and herbs are examples of this.

These plants, which are equally important in the pharmaceutical industry and in gastronomy, have been used by mankind for a long time. They have a strong, concentrated smell and taste, so consuming large amounts of herbs may even have an adverse sensory effect [5]. The spices we used and their active ingredients are applied in the treatment of several diseases. Many scientific books and studies have reported on their use in this area.

A detailed description of the spices used and measured in our experimental program can be found in the following sources: Pushpagadan [6], Kurian [7], Peter [8], Charles [9], Gupta [10], Kintzios [11], Chen [12], Sasikumar [13], Pandey [14]. These works describe the origin of the spices, their physiological effects on humans, and their history.

Spices that contain compounds with potent antioxidant and disease-preventing effects have a high element content, which is important for a balanced diet and lifestyle. Table 1 contains the measurement results of other authors for these parameters.

While Barin et al. [15] and the USDA [21] measured a calcium concentration of 22,000 mg/kg in basil, the values reported by other authors were between 10,000 and 15,000 mg/kg. In the case of dill, the results of Rahmatollah and Mahbobeh [17] and the USDA [21] were similar, while lower concentrations were measured by Özcan [16]. The USDA database [21] had a higher calcium content for oregano than Barin et al. [15] and Öczan [16]. The calcium content of caraway was similar [16, 21], while in the case of chives, there was a 1,000 mg/kg difference [15, 21]. When looking at rosemary, the results show that two authors obtained similar results of about 8,000 mg/kg [18, 19], but in the other two cases, higher calcium contents were determined [16, 21]. In the case of garlic granules, there was no significant difference between the measured concentrations [20, 21].

In the case of potassium content, the highest concentration was measured in dill. Potassium contents close to 34,000 mg/kg were determined by two authors [16, 21], but the values of Rahmatollah and Mahbobeh [17] were twice as high. In the case of basil, concentrations above 24,000 mg/kg were measured by three authors [16, 18, 21], however, Ozygit et al. [19] only measured a potassium content of 8,000 mg/kg. In the case of oregano, the measured values of this parameter were between 12,000 and 19,000 mg/kg. In the case of caraway, the results obtained differed significantly. The USDA database [21] described a potassium content of more than 26,000 mg/kg in chives. The potassium content of rosemary was similar in two cases [16, 21]. Ozygit et al. [19] measured a lower value than this, while the value measured by Özcan [16] was approximately 2,000 mg/kg higher. There was no significant difference in the results of garlic granules.

Looking at the magnesium content results, there were significant differences for all spices between the concentrations measured and published by the researchers. The determined values were in the order of thousands, with the exception of garlic granules.

A similar trend can be observed for the sodium content, as the measured concentrations differ significantly in the various studies.

In the case of phosphorus, similar values were obtained by the authors for oregano and garlic granules. In the case of the other herbs, there were significant differences between the results of the authors, often in the order of thousands.

Regarding the sulfur content of the spices, it can be seen that very different concentrations were determined in dill, and very high values were obtained for the garlic granules.

3. Materials and methods

3.1. Preparation of the breads

In this study, the macroelement content of seven dried spices (basil, dill, oregano, caraway, chives, rosemary and garlic granules) and 42 fortified breads was determined.

The raw materials for the products were purchased in a supermarket in Debrecen. After the analysis of the spices, the breads were prepared based on the recipe of Varga-Kántor et al. [4] and Kántor et al. [20].

These samples contained different concentrations of dried spices (0, 2, 4, 6, 8, 10 and 12 g). The additional ingredients were wheat flour (BL 55, 500 g), 10% vinegar (8 g), sunflower oil (44 g), salt (5 g), granulated sugar (5 g), yeast (30 g), milk (2.8% fat, 150 ml) and 25 °C water (100 ml). The ingredients were stored at room temperature, in their original packaging, in the dark or in a refrigerator until the products were prepared. After kneading, the leavening time was 1 hour at room temperature. The next step was the shaping of the loaves, followed by resting for 10 minutes. The breads were baked in a convection oven at 210 ⁰C and 95% humidity for 15 minutes (RXB 606, convection oven, Budapest, Hungary). After baking, the products were left in the oven for 6 minutes.

3.2. Determination of element content

In the case of spices, the samples purchased in the store were not dried, but the breads were dried according to standard MSZ 20501-1 [22]. Sample preparation was carried out based on the method of Kovács et al. [23]. After measuring the bread into a digestion tube, 10 ml of nitric acid (69% v/v; VWR International Ltd., Radnor, USA) was added to the sample and it was left to stand overnight. Predigestion was carried out at 60 °C for 30 minutes. After cooling, before the main digestion, 3 ml of hydrogen peroxide (30% v/v; VWR International Ltd., Radnor, USA) was used, and then the sample was kept at 120 °C for 90 minutes. After cooling, it was diluted with high purity water (Millipore SAS, Molsheim, France) and the mixture was filtered on filter paper (388, Sartorius Stedim Biotech SA, Gottingen, Germany). The element content was determined using an ICP-OES (Inductively coupled plasma optical emission spectrometer, Thermo Scientific iCAP 6300, Cambridge, UK) instrument. The wavelengths used were 315.8 nm (Ca), 769.8 nm (K), 280.2 nm (Mg), 818.3 nm (Na), 185.9 nm (P) and 180.7 nm (S).

3.3. Statistical analysis

To determine the mean, standard deviation and statistically verifiable differences, one-factor analysis of variance (Tukey and Dunnett’s T3 test) was used with SPSS statistical software (version 13; SPSS Inc. Chicago, Illinois, USA). Measurements were carried out in triplicate.

3.4. Calculation of the daily intake value from the nutrient reference value (NRV)

NRV values are contained in Regulation (EU) No 1169/2011 of the European Parliament and of the Council [24] and the EFSA scientific bulletin [25]. Data are presented as a percentage for 100 g of product, which means the consumption of approximately 1.5 slices of bread.

NRV (%) = (element content of the bread/daily reference intake) x100

In the case of sodium, the daily reference intake is 2,000 mg [25], while no relevant data were found for sulfur.

4. Results and evaluation

4.1. Measurement results of the element content of the spices

The results of the macroelement content measurements of the herbs examined by us are presented in Table 2. The values are given on an as received basis.

The highest calcium concentration was measured in the case of basil, followed by chives. The measured values were similar in dill and oregano. A value of more than 10,000 mg/kg was measured in rosemary, while in the case of caraway, the concentration was higher than 6,000 mg/kg. The lowest calcium content was measured in garlic granules.

In the case of the potassium content, basil and dill exhibited outstanding values. In caraway, chives and garlic granules, the concentrations were above 10,000 mg/kg. The herbs oregano and rosemary showed the lowest values among the plants analyzed.

The highest magnesium content was measured in basil, which had twice the concentration of dill, which also had a high value compared to the other samples tested. The values for oregano, caraway, chives and rosemary were between 2,000 and 3,000 mg/kg. The lowest magnesium content was found in garlic granules.

An outstanding sodium content was measured in dill, but the concentrations were very low on the other samples. Values higher than 100 mg/kg were obtained for the basil and garlic granule samples. In the other cases, the measured values were below 100 mg/kg.

In case of the phosphorus content, the concentration in caraway was the highest, followed by the garlic granules. Values between 3,000 and 4,000 mg/kg were measured for basil, dill and chives. Rosemary had the lowest concentration.

During the determination of the sulfur content, concentrations of more than 1,000 mg/kg were measured in each sample. Similarly outstanding values were obtained for dill and garlic granules, followed by chives with a concentration of more than 3,000 mg/kg. For the other spices, with the exception of basil, sulfur contents between 1,000 and 2,000 mg/kg were detected.

Comparing the results of Table 1 with the concentration measured by us, it can be stated that in the course of our analyses, higher values were obtained for the calcium content of chives and the sulfur content of dill and caraway, while lower values were measured for the potassium content of dill, oregano and chives and the phosphorus content of chives. However, from the measured concentrations it can be concluded that the results obtained are similar to the values mentioned in the other studies, except for the data regarding the sodium content. In this case, results that are significantly different from the literature data can be seen.

4.2. Measurements results of the breads fortified with spices

The breads were prepared based on a predetermined recipe [4, 20], and samples without spices were also prepared. Based on the results, it was determined that the measured parameters of the control breads were similar to the literature data (Ca: 476; K: 2,200; Mg: 260; Na: 2,585; P: 1,478 and S: 1,008 mg/kg [4]; Ca: 510; K: 2,418; Mg: 285; Na: 3,180; P: 1,512 and S: 948 mg/kg [20]), except for the sodium content.

The results are reported on a dry matter basis (Tables 3, 4 and 5). In the tables, significant deviations from the control samples are marked with the letter „a” in each column.

4.2.1. Calcium content results

The calcium contents of the fortified breads are presented in Table 3. In most cases, the addition of spices increased the element content of the fortified breads. The biggest increase was experienced in the case of basil breads. In this case, the difference compared to the control sample was more than 500 mg/kg. Breads containing dill, chives or rosemary showed a difference of about 300 mg/kg between the control bread and the breads fortified with 12 g of spices. The additional value of breads fortified with oregano and caraway was smaller, around 100-200 mg/kg.

Although the calcium content of oregano exceeded 10,000 mg/kg, the increase experience in fortified breads was not as large as when using spices with similar calcium content.

The lowest calcium content was determined in the bread containing 12 g of garlic granules. The other samples showed significant differences compared to the control sample.

4.2.2. Potassium content results

The potassium contents of the samples analyzed are also shown in Table 3. Based on the results, it appears that the addition of basil and dill increased the potassium concentration of the breads the most. In the case of the breads fortified with 12 g of caraway, a difference of about 300 mg/kg was found, compared to the control sample. In the other cases, the difference was barely more than 200 mg/kg.

In terms of potassium content, the greatest increase was exhibited by the breads with basil and dill, followed by the product samples with caraway, oregano and garlic granules. In all cases, the lowest values were measured in breads with rosemary and chives. This difference is probably due to the difference already present in the control breads.

4.2.3. Magnesium content results

Data on the magnesium content of the breads are presented in Table 4. In the spices, the highest values were measured in the case of basil and dill, which affected the magnesium content of the breads. When examining the samples, the highest magnesium content was determined in the product fortified with basil. This was the largest difference (200 mg/kg) between the control bread and the a sample containing 12 g of spice. This result was followed by breads fortified with dill. Products with caraway and rosemary showed a similar trend, with a maximum difference of 60 mg/kg between the sample containing the most spice and the control bread. In the case of breads with oregano, the increase was 40 mg/kg in the bread containing the most spice compared to the control product. For those spices where the magnesium content was below 2,000 mg/kg, there was no significant difference in the fortified breads. When looking at samples with the same amount of spices, the highest values were measured in the basil breads in all cases. The lowest concentrations were detected in breads with garlic granules and chives.

4.2.4. Sodium content results

Data on the sodium content of the products prepared can be seen in Table 4. Regarding the samples, the measured values were between 2,400 and 3,100 mg/kg. The sodium content of the spices was low compared to the other macronutrients, except for dill. There was no statistically proven difference in the results of the products with basil, oregano, caraway and garlic granules. In the case of the samples with dill, the reason for the increase was probably the sodium content of the spice, which affected the element content of the final products.

In the case of chives and rosemary, the sodium content of the spices was below 100 mg/kg. Therefore, the decrease in one case and the increase in the other cannot be explained. When considering the same amount of spices, the highest sodium content was measured in breads with rosemary, dill and basil. This tendency was also observed in the case of the control breads. Since the breads were made by hand, it is possible that the distribution of table salt was not uniform in all cases, and this may also cause differences.

4.2.5. Phosphorus content measurement results

The phosphorus content results of the samples are presented in Table 5. Based on the results, the phosphorus content of the breads was similar. In most cases, there was no statistically verifiable difference between the samples. Smaller differences were measured in the products with caraway and garlic granules, which is due to the phosphorus content of the spices. The phosphorus content of these spices exceeded 4,000 mg/kg. For the other spices, concentrations below 4,000 mg/kg were determined in all other cases.

The highest phosphorus content was measured in the products with caraway, followed by breads with dill, garlic granules and basil. The lowest concentration was measured in the products flavored with chives, however, low phosphorus contents were measured in the breads with rosemary and oregano as well.

4.2.6. Sulfur content results

Table 5 shows the sulfur content of the breads. Based on the concentrations obtained, larger differences were measured in the products with basil and garlic granules, and smaller differences were measured in the case of the other fortifications when increasing the amount of spices. Analyzing the spices, the highest sulfur content was determined in dill and garlic granules (more than 7,000 mg/kg), however, even the addition of larger amounts of spices to the breads did not increase the measured concentrations. It can be seen that in the other cases the value of the measured parameter did not increase with the increase in the amount of spices. Minor differences could be observed, but the sulfur content of the spices had no significant effect on the sulfur content of the final products.

Daily intake contribution results calculated from the nutrient reference value (NRV)

Tables 3, 4 and 5 show the daily contribution values for (Ca, K), (Mg, Na) and (P) per 100 g of product, respectively.

In the case of calcium content, the consumption of 100 g of control bread per day covers 5 to 6% of the daily calcium intake. By increasing the amount of spices in the samples, these values also increased. The highest contribution was calculated for the breads with basil, followed by the products with rosemary, dill, and chives.

In the case of the potassium content, the contribution of the control breads was between 10 and 11%. When different amounts of spices were added, a smaller increase was calculated tan in the case of the calcium content. For the breads with the most spices, the increase in daily contribution was even as high as 3% (samples with basil and dill) compared to the control products.

The magnesium content of the control breads is responsible for approximately 7% of the daily magnesium intake. In this case, once again, the most significant differences were observed in the bread with basil. With 12 g of spice, the increase was more than 5% compared to the control sample.

The sodium intake values of all samples were around 12 to 13%. In the case of the products with dill and rosemary, the values were higher.

In terms of phosphorus content, all of the reads covered more than 20% of the daily phosphorus intake. The contribution of the samples with caraway showed a minimal increase. For the breads fortified with 12 g of spices, the increase was around 2% compared to the control products.

5. Conclusions

As the results show, the spices themselves have a high macronutrient content. In terms of calcium, potassium, magnesium and sodium, basil exhibited outstanding values. High values were also measured in dill, chives, oregano and garlic granules.

The content of calcium, potassium and magnesium in fortified breads increased. In the case of calcium, the biggest difference was found in the products with basil. A clear increase was also observed for the other samples as well, except for the application of garlic granules.

Outstanding results were also achieved in terms of the potassium content of products with basil and dill. A difference of almost 600 mg/kg was measured between the control sample and the bread with 12 g of spice.

There was no significant difference in the magnesium content. A greater increase in concentration was only observed for the products with basil.

The samples with rosemary and dill showed a slight increase in sodium content, which can also be observed with the same amount of spices.

No significant differences were found in the phosphorus and sulfur contents; similar values were measured.

Based on the results, the largest daily contribution of macronutrients was provided by the breads with basil, followed by the breads with dill. In the case of the sodium content of the breads, the daily intake contributions of the products with dill and rosemary were the largest.

Overall, it was possible to prepare products whose element content in most cases differed significantly from that of the control breads, so the contribution of the products to the daily reference values also increased.

6. Acknowledgment

This research was financed by the Higher Education Institution Excellence Program of the Hungarian Ministry of Innovation and Technology (NKFIH-1150-6/2019), within the framework of the 4th thematic program of the University of Debrecen.

7. References

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DOI: https://doi.org/10.52091/EVIK-2021/4-1-ENG

Received: September 2021 – Accepted: November 2021

¹ Hungarian University of Agriculture and Life Sciences, Buda Campus, Institute of Food Science and Technology, Food Science Research Group

Keywords

microalgae, protein, composition, feeding, food use, climate change, carbon footprint

2. Introduction

By 2050, the population of Earth will grow to nearly 10 billion, an increase of about 25% compared to today’s population. In addition, the significant depletion of our Earth’s water resources also makes it necessary to restructure our diet, as the amount of water needed to produce 1 kg of food is 13,000 liters for cattle, 5,520 liters for chicken, while only 50 liters for peas or lentils. All this means that we can expect a significant increase in the price of foods of animal origin, which will mean that we will have to reduce the proportion of them in our diet significantly.

The different plant protein sources make a positive contribution to the protection of the environment and the fight against climate change due to their more efficient use of water. On the other hand, legumes require 30 to 70% less synthetic fertilizers due to their nitrogen-binding properties, they increase soil power and have a positive effect on soil biology. It is also a known fact that due to nutrient transformation losses, the production of 1 kg of animal protein requires at least 6 to 16 times more cultivable area. In addition, the carbon dioxide footprint of the production of foods, especially of those based on beef, is about 10 times more than that of plant-based foods.

The structure of food consumption in Europe is characterized by a 59% proportion of the daily protein intake coming from protein sources of animal origin (meat, fish, milk), with proteins of plant origin representing only 41% of the total. More than 50% of the latter is wheat protein. As a result, a few cereals (wheat, corn, rice) could become staple foods, leading to geographical homogeneity of foods, dietary monotony and nutritional imbalances, increasing the risk of micronutrient deficiencies, overweightness and chronic obesity, as well as NCDs (Non Communicable Diseases), including cardiovascular diseases, stroke, cancer and diabetes.

Based on all this, it is becoming increasingly important to identify and investigate alternative plant and other protein sources, in addition to the protein sources mentioned above, which can contribute to meeting the protein needs of an increasing human population, as well as to addressing the unbalanced nutritional status of them.

An important group of protein crops are legumes (e.g., dried beans, runner beans, chickpeas, horse beans, lentils, grass peas, black-eyed peas, dried peas, autumn and spring peas) with a high protein content of 20 to 40% on average. Dry leguminous seeds are rich in protein and lysine, but low in sulfur-containing amino acids. At the same time, field crops with good nutritional values but low protein content (e.g., sunflower, canola, corn, sorghum, rice, wheat) are low in lysine and rich in sulfur-containing amino acids. Taking into account the positive nutritional values of the two plant groups, products containing complete plant protein can be developed by their combined use.

The protein content of various crops (Table 1) may show significant variability not only between species but also within a given species. In addition, protein content can also be altered by environmental factors and the food processing technology.

Other alternative protein sources include single cell proteins (SCPs) produced by fermentation technologies, seaweeds living in saltwater, duckweed species living in freshwater and various insect species. Protein content values of the different sources can vary widely depending on the species, the cultivation technology and the nutrient supply (Table 2).

3. Characterization and occurrence of microalgae

Algae, also called seaweeds, are eukaryotes capable of photosynthesis. Algae represent one of the oldest life forms on Earth, having existed on our planet for about 3 billion years. They produce one-third of Earth’s living matter and about 50% of its organic carbon [1]. These plants have survived all geological epochs and climate changes. Algae still account for 90% of Earth’s oxygen production. These organisms have allowed life to form on Earth, and they use the power of sunlight to produce organic food from inorganic materials through photosynthesis. In many respects, algae are the most diverse living things in the world. They have the simplest structure and are closely related to bacteria. The most complex ones, Charophyceae species, resemble kelp to the point of confusion. The smallest algae are picoalgae with a size of 0.5 μm, while the largest are 50 to 100 m long Macrocystis species (Phaeophyceae). They occur under the most extreme conditions in fresh and salt water, hot springs, on snow and ice surfaces, in the soil and in the upper layer of some rocks [2]. Algae are mostly eukaryotes, typically classified as „lower” plants that have no true stems, roots and leaves, and are generally capable of photosynthesis. Algae are widely classified into Rhodophyta (red algae), Phaeophyta (brown algae) and Chlorophyta (green algae) and are classified as macroalgae and microalgae by size. Macroalgae (seaweeds) are multicellular, large size algae that are visible to the naked eye, while microalgae are microscopic single-celled organisms and may be prokaryotes, similar to cyanobacteria (Chloroxybacterium), or eukaryotes, similar to green algae (Chlorophyta).

Microalgae, as excellent sources of various organic carbon compounds, can be used in the manufacture of health supplements, drugs and cosmetics [2]. they can also be used in wastewater treatment, atmospheric CO₂ reduction and the production of biofuels. A wide range of bio-products can be extracted from microalgae, such as polysaccharides, lipids, pigments, proteins, vitamins, bioactive compounds and antioxidants [3]. In addition to all this, they are playing an increasingly important role in the feed and food industries (Figure 1).

4. General composition of microalgae

As with all other higher plants, the chemical composition of algae varies depending on the type of cultivation, such as environmental parameters, temperature, illumination, pH and mineral content of the medium, CO₂ supply and mixing speed: 9-77% protein, 6-54% carbohydrate, 4-74% lipid (Table 3)

4.1. Protein and amino acid content of microalgae

Based on research results it can be stated that algae are a source of protein with an amino acid composition similar to that of plant proteins. Examination of the net protein utilization, i.e., of the proper amino acid composition, digestibility and biological value, also led to a similar result.

Several microalgae species produce large amounts of various essential amino acids and proteins that can be used in foods and feeds, one of the main reasons they occupy a prominent place among alternative proteins. Certain species of microalgae can produce as much protein as other rich protein sources, e.g., eggs, meat and milk [6].

In addition, the amino acid pattern of almost all algal species is very similar to the protein pattern of many foods. Of the amino acids, they are only low in cysteine and lysine. Since cells are able to synthesize almost all amino acids, they can be used to provide the essential amino acid needs of both humans and animals [7]. The composition of amino acids synthesized by microalgae, especially the amount and composition of free amino acids, varies greatly, depending on the species, growth conditions and the growth phase [8].

In addition, microalgae proteins are easily digestible and have a relatively high nutritional value. Microalgae produce 2.5 to 7.5 tons/ha/year protein [9], for example, the green microalgae Chlorella is a rich source of different types of marketed proteins. Another protein-rich microalga is Arthrospira. Proteins from microalgae lower cholesterol levels by activating cholecystokinin. They also have other important enzymatic effect [10]. For example, the microalga called Lyngbya majuscula produces microcholine-A, a protein with immunosuppressive effects [11]. The microalga Nostoc produces a protein called cyanovirin, which is known to have an antiviral activity against both HIV and the influenza virus [12]. At the same time, Anabaena and Porphyridium species produce the enzyme SOD (superoxide dismutase), which protects against oxidative damage, while Isochrysis galbana produces the enzyme carbonic anhydrase, which plays a key role in the conversion of CO₂ to carbonic acid and bicarbonate. Microcystis aeruginosa produces a number of amino acids, including proline, serine, glycine and valine.

4.2. Fatty acids

Polyunsaturated fatty acids play an important role in tissue protection and have a beneficial effect on health. Omega-3 and omega-6 fatty acids are especially important for humans, but the human body is unable to produce these fatty acids. Therefore, intake from an external source, such as various foods, is essential. Docosahexaenoic acid (DHA), linoleic acid, eicosapentaenoic acid (EPA), arachidonic acid and gamma-linolenic acid have been shown to lower cholesterol levels, delay aging, protect membrane integrity and prevent cardiovascular diseases [13,14]. Several species of microalgae capable of synthesizing these valuable fatty acids have been studied. These studies have shown that Pavlova lutheri produces large amounts of polyunsaturated fatty acids [15], Arthrospira platensis mainly produces and accumulates stigmasterol, sitosterol and γ-linolenic acid [16], while Porphyridium produces arachidonic acid, Nannochloropsis, Phaeodactylum, Nitzschia, Isochrysis and Diacronema species produce eicosapentaenoic acid and Crypthecodinium and Schizochytrim microalgae produce docosahexaenoic acid in significant amounts [17, 18, 19].

The polyunsaturated EPA and DHA are also pharmaceutically very important omega-3 fatty acids. They play a key role in the treatment of inflammatory diseases, heart problems, arthritis, asthma and headaches, among other things [20, 21, 22].

4.3. Polysaccharides

Polysaccharides are widely used in the food industry, primarily as gelling and thickening agents. Many polysaccharides used in the food industry, such as agar, alginates and carrageenans are extracted from macroalgae, e.g., Laminaria, Gracilaria and Macrocystis species [8]. One of the most promising microalgal species, the unicellular red alga Porphyridium cruentum produces a galactan exopolysaccharide, that can replace carrageenan in many cases. Chlamydomonas mexicana also produces significant amounts of polysaccharides, and it is used in the United States as a soil improver. Sulfated algal polysaccharides also have pharmacological properties and they play a prominent role in stimulating the human immune system [23].

4.4. Photosynthetic pigments

It can be said in general that each species of algae has a specific pigment combination that creates its characteristic colour. In addition to chlorophylls, the primary photosynthetic pigments, supplementary or secondary pigments, such as phycobilins and a number of carotenoids are also produced by microalgae. These natural pigments have the ability to improve the efficiency of light energy utilization and provide protection for algae from the harmful effects of solar radiation. They are used preferentially when added to foods and feeds as natural antioxidants and colourants [24].

4.4.1. Carotenoids

Carotenoids are naturally occurring pigments that play a role in the formation of the colour of fruits, vegetables and other plants [25]. They are typically isoprenoid polyene pigments derived from lycopene, ranging in colour from yellow to red, and are produced by de novo photosynthetic organisms and some other microorganisms [8]. Carotenoids ingested with foods or feeds are either accumulated or metabolized by the body. Carotenoids can be found in the meat of various animals, eggs, fish skin (trout, salmon), crustaceans (shrimp, lobster, Antarctic krill, crab) and subcutaneous fat, skin, egg yolk, liver and the feather of birds (e.g., poultry) [26].

In algae, carotenoids primarily play the role of sunscreen and light collector, i.e., they protect the photosynthetic apparatus from light damage [24]. They also play a role in phototropism and phototaxis. Some microalgae undergo carotenogenesis in response to various environmental effects (e.g., light, temperature, salts, nutrients). During this, algae stop their growth dramatically alter their carotenoid metabolism, resulting in the accumulation of secondary carotenoids [27].

There are more than 600 carotenoids in nature, about 50 of which show provitamin A activity. These include α-carotene, β-carotene and β-cryptoxanthin [28]. A β-Carotene protects membrane lipids from peroxidation, thus preventing and reducing the development of many serious and fatal diseases, such as cancer, cardiovascular disease, Parkinson’s disease and atherosclerosis [29, 30, 31].

Relatively few carotenoids are used in the food and feed industries: β-carotene and astaxanthin, lutein, zeaxanthin, lycopene, etc. Among microalgae, the main carotenoid-producing species are Dunaliella salina and Haematococcus pluvialis which produce significant amounts of β-carotene and astaxanthin, respectively. The microalga Dunaliella salina produces β-carotene in an amount that accounts for 10 to 14% of its dry matter content [32].

β-Carotene serves as an essential nutrient, mainly as a food colouring, and is increasingly used in various dietary supplements due to its health-protective effects, but is also used preferentially by the cosmetics industry [33].

In the food industry, β-carotene is also used regularly in various soft drinks, cheeses, butter or margarines due to its beneficial physiological effect, as it possesses a provitamin activity [34].

Astaxanthin has a number of beneficial properties, including improving eye health, muscle strength and endurance, protecting the skin, reducing premature aging, inflammation and damage caused by UVA radiation. It also plays an important role in animal feeding, as it promotes growth and reproduction, improves vision, has an immunostimulatory effect and also aids in post-injury regeneration [35, 36].

Numerous studies have shown that daily intake of astaxanthin protects cells and tissues from oxidative effects and that its effect on free radicals is significantly, about 500 times more intense than that of vitamin E. The microalga Haematococcus pluvialis produces 4-5% astaxanthin on the basis of dry biomass [37], so its dried biomass is marketed as a rich source of astaxanthin and is sold on the market at a price of about 2,500 US $/kg.

4.4.2. Chlorophyll

Each alga contains one or more chlorophylls. Their primary photosynthetic pigment is chlorophyll-a, and it is also the only chlorophyll in Cyanobacteria (blue-green algae) and red algae (Rhodophyta). Like all higher plants, Chlorophytes (true green seaweeds) and Euglenophytes (flagellate seaweeds) also contain chlorophyll-b; chlorophyll-c, -d and -e are found in many other marine algae. The quantity of chlorophylls is usually up to 0.5-1.5% of the dry matter content [38].

In addition to being used as a food and pharmaceutical colourant, chlorophyll derivatives also have a health-protective effect. They are also traditionally used because of their wound healing and anti-inflammatory properties [39]. Epidemiological studies conducted in the Netherlands (Cohort Study) have shown a significant correlation between chlorophyll consumption and a reduction in the risk of colon cancer [40].

4.4.3. Phycobilins

In addition to chlorophyll and carotenoid lipophilic pigments, Cyanobacteria (blue-green algae), Rhodophytes (red algae) and Cryptophyta algae also contain so-called phycobilins, which are coloured, fluorescent pigments. Like chlorophylls, they bind to proteins (phycobiliproteins) which, contrary to chlorophyll-protein complexes located in the membranes, are water-soluble proteins and are important components of the photochemical system. Significant amounts of phycobilins, mainly blue phycocyanin and red phycoerithrin are found in Spirulina algae and Porphyridium, respectively.

The use of phycobilins is quite widespread. In addition to being widely used in clinical immunofluorescence studies to detect fluorescently labeled antibodies as a fluorescent marker [38], phycocyanin is currently used in Japan and China as a natural colourant in foods, such as chewing gums, candies, dairy products, jellies, ice creams and soft drinks. It is also widely used in the cosmetics industry, for example, in lipsticks, eyeliners and eyeshadows [41].

According to a study, phycocyanin is one of the most versatile blue colourant, providing a bright blue colour to various jellies and coated soft candies [42], while a number of pharmacological properties are also attributed to phycocyanin, including antioxidant, anti-inflammatory, neuroprotective and hepatoprotective effects [43, 44, 45].

4.5. Tocopherols and sterols

Tocopherols are widespread in nature, occurring in both lower and higher plants as parts of the photosynthetic system.

Research in this area has revealed that Euglena has the highest content of tocopherols among the various microalgae species [46].

The sterols produced by plants are called phytosterols. Microalgae can make a major contribution to the production of phytosterols, they can be considered as efficient and promising sources for their large-scale production. Some microalgae contain high levels of sterols. Microalgae sterols have health-protective, cholesterol lowering and anti-inflammatory properties, and are effective in the treatment of certain neurological diseases, such as Parkinson’s disease [47, 48], and are increasingly used in the food industry as dietary supplements and food ingredients [49, 50]. Some microalgae, such as species belonging to the genera Pavlova and Thalassiosira, are rich in sterols [51, 52].

4.6. Vitamins, minerals

Microalgae biomass is a valuable source of almost all essential vitamins, as it contains vitamins B₁, B₂, B₃, B₅, B₆, B₁₂, C, E and H, among other things, and its mineral content (e.g., Na, K, Ca, Mg, Fe, Zn and trace elements) is also significant [53]. The vitamin B₁₂ and iron content of some microalgae, such as Spirulina species, is particularly high, therefore, they are often used in foods and dietary supplements made for vegetarians.

The vitamin content of algae depends on the genotype, as well as the stage of the growth cycle, the nutrition of the algae and the light intensity. Thus, their vitamin content can be increased by selecting the right species, choosing the right culture conditions and/or by genetically modifying them. However, the vitamin content of the cells can be significantly reduced by using inappropriate environmental conditions, harvesting and biomass drying methods [54].

4.7. Antioxidants

Microalgae are photoautotrophic organisms, that is, organisms that depend on light as the energy source to produce organic molecules from inorganic molecules. This process is known as photosynthesis, and the food chain is usually based on these organisms. During their development, these organisms have developed an effective defense system against various abiotic effects affecting them, such as high levels of free radicals and reactive oxygen species [23]. Due to the high antioxidant content of certain algae species (e.g., Isochrysis galbana, Chlorella vulgaris, Nannochloropsis oculata, Tetraselmis tetrathele, Chaetoceros calcitrans), their use has been increasing in some cosmetics (e.g., sunscreens) and in functional foods.

The research of Natrah et al. [55] has shown that the methanolic extract of some fresh/untreated microalgae exhibits an antioxidant activity that is higher than that of α-tocopherol, but lower than that of the synthetic antioxidant BHT (butylhydroxytoluene). However, the latter and BHA (butylhydroxyanisole) being synthetic antioxidants, their safe use if questionable, as their use in high doses may be carcinogenic and tumorigenic [56, 57].

4.8. Other biologically active components

Microalgae are undoubtedly a large repository of versatile compounds with significant biological activity, as well as unique and interesting structure and function [58].

In recent decades, marine microorganisms, especially Cyanobacteria, have become the focus of medical research aimed at developing new drugs and antibiotics. Data published up to 1996 revealed about 208 Cyanobacterial compounds exhibiting biological activity. By 2001, the number rose to 424. The compounds identified include various lipoproteins (40%), alkaloids, amides, etc. [59], many of which have cytotoxic, antitumor, antimicrobial (antibacterial, antifungal), antiviral (e.g., anti-HIV) activities, as well as biomodulatory, for example, immunosuppressive and anti-inflammatory effects [59, 60].

Numerous studies have shown that microalgae may also contain compounds that are effective in treating cancer and tumors by inhibiting angiogenesis. Angiogenesis is a physiological process during which new blood vessels emerge from existing blood vessels. Although angiogenesis is a normal process, pathological conditions can develop under certain conditions, such as cancer, atherosclerosis, arthritis, diabetic retinopathy and ischemic stroke. Pathological angiogenesis promotes the development and growth of tumors [61, 62]. Fucoxanthin and fucoxanthinol, found in many species of microalgae, have been shown to inhibit the process of angiogenesis in the aortic ring of rats by reducing the formation and growth of microvessels [63]. Fucoxanthin has also been shown to protect DNA from photooxidation [64]. Microalgae, especially blue-green algae, are currently considered to be potential sources of active ingredients that can be used in the treatment of cancer, as several studies have demonstrated their anti-cancer effects [65].

5. Some major microalgal species

Although many indigenous microalgal populations have been used for various purposes for centuries, their large-scale cultivation has only begun in the last few decades [66]. Of the assumed roughly 30,000 species of microalgae, only a few thousands are kept in stock collections [67, 68], of which a few hundred are considered more important due to their chemical composition, and very few are grown in industrial quantities [69].

The biotechnologically most relevant microalgae include green algae (Chlorophyta), such as Chlorella vulgaris, Haematococcus pluvialis, Dunaliella salina and Spirulina maxima, which belongs to the phylum of Cyanobacteria. Their cultivation, marketing and use are very significant, mainly as dietary supplements and animal feed additives.

5.1. Spirulina species

Spirulina (Arthrosphira) algae are a tiny, filamentous, freshwater, spiral-shaped, blue-green algae species that is abundant in the alkaline lakes of Mexico and Africa and has been consumed by the local population since ancient times [59]. Its characteristic feature is that its cell membrane is very weak, and this makes it easy to utilize. It is also one of its important physiological features that it can easily become a colloidal solution when exposed to moisture and is very easy to digest. Spirulina is widely grown all over the world (3,000 tons/year) and is used as a food and feed supplement due to its high protein content (60 to 70%, including 18 amino acids, 8 of which are essential) and its excellent nutritional value. For example, its γ-linoleic acid content is remarkably high [70, 71]. Its digestibility and absorption are superior to both animal and plant proteins. It contains the vitamins important to the body (C, B₁, B₂, B₅, B₆, B₉, B₁₂, A, E), trace elements, of which iron, iodine, calcium, sodium, potassium, copper, magnesium, manganese, zinc, phosphorus, selenium, chromium and vanadium are the most significant. It is a particularly good source of iodine and potassium. It stimulates the immune system greatly due to its high content of β-carotene, chlorophyll and γ-linolenic acid. Its polyunsaturated fatty acid content is significantly higher than that of marine fish. Spirulina contains high levels of GLA (gamma-linolenic acid), which is found in greater amounts only in breast milk. Spirulina has a number of health-protecting effects: it lowers high blood fat levels, cholesterol levels, high blood pressure, elevated blood sugar levels, is suitable for treating kidney failure, and promotes the growth of probiotics, such as Lactobacilli, in the gut [19]. It is the main source of natural phycocyanin, used as a natural blue colourant in foods and cosmetic products, and also as a biochemical tracer in immunoassays [70, 71, 72].

5.2. Chlorella vulgaris

This species of algae is one of the oldest, simplest plants on Earth. Its nearly 4% chlorophyll content, strong cell wall and high pigment and cellulose content make the detoxifying effect of Chlorella unique. It binds and removes heavy metals from the body, cleanses the intestinal flora. By improving liver function, it helps to remove other contaminants, in addition to heavy metals, and to detoxify the body.

Its physiological effects are similar to those of Spirulina: it is high in protein, contains all the essential amino acids, and it is a storehouse of various vitamins, trace elements and minerals. Chlorella vulgaris has been used in the Far East since ancient times in alternative medicine, as well as in the preparation of various traditional foods. It is widely cultivated and used, primarily in animal feeding, aquaculture and as a dietary supplement, in many countries, including China, Japan, Europe and the United States. The health-protecting effects of Chlorella are manifested, for example, in the rapid healing of stomach ulcers and other wounds, and it is useful in the treatment of constipation, anemia, hypertension, diabetes, infant malnutrition and neurosis. The prophylactic role of the glycolipids found in Chlorella against the development of atherosclerosis and hypocholesterolemia has been demonstrated by research [58]. However, one of the most important substances in Chlorella is ß-1,3-glucan, which is an active immunostimulant, it binds free radicals and reduces the amount of blood fats [19].

The γ-linolenic acid (GLA) content of Spirulina and Chlorella is very high. The role of GLA in the functioning of the body is extremely diverse. On the one hand, it is important for the proper functioning of the immune system, and on the other hand, it has an anti-inflammatory effect, lowers blood pressure and improves blood circulation. It prevents platelets from sticking together, thus reducing the risk of formation of blood clots. It has a positive effect on cholesterol levels, thus reducing the risk of atherosclerosis. It improves nervous system function and eliminates excess fluid from the body.

5.3. Haematococcus pluvialis

This freshwater microalgae, with a size of barely 0.1 mm, attracted the interest of researchers early on. Haematococcus pluvialis is the plant with the highest astaxanthin content (1.5-3.0% dry weight) based on previous research. This carotenoid pigment has a very strong radical scavenging effect that exceeds the antioxidant properties of β-carotene, vitamin C and vitamin E. The astaxanthin production of the algae is a natural reaction to environmental stress. Thanks to the protective functions of astaxanthin, in a state of deep sleep, these algae can survive without food and water for more than 40 years, so they can easily survive the heat of summer or the icy cold of winter. They will only wake up again and regain their original green, active state when living conditions are right again. As a result, algae defied the harshest environmental conditions even at the early stages of Earth’s history. The ability of certain algae species to survive both droughts and ice ages is due to their astaxanthin shield. Astaxanthin is a bioactive antioxidant that has been shown to be effective against Alzheimer’s disease and Parkinson’s disease, as well as macular degeneration in both animal and human experiments. In some cosmetics, the astaxanthin used can slow down the aging process of the skin. In addition, the immune-boosting and anti-inflammatory effects of astaxanthin have been reported, as well as its beneficial effects on the development of cardiovascular diseases and atherosclerosis.

Haematococcus pluvialis is currently a natural source of this pigment, its commercial utilization is outstanding, especially in aquaculture (salmon and trout farming) [73]. There is another natural source of astaxanthin, however, the yeast Xanthophyllomyces dendrorhous requires large amounts of expensive nutrients for proper pigmentation [36].

5.4. Dunaliella salina

Dunaliella salina is a halotolerant microalgae, its natural habitats being salt lakes. It is able to accumulate large amounts of β-carotene, which is why this species of algae is sought after mainly as a food colourant. Research has shown that the Dunaliella salina community in Pink Lake, Victoria, Australia, can produce up to 14% carotenoids [74] and some Dunaliella algae can contain up to 10% in cultivated cultures.

Higher β-carotene content can be achieved with adequate nutrient supply under high salt and light conditions [75, 76]. Similarly to Haematococcus algae, Dunaliella contains significant amounts of astaxanthin. However, Haematococcus is a freshwater algae that is difficult to grow in outdoor cultures because it is easily infected, requiring a closed system, and the extraction of astaxanthin is more complicated than in the case of Dunaliella, as Haematococcus has a thick cell wall that has to be disrupted by physical methods.

6. Use for animal feed

Today, many species of microalgae (e.g., Chlorella, Tetraselmis, Spirulina, Nannochloropsis, Nitzchia, Navicula, Chaetoceros, Scenedesmus, Haematococcus, Crypthecodinium) are used to feed farm animals, pets and fish.

Even a small amount of microalgae biomass has an immunostimulatory effect, which results in growth stimulation, disease resistance, has antiviral and antibacterial effects, improves absorption and the colonization stimulation of probiotic cultures such as Lactobacilli, and thus results in an increase in reproductive performance and weight [77]. By providing feeds that contain algae, the appearance of the animals improves visibly, which is manifested in a healthy skin and a shiny coat, both in the case of farm animals (poultry, cows, breeding bulls) and in the case of pets (cats, dogs, rabbits, ornamental fish and birds) [78].

Feed is the main exogenous factor influencing animal health and accounts for a significant part of the major cost of animal husbandry, and so it is very important to identify high quality, chemical and toxic substance free alternative protein sources that can replace or complement traditional protein sources [26]. The results of a large number of nutritional and toxicological evaluations have demonstrated the suitability of algal biomass as a valuable feed supplement [38]. Currently, about 30% of the global algae production is sold for feed purposes [53].

Becker et al [53] performed feeding experiments on broiler chickens, in which conventional proteins were replaced with species of different microalgae, namely Chlorella, Euglena, Oocystis, Scenedesmus, Spirulina, usually at a rate of 10%. In laying hens, no differences were found in egg production and egg quality (size, weight, shell thickness, egg solids, albumin index, etc.) and in feed conversion efficiency between birds fed with algae-containing feeds and the control birds.

However, Haematococcus microalgae can also be used as a natural colourant in the feeding of broiler chickens, making the skin of the birds yellower and the egg yolk more orange [79]. Studies were performed on chickens that were fed the biomass (5% or 10%) of red microalgae (Porphyridium species). Although no difference was found in the body weight and egg count of the chickens, the composition of the meat and eggs showed decreased cholesterol levels (by 10%) and healthier fatty acid composition and increased linoleic acid and arachidonic acid levels (by 29% and 24%, respectively). In addition, the colour of the egg yolk was darker due to a carotenoid content that was 2.4 times higher than average [80]. At the same time, it was observed that chickens fed with algal biomass consumed 10% less food in the case of feeds containing either 5% or 10% algae, and had serum cholesterol levels significantly lower (by 11% and 28%, respectively) than that of the control group.

Microalgae biomass is a feed with excellent nutritional values and is eminently suitable for breeding pigs. They can be used to replace traditional proteins such as soy flour or fish meal, and their acceptance is not difficult for the animals [38].

It is hypothesized that algae may be an excellent food source for ruminants, as these animals are able to digest even the cell wall of unprocessed algae. However, only a limited number of experiments have been performed with these animal species, as these procedures are expensive and large amounts of algae are required to perform appropriate feeding experiments. However, some experiments have shown that sheep, lambs and cattle were unable to digest the carbohydrate fraction efficiently when fed certain algal species (e.g., Chlorella, Scenedesmus obliquus and Scenedesmus quadricauda) [81, 82]. Better digestibility was achieved when Spirulina accounted for 20% of the total sheep feed, and it was observed that in calves fed a diet containing Scenedesmus algae, there were only minimal differences when compared to animals fed the control feed [83].

Microalgae feeds are currently used mainly to supplement and replace zooplankton used for the breeding of fish, fish fry and other aquatic animals (crustaceans, etc.) [84, 85]. The species most often used in aquaculture are Chlorella, Tetraselmis, Isochrysis, Pavlova, Phaeodactylum, Chaetoceros, Nannochloropsis, Skeletonema and Thalassiosira [86, 87].

Microalgae contain nutrients that are essential for aquatic animals, and these determine the quality, growth, health and disease resistance of the farmed animals. Mixed microalgae cultures have been shown to be useful for animal growth to provide adequate protein composition, vitamin content and high levels of polyunsaturated fatty acids (mainly EPA, AA and DHA), which are vital for the survival and growth of many freshwater and marine animals in the early stages of life [88]. One of the beneficial effects of algae is attributed to the fact that they increase the food intake of marine fish offspring, which enhances their growth and survival, and also improves the quality of fish meat [89]. In addition, the presence of algae in the breeding tanks of European sea bass larvae has been shown to increase digestive enzyme secretion [90]. When many aquatic species, such as Salmonidae (salmon and trout), shrimp, lobster, marine vertebrates, goldfish and koi carp are kept under intensive conditions, carotenoid colourants are added to their feed to achieve their characteristic muscle colour. Carotenoids, such as astaxanthin and canthaxanthin, have beneficial effects on animal health, growth and reproduction, by promoting the development of larvae [33].

7. Food use

In the early 1950s, microalgae were used to replace certain foods and were often used as single cell proteins in the diets of malnourished children and adults. Today, in human nutrition, microalgae are marketed in the form of various dietary supplement pills, capsules and liquids [91].

Gross et al conducted research in which Scenedesmus obliquus algae was added to the normal diet of children (5 g/day) and adults (10 g/day) during a four-week test period. Blood panel, urine composition, serum protein and uric acid concentration and weight change were measured, but the parameters analyzed did not deviate from normal values, only a slight increase in body weight was observed.

The same authors subsequently performed a three-week study in both slightly (Group I) and severely (Group II) malnourished four-year-old children. Four-year-old children in Group I (10 g algae/day) showed a significant weight gain (27 g/day) compared to children in the control group who received a normal diet, and no adverse symptoms were experienced. Group II was fed a diet enriched with 0.87 g algae/kg body weight, replacing only 8% of the total protein with algae protein, and the daily weight gain was approximately seven times that of children in the control group, while all anthropogenic parameters were normal. The authors concluded that the significant improvement in health can be attributed not only to algae protein but also to other important health-protective and immune-enhancing components [92].

Large-scale production of microalgae suitable for human consumption has been growing worldwide. There are many forms of microalgae and other health-protective dietary supplements on the market, such as various pills, powders, capsules, lozenges and liquids [23, 93]. Microalgae are also used in the preparation of various foods, such as algae pastries, biscuits, breads, snacks, candies, yogurts and soft drinks, which also provide the health and immunomodulatory effects associated with microalgae biomass [94].

Despite some reluctance on the part of consumers over novel foods in recent decades, there is a growing consumer demand for natural, health-enhancing foods today. Thus, functional foods containing microalgae biomass are also becoming increasingly popular. These products are also proving to be very attractive and varied from a sensory point of view, and their consumption also brings health benefits, satisfying consumer needs in all respects [23].

8. Acknowledgement

We are grateful for the funding of the NKTH, TKP2020-NKA 24 "Thematic Excellence Program”.

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DOI: https://doi.org/10.52091/EVIK-2021/4-3-ENG

Received: May 2021 – Accepted: September 2021

¹ University of Debrecen, Institute of Food Science

Keywords

bread, basil, total polyphenol content (TPC), flavonoid content, element content, functional food

2. Introduction

Basil is mostly grown in Mediterranean countries. Its leaves, fresh or dried, are used as a seasoning. It is also known as an herb, its use is recommended against headaches, coughs, diarrhea, constipation, warts, worms and kidney problems, among other things [1]. Rosmarinic acid is responsible for its antioxidant effect, as it binds free radicals. It cannot be used against fungi, but its antibacterial and antiviral activities are known [2]. It has outstanding values in terms of element content, which has been studied by Ghanjaoui et al. [1], Özcan and Akbulut [3], as well as Özcan [4], among others.

Baking is one of the oldest human occupations related to food preparation. When prehistoric man settled down and switched to gathering and farming, cereals became the most important sources of food. With continuous learning and technological development, it has been possible to process the collected seeds into various products [5]. One such product was bread, which was very different from the product that is being consumed today. Nevertheless, many types of bread are still made today. In the Middle East, flat bread dominates, in China, steamed bread, while in America, corn-based product dominate. Bread is mostly made from wheat and some other commonly used cereals, as the proteins of these are best suited to make the right product [6].

According to papers in the literature, there have been several attempts to enrich bread with different substances. Raba et al. [7] prepared and tested bread enriched with garlic and basil. Suleria et al. [8] used aqueous garlic extract, but breads enriched with yellow pepper flour [9], ginger powder [10], turmeric [11], waste onion powder [12], brown algae powder [13], horseradish leaf powder [14], raspberry and strawberry oil cake [15], as well as garlic and its preparations [16] have also been made.

Since, to the best of our knowledge, there have been few bread fortification experiments with spices and then few studies were carried out, we first used basil in our experiment and examined what measurable changes were caused in the baked product by the addition of the spice.

3. Materials and methods

3.1. Bread preparation

In our experiments, different parameters of 7 bread samples were examined. The ingredients were selected and the breads were prepared according to the method of Kántor et al. [16]. The ingredients used to make the products were purchased at retail stores. Basil was added to the bread dough before kneading in the amounts of 0.00 g; 4.25 g; 8.5 g; 12.75 g; 17.0 g; 21.25 g and 25.5 g (Table 1).

During the experiment, the total polyphenol (TPC), flavonoid and element content of basil was examined, and then the dry matter, total polyphenol (TPC), flavonoid, crude fat, cruse protein and macroelement contents of the breads were determined. In the case of the breads, the measured values were reported on a dry matter basis.

3.2. Determination of total polyphenol content (TPC)

For the analysis, the method of Singleton et al. [17] was used for both basil and the breads. The samples were soaked in methanol (Scharlab S. L., Spain): distilled water (80:20), then the mixture was filtered through 292 pleated filter-paper (Sartorius Stedim Biotech S.A., Gottingen, Germany). 1 ml of the resulting solution was pipetted into a test tube, to which 2.5 ml of Folin-Ciocalteu reagent (VWR International S.A.S., France) was added. After 5 minutes, 2 ml of 75 g/l sodium carbonate (Scharlab S. L., Spain) was added to give a colored compound, the absorbance of which was measured with a spectrophotometer (Evolution 300 LC, Thermo Electron Corporation, England) at 760 nm. For the determination of the total polyphenol content, calibration solutions were prepared from a gallic acid (Alfa Aesar GmbH&Co. KG, Karlsruhe, Germany) stock solution. The absorbance of the calibration solution series was also measured, from the results of which a calibration curve was constructed. The total polyphenol content of our sample solution was determined using this curve. Results are given in mg GAE/100 g.

3.3. Determination of flavonoid content

Flavonoid content analytical results for both the spice and the breads are expressed in mg catechin equivalent per 100 g (mg CE/100 g). As a result of the added reagents, the solutions turned pink. Absorbances were measured at 510 nm with a spectrophotometer (Evolution 300 LC, Thermo Electron Corporation, England). The following reagents were used for the analysis: catechin (Cayman Chemical Company, USA), aluminum chloride (Scharlab S.L., Spain), sodium nitrite (Scharlau Chemie S.A., Spain), sodium hydroxide (Sigma-Aldrich Chemie Gmbh, Germany) and methanol (Scharlab S.L., Spain) [18].

3.4. Determination of element content

Sample preparation was carried out by the method of Kovács et al. [19] prior to measurement with ICP-OES (Inductively Coupled Plasma Optical Emission Spectrometer; Thermo Scientific iCAP 6300, Cambridge, UK). Samples were placed in digestion tubes and 10 ml of nitric acid (69% v/v, VWR International Ltd., Radnor, USA) was added to the tubes. The mixtures were allowed to stand overnight, and the next day pre-digestion was performed at 60 °C for 30 minutes. After cooling, 3 ml of hydrogen peroxide (30% v/v, VWR International Ltd., Radnor, USA) was added to the samples and the main digestion was performed at 120 ⁰C for 90 minutes. After this, the cooled samples were diluted with Milli-Q distilled water (Millipore S.A.S., Molsheim, France) and filtered through 388 filter paper (Sartorius Stedim Biotech S.A., Gottingen, Germany). The following element contents were determined by ICP-OES in the resulting digestates:

• Ca 315.8 nm,
• K 769.8 nm,
• Mg 280.2 nm,
• Na 818.3 nm,
• P 185.9 nm,
• S 180.7 nm.

Measurement wavelengths are indicated after the chemical symbols of the elements.

3.5. Determination of dry matter, crude fat and crude protein contents

Breads were tested for these parameters according to standard MSZ 20501-1:2007 [22].

3.6. Statistics

The experiments were performed in triplicate. SPSS statistical software (version 13; SPSS Inc. Chicago, Illinois, USA) was used to evaluate the results. This was also used to determine the mean and standard deviation. To determine statistically significant differences between the results, one-way analysis of variance (Tukey and Dunnett’s T3 test; P<0.05) was used.

4. Results and evaluation

4.1. Results of the examination of basil

Average values of basil analyses are shown in Table 2. This table shows the total polyphenol, flavonoid and element contents of the spice as determined by three replicate measurements.

The polyphenol content values of basil were higher than the amounts (7.15-107 mg GAE/100 g) reported in the study of Moghaddam and Mehdizadeh [20]. However, in the dissertation of Kwee and Niemeyer [21], in which a study of 15 basil varieties was reported, the total polyphenol content ranged from 347 to 1,758 mg GAE/100 g. Based on our results, it can be stated that the value measured by us was high. In the case of basil, outstanding flavonoid content values are to be expected.

Based on our measurements, the spice has primarily high calcium and potassium contents, the values of which were over 20,000 mg/kg. A similar potassium content was measured by Özcan [4] in the case of dried basil (24,811 mg/kg), but the calcium content was much lower than the concentrations determined by us (12,363 mg/kg). The magnesium content of the plant is not negligible either, as a value of almost 8,000 mg/kg was measured from the alkaline earth metal. This is much higher than the values obtained by Özcan [4] and Özcan and Akbulut [3] (5,738 mg/kg and 3,130 mg/kg, respectively).

The analytical results of phosphorus and sulfur were also in the order of thousands in the given sample. Higher P (4,960 mg/kg) and lower S (1,923 mg/kg) contents were measured by Özcan [4] in dried basil of Turkish origin. Of the macroelements, sodium was found to have the lowest value, compared to the values reported by Özcan and Akbulut [3] (2,895 mg/kg).

4.2. Results of bread analyses

The results of bread nutritional value measurements are shown in Table 3.

4.2.1. Results of dry matter content measurements

Dry matter contents of the bread samples were found to be similar. The dry matter content values of the samples ranged from 68.3% to 70.5%. Sample 4th had the lowest value, while the highest value was measured for sample 6^th. Similar results were obtained for samples 1^st, 2^nd and 7^th. For these values, no statistically verifiable differences were found. Compared to the other samples, however, the results of the before mentioned breads were significantly different. The sample with the highest dry matter content, which was sample 6^th, differed significantly from all other samples. Nearly identical dry matter content values were measured for breads 4^th and 5^th, as well as for breads 3^rd and 5^th.

4.2.2. Results of total polyphenol content measurements

In terms of the total polyphenol content, it was found that even the control bread contained a certain amount of antioxidant compound, which was naturally the lowest of all samples. Kántor et al. [16] also found antioxidant compounds in their control bread. As the spices were added to the breads, the quantity of antioxidants increased steadily. The highest amount was measured in the case of sample 7^th. The analytical results of all samples showed significant differences when compared to each other.

4.2.3. Results of total flavonoid content measurements

The quantity of flavonoids increased in proportion to the amount of spice added to the bread dough, similar to the polyphenol content. The lowest value was measured in the control sample, from which the flavonoid content of sample 2nd did not differ statistically, but in all other cases a significant difference was observed. Sample 7th, with a spice content of 25.5 g, had the highest flavonoid content.

4.2.4. Results of crude fat content measurements

In terms of crude fat content, the results were mixed. Values between 5.10% and 6.33% were measured. The highest fat content was determined in the control sample, while the lowest values were found in samples 4^th, 5^th and 6^th. For breads 2^nd and 3^rd, the difference was only 0.1%. The fat content of bread sample 7^th was higher than the previous results, but not more than that of the control sample. None of these results differed from each other in a statistically verifiable way, so it was found that there were no statistically verifiable differences in the fat contents of the breads made by us.

4.2.5. Results of protein content measurements

In terms of protein content, values that differed from each other were measured. The highest value was measured in the case of sample 6^th, while the lowest was obtained in the case of the control sample. With the addition of basil, a minimal increase in the protein content was observed. There were no statistically verifiable differences in the case of the first 3 samples, however, the result of our control sample differed from the values of samples 4^th, 5^th, 6^th and 7^th. Samples with the highest protein content (5^th and 6^th) showed significant differences from samples 1^st, 2^nd, 3^rd and 4^th. The protein content of the bread enriched with the highest amount of spice differed only from the values obtained for samples 1st and 3^rd, as in this case the protein content of the product was lower.

4.2.6. Results of macro element content measurements

Results of the macro element content measurements of the breads are summarized in Table 4.

In the case of the control bread (sample 1^st), our macro element content results were similar to those reported by Kántor et al. [16], with the exception of sodium (Ca: 510 mg/kg; K: 2,418 mg/kg; Mg: 285 mg/kg; Na: 3,180 mg/kg; P: 1,512 mg/kg; S: 948 mg/kg).

Regarding the amount of macroelements that could be measured in the samples, it was found that their amount increased in each case with increasing spice concentration. Sodium and sulfur are exceptions to this, as although slightly different results were obtained, this difference could be verified statistically. The results of basil show that these two elements have the lowest amounts in the plant. While the amounts of calcium, potassium, magnesium, phosphorus and sulfur in the bread showed lower values than in the spice itself, the sodium content increased significantly due to the addition of the same amount of table salt to the samples.

The calcium content of the samples ranged from 476 to 1,614 mg/kg. With the addition of basil, the calcium content increased gradually, in most cases by 200 mg/kg between the individual concentrations. Significant difference could not be detected only between samples 4^th and 5^th.

When determining the potassium content of the bread, values higher than those obtained for calcium were measured. Compared to the control sample, which contained 2,200 mg/kg of potassium, even the product with the lowest amount of spice showed a significant difference. The highest element content was measured in the case of sample 7^th, which contained more than 1,200 mg/kg more potassium than sample 1^st. There were no statistically verifiable differences between samples 4^th and 5^th, or 5^th and 6^th, however, there were significant differences in all other cases.

The magnesium content also increased, as shown by the results. Once again, the control sample had the lowest value, while sample 7^th had the highest value. In terms of macro element content, the lowest values were measured for this element, as even the bread with the highest basil content did not reach a value of 1,000 mg/kg. Based on our results, it can be stated that there were statistically verifiable differences between the measured values in all cases.

The phosphorus content in the samples analyzed was between 1,478 and 1,623 mg/kg, these results being measured in samples 1st and 6th. As the amount of spice increased, the phosphorus content also increased. There were statistically verifiable differences between samples 1^st-5^th, 1^st-6^th, 1^st-7^th, 2nd-5^th, 2^nd-6^th, 2^nd-7^th and 3^rd-6^th. In the other cases there were no differences in the phosphorus content.

5. Conclusion

At the beginning of the experiment, basil itself was examined, and its antioxidant compounds and macro element content were determined. As the results show, the spice itself has a very high total polyphenol and flavonoid content. In addition to these parameters, its macro element content is also significant, as it has outstanding calcium and potassium contents, which is also supported by the studies mentioned in the literature. Magnesium, phosphorus and sulfur were also measured in non-negligible amounts.

During the preparation of the breads, all the ingredients were added in the same amount, except for the spice, so it was expected that there would be differences as the amount of basil increased.

We cannot draw a clear conclusion as to why the dry matter content has changed this way. The expected result was that as the amount of spice increases, the dry matter content of the bread increases as well. Differences may have been due to the nature of the convection oven.

Both the total polyphenol content and flavonoid content results were in line with our expectations, as the addition of basil, containing a large amount of antioxidant compounds, to the bread significantly increased the values of these parameters, despite the fact that most of these compounds are heat sensitive.

No difference was found in the cruse fat content of the samples analyzed, so enrichment does not affect this parameter.

However, differences were observed in the protein content, as an increase in protein content was achieved by the enrichment. Further research is needed to answer the question as to why this value has increased.

In the case of the macro element content, with the exception of sodium and sulfur, a significant increase was achieved, which may be due to the high element content of basil.

Based on our studies and results, it can be said that enrichment with basil had a positive effect on most of the measured parameters. Higher intakes of antioxidant compounds and macronutrients are also beneficial, because these compounds are required for the normal functioning of the human body.

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DOI: https://doi.org/10.52091/EVIK-2021/3-1-ENG

Received: June 2021 – Accepted: August 2021

¹ University of Debrecen, Faculty of Economics and Business, Institute of Marketing and Commerce

Keywords

netnography, functional foods, Consumer Style Inventory Test (CSI test), transtheoretical model

2. Introduction, literature review

2.1. Risk factors for health loss

Parts of health behavior are all health-related behaviors that manifest themselves as components of a healthy lifestyle, and as behaviors resulting from health motivations and health needs [1]. In Hungary, according to the NEFI (National Institute for Health Development) [2] 80% of the risks of health loss can be attributed to behavioral factors, of which a sedentary lifestyle and inadequate nutrition stand out.

Physical inactivity is responsible for 10% of cancers, has a serious impact on coronary heart disease, type 2 diabetes and osteoporosis, results in depressive symptoms, and last but not least, is responsible for 5 million deaths worldwide each year [3,4]. Physical activity and active sporting activities are separate conceptual categories. Activities related to physical activity can be divided into four groups according to their medium and way of implementation. Based on this, work-related, transport-related, household-related and leisure-time physical activities can be distinguished [5]. 53% of Hungarian never participates in any sporting activity and roughly half of the population does not engage in even moderate physical activity [6].

The concept of a sedentary lifestyle is important in the study of health behavior, as it has become a typical way of life in developed societies in recent decades. Any activity during waking hours where the metabolic equivalent (1 MET = 3.5 ml/min/kg body weight oxygen consumption) is less than 1.5 is considered sitting. A sedentary lifestyle has extremely negative effects on health in the long run [7,8]. Nearly half of the adult population of Hungary spends more than 5 hours 31 minutes sitting daily, and 10% work more than 8 hours 31 minutes sitting [9].

It si well known that obesity is a risk factor fro many chronic diseases. In 2008, there were 1.5 billion overweight people [10], in 2014, the number was more than 2.1 billion, and half of humanity is projected to be overweight by 2030 [11]. Results of the latest surveys are depressing, as they indicate that 58% of the adult population is overweight or obese based on their body mass index [12].

2.2. Health foods

The problems outlined are global and pose significant challenges for the food industry, among others. Improvements are needed which, due to their beneficial health effects, can slow down the spread of diseases of civilization and increase life expectancy spent in health [13]. Health foods have been created to treat the deficiencies caused by an unbalanced diet, to restore energy balance and to maintain health. Their names are extremely varied (e.g., healthy food, designer food, functional food, pharmafood), and the term functional food is most commonly used in the literature [14].

Foods with special health protection effects are not officially categorized and defined in Hungary, but the term functional food is widely used in the international literature. Based on the internationally accepted definition of ILSI², this includes foods that, due to their bioactive ingredients, in addition to normal nutrition, have health benefits [15]. The main groups of functional food ingredients are vitamins and minerals, proteins, peptides, antioxidants, fatty acids and phytochemicals, and pre- and probiotics [14]. In the early 2000s, the most popular functional foods were energy and sports drinks, probiotic dairy products, ”heart-friendly” products and ready-to-eat cereals [16]. According to the 2016 statistics of Google Food Trends, within the category of functional foods, ”healthy ingredients”, such as turmeric, apple cider vinegar and avocado oil, as well as bitter melon and kefir proved to be the most popular among consumers [17]. Between January 1990 and June 2018, the most studied functional foods and ingredients were prebiotics, probiotics and antioxidants, according to the bibliometric assessment of Yeung et al. (2018), who analyzed the most cited and sought for items in the literature [18] Among the factors influencing consumers’ willingness to buy, the most significant are health effects, taste, quality, value for money, and their knowledge about functional foods [19,20]. Consumption of health foods and a healthy diet can be considered cornerstones of health behavior.

2.3. Examining health behavior

To study health behavior, a number of models are used by researchers. The transtheoretical model of behavior change, hereinafter TTM³, was originally introduced as an integration of different theoretical concepts in clinical psychology [21, 22]. Prochaska and Prochaska [23], in order for professionals to be able to have a significant and lasting impact on health-threatening behaviors, have developed a model that can be applied to study the health behavior of not only the minority who is motivated for change, but the entire population. TTM encompasses process-oriented variables to predict and explain how and when subjects change their behavior [24]. Behavior change is a process that takes place over a long period of time and goes through a defined series of stages [25]. The model can be used to examine exactly where a person/group is in the transition to sustainable health behavior. Based on this, five stages are distinguished [26]:

• Precontemplation,
• Contemplation,
• Preparation,
• Action and
• Maintenance.

In the precontemplation stage, the individual is unaware of the consequences of risk behavior, does not seek information and is not interested in changing health behavior in a positive direction. In the contemplation stage, the individual weighs the benefits of the change and compares them to the costs of change. They are aware of the need for change, but if the costs are considered to be excessive, further steps are not taken. In the preparation stage, the individual is already prepared to take certain steps and possesses an action plan. In the action stage, he individual takes specific steps to protect their health. As awareness increases, the chances of returning to past behavior decrease. Real behavior change can be achieved in the maintenance stage, after at east six months. At this point, the new form of behavior becomes a natural part of the individual’s life and there is no need for reinforcements from the environment either [14, 26].

In the primary research outlined in the present study, TTM was used by our group to investigate the transition to a healthy diet.

From an economic point of view, the elements of health behavior that manifest themselves in behavior are shopping and consumption. Consumers approach the market with basic decision styles. These can be defined as mental shopping orientations that characterize consumer choices [27]. To measure the diversity of decision styles, the Consumer Style Inventory, hereinafter referred to as the CSI test [28]. CSI has been validated in many countries around the world (e.g., the United Kingdom, New Zealand, South Korea, Germany, Singapore, China, Malaysia, India, Turkey, USA) and is widely used [29]. CSI has been used in the past in general commerce [30, 31], in the monitoring of online behavior [28], and in organic food buying [32], among others. With regard to health/functional foods, no research has been carried out so far in which CSI has also been incorporated, and this was attempted by our group in our quantitative studies. In addition to exploring the decision-making styles and attitudes related to the purchase and consumption of health foods, it was also considered important to carry out a study in the online space, as this is one of the most relevant information source and communication interfaces today.

³ Transteorethical Model

2.4. Health communication nowadays – obtaining information online

According to 2021 data, there are approximately 5.16 billion active Internet users worldwide [33], and 4.48 billion of them use social media [34]. In recent years, social media has changed people’s interactions, including health-related communication [35]. Benetoli et al. [36] identified convenient and quick access, improved health knowledge and a sense of social and emotional support as the benefits of obtaining health information through social media. Disadvantages of social media included questionable credibility, information overload and the increased time spent online, among others. Johns et al. [37] classified studies published between 2000 and 2016 in terms of changes in health behavior and the impact of social media. As a result of their research, it was found that social media had no effect on giving up smoking or weight loss, but had an effect on increased physical activity.

As an axiom, it can be stated that digital communication is an integral part of of the advanced societies of today. Research in the online space is a useful addition to a type of marketing research that are group has been working on. Netnography is a qualitative research method that adapts the techniques of netnographic studies to examine the culture of online communities [38]. It can be used to understand the mindset and decision-making mechanisms of online consumer groups [39]. Ten years ago, Dörnyei and Mitev [40] recorded the basic forms of online communication: instant messengers, e-mail lists, game interfaces, chat applications, blogs, search engines, forums, social media sites. In terms of their usage, these channels have undergone a radical change. While blogs and forums flourished in 2010, today consumers barely use these platforms at all. Today’s most popular, almost exclusive virtual communication interfaces are content and video sharing sites under the umbrella of social media, of which Facebook stands out, with 2.853 billion users wolrdwide [34].

3. Materials and methods

In our research, the goal was to examine consumer attitudes towards health foods, which we analyzed in the common subset of physical activity and the diet. The research took place between April and November of 2019, and then a follow-up was performed in April 2021 by repeating our netnographic analyses. Our studies were carried out in three stages.

In the first step of primary data collection, a netnographic research was conducted with data recorded in a search engine on the one hand and the content analysis of of posts and comments made groups of publicly available social sites on the other hand. The interests of consumers present in the online space, as well as changes in them were detected in terms of health-conscious eating and physical activity.

In the second stage of our research, two focus group interviews were conducted. The impact of regular exercise on the purchase and consumption of health foods was examined, and the implications of this in developing and maintaining a diet considered to be healthier by consumers. 7 people each were included in the studies, based on preliminary criteria. The conditions for inclusion in the groups were as follows:

• The subject is over 18 years of age;
• The subject does not work in the fields of journalism, marketing, advertising, PR or market research;
• The subject has not participated in a market research survey related to the topic of physical activity and/or health-conscious eating in the previous year;
• The subject has not participated in a focus group discussion in the previous year;
• The subject does not have a milk protein allergy;
• First group: The subject regularly engages in physical activity;
• Second group: The subject does not engage in physical activity.

The differentiating factor for inclusion in the groups was performing regular physical activity, so an active and a passive group were created. At the start of the study, participants introduced themselves one by one, and then they had a conversation for a few minutes under the guidance of the moderator, creating group cohesion and an atmosphere of trust. The first part of the scenarios examined the factors that play a role in the development of a healthy lifestyle. In the second block, buying and consumption habits related to health foods were explored. The transtheoretical model of behavior change was incorporated in the scenarios, and it was examined with respect to the topic of healthy eating. In both cases, group discussions took place in an informal style and lasted an hour and a half. Minutes and audio recordings of the discussions were taken, which allowed for accurate analysis.

In the third stage of our research, an online questionnaire survey was conducted, which was shared several social media groups with the help of dietitians. A total of 378 people completed the questionnaire. To ensure representativeness, the sample was adjusted so that it reflects the composition of the Hungarian population in terms of gender distribution. As a result, mathematical-statistical analyses were performed on a sample of 300 people. In addition to key demographic data, based on Eurobarometer [9] surveys, statements related to physical activity and sedentary lifestyle were formulated, which were classified as background variables in the analysis. The questionnaire included the Hungarian translation of the Consumer Style Inventory (CSI), which was adapted and modified for health foods based on the research of Prakash et al. [32]. Items with Cronbach’s alpha values above 0.7 were included in our own research, and one dimension, statements related to environmentally conscious consumption, was omitted. As a result, 25 statements were formulated that respondents had to rate on a Likert scale of 1 to 5. The transtheoretical model of behavior change was incorporated into the questionnaire and it was examined in relation to healthy eating. Based on the focus group discussions, expansion of the TTM statements was considered to be justified, so the 6-point ordinal scale of Szabó [41] was incorporated in the questionnaire. This essentially separates the 5 stages defined in the literature by dividing the action phase into two subcategories. The main goal of our quantitative research was to identify consumer attitudes in the CSI adapted to health foods. To achieve this, in the first step the normal distribution of the variables was tested, and then the reliability of the scales was analyzed, in each case obtaining good or excellent reliability. Following this, factor analysis was performed using the CSI variables. After running several possible procedures, principal component analysis was finally applied with Varimax rotation and Kaiser normalization. The KMO criterion of factor analysis was met, exhibiting an almost excellent value (0.853). During the analysis, three variables were excluded from the CSI scale, as they distorted both reliability and the results of factor analysis (”Typically, I buy health foods at a discount price.”, ”I usually choose lower priced products.”, ”I’d rather buy well-known, domestic brand products.”). As a result, the explanatory power of the model has improved. A total of four factors were created to form differentiated attitude structures. In the next step, the reliability of the factors obtained was checked by calculating Cronbach’s alpha values, and then sample segmentation was carried out. The analysis was performed using the K-means clustering procedure, during which four well-separated, homogeneous groups were identified, based on consumer attitudes in the CSI. Characterization of each cluster was performed by cross-tabulation analysis and analysis of variance.

4. Results and their evaluation

4.1. Results of the netnographic study

Our study was conducted in line with today’s trends, using a search engine on the one hand and by content analysis of social media sites on the other hand. Different search engines and browsers have been optimized for different terms, and so search results and and hit lists may differ from each other. We used the Google search engine through the Google Chrome browser both in 2019 and 2021. As a first step, our results were compared to those of Gál et al. [42] in terms of search hits of nutrition-related keywords. Then the changes in search hits were identified for terms related to healthy eating and regular exercise over the intervening two years. Changes in nutrition-related keyword search results are shown in Figure 1.

Gál et al. [42] included the terms “healthy diet” and “healthy eating” as synonyms for ”health-conscious eating”. It can be seen that both in 2017 and 2019, the leading hits were generated by the term healthy eating, but in 2021 an explosive growth of the term “healthy diet” can be observed. The number of hits has increased nearly tenfold in four years. Search results for terms related to both nutrition and sports are shown in Figure 2.

The number of hits for search engine terms shows a variable rate increase. Of the key terms provided by us, ”regular sporting activities” and ”healthy eating and sports” proved to be the most sought after. ”Healthy eating and sports”, although the second most common content among search terms, shows a declining trend compared to 2019. At the same time, content on the topic ”healthy eating and sports” has tripled, yielding more than 6.5 million hits in 2021.

After defining the keywords, posts from publicly accessible pages were analyzed, and this was followed up by monitoring in 2021. The popularity of forum portals continued to show a declining trend, so those sites were not investigated. However, it is important to note that there was a periodic activity in the case of forum portals (e.g. hoxa.hu, gyakorikerdesek.hu) in 2020. It is assumed that this can be attributed to the quarantine caused by the pandemic. But the explosive growth of social media groups has now almost completely overridden the activity of forum portals.

In Hungary, of the social media sites, currently the trinity of Facebook, Instagram and YouTube is the most popular among active internet users. The “hashtag” is an international communication tool for navigating between and orientation on the surfaces. Hashtags allow us to get to the type of content that interests us on any of the social media giants’ websites (YouTube videos; photo-based short Instagram posts / users / pages; typed text based Facebook posts / users / groups / pages). In addition to the three basic pillars, also appearing are Tik-Tok, which is mainly used by young people, and Twitter, which is less popular in Hungary but more popular internationally. Of social media, the analysis of Facebook pages and groups was chosen, because nowadays most of the internet user community communicates on this interface. All open and closed groups, as well as pages, with at least 3,000 members and followers were examined. Only Hungarian groups and pages were analyzed. In addition to keywords, their hashtag variations were also used (e.g., #regularsports; #healthyeating) to facilitate more accurate content analysis. Four main topics were identified during the analysis of the groups and pages, and these are illustrated in Figure 3.

Based on our analyses, when weighing nutrition and exercise, the topic of healthy eating clearly appears in a more pronounced way in the interest of consumers. Healthy foods and dishes are most commonly associated with the terms ”free” and ”reduced” in public awareness, such as the terms sugar-free, chemical-free, reduced salt, reduced carbohydrate. In addition, there is an increasing emphasis on gluten- and dairy-free eating and different types of diets. This confirms the previous research results of Gál et al. [42], according to which a health-conscious diet and lifestyle is associated with some kind of diet or weight loss program by the majority of people.

The most common content in Facebook groups or pages in the common subset of diet and exercise is a combination of workout plans and recipes. These include short videos or photos that offer some kind of recipe along with a form of exercise or workout plan, typically using ”reduced” or ”free” ingredients. Workout plans are typically “challenges” over a longer period of time (e.g., broken down for a month), or forms of movement presented in a short video. Particularly popular contents are home exercises that can be performed without any aids or with minimal use of aids (e.g., dumbbells).

The content encountered second most frequently is requests for suggestions on a health problem or a change in lifestyle or diet. This is most noticeable regarding the topic of eating, less content requesting suggestions is found on exercise and physical activity.

The third most common content is providing advice or information. In the case of this type of content, mainly articles and stories, often with questionable authenticity, from associated sites appear on the pages, as well as short videos and infographics. Advice on the topic of physical activity is typically about how to start exercising regularly, what pitfalls and difficulties one might encounter. In the field of nutrition, the most common are discussion initiating contents related to gluten and sugar consumption, as well as dairy products and caffeine. This is followed by the presentation and promotion of “healthy products” and posts emphasizing the importance of fruit and vegetable consumption.

Other major type of content is the presentation of motivational examples. In this content, photos of “transformations” that occur as a result of some diet, dietary change or regular exercise are typically uploaded by users. Motivational examples often include presentations of “own stories” about restoring health. In these stories, people who share the content report a positive change in health as a result of a diet considered to be healthy and/or regular physical activity.

Overall, it can be stated that healthy eating and physical activity are popular activities among internet users. In the common subset of diet and exercise, the emphasis was typically on issues related to nutrition in the media examined by us. The most popular types of content are personal in nature and have a community-building power.

4.2. Results of focus group studies

4.2.1. Factors that play a role in the development of a healthy lifestyle

When describing the results of our focus group research, groups are referred to as ”active” and ”passive” ones, revealing the attitudes and peculiar characteristics of the given group. In the first block of the scenario, we sought to answer the question what similarities and differences could be detected between the groups in the topic of health. Regarding the groups, it can be generally said that in the subjective assessment of health, the active group considers their lifestyle to be healthy, while the passive group considers it to be unhealthy. In the development of a healthy lifestyle, the groups studied unanimously thought that the right amount and quality of sleep, proper nutrition, mental health and regular exercise were vital.

Following this, the groups had to rank 15 factors according to the influence of each component. Based on the ranking thus developed, the 5 most important factors according to each group are listed in Table 1.

Regarding the second most important features of the groups, 2 factors were raised to the same level by each group, as in neither case were they able to reduce the ranking to 1 component at this level. Nutrition and exercise were considered to be relevant by both groups, but it should be emphasized that information, accessibility and adequate financial situation were the most important for the passive group. Regarding the transition to a healthy lifestyle, the groups studied had to make arguments as to why it could be easy, as well as what would be difficult in the process. Overall, the same factors were listed both as pros and cons. The groups attached similarly great importance to the influence of the social environment, which they believed had a strong impact on the individual’s health behavior.

4.2.2. Customer and consumer habits and motivations related to health foods

There are differences between the categories of food most often purchased and consumed by the two groups. The passive group consumed a higher proportion of meat products, quick-frozen and processed foods. The active group preferred seasonal fruits and vegetables, dairy products and fresh bakery products. Members of the active group, according to their own statements, plan their purchases in advance, while impulse buying is more common among the passive group.

Prior to examining consumer attitudes towards health foods, their concept as clarified with group members: ”They are foods that have one or more nutritional biological benefits in addition to excellent taste. These advantages include lower energy content, mainly through the reduction of fat content or the omission of sugars, enrichment in certain minerals (Ca, Se, Mg), depletion in others (Na), addition of multivitamins or the use of probiotic lactic acid bacteria in different foods.” All of the subjects in the study bought and consumed health foods. For members of the active group, ”being free of something” was important, which manifested itself mainly in the avoidance of fat, salt and sugar. Members of the passive group typically preferred products ”fortified with something”. The active group bought more types of health food more often than the passive group.

It was characteristic of both groups that subjects had changed their eating habits over the previous year. The reason for this was the development of some kind of sensitivity/allergy, as well as the need to change lifestyles and to try new diets. Nutritional trends affected the active group, but they usually researched a diet before trying it. Members of the passive group are generally said to be uninterested in different trends, as well as dietary recommendations.

Regarding the purchase and consumption of food, the active group considers the healthiness of food to be the most important factor, while for the passive group it is value for money. Members of the active group attach particular importance to nutrient composition, to products that are ”free of something”. For the passive group, in addition to easy availability, previous positive experience has an impact on food buying habits.

4.2.3. Differences in the transition to a healthy diet based on the TTM

Using the statements translated by Soós et al. [26] based on the TTM, it was examined where the groups were in the transition to what they considered to be a healthier diet. Stages in the behavior change are described below, with examples of the statements made:

• Precontemplation: In the next six months, I do not intend to switch to a diet I consider healthier;
• Contemplation: I feel a strong urge to switch to a diet I consider healthier;
• Preparation: Over the next month, I will be taking steps to switch to a diet I consider healthier;
• Action: Over the past six months, I have switched to a diet I consider healthier;
• Maintenance: I have been eating healthier for over six months now.

In the transition to a diet that is considered healthy, 30% of the active group was in the action stage, while 70% was in the maintenance stage. In contrast, 70% of the passive group was in the precontemplation or contemplation stage, while 30% was in the preparation stage. Based on this, it can be concluded that the passive group is less open to developing and maintaining a health-conscious diet.

Overall, it can be stated that great emphasis is placed on the consumption of health foods among the group who perform physical activity regularly. Purchases are planned more purposefully by the active group and, according to their own statements, their diet is more based on awareness.

4.3. Results of the questionnaire survey

4.3.1. Presentation of the sample

Our quantitative study was conducted in the online space. The gender distribution of our sample reflects the composition of the Hungarian population, however, our results are more exploratory, as the sampling took place in a specific medium. The sample was made up of people who follow the online work and activity of dietitians, and themselves spend time regularly in the online space. The distribution of the sample according to different background variables is shown in Tables 2 and 3.

Examining the age distribution, it can be stated that our sample is representative of internet users, i.e., the 18 to 49 age group is typically represented. Compared to the demographic composition of the Hungarian population, the proportion of people with higher education is much higher in our sample. Nearly half of the respondents consider themselves mostly health-conscious, engage in physical activity regularly, and 41.7% of these people spend 31 to 60 minutes with exercise daily. Two thirds of the sample spend between 2 hours 31 minutes and 8 hours 30 minutes a day sitting, with an additional 17% spending even more. This rate is higher than the Hungarian data measured by the Eurobarometer [9].

The transition to a healthier diet shows a more positive picture compared to the overall data of the Hungarian population. It must be added that, based on our representative national surveys, the proportion of people in the precontemplation stage is decreasing, while the proportion of people in the preparation, action and maintenance stages is increasing. In 2014 and 2019, 48% and 41% of the population was in the precontemplation stage, respectively, while the proportion of people maintaining a diet considered to be healthier has increased from 17.4% to 23.6% [43]. Development of the transition to a diet considered to be healthier in our sample is shown in Table 4.

4.3.2. Results of the factor analysis

According to our results, CSI’s attitudes towards health foods are determined by four factors in our sample. The factor structure of the CSI test is illustrated in Table 5.

The second factor is the Recreational, hedonistic value dimension, where the explained variance is 11.834%. This attitude is driven by the joy of of shopping, which has a decisive influence. Once again, high factor weights can be seen in the analysis, so this attitude is significantly separated from the other factors. This factor is skewed to the right (Skewness = 0.275), which means that in the whole sample, respondents do not really consider this attitude to be characteristic of themselves.

The third factor is the Uncertain, confused value dimension, in which the explained variance is 11.308%. It is characteristic of this attitude that the individual has difficulties making a decision about where to buy, as well as about what brand to choose, they feel that purchases should be planned more carefully. This factor is slightly skewed to the right (Skewness = 0.049), suggesting that the respondents who completed the questionnaire consider this attitude to be less characteristic of themselves.

The fourth factor is the Devoted, brand-loyal value dimension (explained variance: 10.872%). This attitude is characterized by brand loyalty and identifies quality with a higher price. The factor is significantly skewed to the left (Skewness = -0.882), so this type of behavior appears with a positive sign in the mindset of the respondents in the sample.

Method: Principal Component Analysis, Rotational method: Varimax with Kaiser Normalization. KMO=0,849

Prior to the cluster analysis of the CSI adapted for health foods, it was considered necessary to validate the list of claims. The Health- and self-conscious value dimension includes nine items, with a Cronbach’s alpha index of 0.922. There are four elements in the Recreational, hedonistic value dimension. Two of these items are considered inverted (”Shopping is not a pleasant activity for me”, ”I make my shopping trips fast”), after the recoding of which the scale has a Cronbach’s alpha index of 0.720. The Uncertain, confused value dimension contains five elements, with a Cronbach’s alpha index of 0.701. The fourth value dimension is Devoted, brand-loyal, which contains four statements. The Cronbach’s alpha index of the scale is 0.673. By deleting elements, there was no significant improvement in the Cronbach’s alpha index in any of the value dimensions. Based on our results, the list of statements is suitable for the characterization of the examined dimensions.

4.3.3. Segmentation results

The results of the factor analysis confirmed that the obtained factors are suitable for the cluster analysis, so in the next step the segmentation of the sample was performed. The analysis was performed using the K-means clustering procedure, and four groups were separated along the 22 factors. The value dimensions characteristic of the clusters are illustrated in Figure 4.

Following this, the socio-demographic background of each segment was characterized by cross-tabulation analysis and the deviations from the mean were examined by analysis of variance. Finally, the differences between the groups in the areas of physical activity, sedentary lifestyle and the transition to a diet considered to be healthier were examined.

4.3.3.1. Uninterested (cluster 1)

For those in the Uninterested group, the impact of foods on health is less important, they make no effort to buy good quality health food. Of the clusters, they show the least propensity to consume health foods, but cannot be considered dismissive. They do not identify the price of products with quality. They do not have favorite brands, and if they find a brand they like, they are not loyal to it. They want to finish shopping as soon as possible, since it is not a pleasant activity for them at all, they make quick decisions in the choice of both the store and the product. Typically, all statements are undervalued by Uninterested people compared to the other clusters.

The first cluster is the smallest group, making up 14.3% of the sample. In this cluster, men are inordinately overrepresented (72.1%), and the youngest age group, 18-29 is prominent (41.9%). This group has the highest proportion of people with high school diplomas (37.2%). Uninterested people has the highest proportion of respondents who are not health-conscious at all (16.3%) or mostly not health-conscious (34.9%). 60.5% of the group do not intend to switch to a diet they consider healthier in the next 6 months. They perform physical activity occasionally (32.6%) or infrequently (27.9%), spending on it 30 minutes or less (55.8%). Members of the group typically spend between 5 hours 31 minutes and 8 hours 30 minutes sitting daily (41.9%).

4.3.3.2. Health-oriented (cluster 2)

For Health-oriented people, it is extremely important to buy high quality health foods, and they are making a special effort to do so. They believe less that the price of a product determines its quality. For the sake of variety, they shop in several stores and always have health foods in their kitchen. Shopping is not one of the favorite activities in their lives, they like to get it done quickly. They have a few favorite brands and typically buy these. When they buy health foods, they usually do so in the same store. Of the clusters, they consider themselves the most health-conscious. In order to maintain their health, they choose foods very carefully and intend to make efforts in the near future to buy health foods.

The second cluster accounts for nearly one-third of the sample (32.6%). In this group, we find almost equal numbers of women (49%) and men (51%). Based on age, the majority belong to the 30-39 (25.3%) and 40-49 (18.4%) age groups, who typically have college degrees. Those in the group consider themselves mostly (54.1%) or very health conscious (27.6%). According to their own statements, almost one-third of the cluster (29.6%) have been eating healthily for at least six months, while one-fifth (20.4%) have always been eating healthily. This group contains the highest proportion of those who engage in physical activity regularly (76.5%). They typically spend 31-90 minutes on active movement. Members of the cluster mostly spend between 2 hours 31 minutes and 5 hours 30 minutes sitting daily (40.8%).

4.3.3.3. Variety seekers (cluster 3)

Variety seekers are less likely to identify product quality with high price. A higher than average proportion of them is reported to be looking for new types of health foods for purchase. Shopping is a decidedly pleasant and fun experience for them, members of the group believe that it is one of the really enjoyable activities of their lives that they spend a significant amount of time on. Compared to the sample average, they are more likely to have health foods at home. They are less loyal to brands, much more interested in novelty and variety.

The third cluster makes up 28% of the sample. Two-thirds of the group are women (64.3%). In this cluster, people in the 18-29 and 30-39 age groups make up two-thirds of the group (67.8%). A quarter of the group (26.2%) say they are partially health-conscious, while 20.2% feel a strong urge to switch to a diet they consider to be healthier. Two-thirds of the cluster performs physical activity on a regular basis, spending on average 31-60 minutes on it daily. Variety seekers typically (38.1%) spend between 5 hours 31 minutes and 8 hours 30 minutes sitting daily.

4.3.3.4. Uncertain brand choosers (cluster 4)

It is important for Uncertain brand choosers people to buy high quality health foods, they are the ones who clearly identify product quality with a high price. Compared to the sample average, they are more likely to look for new types of health foods to buy, but these are not accumulated in their homes. They consider shopping less enjoyable and usually do this activity quickly. They have a few favorite brands that they are loyal to. They believe they should plan their shopping more carefully. For them, it takes time to choose carefully for the best possible purchase, many times it is even difficult to choose the store where they want to shop. It presents a great difficulty for them when they have to choose from a number of brands, a large selection confuses them. Compared to the sample average, they consider themselves less health-conscious, but they would like to make an effort to buy health foods in the near future.

The fourth cluster makes up one-fourth of our sample (25%). This group is also characterized by the majority of women (54.7%). Age groups 50-59 (16%) and over 60 years (18.7%) dominate this group. This cluster contains a higher proportion of partially health-conscious respondents (28%) who would like to take steps in the near future to switch to a diet they consider healthier (22.7%). Uncertain brand-loyal people tend to engage in occasional physical activity (32%), spending less than 30 minutes on it. On average, they spend between 2 hours 31 minutes and 5 hours 30 minutes (32%) sitting daily.

The four clusters show distinctly different socio-demogpraphic characteristics and represent different value dimensions in relation to attitudes towards health foods. They are at different stages in the transition to a diet that is considered healthy. Significant differences can be detected in our sample regarding the regularity and duration of physical activity and the diet considered to be healthier. However, a sedentary lifestyle cannot be considered determinant of attitudes towards health foods.

5. Summary

In the online space, when balancing nutrition and exercise, consumers clearly place more emphasis on healthy eating, based on our netnographic survey. Search engine hit lists show a variable rate growth from year to year in the area of healthy eating and exercise. For the keyword healthy eating, a nearly tenfold increase was observed between 2017 and 2021. In social media, in the common subset of diet and exercise, four main types of content can be distinguished, the most popular of which are posts dealing with a combination of training plans and recipes. Our focus group studies have highlighted that different consumer preferences can be observed for health foods depending on whether we are talking about active or passive groups. According to the active group, the most important feature of a healthy lifestyle is the consumption of health foods, while the passive group believes that the most important thing to be informed about what is healthy and what is not. The active group characterizes health foods with being free of something, while the passive group characterizes them with fortification with something. Those who engage in physical activity regularly are more open to consuming health foods and are more affected by diet-related trends. Based on our quantitative research, attitudes towards health foods are determined by four value dimensions. In the Health- and self-conscious attitude, consumption of health foods is of paramount importance. The Recreational, hedonistic attitude is characterized by the joy of shopping. In the Uncertain, confused value dimension, uncertainty and indecisiveness stand out, which is reflected in both store and brand choices. The Devoted, brand-loyal attitude identifies quality with a high price, and both store and brand selection are carried out along definite ideas. Following factor analysis, sample segmentation was performed, resulting in the identification of four major groups. Among the clusters, Uninterested people represent the smallest proportion of our sample, and they undervalue all statements. The Uninterested group does not want to switch to a diet they consider healthier, for them it is not important to consume health foods. They perform physical activity infrequently and for a short period of time. The Health-oriented cluster is characterized by the exact opposite set of values. For them, it is important to buy health foods and they make an effort to do so. Shopping itself is not a pleasant activity for them. Members of the cluster perform physical activity regularly and for longer periods of time, and they have the highest proportion of actors and maintainers among the stages of the diet they consider healthy. For Variety seekers, quality is not related to a high price. They are looking for new types of health foods, however, they are more motivated by the joy of shopping. they are not loyal to a particular store or brand type. They engage in physical activity regularly and feel a strong urge to switch to a diet they consider healthier. In contrast, Uncertain brand choosers people stick to one brand type, but are confused by a large selection of brands. They consider themselves less health-conscious, but in the future they would like to strive to buy health foods and also to switch to a diet they consider healthier. Typically, they perform physical activity occasionally and for shorter periods of time.

According to the results of our research, the purchase and consumption of health foods and attitudes towards health foods are related to the diet and physical activity of the individuals, however, independent of the time consumers spend sitting daily.

6. Acknowledgment

This publication was supported by the project titled Debrecen Venture Catapult Program, No. EFOP-3.6.1-16-2016-00022. The prject was supported by the European Union and co-financed by the European Social Fund.

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DOI: https://doi.org/10.52091/EVIK-2021/3-2-ENG

Received: March 2021 – Accepted: June 2021

¹ University of Debrecen, Faculty of Economics and Business, Institute of Marketing and Commerce

Keywords

food industry developments, food nanotechnology, consumer acceptance, willingness to buy, food industrial use of titanium dioxide

2. Introduction – nanotechnology

One of the most dynamically developing disciplines today is the research of nanoscale materials. Research and application of nanotechnology is one of the great scientific, developmental and technical challenges of the 21^st century.

Nanotechnology means the production, of materials, devices and systems that use artificially formed nanoparticles, i.e., particles of material that do not exceed 100 nanometers in size [1]. Nanostructured materials are also found in nature (e.g., clays, zeolites), but can be produced artificially as well.

Many nanotechnological applications are known in practice. Examples include highly resistant materials used in construction; lightweight, elastic clothing and sports equipment made of materials resistant to physical stress; self-cleaning paints that protect buildings from, for example, the harmful effects of smog and other contaminants; nanosensors that enable efficient and economical quality control in the food industry; highly miniaturized electronic devices; antibacterial coatings for industrial equipment and household appliances; selective release and high bioavailability drugs; innovative tools for the remediation of contaminated soils and waters. However, in addition to the benefits, nanotechnology poses risks to the environment and human health that are difficult to assess. Scientific research, while still proving to be scarce, suggests that nanoparticles are more reactive and mobile than larger particles and can therefore be toxic to humans and the environment. Little is known about the fate of nanoparticles in the environment. In the human body, nanoparticles may be able to cross the cell membrane and reach internal organs. Some studies have shown that many types of nanoparticles cause greater oxidative stress at the cellular level, increasing the risk of degenerative diseases [1].

2.1. Nanotechnology in the food industry

Due to their special properties, the use of nanostructured materials can also be promising in many food applications [2].

Foods containing nanoparticles should be considered as novel foods under Regulation (EC) No 258/97, as foods or food ingredients produced by such technology were not consumed in significant quantities in the European Union before May 15, 1997; thus, their placing on the market is preceded by an authorization procedure accompanied by a rigorous safety assessment [2]. As part of the authorization process, EU regulation has recently required food ingredients derived from the use of nanotechnologies to undergo a safety assessment before they can be placed on the market, and only then can they be authorized [3]. Related to this, the term nanofood has emerged to refer to foods that are produced, processed or packaged using a nanotechnology technique or device, or to which a nanomaterial is added and/or is enriched with a nanomaterial [4].

Nanotechnologies aimed at improving food quality or safety can theoretically be diverse, but their practical application is still in its infancy. Since food nanotechnology is also a new field for food science, nanotechnology is also a major challenge for the food economy, including food security and safety, traceability, certain areas of food processing and packaging, some new opportunities for nutrient intake, longer food shelf life and many other aspects of consumer protection, from agricultural production to the consumers’ tables [2].

The use of nanoparticles in food processing can contribute to the improvement of nutritional quality, taste, color and stability or to increasing shelf life and, in the case of liquid foods, to the improvement of flow properties. An additional benefit of nanotechnology may be that it can contribute to the development of foods with lower fat, sugar and salt content, thereby reducing the incidence of food-related diseases [5].

Currently, these products are available in four categories:

• nanostructured food ingredients and substances, such as nano-titanium dioxide, which is used as an anti-caking agent or pigment;
• nanostructured delivery systems that improve the bioavailability of bioactive compounds in fortified foods and supplements;
• novel packaging materials designed to strengthen the protective function of the product;
• and the use of food contact materials for food processing and storage, such as nano-silver, which is used for its antimicrobial properties [6, 7, 8, 9].

Nanotechnology is currently considered to be the most widespread among food industrial commercial applications in the packaging process [2, 10]. Several types of use of nanomaterials in packaging materials can be distinguished. In the case of nanocomposites, advantageous properties (mechanical or functional, e.g., gastightness, temperature / humidity stability) are achieved by adding nanoparticles to the plastic.

A similar effect can be achieved with nanocoatings applied to the surface of the packaging material. Aluminum coatings applied with the help of vacuum are now widespread mainly in the packaging of snacks, confectionery and coffee. For example, if the thickness of the aluminum layer applied as a coating does not exceed 50 nm, the coating metal can be considered a nanomaterial [11]. In addition to the above, there are several applications that are still in the research phase [12, 13, 14, 15], such as newly developed food packaging capable of detecting the presence of pathogens and contaminants.

Although this technology offers consumers unprecedented benefits such as higher added value, longer shelf life and increased food safety, nanofoods also pose health, environmental, economic, social and political risks [16, 17]. According to Berekaa, despite the huge benefits that nanoparticles can bring to the food industry, the public is very concerned about their toxicity and potential negative environmental impact. Due to the health consequences of the nanoparticles entering the human body, their potential risks to human health need to be assessed without delay [5]. In his paper, Halliday points out that EU regulations on food and food packaging require a specific risk assessment before nanomaterials are placed on the market [18].

In the course of our research, it was examined to which extent the concept of nanotechnology in the food industry has spread in the public consciousness, i.e., presumably how many people are aware of this technology and its potential application in the food industry. Following this, it was assessed how receptive consumers were about the technology, how they saw its future, and whether they would be willing to buy nanofoods. In our work, the potential dangers of nanotechnology were analyzed, and also the areas in which they may occur, as well as how attitudes, consumer acceptance and willingness to buy change in the light of this.

2.1.1. Foods and packaging materials produced using nanotechnology – some examples [1]

2.1.1.1. Creamier ice cream with unchanged fat content

When making ice cream that is creamier than traditional ones, titanium dioxide consisting of nano-sized grains is added to the raw material of ice cream to increase its creaminess and improve its taste, while keeping its fat content the same as that of traditional ice creams. In its nano form, titanium dioxide is thought to be cytotoxic, however, no data have been found in the scientific literature on the mechanism of absorption of nano TiO2 from the intestinal tract.

2.1.1.2. Table salt and sugar that do not form lumps with moisture

Nano-sized particles of titanium dioxide are added to table salt and sugar as anti-caking agent. For toxicological aspects see Section 2.1.1.1.

2.1.1.3. Fruit juices enriched with bioactive molecules

Bioactive molecules such as phytosterols, vitamins and antioxidants are added to fruit juices by the way of nanoencapsulation to improve them. Nanoencapsulation is not known to have adverse health effects.

2.1.1.4. Bread enriched with omega-3 fatty acids

Omega-3 fatty acids are added to bread by nanoencapsulation; this way the unpleasant taste of the fatty acids is not felt, and thus the fortified bread retains its traditional taste. Nanoencapsulation is not known to have adverse health effects.

2.1.1.5. Plastic bottles for beer

Beer bottles with a modified composition are produced by adding a nanocomposite material containing clay particles. The purpose of clay-polymer nanocomposites is to minimize carbon dioxide loss and oxygen uptake to extend the shelf life of carbonated beverages. The toxicological effects of the nanolayer are unknown; it has not yet been demonstrated that nanoparticles can be released from the packaging material.

2.1.1.6. Antimicrobial food packaging for meat and other foods

Food packaging materials containing active nano-silver inhibit the growth of microbes and help to prevent possible bacterial contamination. Nanoparticle-sized silver is presumably cytotoxic. It has not yet been demonstrated that nanoparticles can be released from the packaging material.

3. Materials and methods

To answer the research questions, online questionnaire interviews involving 200 people were conducted. During the sampling, the snowball method was used, i.e., the selection of the sample was not random, but in this way we were able to reach a wide range of respondents. Under these conditions, the survey cannot be considered representative, the results obtained can only be applied to the actual respondents. Background variables of the questionnaire included gender, age, place of residence, education and average income.

In the course of the questionnaire survey, consumer attitudes towards nanotechnology in the food industry were assessed using 17 closed-ended questions. Then, in order to be able to analyze them in depth, two focus group studies were conducted. Consumers’ attitudes towards the topic were determined in advance by screening questions, based on which they were classified into one of the two focus groups. The first group included consumers who rejected nanotechnology based on the screening questions, while participants in the second group viewed this technology favorably. During the formation of the two groups, we sought to ensure that the consumers interviewed were included in the research in an equal distribution with regard to gender. In terms of age, people between the ages of 20 and 65 participated in the interviews.

Due to the pandemic situation at the time of the research, the two groups of eight people each were interviewed via an online platform.

At the beginning of the focus group interviews, participants were asked to briefly introduce themselves, and then two passages, taken from Sodano et al.’s communication and translated into Hungarian [1], were read aloud in the first half of the discussions. The first text introduced nanotechnology in general, while the second part described six products that had been made by some kind of nanotechnological process, but only the advantageous properties of the products have been emphasized in the description. The first half of the interview questions concerned the awareness and acceptance of nanotechnology in the food industry, but group members also had to answer questions related to the texts they had heard.

In the second half of the focus group discussion, the part of the text that highlights the potential risks and negative impacts associated with the technology and, thus, the products was read aloud. Following this, once again participants were asked questions, this time focusing on the risks, and it was examined how much their attitude towards the topic had changed.

4. Results and evaluation

In this chapter, the most important results of the primary research are presented, in the order they took place.

4.1. Results of the questionnaire survey

The first question of the questionnaire focused on factors considered important when purchasing food. This was important because, after this, the backbone of the research was the examination of the acceptance of nanotechnology in the food industry, taking into account the categories mentioned here. As can be seen from Figure 1, of the factors listed, taste was mentioned first, i.e., for 76.0% of the respondents taste was the most important consideration when purchasing or selecting a food. Based on the comparison with the background variables, it was revealed that men in the sample had a significantly (p=0.014) higher proportion (80.0%) who considered taste important than women (61.2%), and also that consumers who, according to their own statements, live in better-than-average financial conditions (live well on their income and can also save some money) also consider taste to be an important criterion when choosing (88.9%).

Slightly behind, high quality (68.5%) and price (63.5%) was second and third in terms of purchasing considerations. As had been expected previously, for these categories, 86.2% of those with a sound financial background rated high quality as an important aspect, while in terms of price, this proportion fell to 47.3%.

High food safety was considered important even less than one half of the respondents (47.0%), which may be due to the fact that they were not aware of the specific meaning of the term.

Respondents considered added value (for example, higher omega-3 fatty acid content) to be the least important aspect, with this factor ranking last of the listed ones with 17.5%. Only 20.0% of men and 16.4% of women consider this category when purchasing food. In terms of financial status, this criterion was least important for consumers with below-average income (7.0%).

In the following, the proportion of respondents with knowledge on nanotechnology in the food industry (spontaneous recall) was examined. The innovative and novel nature of the technology is also supported by the fact that only a quarter of respondents have heard of it.

When the four categories of nanotechnology currently available in the food industry were also listed [6, 7, 8, 9] (supported knowledge), only 62.0% of consumers still answered that they had not yet heard of the new technology in question (Figure 2). Of the entire sample, there was only one person who had heard of all the categories listed. Of the four categories, packaging materials made using nanotechnology were the best known (28.5%). 11.5% each of the participants in the survey have already heard of nanostructured food ingredients and materials, as well as the use of food contact nanomaterials, respectively. Respondents were least familiar with nanostructured delivery systems, the proportion in this case was not even 5.0%. Consumers who have heard of this category had some kind of college degree.

In the following, the acceptance of nanotechnology in the food industry was examined using the aspects listed in the first question that were considered important at the time of purchase. The results are shown in Table 1.

Based on the results obtained (for the sample), it can be said in general that the majority is open to the new technology if it has a beneficial effect on one of the properties of the food purchased. 71.9% of respondents would buy food made with nanotechnology if its organoleptic properties were better. Of the aspects considered important when buying food, taste finished first: 76% of respondents chose this factor. It should be noted that a greater willingness to buy due to more favorable sensory properties was an expected outcome. The older age group gave the highest proportion of affirmative answers to this characteristic (89.5%, p=0.047), and there was no significant relationship to the other background variables. In order to have a positive effect on the texture of foods, 68.8% of the consumers in the sample would buy a product made with a nanotechnological process. In the hope of better texture, 85.2% of respondents aged 56-65 would be open to buying products made with the new technology. A significant increase in the shelf life and use-by date of foods due to the nanotechnology process had an incentive effect on shopping for 62.5% of respondents. In the case of this question, a significantly higher proportion of women answered yes than men (women: 70.0%, men: 46.3%). 78.6% of the respondents to the questionnaire would buy food made with some kind of nanotechnology process if it increased food safety. 90% of the older age group and 78.6% of women were represented in the „yes” answers in this regard. 63.0% of respondents answered „yes” to the question of whether they would buy a food produced with nanotechnology development if it has added value such as a higher omega-3 fatty acid content. This represents an exceptionally high proportion considering that added value as a purchase criterion finished last in terms of importance with 17.5%. Thus, although it is typically not an important factor for the consumers in the sample that the food has some added value, they would still choose a product manufactured with nanotechnology that is richer in omega-3 fatty acids. Finally, 78.1% of respondents were open to food packaging produced with a new method that guarantees safer storage. In this case as well, women and those aged 55-65 had the highest proportion of „yes” answers.

Figure 3 illustrates how many percents more respondents would be willing to pay for a food that has been produced or modified using some kind of nanotechnology process. Typically, the additional cost consumers in the sample considered most acceptable was between 0% (i.e., they would not pay more at all for a product manufactured with this technology) and 5-10% (30.7% and 30.7% of respondents, respectively). 22.4% would pay 0-5% more and 15.6% would pay 10-20% more for this type of food. The proportion of respondents willing to assume an additional cost of more than 20% did not even reach one percent. Consumers who would be willing to pay 0-5% more for a product manufactured with nanotechnology are those who have a lower-than-average monthly net income, while respondents who say they live in better-than-average financial conditions would be willing to pay 5-10%, 10-15%, 15-20%, or even more than 20% more for such foods.

In our research, it was also addressed how respondents felt about the possible adverse consequences of nanotechnology in the food industry. Based on the results obtained, it was found that more than half of the questionnaire respondents (53.6%) believed that foods made with the nanotechnology process carry unknown hazards. In this case, in terms of proportions, men can be said to be the most skeptical, with 74.3% saying that nanotechnology in the food industry could pose a risk.

Figure 4 shows the probability of the occurrence of the different hazards in the opinion of the respondents in percentage distribution. 71.4% of respondents who consider the technology to be risky believed that foods made with the nanotechnology process pose mainly health risks. This was followed by environmental risks (56.3%). In this case, almost twice as many women believed that nanotechnology in the food industry could cause environmental damage (p=0.020). Consumers in the sample considered negative economic and social impacts to be the least probable. For these two categories, typically women were also in the significant majority (p=0.001). However, it can be said for all categories that respondents with higher education represented a higher proportion.

4.2. Results of the focus group studies

Since the main objective of our research was to examine nanotechnology in the food industry from a consumer perspective and to explore the expected rate of acceptance and possible rejection of the technology, after examining the quantitative results of the online questionnaire, it was considered appropriate to analyze the responses received in more depth using a qualitative method, therefore, focus group interviews were conducted to facilitate interpretation.

4.2.1. Results of the focus group study of people accepting nanotechnology in the food industry

Our discussion began with an association game designed to resolve any anxieties of the interviewees. Members of the group were asked to say positive and/or negative words and phrases that come to mind in connection with the topic. The following words were mentioned: innovation, invention, new opportunities, interesting, sci-fi, foods of the future, possible solutions to many problems.

The next question was whether they had already encountered any of the listed categories of nanotechnology applications or something similar (creamier ice cream with the same fat content; salt and sugar that do not form lumps with moisture; fruit juices enriched with bioactive molecules; bread enriched with omega-3 fatty acids; plastic bottles for beer; antimicrobial food packaging for meat and other foods). All of the respondents had already met soft drinks and beers packed in special PET bottles. Fruit juices enriched with various vitamins, minerals and antioxidants were mentioned by several people, and one person saw bread enriched with omega-3 fatty acids in a store while shopping (he didn’t remember exactly which store it was). In addition to the categories read aloud, they have seen eggs that contained excess omega-3 fatty acids, known various dietary supplements to which vitamins, minerals or antioxidants were added, and a participant had read on the internet about an intelligent packaging material that recognizes contaminants. He did not remember whether the packaging material had been made with nanotechnology in the food industry, but he believed that this category fit exactly into this topic.

Following this, those present were asked to express their views and evaluate how they perceived the six categories described above. Positive thoughts were associated with the products by everyone. They were thought to be useful in many ways, and it was thought to be a good idea to add such extra values to foods that allow people to get vitamins and other minerals without having to take separate capsules into their body. According to the participants, the facts that the use of nanotechnology can make food storage safer and increase shelf life can also be advantages. When asked if they would like to buy this type of food, all participants answered in the affirmative. One person stated that he was somewhat averse to nanotechnology-modified ice cream, while two people said the same thing about bread enriched with omega-3 fatty acids, but they could not specifically explain why.

This was followed by solving a task together, in which members of the group were asked to jointly establish an order for the six products based on which they considered to be the most sympathetic and which the least. The popularity of the products is illustrated by the data in Table 2.

The group unanimously agreed with the assumption that in the future we would encounter many of these or similar products on store shelves. It was thought that foods produced with nanotechnology were likely to become more widespread if the pace of food industry developments remained the same. One of our interviewees said that due to the overpopulation of the Earth and the constant decline of arable land, it will be necessary to deploy such tools in order to avoid an increasing rate of hunger and malnutrition, and to prevent people from suffering from the lack of certain nutrients. Everyone has accepted the vision that foods produced with such technology and other similar developments will become more popular and accessible, provided, of course, that they will be available at affordable prices. Intelligent food packaging that recognizes bacteria and contaminants has been found to be especially useful and practical.

According to them, basic foods (dairy products, pasta, flours, cereal flakes) could also be enriched with added values (vitamins, minerals, antioxidants).

In the second half of the focus group interview, the part of the texts was read aloud that described the potential risks involved in using the technology. Following this, it was assessed whether participants’ opinions, attitudes and willingness to buy changed as a result of what they had heard. The majority believed that if it were not safe to consume a product, it would ultimately not be able to be marketed. According to another opinion, while it sounded a little scary, and so he would think twice about buying this type of product, he still would not reject the technology.

Finally, participants were asked to reconsider, in light of the information they had learned, the order established above, as to which category they would be most likely to purchase. For better comparability, the order before describing potential hazards and the new order are listed in the same table. The results are shown in Table 3.

Although the final order was changed at several points, the opinions and willingness to buy of the group members did not change significantly after the exploration of possible dangers.

4.2.2. Results of the focus group study of people rejecting nanotechnology in the food industry

The study scenario in this case was the same as it was for the previous group. Presentation of the first part of the text was followed by an association game, the essence of which was that participants had to say adjectives and expressions, whether positive or negative, that came tom mind about nanotechnology. This time, compared to the interviews with the accepting group, the opinions (answers) were much more mixed: innovative, dangerous, bizarre, this is the future, foods made in a laboratory, unnatural. One of our interviewees also noted that these products were likely to be very expensive.

Of the six products made with nanotechnology in the food industry, half of the group had already encountered fruit juices enriched with bioactive molecules, and everyone was familiar with the special PET bottles. As similar products, sports drinks and dietary supplements fortified with vitamins and minerals were mentioned, which had already been encountered by them in retail trade, and one person had already read online about packaging materials made with nanotechnology, and another participant cited a scientific paper on artificial meat as an example.

Following this, once again, members of the group were asked to share their views on the six products which had been introduced at the beginning of the interview. Someone thought it was extremely scary to hear about these, while others thought that they would be very unhealthy for sure. Many people felt that it was unnecessary to enrich fruit juices with such substances when they were already full of vitamins anyway. The idea of bread enriched with omega-3 fatty acids was specifically thought to be „crazy”. One participant did not consider packaging to be a bad idea, and two of them also commented favorably on PET bottles.

When asked whether they would buy these products, the answer was clearly no. The group was less prone to rejection in the case of the PET bottles, with 4 people inclined to buy, and one person said the same about antimicrobial packaging.

Subsequently, the group rejecting nanotechnology also had to jointly establish an order for the six products, based on acceptability (in this case, we cannot speak of popularity, as the members of the group reject nanotechnology in the food industry). The results are shown in Table 4.

Regarding the vision for the future, participants believed that the trend of developments suggests that more and more products of this kind will be available commercially. There was also a remark in this regard that „the world is not moving in the right direction”. One person added that he was confident that we would stick to natural food sources. Several people agreed with the statement that if it is not the food industry that works with such technology, but the construction or textile industry, it may even be useful. When asked whether they would like more of these products to be available in the future, the group’s response was a clear and consistent no.

The final chapter of the focus group interview concentrated on the potential risks of nanotechnology. After discussing the potential dangers of nanotechnology with participants, their opinions were asked. Their position did not change much after what they had heard, since, as they said, they had not considered it to be a good idea, and it only strengthened their belief that such a technology could have negative consequences. The unanimous opinion of the group was that they would continue to not buy such products as they are sure that they are harmful not only to human health but also to the environment.

As a final task, participants were asked to, in possession of all the information, jointly establish a new, final order as to which category they would consider most acceptable and which least acceptable. Compared to the previous one, the order did not change much, and the result was as follows. The orders before and after the description of the risks (new order) are shown in Table 5.

5. Conclusions

Despite the fact that 74.5% of the respondents were not previously familiar with nanotechnology and its application possibilities, and almost half of the respondents believed that it involved some risk, the survey of knowledge of nanotechnology and the examination of consumers’ willingness to buy revealed that the degree of acceptance of the technology and the willingness to buy can be said to be very favorable. If, through this technology, food quality is expected to change in a positive direction, acceptance exceeds 60%.

The most important aspect when buying foods was taste, while added value finished last with 17.5%. Nevertheless, 63.0% of those who completed the questionnaire replied that they would buy a product made with a nanotechnology process if the product thus contained some kind of added value.

The focus group interview revealed that the group of acceptors, as expected, was extremely positive about the technology, and even after the description of the potential risks, neither their opinion, nor their willingness to buy typically changed.

Reaffirming Berekaa’s claim that the public is very concerned about toxicity and potential negative environmental effects [5], in the case of the group of rejectors, participants unanimously stated that the technology is extremely risky and dangerous to both the environment and humans. However, they also added that in their view and based on the trends, the proliferation of commercially available such products will be inevitable in the future. In their case it can be said that although they do not prefer the possibilities of using nanotechnology, their rejection was less pronounced for those categories of application of the technology that do not specifically change the properties of foods, but their peripherals (such as packaging).

In Chapter 3 of our paper, the statements of Sodano were already quoted, according to which the willingness to buy nanofoods for the six categories examined (creamier ice cream with the same fat content; salt and sugar that do not form lumps with moisture; fruit juices enriched with bioactive molecules; bread enriched with omega-3 fatty acids; plastic bottles for beer; antimicrobial food packaging for meat and other foods) depends to a large extent on the assessment of the perceived risks and benefits [1]. Our results obtained in the course of our research support this, as the willingness to buy of consumers who already had a positive attitude towards the technology is also very favorable, while rejectors showed the opposite consumer behavior.

6. Acknowledgment

This publication was prepared with the professional support of the New National Excellence Program of the Ministry of Innovation and Technology, code number ÚNKP-20-3-I-DE-404, financed from the National Research, Development and Innovation Fund.

7. References

[1] Sodano, V., Gorgitano, M.T., Verneau, F. (2015): Consumer acceptance of food nanotechnology in Italy. British Food Journal 118 (3) pp. 714-733

[2] Zentai A., Frecskáné Csáki K., Szeitzné Szabó M., Farkas J., Beczner J. (2014): Nanoanyagok felhasználása az élelmiszeriparban. Magyar Tudomány 175 (8) pp. 983-993

[3] Cubadda, F., Aureli, F., D Amato, M., Raggi, A., Mantovani, A. (2013): Nanomaterials in the food sector: new approaches for safety assessment. Rapporti ISTISAN 13/48.

[4] Joseph, T. and Morrison, M (2006): Nanoforum report: nanotechnology in agriculture and food. (Hozzáférés: 2014. 06. 12.).

[5] Berekaa, M. M. (2015): Nanotechnology in food industry; Advances in Food processing, Packaging and Food Safety. International Journal of Current Microbiology and Applied Sciences 4 (5) pp. 345-357

[6] Chaudhry, Q., Scotter, M., Blackburn, J., Ross, B., Boxall, A., Castle, L. y and Watkins, R. (2008): Applications and implications of nanotechnologies for the food sector. Food Additives and Contaminants 25 (3) pp. 241-258

[7] Cushen, M., Kerry, J., Morris, M., Cruz-Romero, M. and Cummins, E. (2012): Nanotechnologies in the food industry. Trends in Food Science & Technology 24 (1) pp. 30-46

[8] Weir, A., Westerhoff, P., Fabricius, L., Hristovski, K. and von Goetz, N. (2012): Titanium dioxide nanoparticles in food and personal care products. Environmental Science & Technology 46 (4) pp. 2242-2250 DOI

[9] Mura, S., Seddaiu, G., Bacchini, F., Roggero, P.P. and Greppi, G.F. (2013): Advances of nanotechnology in agro-environmental studies. Italian Journal of Agronomy 8 (18) pp. 127-140

[10] Chaudhry, Q., Castle, L., Watkins, R. (2010): Nanotechnologies in Food. Royal Society of Chemistry Publishers, Cambridge, UK.

[11] Bradley, E. L., Castle, L., Chaudhry, Q. (2011): Applications of Nanomaterials in Food Packaging with a Consideration of Opportunities for Developing Countries. Trends in Food Science & Technology 22 pp. 604-610

[12] Sozer, N. and Kokini, J.L. (2009): Nanotechnology and its applications in the food sector. Trends in Biotechnology, 27 (2) pp. 82-89.

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[14] Cushen, M., Kerry, J., Morris, M., Cruz-Romero, M., Cummins, E. (2012): Nanotechnologies in the food industry. Trends in Food Science & Technology 24 (1) pp. 30-46

[15] Qureshi, M.A., Karthikeyan, S., Karthikeyan, P., Khan, P.A., Uprit, S. and Mishra, U.K. (2012): Application of nanotechnology in food and dairy processing: an overview. Pakistan Journal of Food Sciences 22 (1) pp. 23-31

[16] Cockburn, A., Bradford, R., Buck, N., Constable, A., Edwards, G., Haber, B., Hepburn, P., Howlett, J., Kampers, F., Klein, C., Radomski, M., Stamm, H., Wijnhoven, S. and Wildermann, T. (2012): Approaches to the safety assessment of engineered nanomaterials (ENM) in food. Food and Chemical Toxicology 50 (6) pp. 2224-2242

[17] Hubbs, A.F., Sargent, L.M., Porter, D.W., Sager, T.M., Chen, B.T., Frazer, D.G. and Battelli, L.A. (2013): Nanotechnology toxicologic pathology. Toxicologic Pathology 41 (2) pp. 395-409

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DOI: https://doi.org/10.52091/EVIK-2021/3-4-ENG

Received: 2020. November – Accepted: 2021. March

¹ South Ural State University (national research university), Chelyabinsk, Russian Federation

Keywords

semi-finished products from meat of broiler chickens, freeze-dried ground apples, Brazil nuts

2. Introduction

Poultry meat is a dietary product with a high content of easily digestible proteins, low content of fat and cholesterol, it costs less than other meat, takes little time to cook and suits well for daily consumption [1]. However, today consumers tend to prefer “healthy” products, which makes producers expand the range of foods enriched with nutrients. This explains the relevance of using plant-based natural additives in meat processing industry, because they improve the quality characteristics of raw meat, and also increase nutritional and biological value of finished products [2].

It is a known fact that apple powder is rich in vitamins, organic and phenol carboxylic acids, monosaccharides, pectins, and dietary fiber, while the Brazil nut is considered a great source of complete protein, such mineral nutrients as Se, Cu, Mn, I, and fatty acids [3, 4, 5, 6, 7]. That is why, these plant raw materials are separately used in cakes, bread, chocolate, cutlets, curd cheese, cereal bars, nut and seed butters [8, 9, 10, 11, 12, 13, 14] to increase their nutrient density. The aim of our research was to study the possibility of combined use of freeze-dried ground apples and Brazil nut kernels in the technology of stuffed meat products with increased nutritional value.

3. Materials and methods

The following was used as materials of the research:

• Chilled broiler chicken legs manufactured by OAO Turbaslinskiye Broilery (Republic of Bashkortostan, Blagoveshchensk) in accordance with GOST 31962-13;
• Freeze-dried ground apples manufactured by PAO Sibirskiy Gostinets (Pskov Region, Moglino) in accordance with TU 10.39.25-001-34457722-18;
• Kernels of Brazil nuts of Bolivian origin manufactured by OOO Komservis (Moscow Region, Mytishchi) in accordance with TU 9760-002-76440635-16;
• Letniy Sad food additive manufactured by OOO Kulmbakh-D (Moscow Region, Krasnoarmeysk) in accordance with TU 10.89.19-008-58251238-20. Ingredients: dill, garlic, mustard, table salt, maltodextrin, dextrose, E621, dill extract, caraway extract, E100;
• Chicken pockets with butter and herbs cooked according to TU 9214-013-64474310-12 by way of baking stuffed broiler chicken legs at 200 ˚C for 20 minutes.

Control samples were cooked according to a traditional recipe (Table 1), test samples were cooked adding 7% dried ground apples, 5% crushed Brazil nut kernels and 4% less butter.

The dosages of the plant raw materials were chosen taking into account the known data published in a number of scientific papers [8, 9, 10, 11, 12, 13, 14] The test samples of chicken pockets were cooked using deboned chicken legs with skin, flat in shape, with a longitudinal cut in the form of a pocket filled with butter, mixed herbs, ground dried apples, and Brazil nut kernels. The cut was joined with skewers.

The plant raw materials were tested for the content of protein and fat according to MU 4237-86, sugar – GOST 8756.13-87, table salt – GOST 15113.7-77, starch – using standard approach [15]. The meat and meat products were tested for protein according to GOST 25011-2017, fat – GOST 23042-2015, moisture – GOST 9793-2016, table salt – GOST 9957-2015. Sensory evaluation of the laboratory samples was carried out according to GOST 9959-2015. The content of dietary fiber in all samples was determined using the traditional approach [15], content of organic acids – according to М 04-47-12, mineral elements – using iCAP 7200 DUO emission spectrometer.

All measurements were carried out in three replications. Statistical analysis was performed using Microsoft Excel XP and Statistica 8.0 software package. The statistical error of the data did not exceed 5% (at 95% confidence level).

4. Results and discussions

Analyzing the nutritional composition of the non-traditional plant raw materials in comparison with poultry meat (Table 2), it was found that Brazil nut kernels contained a relatively high amount of lipids (11 times more), which made it possible to reduce the amount of butter in the recipe, and hence to decrease cholesterol content in the test samples.

Apple powder proved to have relatively high levels of sugars, dietary fiber, and organic acids, in comparison with both raw meat and other plant components. It is well known that non-volatile acids in fruits not only determine taste and aroma of finished products, but also contribute to the production of gastric juice and have a choleretic effect [16], while insoluble (lignin, cellulose, chitin) and soluble (pectin, inulin) dietary fiber is able to effectively bind heavy metal ions and organic substances [17]. All these factors a priori suggest that this new component in the chicken pockets recipe should have a positive effect on the human organism.

The amino acid content in Letniy Sad food additive was due to sodium glutamate (E621) in its composition, while the presence of table salt at the level of 34.9 ± 2.2% allowed not to introduce any more of it.

The mineral composition of all plant components turned out to be richer than that of broiler chicken legs in terms of the number of elements (Table 3). In terms of the content of micronutrients, which have great physiological importance for the human organism, the Brazil nut contained 12 times more Ca, 7.4 times more Fe, 7.2 times more Se, 6.3 times more Mg, 3.6 times more P and Zn, but the Cu, Mn and Co content were also higher than in the poultry meat. Similarly, the dried ground apple powder contained 2.4 time more Fe, 2 times more Ca and 2.7 times more Si, additionally it’s Ag, Au, B, Be, Cu, Ga, Mn, Mo contain were also higher, than the content of poultry meat. Considering 0.5% dosage of Letniy Sad food additive as per the recipe, its contribution to the total mineral value of ready chicken pockets can be considered significant only in terms of Na content, which was 38 times more than in raw meat.

The levels of heavy metals in nuts – As, Cd, Pb, not found in semi-finished meat products, did not exceed the regulated norms of TR CU 021/2011.

Chilled chicken legs had a relatively high content of K, Si, as well as Na.

Thus, it was proved efficient to use such plant components in the technology of baked meat products in order to increase their nutritional value.

Tasting of the laboratory samples of chicken pockets established that apple and nut raw materials in the specified ratio had a positive effect on the consumer characteristics of the product. At the same time, the control sample did not have outstanding taste and aromatic properties, with creamy tones predominant, leveling the characteristics of a meat product. The mixture of the plant materials accounted for the formation of apple and nut notes in the smell and a slight sour-sweet tone in the taste of the products. The color on the cut acquired a caramel shade. The appearance, consistency, and juiciness of all samples were consistently high.

When testing physical and chemical indicators, it was found that the samples under study did not differ significantly in moisture, fat, and sodium chloride content (Table 4). However, the test samples contained slightly more protein (by 2.1 %), as well as dietary fiber and organic acids, which is a benefit from the standpoint of modern nutritional science.

The study of the mineral composition of the laboratory samples revealed that the test samples exceeded the control ones in terms of the amount of most macro- and microelements (Figures 1, 2). Specifically, as for macronutrients, baked samples with a modified recipe contained more Ca (1.7 times), Mg (35.4 %), and P (20 %); as for microelements – more Mo (473 times), Au (132 times), Cu (56 times), B and Mn (28 times), W (20 times), Be (17 times), Sn (15.8 times), Fe and Ti (1.5-1.6 times), Se (1.4 times), Zn (23.1 %), etc.

Furthermore, the amounts of microelements established according to MR 2.3.1.2432-08 satisfy the daily demand of an adult in Mo by 30.4 %, Cu - by 4.3%, Mn - by 2.1 % if one eats 100 g of baked poultry meat products with the added apple powder and Brazil nut.

Minerals are essential for the human body. They are a part of tissues, hormones, enzymes, intracellular fluid. They are needed for the formation of blood and bone cells, functioning of the nervous system, regulation of muscle tone, processes of energy generation, growth and recovery of the body [18, 19].

5. Conclusions

The nutrient composition of the raw materials and finished products was studied. We found that it is possible to use freeze-dried ground apples (in an amount of 7%) and Brazil nut kernels (in an amount of 5 %) together in the recipe of stuffed meat products. Modifying the recipe for chicken pockets, we obtained a product with improved consumer properties, increased nutrition value, and a decrease in the amount of butter by 4%.

6. Acknowledgement

The work was supported by Act 211 of the Government of the Russian Federation, contract № 02.A03.21.0011.

7. References

[1] Denisyuk, E. A., Tyurina, E. O. (2019): Effect of spinach on food value and economic efficiency of poultry meat semi-finished products production in conditions of LLC “Pervy Myasokombinat”. Bulletin of the Nizhny Novgorod State Agricultural Academy, 4 (24), pp. 28-32.

[2] Asfondyarova, I. V., Sagaidakovskaia, E. S. (2018): Meat semi-finished products of high nutritional and biological value. XXI Century: Resumes of the Past and Challenges of the Present, 7(43), pp. 87-92.

[3] Kishtikov, Kh. B., Dzhappueva, Zh.R. (2017): Chemical composition and curative, dietary, and preventative functions of fruit and vegetable powders added to bakery goods made of wheat flour. Alley of Science, 4(9), pp. 789-796.

[4] Pyanikova, E. A., Cheremushkina, I.V., Kovaleva, E.A., et al. (2020): The effect of apple powder on the consumption of crispbread. Bulletin of Voronezh State University of Engineering Technology. 82(1), pp. 157-163. doi.org/10.20914/2310-1202-2020-1-157-163

[5] Kantoroeva, A. K. (2019): Analysis of the development of the world market for nut crops. Economics and Management: Problems, Solutions. 2(3), pp. 147-154.

[6] Klimova, E. V. (2008): Comparative study of total oil content, fatty acid profile, peroxide value, concentration of tocopherol, phytosterol and squalene in the kernels of Brazil nuts, pecans, pine nuts, pistachios and cashews. Food and processing industry. Abstract journal. 2, p. 369.

[7] Martins, M., Klusczcovski, A.M., Scussel, V.M. (2014): In vitro activity of the brazil nut (bertholletia excelsa h. b. k.) oil in aflatoxigenic strains of aspergillus parasiticus. European food research and technology. 239(4), pp. 687-693.

[8] Nurgalieva, A. A., Pusenkova, L. I. (2017): Use of apple powder in baked confectionery products. Alley of Science. 3(10), pp. 241-248.

[9] Perfilova, O. V. (2019): Development of a new method for preparing white flour dough using apple and pumpkin powder. New Technologies. 1(47), pp. 141-148. doi.org/10.24411/2072-0920-2019-10114.

[10] Linovskaya, N. V. (2019): Development of chocolate with finely ground additions. Scientific works of the Kuban State Technological University” electronic network polythematic journal. 9, pp. 114-123.

[11] Mogilniy, M. P. (2017): Evaluation of the biological value of minced meat products with fruit fillings. Modern Humanities Success). 2(6), pp. 57-62.

[12] Ukkonen, T. I., Belozerova, M. S. (2017): Development of curd cheese with increased selenium content. Materials of the VIII International Scientific and Technical Conference «Low-temperature and food technologies in the XXI century». pp. 264-267.

[13] Patent No. 2706159 RF. Cereal bar for nutrition of those working with harmful compounds of arsenic and phosphorus. Kazan National Research University. Gumerov T. Yu., Gabdukaeva L. Z., Shvink K. Yu. Application dd. 14.05.2019; published 14.11.2019.

[14] Patent No. 2603892 RF. Method for preparing nut-like mass. Rodionova N. S., Popov E. S., Alekseeva T. V., Sokolova O. A., Shakhov A. S. Application dd. 01.07.2015; published 10.12.2016.

[15] Skurikhin, I.M., Tutelyan, V.A. (1998): A guide to the methods of analyzing food quality and safety. Moscow, Brandes, Medicine, p. 342.

[16] Nechaev, A. P., Traubenberg, S. E., Kochetkova, A. A., et al. (2012): Food Chemistry: 5th edition, revised and expanded. – SPb.: Giord, p. 670.

[17] Nikiforova, T. E., Kozlov, V. A., Modina, E. A. (2010): Solvation-coordination mechanism of sorption of heavy metal ions by cellulose-containing sorbent from aqueous media. Chemistry of plant raw material. 4, pp. 23-30.

[18] Dydykina, I. S., Dydykina, P. S., Alekseyeva, O. G. (2013): Trace elements (copper, manganese, zinc, boron) and healthy bone: prevention and treatment of osteopenia and osteoporosis. Effective Pharmacotherapy. 38, pp. 42-49.

[19] Krutenko, V. V. (2013): A close look at the role of gold trace element in the human body. Bulletin of problems of biology and medicine. 2(3), pp. 19-24.

DOI: https://doi.org/10.52091/JFI-2021/1-4-ENG

Received: June 2020 – Accepted: October 2020

¹ South Ural State University (national research university), Chelyabinsk, Russian Federation

Keywords

mayonnaise sauce, pine nut oil cake, protein concentrate, functional food product, β-carotene

2. Bevezetés

Mayonnaise sauces, like all mayonnaise products, are among the most popular everyday consumer goods. Main components for mayonnaise sauces involve natural products with high biological value and health-promoting properties. In this regard, the development of mayonnaise product formulas can be viewed as a promising line of research [1, 2].

Hydrocolloids and protein-polysaccharide complexes, plant extracts, vitamin and mineral complexes, dietary fiber, polyunsaturated fatty acids and protein concentrates are the most valuable functional ingredients in the production of emulsion foods for particular nutritional uses. These biologically active components allow you to structure a person’s diet in order to improve metabolism, immunity, nervous and endocrine systems, functioning of individual organs and body systems [3, 4].

Currently, protein concentrates are widely used in production of various sauces, pastes, dairy, and confectionery products. Such popularity of protein concentrates is due to the protein deficiency that more than 60% of people suffer from to a varying degree [5].

At the same time, every year scientists all over the world come up with new sources and methods of protein isolation to create new functional foods enriched with protein concentrates. It has been established that regular consumption of such products helps to increase body’s resistance to harmful factors, strengthen immunity, and improve metabolism [6].

Pine nut oil cake, obtained by extracting oil from pine nut kernels, is a secondary raw material, but it is of great importance as an additional source of complete protein, easily digestible carbohydrates, vitamins, and minerals. With the right choice of the method of its extraction and purification, it is possible to obtain a protein-rich concentrate that can be added to various foods in order to give them functional properties.

The composition of the protein of pine nut oil cake is determined by the composition of the protein of the kernels of pine nuts.

The content of essential amino acids in the protein composition of pine nut kernels ranges from 36 to 40%.

The content of some individual essential amino acids in pine nut protein is specific, which is characteristic of all types of plant materials. It should be noted that in terms of amino acid composition, namely, the content of phenylalanine, tyrosine, histidine, tryptophan, arginine, pine nut oil cake protein is as good as the protein of the major grain and oilseed crops. It is close to dairy protein in terms of tryptophan content, surpassing it in terms of arginine and histidine content.

The composition of the lipid fraction of pine nut oil cake is characterized by a quantitative predominance of polyunsaturated fatty acids – linoleic and γ-linolenic –, belonging to the ω-6 family.

The vitamin and mineral value of pine nut oil cake depends both on the initial chemical composition of the processed nut and on the residual oil content in the oil cake after pressing.

Pine nut oil cake has a high content of tocopherols (11.8 mg/100 g of product), thiamine (0.6 mg/100 g of product), and riboflavin (1.83 mg/100 g of product).

Pine nut oil cake is a concentrate of biologically valuable food substances like proteins, lipids, carbohydrates [7].

3. Materials and methods

The following was used as the material for research:

• ground cake of pine nut kernels, produced in accordance with TU 9146-001-53163736-06 (by “Siberian Product”, supplied by “Altai Dar LLC”, Altai Territory, Barnaul, Russia) [14];
• Beta-carotene 30% os, plant-based, liquid, oil-soluble (by “NATEC”, Moscow);
• reference and test samples of mayonnaise sauces.

Organoleptic characteristics of mayonnaises and mayonnaise sauces must comply with the requirements of GOST 31761-2012 “Mayonnaises and mayonnaise sauces. General specifications” [8]. Testing of organoleptic characteristics was carried out at (20±2) °C after at least 12 hours after production.

Organoleptic indicators were determined in the following sequence: texture, appearance, colour, smell, taste.

The mass fraction of protein was determined by the Kjeldahl titration method.

The stability of the emulsion was determined by centrifugation.

The intact emulsion stability was determined by centrifuging the emulsion for 5 minutes at 1500 rpm.

The dynamic viscosity of the samples was determined using “Reostat-2” rotational viscometer (Germany) at 20 ºС.

The degree of oxidative deterioration was determined by the peroxide number of the oil phase using the iodometric method and calculating the degree of oxidative deterioration of the product [9-11].

All measurements were carried out in three replications. Statistical analysis was performed using Microsoft Excel XP and Statistica 8.0 software package. The statistical error of the data did not exceed 5% (at 95% confidence level).

4. Results and discussion

Mayonnaise sauce is a finely dispersed emulsion product with a fat content of not less than 15%, produced from refined deodorized oil, water, with or without dairy by-products, food additives and other food ingredients (GOST 31761-2012 “Mayonnaises and mayonnaise sauces. General specifications”) [8].

The ingredients of the obtained mayonnaise sauce included refined deodorized cooking oil, egg powder, mustard flour, granulated sugar, table salt, 80% acetic acid, as well as a protein concentrate made of pine nut oil cake, natural β-carotene and water. Introducing β-carotene to the formula of the mayonnaise sauce increased the stability of its fatty phase to oxidation and extended its shelf life [12].

The production technology of the functional mayonnaise sauce was based on the “classic” mayonnaise sauce production technology.

The specified amount of water of 35–40 °C (not taking into account the water used to prepare the acetic acid solution) was poured into the mixer with a steam-water jacket. The mixer was turned on, and dry components – granulated sugar, salt, pine nut oil cake – were heated and added to the mixer. The mass was mixed intensively at 70-80 rpm and heated to 80-85 °C for 25-30 minutes. Then, the resulting suspension was cooled to 35-40 °C, egg powder and mustard flour were added, after which the emulsion was heated to 55-60 °C during 15-20 minutes.

After heating, the emulsion was again cooled to 25-30 °C, the number of revolutions was reduced to 30-40 rpm, and oil with pre-dissolved β-carotene was introduced. Following that, after adding acetic acid solution into the sauce, it was subject to stirring for another 3-5 minutes and subsequently homogenized at a pressure of 0.9-2.5 MPa.

The use of pine nut oil cake made it possible to reduce the content of egg products in the sauce formula, to lower the cholesterol, and to increase the protein content in the finished product.

The use of β-carotene in the sauce formula enhanced the color of natural egg products.

The use of pine nut oil cake not only simplified the mayonnaise sauce production process, but also allowed to obtain a colloidal system consisting of finely dispersed particles of cell walls. Intensive mixing ensured a complete interaction of proteins, fats and carbohydrates with other components, which facilitated emulsion stability, as finely dispersed cell walls of pine nut oil cake formed a solid three-dimensional structure, enhancing the emulsifying and stabilizing effect.

Reducing the mass fraction of egg powder in the formula to less than 1% made it difficult to obtain a stable emulsion, which led to a decrease in the viscosity of the finished product. The consistency of the finished product became watery, its organoleptic characteristics were low [13]. Therefore, we chose 1%, 2% and 3% dosages of pine nut oil cake to be introduced instead of egg powder.

Formulas of mayonnaise sauces are given in Table 1.

Test samples of mayonnaise sauces with pine nut oil cake were tested for organoleptic characteristics (Table 2).

The appearance of mayonnaise sauces is shown in Figure 1.

Physical and chemical indicators are given in Table 3.

The use of pine nut oil cake increased the overall protein content in the finished product. Pine nut oil cake is an effective emulsifier, and in combination with a conventional emulsifier – egg powder – ensured a good, smooth consistency of the sauce and high stability of the emulsion. It allowed to obtain a finished product with a viscosity that meets consumer requirements for compatibility with other ingredients of a dish or food systems.

In the next stage of research we studied how the quality of the mayonnaise sauce changed during storage.

Storing the samples at 20 ºC provoked the oxidation without changing the mechanism of the process and violating the colloidal stability of the product. The dynamics of the peroxide number of the oil phase of mayonnaise sauce samples during the storage at 20 ºC is shown in Figure 2.

The oxidation of the samples during storage was caused by exposure to light. When stored for more than four weeks, the peroxide number of the reference sample exceeded the level of 11 mmol of active oxygen/kg, and as for the test samples, it did not reach the level of 6 mmol of active oxygen/kg.

The use of β-carotene (0.2%) in the mayonnaise sauce can significantly increase the oxidation stability of the product without adding a preservative, as well as enrich the mayonnaise with biologically active substances of plant origin.

Based on all types of studies, it can be concluded that 3% was the most preferable dosage of pine nut oil cake in the formula of the mayonnaise sauce.

5. Conclusions

Using pine nut oil cake, which possesses good emulsifying properties, in the amount of 3%, alongside a conventional egg powder emulsifier, increased the viscosity of the finished product, ensured a smooth texture of the sauce and high stability of the emulsion. The use of pine nut oil cake made it possible to reduce the content of egg products in the sauce formula and lower the amount of cholesterol in the finished product. Besides, the introduction of pine nut oil cake to the sauce formula increased its protein content. The use of β-carotene (0.2%) in the mayonnaise sauce can significantly increase the oxidation stability of the product without adding a preservative and enrich the mayonnaise with biologically active substances of plant origin.

Thus, pine nut oil cake obtained by processing kernels of pine nuts is a promising functional additive. This material is a suitable to produce fat emulsions, including the reduced fat content products, due to the protein and carbohydrate content of oil cake, thus can provide the necessary rheological structure for low-fat products too.

6. Acknowledgement

The work was supported by Act 211 of the Government of the Russian Federation, contract № 02.A03.21.0011.

7. References

[1] Chung, C., Degner, B., McClements, D. J. (2014): Development of reduced-calorie foods: Microparticulated whey proteins as fat mimetics in semi-solid food emulsions. Food Research International, 56, pp. 136–145. http://doi.org/10.1016/j.foodres.2013.11.034.

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[4] Ma, Z., Boye, J. I. (2013): Advances in the design and production of reduced-fat and reduced-cholesterol salad dressing and mayonnaise: A review. Food and Bioprocess Technology, 6 (3), pp. 648–670.

[5] Sikora, M., Badrie, N., Deisingh, A. K., Kowalski, S. (2008): Sauces and Dressings: A Review of Properties and Applications. Critical Reviews in Food Science and Nutrition, 48 (1), pp. 50-77. http://doi.org/10.1080/10408390601079934.

[6] Diftis, N. G., Biliaderis, C. G., Kiosseoglou, V. D. (2005): Rheological properties and stability of model salad dressing emulsions prepared with a dry-heated soybean protein isolate–dextran mixture. Food Hydrocolloids, 19 (6), pp. 1025–1031. http://doi.org/10.1016/j.foodhyd.2005.01.003

[7] Gómez-Ariza, J.L., Arias-Borrego, A., García-Barrera, T. (2006): Multielemental fractionation in pine nuts (Pinus pinea) from different geographic origins by size-exclusion chromatography with UV and inductively coupled plasma mass spectrometry detection. Journal of Chromatography, 1121 (2), pp. 191-199. http://doi.org/10.1016/j.chroma.2006.04.025.

[8] GOST 31761-2012. Mayonnaises and mayonnaise sauces. General specifications. Moscow, 2013. pp. 1-13.

[9] Skurikhin, I.M., Tutelyan, V.A. (1998): A guide to the methods of analyzing food quality and safety. Moscow, Brandes, Medicine, pp. 110–115.

[10] Karas, R., Skvarča, M., Žlender, B. (2002): Sensory quality of standard and light mayonnaise during storage. Food Technology and Biotechnology, 40, pp. 119–127.

[11] Calligaris, S., Manzocco, L., Nicoli, M. C. (2007): Modelling the temperature dependence of oxidation rate in water-in-oil emulsions stored at sub-zero temperatures. Food Chemistry, 101 (3), pp. 1019–1024. http://doi.org/10.1016/j.foodchem.2006.02.056

[12] Cortez, R., Luna-Vital, D. A., Margulis, D., Mejia, E. G. (2017): Natural pigments: stabilization. 6th International Conference on Agriproducts processing and Farming. IOP Conf. Series: Earth and Environmental Science. 422, (20), IOP Publishing. http://doi:10.1088/1755-1315/422/1/012090.

[13] Kishk, Y. F. M., Elsheshetawy, H. E. (2013): Effect of ginger powder on the mayonnaise oxidative stability, rheological measurements, and sensory characteristics. Annals of Agricultural Sciences, 58 (2), pp. 213–220. http://doi.org/10.1016/j.aoas.2013.07.016.

[14] Oil industry by-products. TU catalog. Number: TU 9146-001-53163736-2006. Name: Pine nut kernel cake. Siberian product "; 656055, Altai kr., Barnaul, st. A. Petrova, 1886. http://92.243.65.78/techdocs/kgs/tu/679/info/126955/ (Hozzáférés: 2020. 06. 11.)

DOI: https://doi.org/10.52091/EVIK-2021/1-5-ENG

Received: November 2020 – Accepted: January 2021

¹ University of Debrecen, Faculty of Agricultural and Food Sciences and Environmental Management, Institute of Food Technology
² University of Debrecen Doctoral School of Nutrition and Food Sciences

Keywords

brewer’s spent grain, inactive malt, byproduct, sustainability, fiber

2. Introduction

Brewer’s spent grain is a byproduct of the brewing technology (Figure 1), which is usually utilized as animal feed, but in many cases it is transported from manufacturing plants as waste. With our experiments, we were looking to answer the question whether brewer’s spent grain can be reintroduced into the food industry, and whether enrichment with it has a proven positive effect on the nutrition characteristics of medallions made from wheat flour.

3. Brewery byproducts

The brewing industry uses various grains to produce malt. In addition to the usual and most often used barley (Hordeum vulgare L.) and wheat (Triticum aestivum L.), other grains such as maize (Zea mays L.), rice (Oryza sativa L.), oats (Avena sativa L.), millet (Pancium miliaceum L.), rye (Secale cereale L.), sorghum (Sorghum bicolor L.), spelt (Triticum spelta L.), quinoa (Chenopodium quinoa Willd.), buckwheat (Fagopyrum esculentum Moench) and amaranths (Amaranthaceae) are used more and more often as sources of starch [2, 4, 36, 37, 39].

In order to achieve the targeted organoleptic and chemical properties of the different recipes, malt blends are commonly used, which not only affects the properties of the final product, i.e., beer, but at the same time also affects the byproducts [5, 26, 29].

In the brewing process, the goal is to obtain the maximum extract content from malt and the additives during mashing. The byproduct left behind after the filtration of the mash is called brewer’s spent grain, also known as inactive malt [3, 10, 38, 40].

Brewer’s spent grain accounts for about 85% of the byproducts generated during the brewing process [25, 34]. According to some studies, the disposal of brewer’s spent grain as waste may be of environmental concern, which is why one possible use of brewer’s spent grain in aquaculture feed is being addressed. In feed intended for fish, it can effectively replace soybean meal at a rate of 50% as a potential source of protein [8, 12, 13].

Other brewery byproducts include malt germ, hot lees, brewer’s yeast and other gases, such as carbon dioxide [11, 33, 35].

3.1. Nutritional parameters of brewer’s spent grain

Brewer’s spent grain is a valuable source of nutrients. Data on the average nutrient content per 1000 g dry matter are shown in Table 1. It is a useful source of protein and fiber, rich in vitamins (mainly B1, B2 and B6) and minerals, especially calcium, phosphorus, magnesium, potassium and sodium [1].

3.2. Enrichment possibilities of bakery products with brewer’s spent grain

The regulations for bakery products can be found in prescription 1-3/16-1 of the Hungarian Food Codex (HFC) [17]. According to the definition of the HFC, enriched foods are products that contain a significant amount of one or more complementary food components. These products are not necessarily developed for general consumption, but are targeted at a specific target group [7, 32]. In the case of brewer’s spent grain added to the dough of bakery products, it can also be called an enrichment, since following drying and crushing/grinding, brewer’s spent grain can also be used in bakery products in the form of flour.

According to the literature, one of the most practical uses of brewer’s spent grain is composting, but it can also be used as an enrichment agent in the production of foods, such as baking bread, in a proportion of 5 to 10% [40]. When brewer’s spent grain is used in a higher proportion, the crumb of the bread may be sticky [15]. As a result of the enrichment, the dietary fiber content of the finished product increases. Dietary fiber has a beneficial effect on the functioning of the stomach, the small intestine and the colon [14, 41]. According to literature data, the consumption of dietary fiber by the Hungarian population is only 20-25 grams, compared to the recommended 30-35 grams per day. Enrichment with brewer’s spent grain would not only increase fiber intake, but also protein intake [9, 28, 30]. Due to its easy digestibility, barley malt in the inactive form is also used in many cases in products for small children, and its infusion has a digestion stimulating effect [27, 31].

4. Materials and methods

4.1. Preparation of products enriched with brewer’s spent grain

During our experiments, our control medallion recipe was compiled as defined in the Hungarian Food Codex [17]. In the case of the enriched products, light (barley) and dark (a 1:1:1 mixture of Chateau black dyeing malt and barley malt roasted to chocolate color and to black color) malts were used in different concentration relative to the weight of the flour. The medallions were prepared with both malts at 10%, 25% and 50% enrichment levels. After adequate mixing, the dough of the medallions was prepared from the ingredients in the recipe (Figure 2), the balls of 4-5 cm diameter were formed from the dough, and the products were prepared by baking the balls at 150 oC for 45 seconds using an electric medallion oven (Figure 3).

Names and abbreviations of the samples prepared:

• C: Control malt
• LM 10%: Light malt, 10% enrichment
• LM 25%: Light malt, 25% enrichment
• LM 50%: Light malt, 50% enrichment
• DM 10%: Dark malt, 10% enrichment
• DM 25%: Dark malt, 25% enrichment
• DM 50%: Dark malt, 50% enrichment

4.2. Chemical characteristics of the medallions enriched with brewer’s spent grain

Laboratory analyses were carried out in triplicate in the laboratories in the Institute of Food Technology and the Institute of Food Science of the Faculty of Agricultural and Food Sciences and Environmental Management of the University of Debrecen. The test were performed according to the relevant standards and methods (Table 2).

4.2.1. Total polyphenol content

In terms of the total polyphenol content of medallions enriched with brewer’s spent grain, higher values were recorded in each case compared to the control sample (Figure 4). The medallion enriched with light malt at a concentration of 50% (LM 50%) had the highest total polyphenol content of 85.17 mg GAE/100 g. Of the raw materials, the test was also performed on the light and dark malt. Dark malt had a higher total polyphenol content (132.18 mg GAE/100 g) than light malt (102.22 mg GAE/100 g).

MSZ: Hungarian Standard; MÉ: Codex Alimentarius Hungaricus

4.2.2. Flavonoid content

Regarding the flavonoid content of the medallions, it was found that enrichment with brewer’s spent grain resulted in an increase in the flavonoid content. Compared to the control sample, higher values were observed in this case (Figure 5). DM 50% medallion enriched with brewer’s spent grain had the highest flavonoid content, with a value of 27.32 mg CE/100 g.

4.2.3. Dry matter content, moisture content

In the case of the dry matter content (Figure 6), only the medallion with the code DM 10% had a higher value, 93.52%, compared to the control sample. We found that in the case of samples made from light and dark malt, the products enriched with smaller amounts of brewer’s spent grain had a higher dry matter content. With respect to the average of dry matter content values, higher values were measured in the samples enriched with dark malt, but the difference of only a few tenths of a percent did not prove to be significant.

4.2.4. Crude protein content

In terms of protein content (Figure 7), higher values were obtained for all of our enriched products compared to the control sample. Medallion with the code LM 50% had the highest protein content (13.04%). The average protein content of the products enriched with light malt, 11.88%, was higher than the average of the medallions enriched with dark malt (11.56%).

4.2.5. Fat content

During the examination of the fat content, higher values were measured in all cases compared to the control sample. As the enrichment concentration increased, the fat content of the medallions increased as well, both in the case of samples enriched with light malt and dark malt (Figure 8). The average value of the products with different light malt enrichment was 21.15%, while in the case of dark malt, the value was 23.76%. For all products enriched with dark malt, a higher fat content was measured compared to the products enriched with light malt (LM 10% - 19.55%; LM 25% - 20.49%; LM 50% - 23.4% and DM 10% - 19.7%; DM 25% - 23.72%; DM 50% - 27.87%).

4.2.6. Carbohydrate content

Of the data for total carbohydrate content (Figure 9), the highest value, 57.7%, was obtained for the control medallion, of which sugar accounted for 0.7%. This characteristic was found to be 57.23% for the sample coded DM 10%. It was true for all products enriched either with light or dark malt that the carbohydrate content decreased with the increasing rate of enrichment. Sample LM 50% had the highest sugar content of 2.62%.

4.2.7. Dietary fiber content

The dietary fiber content of the medallions was higher than that of the control medallion without enrichment for all enriched products (values ranged from 10 to 40%). The dietary fiber content increased with the rate of enrichment for the medallions enriched with both types of malt, however, the values of LM 10% (17.4%) and LM 25% (19.2%), as well as those of DM 10% (15.6%) and DM 25% (18.5%) were similar to each other, as opposed to the medallions enriched with 50% malt. The highest value was obtained for medallion DM 50% (38.9%), followed by the dietary fiber content of sample LM 50% (27.9%). The outstanding value is almost double of the value of the control sample (Figure 10).

4.2.8. Common salt content

When measuring the salt content of each medallion (Figure 11), the highest value was obtained for the control medallion (2.5%). This was followed by the products with an enrichment of 10% (LM 10% 2.28% and DM 10% 2.36%), then the products with an enrichment of 25% (LM 25% and DM 25%), and finally the medallions with an enrichment of 50% (samples LM 50% and DM 50%). The medallions enriched with dark malt always exhibited higher values (2.36%; 1.73%; 1.41%) than their light counterparts (2.28%, 1.52%, 1.18%).

4.2.9. Energy content

During the study, the energy content of the medallions was also determined (Figure 12). The energy content value of the control medallion (1984 kJ/100 g, 474 kcal/100 g) was exceeded in all cases by the enriched medallions. The medallion enriched with 50% dark malt had the highest energy content of 2324 kJ/100 g (555 kcal/100 g). In terms of energy content, the data were almost identical, showing only small differences compared to the control sample and also to each other.

4.2.10. Organoleptic analysis

In April 2019, 20 judges were asked to evaluate, by tasting and completing a questionnaire, the following four organoleptic characteristics: appearance, smell, taste, texture. They were able to express their opinions using a scale from 1 to 5, where 1 meant very bad and 5 meant delicious.

As a result of the sensory examinations, it was found that enrichment with brewer’s spent grain deteriorated the properties of the products in all cases (Figure 13). Irrespective of the malt type, there was only a slight difference between the 10% and 25% enrichments, while the 50% enrichment resulted in a large decrease. All parameters of the products with 10 and 25% enrichment with light malt fell into the good category (with values above 4.0), so we definitely would like to continue our research with these two products.

5. Summary and recommendations

In terms of the total polyphenol, flavonoid, protein, fat, dietary fiber and energy content, higher values were measured in all cases compared to the control sample, whereas a decrease was found for three of the parameters analyzed: dry matter, carbohydrate and common salt content. This effect can be considered advantageous in the case if parameters with reduced values, especially because of the reduced carbohydrate content, while among the chemical components with increased values, the increase in fiber content is particularly important. It is possible to introduce the utilization of brewer’s spent grain in the baking industry, and the enrichment of wheat flour medallions with brewer’s spent grain had a positive effect on their nutritional values. However, as a result of the enrichment, based on the data of organoleptic analysis, a certain unfavorable change in the properties of the medallions (apeearance, smell, taste, texture) could be observed, but the results of the enrichment with light brewer’s spent grain show that an edible product can be prepared by further developments aimed at improving the organoleptic properties.

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