Method for obtaining plant-based analogues of dairy products

Abstract

FIELD: food industry.SUBSTANCE: method for obtaining plant analogues of dairy products is proposed. The method involves obtaining an alkaline extract of peanut cake, its heat treatment at (115±1) °C for 30 min, cooling to (37±1) °C. The resulting extract is inoculated with lactobacilli strains selected from the group: Lacticaseibacillus casei UQM 41618, L. casei UQM 41619, L. casei UQM 41620 and L. rhamnosus UQM 41622 grown on sterile cow's milk at (37± 1) °C for 24 hours, followed by fermentation under static conditions at a temperature of (37±1) °C.EFFECT: invention allows to obtain lactose-free plant-based fermented products using a by-product of peanut oil production – cake.1 cl, 1 tbl, 4 dwg, 1 ex

Description

[0001] With a constantly growing population and changing consumer preferences, traditional dairy production methods are not always able to meet market demands. A promising approach to addressing this issue is the development of plant-based dairy alternatives.

[0002] Among the plant materials, peanuts (Araches hypogea) have great potential, playing an important role in the food security of many countries. The main producers of peanuts are China (18.3 million tons / year), India (10.1 million tons / year) and Nigeria (4.3 million tons / year) [1]. There is a high interest in this crop among the population of the Russian Federation; however, at present, its industrial production is practically non-existent in the country. Every year, more than 150 thousand tons of peanuts are imported to us from abroad [2]. Nevertheless, Russian breeders are creating and researching zoned peanut varieties (Otradokubansky, Dessertny, etc.) for its potential industrial production [3, 4]. It should be noted that peanuts improve soil quality by enriching it with nitrogen, which reduces the use of chemical fertilizers.

[0003] The advantages of peanuts are their high protein content (22-30%), oil (44-56%), of which about 80% are unsaturated fatty acids, the presence of vitamins and microelements, and the absence of lactose [4, 5]. Peanuts contain compounds with high biological activity: tocopherols, polyphenols, flavonoids, paracoumaric acid, sterols, sterols, and therefore, it is advisable to obtain functional products from this crop [6-10]. Peanut consumption reduces the risk of the occurrence and development of cancer, cardiovascular, diabetic, and infectious diseases [11, 12].

[0004] Nevertheless, peanuts are a strong allergen for 15% of the world's population. Thermal or enzymatic processing of raw materials can reduce or eliminate allergenicity [13-15]. Dairy product analogues are produced from peanut kernels, the basis of which is peanut "milk" or peanut protein isolate [16, 17]. Peanut "milk" is obtained by pre-sprouting or soaking the kernels in an alkaline solution, blanching, roasting, extraction, followed by neutralization and filtration

[18] .

[0005] Due to its composition, peanut "milk" is a complete nutrient base for the cultivation of lactic acid bacteria. Studies show that lactobacilli grow faster on peanut "milk" compared to natural cow's milk

[19] . For the fermentation of plant "milks", including peanut milk, starters of lactobacilli Lactobacillus helveticus, L. rhamnosus, Streptococcus thermophiles, L. bulgaricus, L. delbruecki, acetic acid bacteria and dairy yeasts are used [19-21]. During the fermentation process, under the influence of microorganisms, the chemical composition and structure of plant materials changes, anti-nutrients and allergens are inhibited, the nutritional value, as well as the antioxidant and immunomodulatory properties of the resulting product, increase [22-26]. Peanut "milk" and yogurts made from it are high in protein, essential amino acids, unsaturated fatty acids and minerals.Yogurts made from peanuts have increased water-holding capacity and decreased syneresis

[27] .

[0006] After pressing the oil from peanuts, a cake is formed—a byproduct whose protein content reaches up to 50%, significantly exceeding the amount found in the kernels. The cake also contains several biologically active substances, including resveratrol, which has anti-cancer and antimicrobial properties

[28] .

[0007] Previously, due to its coarse texture and beany flavor, peanut cake was used primarily as a feed additive. Currently, peanut cake is used to produce flour, which is used in the production of food products such as halva. The favorable chemical composition and lower cost of peanut cake compared to whole grains demonstrate its potential for use in the creation of functional dairy analogues. The biotechnological potential of peanut cake is currently underestimated, and research in this area is limited.

[0008] The purpose of the present invention is to develop a method for producing lactose-free plant-based fermented products using a secondary product of peanut oil production - cake, and to study the physicochemical properties, chemical composition and organoleptic indicators of the resulting lacto-fermented product.

[0009] The invention can be demonstrated by the following set of essential features:

[0010] A method for producing plant analogues of dairy products, characterized in that the alkaline extract of peanut cake after heat treatment (115 ± 1) °C for 30 min is cooled to (37 ± 1) °C, the resulting extract is inoculated with lactobacilli strains Lacticaseibacillus caseiKMS-1 or L. caseiKMS-2 or L. caseiKMS-3 or L. rhamnosusKMS-5, grown on sterile cow's milk at (37 ± 1) °C for 24 hours, followed by fermentation under static conditions at a temperature of (37 ± 1) °C.

[0011] Example 1

[0012] The object of the study was peanut kernel cake obtained in laboratory conditions on a L'equip Oil Presso LOP-G3 screw oil press (China) from peanut kernel samples (Auchan LLC) harvested in 2023.

[0013] The study utilized the lactobacillus strains Lacticaseibacillus caseiKMS-1, L. caseiKMS-2, L. caseiKMS-3, and L. rhamnosusKMS-5 from the collection of the S.N. Vinogradsky Institute of Microbiology, Russian Academy of Sciences, isolated from kumys in the Republic of Bashkortostan (Russia). The strains are deposited in the Collective Use Center "Collection of Unique and Extremophilic Microorganisms of Various Physiological Groups for Biotechnological Purposes (UNIQEM)" of the Federal Research Center of Biotechnology, Russian Academy of Sciences, under the registration numbers UQM 41618, UQM 41619, UQM 41620, and UQM 41622, respectively.

[0014] The plant base for obtaining the lacto-fermented product (LFP) was an alkaline extract of peanut cake, obtained according to the method described in

[29] . After heat treatment ((115 ± 1) °C for 30 min) and cooling to (37 ± 1) °C, the extract was inoculated with lactobacilli strains grown in sterile cow's milk (OJSC Severnoye Moloko) at (37 ± 1) °C for 24 hours. Fermentation was carried out under static conditions at a temperature of (37 ± 1) °C for different durations. The number of viable lactobacilli cells in the LFP was determined by serial dilutions in a sterile sodium chloride solution (0.9 g / dm 3 ), followed by plating on MRS agar plates, then the grown colonies were counted and the viability value (CFU / cm) was calculated. 3 ).

[0015] The degree of syneresis of peanut LFP was determined after storage for 24 hours at (4 ± 1) °C according to the method

[30] . The degree of syneresis was established by measuring the amount of whey released during 2 hours of free filtration of 100 cm 3 The viscosity of the LFP consistency was determined using a Brookfield DV-II + Pro viscometer (USA) with a 06 spindle at 100 rpm and a temperature of (20 ± 1) °C. The active acidity of the samples was measured using a 150 MI pH meter (Izmeritelnaya Tekhnika LLC). The organoleptic evaluation of the LFP samples was conducted according to GOST R 70650-2023.

[0016] For further studies, the LFP samples were dried in a FreeZone Labconco (USA) lyophilization unit in a vacuum at –80 °C.

[0017] The mass fraction of moisture in the samples was determined according to GOST R 54705-2011; protein (Nx6.25) – according to GOST 10846-91; ash content – ​​according to GOST 13979.6-69; fat – according to GOST 13496.15-2016; carbohydrates – as the difference between 100% and the sum of the remaining components.

[0018] The amino acid composition was determined according to GOST 32195-2013 using a liquid chromatograph from Hitachi (Japan) in the standard mode for analyzing protein hydrolysates with sulfonated styrene-divinylbenzene copolymer and a step gradient of sodium citrate buffer solution with increasing pH and molarity. During sample preparation during acid hydrolysis of the samples, tryptophan was destroyed and was not taken into account in the calculation. The amino acid score of the samples was calculated taking into account the FAO / WHO 2011 "reference protein" scale

[31] .

[0019] The functional and technological properties and fractional composition of peanut cake proteins and LFP were studied using the methods described in [32–34]. Protein digestibility in vitro was determined using the method of A.A. Pokrovsky and I.D. Ertanov

[35] , using a model of digestion in the human gastrointestinal tract with pepsin (JSC LenReaktiv) in an acidic medium (pH 1.8 ± 0.1) and trypsin (OOO Samson-Med) in an alkaline medium (pH 8.2 ± 0.1), the total duration of the process was 360 min.

[0020] Lipids from lyophilized samples were extracted with a chloroform:methanol hydrochloride (SupelcoMethanolic-HCl 0.5 N) mixture (2:1) using the Folch method. The fatty acid composition of lipids was analyzed using a Shimadzu GCMS-QP 2010 Ultra chromatograph with a mass detector (Japan) at 120 °C with a gel carrier at a flow rate of 35.6 cm / s.

[0021] The studies of the amino acid composition and fatty acid profile were carried out at the Industrial Biotechnology Collective Use Center of the Federal Research Center of Biotechnology of the Russian Academy of Sciences and the A.N. Belozersky Institute of Physicochemical Biology of Moscow State University.

[0022] All experiments were performed in triplicate, and data were processed using Microsoft Excel 2007 and additional add-ins. Calculations included the mean and standard deviation for p < 0.05.

[0023] Results and discussion. The experimental products were obtained using a sterile extract obtained by alkaline extraction of peanut cake, which had the following chemical composition: moisture - (4.69 ± 0.12)%, protein - (41.73 ± 0.07)% dry matter (DM), ash - (2.84 ± 0.04)% DM, lipids - (39.41 ± 0.09)% DM, carbohydrates - (16.02 ± 0.20)% DM. Lactobacilli capable of actively fermenting the new substrate were selected from strains of the genus Lacticaseibacillus. All studied lactobacillus strains were fermented with extract, forming curds of varying consistencies. The pH of the suspension decreased from 8.5 to 5.0–6.3 over 4–7 hours of fermentation. The rate of dense curd formation and pH reduction was highest for the L. rhamnosus KMS-5 strain, with a pH of 5.0 achieved within 4 hours of fermentation (Fig. 1). Further studies were conducted with a starter culture containing the L. rhamnosus KMS-5 strain.

[0024] Fig. 1 shows the dependence of the change in pH of peanut LFP depending on the duration of fermentation using different strains of lactobacilli.

[0025] The organoleptic characteristics and physicochemical properties of the resulting experimental LFP sample were studied. The LFP consistency was dense and uniform, with a viscosity of 2200 mPa s and no foreign inclusions. The amount of whey isolated from 100 cm 3 LFP clot after 2 hours of free filtration amounted to 45 cm 3 (the degree of syneresis is 45%), which is somewhat lower or comparable to the degree of syneresis of fermented milk drinks based on cow's milk (43–52%)

[36] . During the fermentation process, the suspension's color changed from light cream to white, while the LFP was distinguished by a pronounced fermented milk and weak peanut aroma and taste (Fig. 2).

[0026] Analysis of the chemical composition of the obtained LFP revealed the content (g / 100 g of product): dry matter - (13.75 ± 0.03), protein - (6.06 ± 0.04), ash elements - (0.65 ± 0.01) g, lipids - (4.71 ± 0.03), carbohydrates - (2.33 ± 0.11). The energy value of LFP per 100 g of product was 318 kJ, or 76 kcal. The number of viable cells of lactic acid bacteria is 1 10 9 CFU / cm 3 No mold or yeast were detected. Therefore, based on its consistency, viscosity, degree of syneresis, lactobacillus count, chemical composition, energy value, and the absence of lactose, peanut LFP can be classified as a functional product of the "plant-based yogurt" type.

[0027] Fig. 2 shows the appearance of peanut cake (a) and LFP (b).

[0028] The LFP contained 17 amino acids (excluding tryptophan), the predominant ones being glutamic and aspartic acids, arginine, leucine, glycine, and proline (Fig. 3). The total amount of essential amino acids per 100 g of product is 2.01 g.

[0029] Fig. 3 shows the amino acid composition of peanut LFP. Values ​​of the amino acid LFP score above 100% were observed for histidine, leucine, lysine, and the sum of phenylalanine and tyrosine (Table 1). The score of the limiting essential amino acid (isoleucine) is 88%, which is a high indicator of protein quality and is comparable to the score of the limiting amino acid (86% for valine) of chickpea protein concentrate with a protein content of over 80%

[37] .

[0030] Table 1

[0031] Content of essential amino acids and amino acid score of peanut LFP

[0032] Substance Content 1 2 Essential amino acids, mg / g protein LFP: Val 37,47 His 25,50 Ile 26,35 Leu 62,40 Lys 49,29 1 2 Met + Cys 21,04 Trh 22,31 Phe + Tyr 89,77 Essential amino acids, mg / g “reference protein” (FAO / WHO, 2011): Val 40 His 16 Ile 30 Leu 61 Lys 48 Met + Cys 23 Trh 25 Phe + Tyr 41 Amino acid score of LFP, % of the “reference protein” values: Val 94 His 159 Ile 88 Leu 102 Lys 103 Met + Cys 92 Trh 89 Phe + Tyr 219

[0033] Analysis of the fractional composition of peanut LFP proteins compared to the original cake revealed an increase in the salt-, alcohol-, and acid-soluble fractions by 4.3, 4.1, and 2.0 times, respectively (Fig. 4). At the same time, a decrease in the water-soluble fraction by 6.8 times was observed. However, the content of poorly soluble (soluble in an alkali solution at pH 12.5 ± 0.1) and insoluble fractions also decreased by 2.5–2.6 times, indicating an increase in the bioavailability of LFP proteins compared to the proteins of the original cake.

[0034] Fig. 4 shows the fractional composition of proteins in peanut cake and LFP.

[0035] The increase in the bioavailability of LFP proteins is confirmed by a study of their digestibility in an in vitro model (Table 2). Compared to the original cake, the proteins of peanut LFP were digested 9.90% more intensively after 6 hours of hydrolysis. Moreover, in an acidic medium with pepsin, LFP proteins were digested 14.33% faster, and in an alkaline medium with trypsin, the proteins of the original cake were digested 15.62%. Adjusted for increased digestibility (83.61%), the biological value of protein (PDCAAS) of LFP for the limiting essential amino acid (isoleucine) is 73.58%, while, according to the literature, the biological value of peanut proteins is 52%

[38] .

[0036] Table 2

[0037] Digestibility of proteins of peanut cake and LFP, %

[0038] Sample Wednesday With pepsin (pH 1.8±0.1) 3 hours With trypsin (pH 8.2±0.1) 3 h Total 6 hours Oilcake 64,81±0,12 11,27±0,20 76,08±0,32 LFP 74,10±0,17 9,51±0,14 83,61±0,31

[0039] The fatty acid composition of LFP is represented by 10 components, the main part of which are omega-6 (linoleic) and omega-9 (oleic) unsaturated fatty acids (Table 3). Linoleic acid is considered essential and enters the body exclusively with food. In general, among the lipids of LFP, the share of unsaturated fatty acids accounts for 74.38%. Among the saturated fatty acids, palmitic acid, which is the main fatty acid of breast milk and is included in the composition of most infant formulas, predominated [39, 40]. Thus, peanut LFP was a yogurt-type product with a high content of biologically valuable protein and unsaturated fatty acids.

[0040] Table 3

[0041] Fatty acid composition of peanut LFP, % of the total amount of acids

[0042] Acid Compound Saturated fatty acids: Myristic C14:0 0,24 Palmitic acid C16:0 15,72 Stearic C18:0 3,92 Arachidic C20:0 1,37 Behenic C22:0 3,37 Lignocerol C24:0 1,00 Unsaturated fatty acids: Linoleic C18:2(9,12) 37,36 Oleic C18:1(9) 33,75 Petroselin C18:1(6) 1,50 Gondoevaya C20:1(11) 1,77

[0043] Due to the high content of valuable protein ((42.83 ± 0.10)% on a dry basis), another area of ​​application of the obtained LFP after drying is its use as a protein additive that improves the structure and functional properties of food products. For this purpose, the functional and technological properties of the LFP were determined and compared with similar properties of the original peanut cake, which also had a high protein content ((41.73 ± 0.07)% on a dry basis). As can be seen from Table 4, the dried LFP had, compared to the cake, a water-binding capacity (WBC) that was 2.2 times higher, a foaming capacity (FC) that was 1.8 times higher, a foam stability (FS) that was 2.1 times higher, a fat-binding capacity (FBC) that was 3.7 times higher, and an emulsion stability (ES) that was 13.46%. A decrease was recorded only in the fat emulsifying capacity (FEC) of LFP by 20.34%.The established properties of LFP are similar to the properties of commercial pea protein concentrate

[41] , and the values ​​of DP and YSS are 3.9–4.3 times higher, which allows us to recommend dried LFP as a protein additive in the technology of obtaining plant “meat” and food products with a foam system (marshmallows, pastilles, mousses, etc.).

[0044] Table 4

[0045] Functional and technological properties of peanut cake and dried LFP Functional and technological properties of peanut buttercake and dried LFP

[0046] Indicator Oilcake LFP VSS, g / g 0,97±0,07 2,15±0,02 POS, % 9±1 16±1 SP, % 33±1 70±1 ZhSS, g / g 1,01±0,03 3,74±0,26 Housing and communal services, % 59±1 47±1 SE, % 52±1 59±2

[0047] Conclusion. Peanut cake is a favorable raw material for lactic acid bacteria in the development of functional lacto-fermented products. The lactobacillus strain L. rhamnosus KMS-5 demonstrated better affinity for the substrate, actively reducing the pH of the medium and forming a dense clot after 4 hours of fermentation. The obtained LFP had a high content of valuable protein (6.06 g / 100 g) and lipids (4.71 g / 100 g). The total amount of essential amino acids in LFP is 2.01 g / 100 g, the amino acid score is 88-219%, and the biological value of the protein is 73.58%. The increase in the bioavailability of LFP protein compared to cake proteins is confirmed by a decrease in poorly soluble and insoluble protein fractions, as well as an increase in its digestibility. The mass fraction of unsaturated fatty acids in LFP lipids was 74.38%, omega-6 (linoleic) acid accounted for 37.36%.The LFP had a dense, uniform consistency with a viscosity of 2200 mPa s and a syneresis degree of 45%. It was white in color, had a fermented milk flavor with a slight peanut flavor, and contained the required number of viable lactic acid bacteria cells. The absence of lactose allows the product to be classified as a functional "plant-based yogurt" and recommended for people with lactose intolerance. The dried LFP possessed high functional and technological properties, particularly fat-binding capacity and foam stability, making it suitable for use as a protein supplement in the production of plant-based "meat" and food products with foam systems.

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Claims

A method for producing lacto-fermented products based on plant material, which involves obtaining an alkaline extract of cake, heat treating the obtained alkaline extract of cake for 30 minutes, cooling, inoculating with a strain of lactobacilli selected from the group comprising Lacticaseibacillus casei UQM 41618, Lacticaseibacillus casei UQM 41619 and Lacticaseibacillus casei UQM 41620, grown on a sterile medium for 24 hours, fermenting under static conditions, characterized in that an alkaline extract of peanut cake is used as the alkaline extract of cake, the heat treatment of which is carried out at a temperature of (115 ± 1) ° C, cooled to (37 ± 1) ° C, when inoculating the alkaline extract of peanut cake, the strain Lacticaseibacillus is included in the group of strains Lacticaseibacillus casei rhamnosus UQM 41622, when growing the indicated strains, cow's milk is used as a medium, grown at (37±1)°C, fermentation is carried out at a temperature of (37±1)°C.

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