RU2791226C1 - Method for obtaining fodder concentrate - Google Patents

Abstract

FIELD: biotechnology.

SUBSTANCE: invention relates to the field of biotechnology. The invention is a method for obtaining a fodder concentrate, which provides for the preparation of a nutrient medium, having settled microorganisms, and conducting enzyme. In the process of preparing the nutrient medium, serum waters of legumes (peas, chickpeas) formed during the production of protein are used, insoluble residue in the amount of 2-10% of their mass is introduced into serum water, and the suspension is subjected to acid hydrolysis with the storage of 1.8 -3.0, temperatures 60-129°C for 10-30 minutes or enzymatic hydrolysis with cellulolytic and amylolytic enzyme preparations in a concentration of 50-170 and 2-5 units. activity per 1 g of dry matter, respectively; processing is carried out 3.0-4.0 hours at 50-60°C and pH 5.0-6.0 when mixed; at the end of the processing of the pH, the suspension is brought to 4.3-5.0, they are applied with Xilanase and gluc-amymilasis enzyme drugs at the rate of 50-70 and 2-4 units. activity per 1 g of dry substance, processing is carried out within 3-4 hours at 55-60°C when mixed, corrected by pH, sowed by the birth of Geotrichum, Saccharomyces, Bacillus or their consortium composed in any ratio in the amount of 5-10% to mass of a nutrient medium.

EFFECT: invention improves the quality of the nutrient medium and the target product, achieved by increasing the degree of conversion (modification) of the insoluble residue containing starch, non-starch polysaccharides and proteins into water-soluble reducing sugars and peptides (amino acids) easily digestible by microorganisms due to acid or enzymatic hydrolysis; use instead of the whey of grain crops more complete whey of leguminous crops - peas, chickpeas, containing more digestible essential amino acids, soluble carbohydrates, minerals; increase the mass fraction of biologically complete protein, soluble fibers, minerals in the composition of the feed concentrate, its digestibility due to a more balanced composition of the nutrient medium and the selection of the type of microorganisms; give the product probiotic properties.

1 cl, 4 dwg, 4 ex

Description

Technical field

The invention relates to biotechnology, microbiological and processing industries, in particular, to the production of feed concentrate by microbial transformation of raw materials.

State of the art

Despite the active development of the market for feed preparations, the agro-industrial complex is experiencing a shortage of cheap complex feed additives containing proteins, amino acids, carbohydrates, and minerals. Most of them, as a rule, have a high cost, to reduce which it is necessary to use an inexpensive, but chemically complete and safe nutrient substrate for fermentation by microorganisms - producers of biologically valuable compounds.

Traditional methods of obtaining fodder biomass, based on the use of nutrient media obtained from hydrolysates of plant materials, oil hydrocarbons and lower alcohols, have a number of environmental disadvantages. Nutrient media require laborious purification from lignohumic impurities, furfural, hydroxymethylfurfural, turpentine and other compounds that inhibit the growth of microorganisms. When obtaining fodder biomass on hydrocarbons, it is also necessary to thoroughly clean the substrate from oil paraffins, use a significant amount of water, including for cooling heat exchangers.

Microbiological productions are characterized by a large consumption of water for the preparation of a nutrient medium and the growth of microorganisms with the formation of a culture liquid after the extraction of microbial biomass. The latter contains organic and mineral substances that are discharged into sewers or water bodies, polluting the environment. Therefore, in order to fill the deficit of feed biomass, new raw materials and technical solutions are needed to save water, reduce costs, increase the feed value of the product, save resources and ensure environmental safety of production.

A known method for producing a protein feed additive, which involves the use of post-alcohol stillage as a nutrient medium, to which starch-containing raw materials are added in an amount of 1-8% (bran or flour or a mixture thereof in any ratio), subsequent hydrolysis of the suspension with keeping the mixture for 1 hour at pH 4.0-5.5 and a temperature of 60-100°C, the introduction of nitrogen, phosphorus, iron and magnesium, the cultivation of Candida retilis yeast, the production of biomass subjected to heat treatment at a temperature of 90-100°C for 1 hour and drying on a spray dryer (Pat. RF No. 2159287, 2000). The disadvantages of this method are the duration of the hydrolysis process, significant energy and water consumption, high fiber content - 11.5-28.6% on dry matter.

A known method for obtaining biomass of microorganisms, including the preparation of a nutrient medium containing starch and cellulose-containing vegetable raw materials - substandard grain and flour mill waste, its enzymatic hydrolysis using a multi-enzyme composition as a biocatalyst at a temperature of 50-70 ° C for 20-180 minutes and The pH of the medium is 4.5-5.5. Seeding of the hydrolyzate is carried out with culture strains of Endomycopsis fibuligera, Aspergillus niger . The disadvantages of the method are the complexity of preparing a multicomponent composition of the nutrient medium, the use of mold fungi of the genus Aspergillus as a biomass producer, which are the causative agents of superficial mycosis and spergillosis , which affects the respiratory tract, lungs and paranasal sinuses. Mortality in disseminated aspergillosis reaches 80%. (Pat RF No. 2112 806, 1998).

A method for obtaining fodder yeast biomass is described, including acid hydrolysis of coffee sludge at a pH of 0.8-1.3, a temperature of 118-123°C for 30 minutes, cultivation on the obtained hydrolyzate of the Saccharomyces cerevisiae yeast, separation of the biomass from the cultural liquid by filtration to a dry content. substances 40-50% and drying of the biomass at a temperature of 105°C. (Pat RF No. 2 393 719, 2010). The disadvantages of the method are the presence in the composition of the feed product of residual components of coffee sludge, indigestible by yeast, and the low content of protein substances - 10.6-15.7 g/l.

A known method for producing fodder yeast, involving the inoculation of rosehip juice with wine yeast Saccharomyces vini , the addition of mineral components, fermentation, production, dilution of the cultural liquid with water, the introduction of mineral salts, an aqueous extract of spruce needles, the cultivation of Candida utilis yeast, the isolation of biomass and its drying (Pat. RF No. 2 580 160, 2016). The disadvantages of this method are the complexity, the multi-stage preparation of the nutrient substrate and the cultivation of yeast, the use of conditionally pathogenic yeast Candida capable of causing candidiasis.

Closest to the proposed method is a method of obtaining a feed microbial-vegetable additive, involving the preparation of a nutrient medium using whey water released after the extraction of starch and protein components from cereals, into which an insoluble residue formed during protein production is introduced in an amount of 2-5 % by weight of whey and pulp remaining after extraction of starch A, in an amount of 2-7% by weight of whey, inoculation of microorganisms, fermentation, heat treatment and drying of the suspension (Prototype Pat. RU No. 2 673 125, 2018). The disadvantage of this method is the use of whey as part of the nutrient medium, which is formed after the isolation of starch and protein from cereals, insoluble residue and pulp, which are of poor quality due to the presence of a large amount of poorly digestible fiber and unextracted starch. This is reflected in the yield of biomass and the physicochemical parameters of the feed concentrate: the mass fraction of protein, biological value and digestibility.

The problem lies in the creation of a method for obtaining a feed concentrate by bioconversion of the whey of leguminous crops and a modified insoluble residue, which are formed as secondary products of their deep processing during the isolation of protein concentrates.

The technical result is achieved by the fact that water or other reagents are added to pea or chickpea flour at a ratio of 1:10÷1:15, soluble protein compounds are extracted from the flour by any method, and the released whey water with a chemical composition containing 3-20% solids (CB), nitrogenous substances 7-22%, reducing sugars: glucose - 4-43%, fructose - 10-15%, galactose 4-5% of the total amount of carbohydrates, seeded with yeast and fermented. After the end of fermentation, the suspension is thermally treated and dried. As a biomass producer for the fermentation of whey water, monocultures of the genera Geotrichum, Saccharomyces, Bacillus or their consortium, composed in any ratio in the amount of 5-10% by weight of the nutrient medium, are used, after fermentation, the suspension is heated to a temperature of 90-100 ° C for 10- 20 min and dry.

The technical result is also achieved by the fact that a solid insoluble residue is introduced into whey water in the amount of 2-10% of their mass, which is formed, along with whey, in the process of extracting protein from the extract, acid hydrolysis of the suspension is carried out at pH 1.8-3.0 with incubation for 10-30 min at a temperature of 60-129°C or sequential enzymatic hydrolysis with cellulolytic and amylolytic enzyme preparations at a concentration of 50-170 and 2-5 units. activity per 1 g of dry matter, respectively, for 3.0-4 hours at 50-60°C and pH 5.0-6.0 with stirring; at the end of the treatment, the pH of the suspension is adjusted to 4.3, aqueous solutions of xylanase and glucoamylase enzyme preparations are added at the rate of 50 and 2.5 units. activity per 1 g of dry matter, treatment is carried out for 3 hours at 55°C with stirring, the resulting nutrient medium is treated at a temperature of 90-120°C for 10-20 minutes, cooled, monocultures or their compositions are sown.

The technical result of the invention is:

- improving the quality of the nutrient medium and the target product, achieved as a result of increasing the degree of conversion (modification) of the insoluble residue containing starch, non-starch polysaccharides and proteins, into easily digestible water-soluble reducing sugars and peptides (amino acids) by microorganisms due to acid or enzymatic hydrolysis;

- the use of more complete whey of leguminous crops (peas, chickpeas) instead of the whey of grain crops, containing a greater amount of digestible essential amino acids, soluble carbohydrates, minerals;

- increasing the mass fraction of biologically complete protein, soluble fibers, minerals in the composition of the feed concentrate, its digestibility due to a more balanced composition of the nutrient medium and the selection of the type of microorganisms;

- introduction into the consortium of microorganisms of sporogenousBacillus subtilis forgiving the product probiotic properties;

- the possibility of obtaining a new line of cheap, but complete in composition, feed products for animals, poultry, aquaculture;

- reducing the cost of the target feed product and the load on the biosphere as a result of a waste-free method for processing a liquid by-product released during the production of food protein.

Whey and insoluble sediment of legumes, formed during the production of food protein concentrate, are superior in composition to whey and insoluble sediment, which are released during the production of starch and protein from cereals (triticale, oats, barley) using a similar technology. The whey from leguminous crops (peas, chickpeas) in total contains more low molecular weight reducing sugars (glucose, fructose, xylose, arabinose, galactose) and, accordingly, less high molecular weight carbohydrate compounds, which include dextrins of various molecular weights (Fig. 1). The latter will be more difficult to transform under the action of microbial enzymes into new synthetic biomass substances. A more active synthesis of biomass will also be facilitated by a 1.35-3.40 times higher amount of nitrogenous substances, including true protein, in the whey and in the hydrolyzate, which are necessary to build biologically complete proteins of the feed product due to their greater content in the raw materials of legumes. , compared with cereals (triticale, oats, barley) (20.1-24.0% vs. 9.9-12.8%). Non-starch polysaccharides (fiber, hemicelluloses), starch and, in part, proteins of the insoluble residue in the process of preliminary acid or enzymatic hydrolysis are hydrolyzed to form smaller reducing sugars (hexose, pentose) and amino acids and serve as more effective nutritional components for microorganisms due to their lower molecular weight. masses.

The feed concentrate obtained by the developed method on the basis of a new nutrient medium surpasses the similar concentrate from grain crops in terms of protein mass fraction (2.1 times), lower lipid content (2-5 times), more soluble fibers (40%), less, respectively, insoluble fibers and a greater mass fraction of ash (1.3-1.8 times), reflecting the content of minerals, which positively affects the nutritional value and digestibility of the concentrate; the latter is higher in the new concentrate by 12-17% compared to the prototype (Fig. 2). Soluble and insoluble fibers of the feed concentrate normalize the functioning of the gastrointestinal tract, promote the detoxification of unwanted compounds, the removal of heavy metals and radionuclides, providing increased feed digestibility.

The introduction of the sporogenous probiotic Bacillus into the composition of the consortium of microorganisms separately or with producers of the genera Geotrichum, Saccharomyces, gives the feed concentrate preventive qualities, since its enzymes contribute to a more complete digestion of feed, and synthesized peptides and non-peptide substances with a wide spectrum of antimicrobial activity stimulate the normal growth of intestinal microflora ( Sevustyanenko A.V., 2016; Feoktistova N.V. et al., 2017).

The method includes the following technological operations:

Water or other reagents are added to pea or chickpea flour at a ratio of 1:10 ÷ 1:15, soluble protein compounds are extracted from the flour by any method, and to the released whey water with a chemical composition, including 3-21% dry matter (DS), from of nitrogenous substances 8.62-22.48% and reducing sugars (glucose, fructose, galactose, arabinose, xylose, maltose) 48.11-67.48% of the total amount of carbohydrates, contribute an insoluble residue in the amount of 2-10% to their mass, acid hydrolysis of the suspension is carried out at pH 1.8-3.0 with keeping for 10-30 minutes at a temperature of 60-129 ° C or enzymatic hydrolysis with cellulolytic and amylolytic enzyme preparations at a concentration of 50-170 and 2-5 units. activity per 1 g of dry matter, respectively; processing is carried out for 3.0-4.0 hours at 50-60°C and pH 5.0-6.0 with stirring; at the end of the treatment, the pH of the suspension is adjusted to 4.3-5.0, xylanase and glucoamylase enzyme preparations are added at the rate of 50-70 and 2-4 units. activity per 1 g of dry matter, processing is carried out for 3-4 hours at 55-60°C with stirring; the resulting nutrient medium is treated at a temperature of 90-120 ° C for 10-20 minutes, cooled and inoculated with monocultures of the genera Geotrichum, Saccharomyces, Bacillus or their consortium, compiled in any ratio in the amount of 5-10% by weight of the nutrient medium, after fermentation, the suspension heated to a temperature of 90-100°C for 10-20 minutes and dried to a moisture content of 5-10%.

Example 1

Water or other reagents are added to pea flour at a ratio of 1:10, soluble protein compounds are extracted from the flour by any method, and to the released whey water with a chemical composition that includes 3-21% dry matter (DM), of which nitrogenous substances 8.62 -22.48%, reducing sugars (glucose, fructose, galactose, arabinose, xylose, maltose) 48.11-67.48% of the total amount of carbohydrates, an insoluble residue is added in the amount of 2% of their mass, acid hydrolysis of the suspension is carried out at pH 3.0 with incubation for 10 minutes at a temperature of 60-129°C or enzymatic hydrolysis with cellulolytic and amylolytic enzyme preparations at a concentration of 50 and 2 units. activity per 1 g of dry matter, respectively; processing is carried out for 3.0 hours at 50°C and pH 5.0 with stirring; at the end of the treatment, the pH of the suspension is adjusted to 4.3, xylanase and glucoamylase enzyme preparations are added at the rate of 50 and 2.5 units. activity per 1 g of dry matter, the treatment is carried out for 3 hours at 55°C with stirring; the resulting nutrient medium is treated at a temperature of 90-120°C for 10-20 minutes, cooled and inoculated with a Geotrichum fermentans monoculture in an amount of 5% by weight of the nutrient medium, after fermentation, the suspension is heated to a temperature of 90°C for 10 minutes and dried.

The dried feed concentrate is a friable powder of light cream color with a pleasant smell of yeast. The concentrate contains on the SV: protein 47%, carbohydrates 35%, of which soluble fibers 17%, lipids 10%, ash 8%.

Example 2

Water or other reagents are added to chickpea flour at a ratio of 1:12, soluble protein compounds are extracted from the flour by any method, and to the released whey water with a chemical composition that includes 3-21% dry matter (DM), of which nitrogenous substances are 8.62 -22.48%, reducing sugars (glucose, fructose, galactose, arabinose, xylose, maltose) 48.11-67.48% of the total amount of carbohydrates, make an insoluble residue in the amount of 4.5% of their mass, carry out acid hydrolysis suspensions at pH 2.5 with aging for 15 minutes at a temperature of 60-129°C or enzymatic hydrolysis with cellulolytic and amylolytic enzyme preparations at a concentration of 90 and 3 units. activity per 1 g of dry matter, respectively; processing is carried out for 3.5 hours at 55°C and pH 5.5 with stirring; at the end of the treatment, the pH of the suspension is adjusted to 4.5, xylanase and glucoamylase enzyme preparations are added at the rate of 60 and 3 units. activity per 1 g of dry matter, treatment is carried out for 3.5 hours at 60°C with stirring, the resulting nutrient medium is treated at a temperature of 90-120°C for 10-20 minutes, cooled and inoculated with Geotrichum candidum monoculture in the amount of 6, 5% by weight of the nutrient medium, after fermentation, the suspension is heated to a temperature of 90°C for 10 min and dried.

The dried feed concentrate is a friable powder of light cream color with a pleasant smell of yeast. The concentrate contains on SV: protein 53%; carbohydrates 29%, of which soluble fiber 14%; lipids 8%; ash 10%.

Example 3

Water or other reagents are added to pea flour at a ratio of 1:13, soluble protein compounds are extracted from the flour by any method, and to the released whey water with a chemical composition that includes 3-21% dry matter (DM), of which nitrogenous substances 8.62 -22.48%, reducing sugars (glucose, fructose, galactose, arabinose, xylose, maltose) 48.11-67.48% of the total amount of carbohydrates, an insoluble residue is added in the amount of 7.0% of their mass, acid hydrolysis is carried out suspensions at pH 2.0 with aging for 20 minutes at a temperature of 60-129°C or enzymatic hydrolysis with cellulolytic and amylolytic enzyme preparations at a concentration of 130 and 4 units. activity per 1 g of dry matter, respectively; processing is carried out for 4.0 hours at 55°C and pH 6.0 with stirring; at the end of the treatment, the pH of the suspension is adjusted to 4.7, the xylanase and glucoamylase enzyme preparation is added at the rate of 65 and 3.5 units. activity per 1 g of dry matter, the treatment is carried out for 3.5 hours at 60°C with stirring; the resulting nutrient medium is treated at a temperature of 90-120°C for 10-20 minutes, cooled and inoculated with cultures of Saccharomyces uvarum and Bacillus licheniformi in a ratio of 3:1 in an amount of 8% by weight of the nutrient medium, after fermentation, the suspension is heated to a temperature of 90°C for 10 min and dry.

The dried feed concentrate is a friable powder of light cream color with a pleasant smell of yeast. The concentrate contains on SV: protein 59%; carbohydrates 24%, of which soluble fiber 7%; lipids 5%; ash 12%.

Example 4

Water or other reagents are added to chickpea flour at a ratio of 1:15, soluble protein compounds are extracted from the flour by any method, and to the released whey water with a chemical composition that includes 3-21% dry matter (DM), of which nitrogenous substances are 8.62 -22.48%, reducing sugars (glucose, fructose, galactose, arabinose, xylose, maltose) 48.11-67.48% of the total amount of carbohydrates, an insoluble residue is added in the amount of 10% of their mass, acid hydrolysis of the suspension is carried out at pH 1.8 with incubation for 30 minutes at a temperature of 60-129°C or enzymatic hydrolysis with cellulolytic and amylolytic enzyme preparations at a concentration of 170 and 5 units. activity per 1 g of dry matter, respectively; processing is carried out for 4.0 hours at 60°C and pH 5.5 with stirring; at the end of the treatment, the pH of the suspension is adjusted to 5.0, xylanase and glucoamylase enzyme preparations are added, at the rate of 70 and 4 units. activity per 1 g of dry matter, the treatment is carried out for 4 hours at 60°C with stirring; the resulting nutrient medium is treated at a temperature of 90-120°C for 10-20 minutes, cooled and inoculated with cultures of Saccharomyces cerevisiae and Bacillus subtilis in a ratio of 1:1 in an amount of 10% by weight of the nutrient medium, after fermentation, the suspension is heated to a temperature of 90°C for 10 min and dry.

The dried feed concentrate is a friable powder of light cream color with a pleasant smell of yeast. The concentrate contains on the SV: protein 65%, carbohydrates 18%, of which soluble fibers 10%, lipids 2%, ash 15%.

The proteins of the concentrates contained 18 amino acids: glutamic, aspartic acids, glycine, proline, lysine, etc. The score for all essential amino acids of both concentrates from the secondary products of processing of leguminous crops was above 100% (Fig. 3), which indicated a high biological value of the concentrates. In the triticale grain biomass concentrate, the score for threonine, lysine, leucine, isoleucine, valine and aromatic amino acids was also above 100%, while for the essential sulfur-containing amino acids (cystine + methionine), histidine and tryptophan of the prototype, the score was less than 100% (51- 90%), which indicates the biological inferiority of the proteins of the last concentrate.

The fatty acid composition of chickpea concentrate was represented by 10 components, pea - 30 compounds, among which the share of fatty acids in the first concentrate accounted for 92.5%, in the second concentrate - 97.0%. In chickpea concentrate, the ratio of the sum of saturated (25.40%) and unsaturated fatty acids (67.09%) was 1:2.6, the content of omega-6 fatty acids (linoleic acid) was 41.26%, in pea concentrate, respectively , the ratio of 1:3 (23.5/71.67) and omega-6 fatty acids - 19.73%, the ratio of saturated: unsaturated fatty acids in cereals (prototype) 27/73:43, equal to 1:2.7 . The content of cis-isomers in concentrates from leguminous crops is 91-93%, trans-isomers - 5-7%, the ratio of saturated:unsaturated fatty acids - 1:2.6-1:3.0. Thus, in terms of the composition and type of fatty acids, both concentrates, as well as concentrates from by-products of cereal processing, approach edible oils and fats, in particular, soybean oil.

The mineral composition of concentrates includes macro- and microelements (Fig. 4). Concentrates from legumes contain significantly more sodium, potassium, magnesium, cobalt, zinc, copper, manganese, cobalt, nickel, and less iron and chromium. A significantly higher content of mineral elements in concentrates from leguminous crops (peas and chickpeas) is indicated by a high ash content of new protein concentrates.

Claims (1)

A method for producing a feed concentrate, involving the preparation of a nutrient medium, inoculation of microorganisms, and fermentation, characterized in that in the process of preparing the nutrient medium, whey water of legumes - peas or chickpeas, formed in the process of obtaining protein, is used, an insoluble residue in the amount of 2 is introduced into the whey water -10% of their mass, the suspension is subjected to acid hydrolysis with keeping it at pH 1.8-3.0, temperature 60-129°C for 10-30 min or enzymatic hydrolysis with cellulolytic and amylolytic enzyme preparations at a concentration of 50-170 units activity per 1 g of dry matter and 2-5 units. activity per 1 g of dry matter, respectively; the suspension is subjected to enzymatic hydrolysis for 3.0-4.0 hours at 50-60°C and pH 5.0-6.0 with stirring; after which the pH of the suspension is adjusted to 4.3-5.0, xylanase and glucoamylase enzyme preparations are added at the rate of 50-70 units. activity per 1 g of dry matter and 2-4 units. activity per 1 g of dry matter, respectively, the suspension is subjected to enzymatic hydrolysis for 3-4 hours at 55-60 ° C with stirring, pH is adjusted, inoculated with monocultures of the genera Geotrichum, Saccharomyces, Bacillus or their consortium, compiled in any ratio in the amount of 5 -10% to the mass of the nutrient medium.

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