RU2391859C2 - Method of protein-vitamin fodder production - Google Patents

Abstract

FIELD: agriculture.

SUBSTANCE: invention is intended for use in fodder production. Method for production of protein-vitamin fodder consists in incubation of lactic acid and propionic bacteria used in pairs on nutrient medium, which contains cobalt salt and source of carbon in the form of processed raw materials to produce target product. Raw materials are represented by ground grain, flour-grinding and/or starch wastes. In order to prepare nutrient medium, raw materials are acidified and moistened with liquid fraction of cultural fluid, then exposed to hot extrusion to make extrudate. Produced extrudate is acidified and liquefied with liquid fraction of cultural liquid in order to make batch. Produced batch is soaked for 10-15 minutes to produce hydrolysate. The following components are added to produced hydrolysate - salt of cobalt with production of nutrient medium, on which Lactobacillus plantarum 578/26 and Propionibacterium freudenreichii subsp. shermanii 103/27 culture strains are incubated. Target product is cultural fluid.

EFFECT: invention makes it possible to produce fodder rich in biologically active components and protein, containing protective substances and live cells of microorganisms, makes it possible to enrich microflora of gastrointestinal track of animals, to simplify technological process, to reduce power inputs and to preserve raw fodder.

3 cl, 1 tbl, 5 ex

Description

The invention relates to biotechnology, to agriculture, to feed production.

By the beginning of the 21st century, the world protein deficit is estimated at 30-35 million tons per year. The main way to reduce this deficit is to produce biomass using microbiological synthesis.

Grain raw materials - rye, barley, wheat, flour milling waste, including substandard waste, are used for livestock feed or for the preparation of animal feed. However, grain raw materials are not a complete feed due to their low protein content, poor digestibility, and an unfavorable ratio of proteins and carbohydrates. More promising is the production of protein feed by growing microorganisms on grain raw materials.

A known method of producing protein-vitamin feed by incubating microorganisms on a nutrient medium containing mineral salts and a carbon source in the form of a waste product from the processing of natural raw materials to obtain the target product in the form of biomass of microorganisms (Patent RU No. 2183666, class C12F 3/10, C12P 7/06, A23K 1/06, publ. 06/20/2002) [1].

As waste products for the processing of natural raw materials in this known method, grain is used, which is crushed, subjected to enzymatic hydrolysis to obtain an enzyme lysate, from which a nutrient medium is prepared for growing Saccaromyces cerevisiae yeast on it to obtain alcohol. The resulting biomass is used as feed.

However, the yeast is grown under aerobic conditions, which requires significant energy consumption for supplying air to the environment for their incubation. To use the resulting biomass in the feed, it is necessary to heat it in order to stop the yeast from functioning. Such processing requires significant energy consumption and is accompanied by the destruction of part of the biologically active components of the feed. This food does not contain living cells that can settle in the gastrointestinal tract of the animal and improve its performance.

A known method of producing protein-vitamin feed by incubating microorganisms in a nutrient medium containing mineral salts and a carbon source in the form of a waste from the production of processing natural raw materials (Patent RU No. 2055881, CL A23K 1/165, publ. 02.27.1996) [ 2].

As waste from the processing of natural raw materials in this well-known case, ground grain and waste from flour milling are used, which are subjected to heat treatment, and then the yeast of the genus Candida and bacteria producing amylolytic enzymes that carry out enzymatic hydrolysis of ground raw materials and waste from flour milling are co-grown. .

The disadvantage of this known method is the use of yeast of the genus Candida for the production of protein feed, which is a conditionally pathogenic microorganism, which requires an additional thermolysis process and special equipment, as well as a complex system for the treatment of wastewater and air emissions, requires significant amounts of air used. Sanitary and epidemiological rules SanPiN 2.3.2.1078-01 yeast of the genus Candida are included in the list of substances that have a harmful effect on humans.

Significant disadvantages of this method should also include a large consumption of air and heat and energy resources, the need for high-performance, complex and energy-intensive equipment and a significant consumption of auxiliary materials.

Closest to the claimed method is a method of preparing protein-vitamin feed from grain, flour and starch waste, pre-treated by incubating lactic acid and propionic acid bacteria, which, when fed to cattle, enriches the intestinal microflora of animals (prototype).

In this method, grain and milling waste are finely ground, diluted with water in a ratio of 1: 3 and then boiled, cooled, and treated with a complex of enzymatic preparations of cellulolytic and amylolytic action. Starch waste is also subjected to the same treatment (Patent RU No. 2243678, class С1 А23К 1/06, C12N 1/16, С12Р 21/00, publ. 10.01.2005, bull. No. 1).

The disadvantages of this method are:

- multistage preparation of grain raw materials, including grinding, kneading, stepwise heating to a temperature of 95-100 ° C, boiling, cooling to a temperature of 45-50 ° C, saccharification using at least two enzyme preparations, cooling to a fermentation temperature. This procedure is associated with high consumption of heat and electricity;

- insufficient formation of simple carbohydrates for the subsequent fermentation process;

- the composition of the carbohydrate part of the feed is not regulated to obtain a favorable ratio of proteins to carbohydrates;

- strains of lactic acid and propionic acid bacteria produce an insufficient amount of protein, due to poor resistance to acid formation and a limited number of simple carbohydrates in the composition of the nutrient medium.

The technical result achieved by the present invention is to obtain food rich in biologically active components and protein, containing protective substances and living cells of microorganisms, enriching the microflora of the gastrointestinal tract of animals, simplifying the process, reducing energy consumption, improving the safety of raw food.

The indicated technical result is achieved in that the raw material processing method aims to prepare a nutrient medium for more complete use of the raw material components by microorganisms with the formation of increased amounts of biomass and protein, as well as to obtain feed enriched with protective substances, probiotics, lactic acid and propionic acid bacteria.

The use of microorganisms with probiotic properties, it is advisable in combination with prebiotic igridientami nutrient medium, which selectively stimulate the growth and catabolic activity of bacteria that live in the intestinal tract.

To achieve these goals, raw materials (grain, bran, waste from flour milling, starch waste) are mixed with a portion of the centrate, containing organic acids and other soluble substances, to obtain a grain mass with a moisture content of 20-25%, which is extruded. In this process, under the mechanical influence, the influence of temperature and a catalyst in the form of organic acids up to 2.5%, the matrix of grain raw materials is degraded, the mass is liquefied, the starch is gelatinized and the enzyme activity of the substrate is increased, as well as the partial hydrolysis of starch and proteins to produce products of incomplete hydrolysis - dextrins, oligisaccharides, peptones, poly and oligopeptides. This composition of the nutrient medium provides a more efficient subsequent fermentation using newly isolated cultures of lactic and propionic acid bacteria due to the fact that in the raw material prepared for fermentation there are already soluble components of the substrate, ready for consumption by microorganisms, and an additional saccharification step is not required.

In this case, the degree of saccharification of the substrate is 5-10% of RV in terms of glucose, but this degree of hydrolysis is due to the dextrins, tri-, tetra-, and oligosaccharides contained in the substrate. An additional breakdown of the high molecular weight components of starch occurs during the fermentation process, and low molecular weight components are consumed by microorganisms without being split into simple carbohydrates. This provides a higher content of soluble solids and reducing substances in the mash, a higher fermentation process with the help of lactic acid and propionic acid bacteria (reduction of the cycle by 6 hours) and a greater formation of biomass and proteins up to 53%.

In addition, the carbohydrate portion of the resulting feed is represented in large part by oligosaccharides and oligopeptides, which are prebiotics, which improves the quality of the feed.

The culture fluid obtained in this way, having all the advantages, can be used directly on livestock feed, since the acids, protective substances, and probiotics contained in it serve to protect against microbiological damage and other adverse effects, including mold.

To obtain feed in concentrated form, the culture fluid is divided into two phases - solid and liquid.

The solid phase is sent for drying to a moisture content of 10-12%.

The liquid phase (centrate or filtrate) is returned to production at the stage of preparation of the batch and to moisten the raw materials before extrusion. The centrate contains soluble reducing substances in the form of oligosaccharides, nitrogen compounds in the form of peptides, amino acids and other components, organic acids, enzyme complexes, live lactic and propionic acid bacteria, bacteriocins.

Due to its composition, the centrate can also be used as liquid feed (kefir) for feeding young animals and for adult animals instead of water.

When the centrate is returned to production, it plays the previously indicated positive role in the preparation of raw materials and its fermentation.

A method of producing protein-vitamin feed consists in incubating microorganisms on a nutrient medium prepared by the indicated method, containing mineral salts and a carbon source in the form of a prepared waste from the production of processing natural raw materials to obtain the target product in the form of biomass of incubated microorganisms, using salt as mineral salts cobalt, as production waste use grain, flour waste, starch waste, which are extruded, and inc iruyut in a nutrient culture medium of lactic acid and propionic bacteria used pair concurrently or sequentially: first incubated lactic acid bacteria, and then continue to the joint incubation of lactic acid and propionic bacteria.

It is advisable to incubate a pair of newly selected strains of lactic acid and propionic acid bacteria: Lactobacillus plantarum 578/26 and Propionibacterium freudenreichii subsp. shermanii 103/27.

A pair of lactic acid and propionic acid bacteria is recommended to be introduced into the nutrient medium in the ratio (0.8-1.2) :( 0.8-1.2) in an amount of 5-20 vol.%, And incubation should be carried out with periodic stirring at a temperature of 37- 50 ° C and pH 5.9-6.0.

During sequential incubation, it is advisable to carry out a joint incubation of lactic and propionic acid bacteria after 1.5-6 hours of preliminary incubation of lactic acid bacteria.

The target product with increased biological value enriched by living cells in the active phase of growth can be obtained by incubating a pair of microorganisms on a nutrient medium for 3-23.9 hours, and the target product with a high protein content can be obtained by incubating them for 24-36 hours .

The essence of the proposed method consists in the development of non-waste production technology, which provides for the production of biomass of microorganisms by incubating them on a nutrient medium containing mineral salts and a carbon source in the form of specially treated solid waste from natural raw materials processing plants to produce high-quality, easily digestible and digestible protein-rich feeds and enriched with microelements, vitamins, biologically useful substances, which has its own protective organisms using the microorganisms of the genera Lactobacillus and Propionibacterium for fermentation of a nutrient medium with the accumulation of biomass, metabolic products, enzyme systems, bacteriocins, as well as prebiotics enriching the feed product.

The value of the microorganisms used by the method according to the invention is that they consume mono- and disaccharides, hydrolysis intermediates, pentosans and other organic substances. In the process of cultivation, they synthesize, in addition to biomass, vitamins, amino acids, complexes of enzyme systems, bacteriocins that have protective and prophylactic properties. The result is a protein feed enriched with biologically active components, prebiotics, as well as protective substances that provide long-term storage of feed in raw form.

The wet food obtained by the method according to the invention can be stored for 3-6 months without microbiological damage due to the presence of protective substances in it (organic acids, their salts, enzyme systems and complexes).

The microorganisms used are not spore-forming, therefore, neutralization of the culture fluid and wastewater is not required. According to SanPiN 2.3.2.1078-01 they are included in the list of substances that do not have a harmful effect on food products.

These microorganisms are anaerobes, therefore, air flow is not required, which makes the process highly economical.

Their valuable property is the ability to develop on various complex environments, without requiring additional nutrients.

By the method according to the invention, it is recommended to incubate a pair of cultures of lactic acid and propionic acid bacteria for 3-23.9 hours or for 24-36 hours. A shorter incubation time (3-23.9) allows you to prepare food in the form of biomass of cells in the phase of exponential growth. When an animal enters the gastrointestinal tract, such living cells of lactic acid and propionic acid bacteria take root in it and are included in the regulation of metabolic processes, increase feed absorption and digestibility, and contribute to an increase in weight gain when feeding them to animals and birds, especially young animals. This food also contains protein, but in a smaller amount than food obtained by a longer incubation of microorganisms.

With the duration of the cultivation of microorganisms up to 24-36 hours, more vitamin B ₁₂ accumulates, which remains in the solid phase of the culture fluid, the volume of biomass increases, that is, the yield of feed enriched with protein.

The target product obtained as a result of the fermentation of microorganisms contains 35–53% protein on a dry matter basis, 30–44% amino acids, enriched with vitamins E and group B, including B ₁₂ , enzyme systems, bacteriocins, and complexes providing protective and preventive properties of the obtained feed .

If the process of growing biomass of lactic acid and propionic acid bacteria is carried out, for example, within 18 hours, then the target product contains 35% s.v. protein, during fermentation 24 hours - 39%, for 36 hours - 45% s.v. During 18 hours of cultivation (exponential phase), protective substances, catalase-peroxidase complexes, accumulate in the culture fluid, therefore, the process takes up to 23.9 hours to obtain a feed product with protective properties, although protein accumulation in this case is less (35%).

Lactic acid and propionic acid bacteria in this case are in an active state of reproduction, which contributes to their favorable introduction from feed into the gastrointestinal tract of animals, and also have the greatest viability and stability when dried.

When bacteria are cultured for 24-36 hours, an intensive accumulation of biomass occurs with an increase in the protein content in the cells. The resulting product is more enriched with vitamins and amino acids.

The method according to the invention can be used to obtain high protein feed directly at the enterprises for the processing of natural raw materials or at the enterprises for feed production and can be used both in dried and in liquid form.

The fermentation process can be carried out both continuously and remotely-refill method.

By the successive introduction of lactic acid bacteria and then propionic acid bacteria into the nutrient medium, the yield of the target product is increased due to the intensification of the growth of propionic acid bacteria on the medium, which contains the vital products of lactic acid bacteria.

The method is simple to execute. The nutrient medium is available, since it consists only of industrial wastes, to which only one cobalt salt is added, which reduces the cost and simplifies the method. The process does not require aeration, which reduces energy consumption. The method allows to utilize large volumes of industrial waste, turning them into a complete protein-vitamin feed.

The production of feed by the method according to the invention is waste-free.

The use of the invention has the following advantages - the exclusion of several stages of preparing raw materials for fermentation - heating, boiling, cooling, processing with enzymes, heating to 95 ° C, holding at this temperature, cooling, saccharification. This provides fuel and energy savings, reduces the duration of the process, as well as the loss of raw materials;

- the concentration of mash increases, which affects the decrease in product flows and allows you to reduce the amount obtained in the subsequent stages of the centrate (filtrate);

- the protective functions of the feed increase due to the presence of prebiotics in the carbohydrate part.

The process is characterized by less metal and material consumption.

The following are examples of the implementation of the method.

Example 1

Grain of medium degree of grinding is mixed with part of the centrate (5-15%) and enters the extruder, where it is heated to 180 ° C for 1 minute, then most of the filtrate is fed into the extruder chamber to obtain a flour: filtrate ratio of 1: 2.5 -3. The temperature of the mass becomes 55-60 ° C and enters the hydrolyzer (curing agent), in which it is held for 10-15 minutes until a degree of destruction of 5-15% of the PB is obtained. Dextrins and oligosaccharides predominate in the carbohydrate part of the nutrient medium, peptides, poly-, oligopeptides in the nitrogen part.

The nutrient medium prepared in this way is fed to anaerobic fermentation using lactic acid and propionic acid bacteria. After 16-24 hours of fermentation at a temperature of 37-40 ° C, the culture fluid contains 49.7% protein, 40% SV carbohydrates, vitamins, trace elements, protective substances in the form of probiotics and prebiotics, living bacteria.

Example 2

The culture fluid prepared according to claim 1 is separated by filtration or centrifugation into a solid phase, which is sent for drying to a moisture content of 10-12%.

The liquid phase is a centrate or filtrate contains 1.2-2% dry matter, including 1-1.2% protein, protective substances, bacteriocins, organic acids 1.5-2.5%. The centrate is divided into two parts in a ratio of 1:15 and is sent to moisturize the raw materials to a moisture content of 20-25% and the majority to kneading.

Example 3

Flour and starch waste enters the extruder together with part of the centrate (filtrate), is heated at a temperature of 180-190 ° C and a pressure of 0.4-0.5 MPa for 1 minute, mixed with most of the centrate (filtrate) and aged in hydrolysis at a temperature of 55-60 ° C to a degree of destruction of 10-15% for 3-5 minutes. Prepared culture medium is fed to fermentation.

Example 4

The nutrient medium prepared according to claim 1, 2, 3 is fed to fermentation using a pair of newly selected strains of the microorganisms Lactobacillus plantarum 578/26 and Propionibacterium freudenreichii subsp. shermanii 103/27 at a temperature of 37 ° C. After 24-36 hours, the culture fluid contains 50-53% protein, 1.5-2.5% organic acids and live bacteria in an amount of 2 · 10 ⁹ -2 · 10 ⁸ CFU / g.

Example 5

A clean culture of Lactobacillus plantarum 578/26 is incubated in the culture medium prepared according to claim 1, 2, 3, and after 1.5-6 hours, a pure culture of Propionibacterium shermanii 103/27 is added to it and the incubation is continued for 22.5-30 hours.

Strains of microorganisms are introduced into the nutrient medium sequentially, first Lactobacillus, and after 6 hours - Propionibacterium. The results indicate an increase in the biomass growth rate and an increase in yield (see table).

Table Change in biomass yield and cycle time during joint and sequential cultivation The name of the strain The biomass yield,% Relative cycle time Pr.freudenreichii subsp. shermanii
Lact. plantarum (applied simultaneously) one hundred 1,0 Lact. plantarum
Pr. freudenreichii subsp.shermanii (introduced after 6 hours) 160 0.58

With the simultaneous introduction of two cultures, the process is characterized by an increase in the amount of biomass formed by 1.6 times, which indicates a greater accumulation of bacterial cells and a reduction in cycle time by 16% compared with the introduction of one culture.

With the successive introduction of crops (first lactic acid, and then propionic acid), the amount of biomass formed increases by 1.8 times, and the cycle time is reduced by 42%.

Thus, the method according to the invention allows to obtain high-quality protein-vitamin feed containing live cells of lactic acid and propionic acid bacteria. The method is simple to implement, does not require increased energy consumption and provides complete utilization of a wide range of waste from industries for the processing of natural raw materials. Due to the high content of protective substances (organic acids, enzyme complexes, bacteriocions, probiotics and prebiotics), the food can be stored raw for a long time.

Claims (3)

1. The method of obtaining protein-vitamin feed, which consists in the incubation of pairs of lactic acid and propionic acid bacteria in a nutrient medium containing a cobalt salt and a carbon source in the form of processed raw materials, which are used as crushed grain, flour and / or starch waste to obtain the target product, characterized in that for the preparation of a nutrient medium, the raw material is acidified and moistened with 20-25% of the liquid fraction of the culture fluid containing soluble substances and organic acids from 2.5%, then subjected to hot extrusion for 1 min to a temperature of 180-190 ° C at a pressure of 0.4-0.5 MPa to obtain an extrudate, the resulting extrudate is acidified and diluted with the liquid fraction of the culture fluid to the extrudate: liquid ratio culture fluid fraction 1: (2.5-3.0) to obtain a batch; the batch obtained is kept for 10-15 minutes at a temperature of 55-60 ° C until 5-15% of reducing substances are destructed to obtain a hydrolyzate from the obtained hydrolyzate prepare the culture medium by adding salt to it Balta further incubated for cooked Lactobacillus plantarum 578/26 and Propionibacterium freudenreichii subsp medium cultures of strains. shermanii 103/27, and the target product is a culture fluid, or to obtain feed in concentrated form, the culture fluid is divided into two phases - solid and liquid, while the solid phase is sent to dry to a moisture content of 10-12%, and then used for food The liquid fraction can also be used as liquid feed. 2. The method according to claim 1, characterized in that for the flour and starch waste for the preparation of the hydrolyzate, the batch is kept for 3-5 minutes until a degree of destruction of 10-15% of reducing substances is obtained. 3. The method according to any one of claims 1 or 2, characterized in that the solid fraction obtained from the culture fluid is dried.