RU2580762C1 - Method of evaluating quality of fodder in in vivo test system - Google Patents

Abstract

FIELD: agriculture.

SUBSTANCE: invention relates to determining quality of feed. Test samples are introduced into incubation medium for cultivation of a strain of Medusomyces Gisevii Lindau tea fungus, microorganism is incubated for 12-14 days, followed by weighing biomass of fungus and based on fungus biomass multiplicity of values in experiment and control (g), determining efficiency factor (CE). For efficiency factor equal to 1.1-1.6, poor quality of feed is determined; 1.7-2.5 - average quality; 2.6-3.0 - good quality and 3.1-4.0 - highest quality of feed.

EFFECT: technical result is reduction of time of sample preparation and analysis of most adequate in vivo test system to obtain full information on integral quality index - biological adequacy of feed.

1 cl, 8 ex

Description

The invention relates to the field of agriculture, in particular, to determining the quality and usefulness of feed at feed mills, bread processing plants, in nurseries, in laboratories of veterinary and agrochemical examination, etc. at any stage of preparation of feed mixtures and feeding agricultural animals.

Various recipes for dry and wet food are being developed and used. At the same time, complete feeds include a wide variety of ingredients: grain of cereals and legumes, poultry, cattle, rabbit, ocean and river fish, meat and bone, fish flour, milk powder, dried vegetables, vitamin and mineral supplements, antibiotics, etc. . Feed and feed additives are most often prepared in the form of granules having a different shape, size, color and smell. All feed and feed additives are usually of high cost.

Known methods for determining the quality of feed, in particular compound feeds by standard methods involving wet ashing of the sample with acid in determining nitrogen; dry ashing - when determining gray ash, calcium, phosphorus; extraction of the sample with ether - when determining crude fat, etc. These methods are laborious to implement. At the same time, it takes 2-3 days to analyze each sample according to the full zootechnical scheme. In addition, the use of chemical reagents, chemical glassware, expensive instruments and equipment is required. And these methods are not safe for performers (Markov D.I., Shumilin I.S., Gorshkov G.I. et al. Guidelines for feed analysis. - M .: Kolos, 1982).

It is also known to determine the quality of feed by diffuse reflection of samples in the near infrared region of the spectrum (750-2500 nm) using IR analyzers (Method for determining the content of crude protein, crude fiber, crude fat and moisture using spectroscopy in the near infrared region. GOST R 50817 -95. Method for determining the content of ash, calcium and phosphorus using spectrometry in the near infrared region. GOST 50852-96. Krishchenko VP. Near infrared spectrometry. - M. 1974. - S. 333, 336-339. RF patent № 2138806. IPC G01N 33/43, A61K 1/00, о UBL. 27.09.1999).

The disadvantage of this method is that the development of calibration equations by which the analysis is carried out requires a large amount of work on the selection of feed samples, preparing them for analysis and the analysis itself using standard chemical methods. The implementation of these works takes from 4 to 12 months (Mullard I. Analysis of feed in the near infrared region. - Agricultural use of spectroscopy in the near infrared region (Sat. Scientific. Proceedings on ICS - M., 1986. - P. 37-42) .

It is known that the biological usefulness of feed is more assessed by the level of protein in it. In addition, when compiling diets for animals, the balance of feed for sugar and protein is important - a ratio that is expressed by the amount of easily fermentable sugars per 1 g of protein. Protein diets are used better in feeds where the sugar-protein ratio is - 1.0: 1.5 (Feeding farm animals: A guide edited by A.P. Kalashnikov and N.I. Kleimenov. - M.: Rosagropromizdat, 1988. - 366 p.).

Therefore, in the development of this method for determining the quality of feed, a method of laboratory modeling of key processes of protein conversion is used both in the cicatricial and post-cicatricial tracts of the digestive tract in animals.

One of the approaches to the study of cicatricial digestion, namely the cultivation of cicatricial microorganisms under laboratory conditions, makes it possible to study the metabolism of feed protein in this area outside the background of digestion along the entire length of the digestive chain.

When developing in-vitro methods, they strive to one degree or another to simulate the living conditions of cicatricial microflora in the animal’s stomachs, where pH is stabilized due to the influx of saliva, which has its own buffer properties (the first condition); mineral composition of the medium (second condition); the wall of the scar provides rhythmic mixing (the third condition); removal of metabolic products (fourth condition); anaerobicity of the medium (fifth condition) and a constant temperature of 39 ° C (sixth condition).

In view of the foregoing, a method has been developed for assessing the biological usefulness of feed in vitro using the “artificial scar” apparatus, which ensures the preservation of the amount of scar microflora introduced into the apparatus and accounting for the ammonia formed as a result of protein breakdown (A. Tyutyunnikov, V. M. Fadeev Improving the quality of feed protein. - M .: Rosselkhozizdat, 1964. - S. 17-19).

The disadvantage of this method is the limited information received on the quality and biological usefulness of feed, since it depends not only on the amount of protein formed, but also on other important components of the feed: enzymes, vitamins, carbohydrates, fatty acids, minerals, etc. In addition, the in-vitro laboratory model is very far from the optimal in-vivo model, since this method does not provide information on the true biological usefulness of the test food - there is no information on the products received, namely on the increase in live weight and, in addition, there is no quantitative assessment of the synthesis of biologically active substances.

The technical result to which the invention is directed is to develop an optimal in-vivo test system for evaluating the biological usefulness of feed and feed additives, simplifying the method of analyzing feed and reducing the time of research.

To achieve the above technical result in the method for evaluating the quality of feed in an in-vivo test system, the samples of the studied feed are pre-crushed, they are placed in gauze bags of 4-5 g, the bags are tied with threads. Evaluation of the quality of the feed is carried out by incubating the microorganism of Kombucha strain Medusomyces Gisevii Lindau in a solution of tea with sugar with the addition of a sample of the studied feed. After 12-14 days, the obtained biomass in the experimental and control banks is weighed and the quality of the feed or feed additive, expressed in the integral indicator - efficiency coefficient (FE), is determined by the ratio of the fungus biomass in the experiment (g) to the weight of the control sample (g). The efficiency coefficient equal to 1.1-1.6, judged by the low quality of feed; 1.7-2.5 - average quality; 2.6-3.0 - good quality and 3.1-4.0 - high quality feed, which provides in-vivo obtaining the most important information, which is the increase in the biomass of the fungus when growing it with samples of tea fungus in the test feed or dietary supplements. The above method, in comparison with the known ones, simplifies the selection, preparation and analysis of feed, bioadditives and their components, reduces the time of research by 15-30 times.

The method is as follows. At the first stage, samples of each of the studied dry feeds (cereals, legumes, dry food additives or compound feeds, hay, straw, etc.) are prepared, they are crushed in a cyclone mill with a hole diameter of 1 mm, mixed thoroughly, placed in jars with ground stoppers and stored in the refrigerator.

The tests are carried out with samples weighing 1, 2, 3, 4, 5, 7, 7, 8, 9 and 10 g, which are placed in gauze bags, tied with threads, leaving a free end of the threads 10-15 cm long (depending on growing conditions and the height of the setter jars).

In vivo simulation of a test system for assessing the biological usefulness of feed is carried out by incubating a kombuch microorganism of the strain Medusomyces Gisevii Lindau, used to produce kombucha beverage (kvass). For the cultivation of Kombucha use 3-liter cans with a wide neck, which are covered with several layers of gauze. For the growth and development of the fungus, a weak tea solution (1.5-2.0 L) with sugar dissolved in it is used. It is prepared on the basis of: 100-120 g of sugar per 1 liter of boiled water at room temperature.

In parallel experimental cans, the fungus is grown in the presence of test samples of dry and liquid feed. To do this, preliminarily, samples of feed, feed additives or their components (prepared as described above) weighing from 1 to 10 g (liquid samples from 1 to 10 ml) are placed on the bottom of the jars, they are filled with incubation liquid (tea and sugar solution) and incubated in identical conditions, like control banks.

Control and experimental banks are kept at 23-25 ° C in a place located away from the window (to exclude direct sunlight) for 12-14 days. After the specified expiration, the resulting biomass of the fungus is weighed, after dehydration is carried out, for this the fungus is removed from the can, placed for 3-5 minutes on the bottom of the sieve (to completely drain the residual liquid), then it is dried with filter paper, placed on a laboratory balance and weigh the resulting biomass of the fungus from the control and experimental cans.

The determination of biomass is carried out by direct weighing, obtaining the weight of the raw biomass. The biomass parameter is a fundamental indicator, reflects the degree of fungal growth intensity, expressed through important integral characteristics - economic and metabolic coefficients. Weighing the fungus gives information about the mass, volume, linear size, mass of the substrate consumed, the mass of the product formed, the metabolic rate and the ratio of fungus biomass in the experiment (g) to the biomass of the control sample (g) determine the efficiency coefficient (CE), which shows how many times the biomass of the fungus in the jar with the test food exceeds the biomass of the fungus of the control sample.

The results of numerous experiments showed that the efficiency coefficient of 1.1-1.6 corresponds to the low quality of the studied feed; an efficiency factor of 1.7-2.5 corresponds to average quality; CE equal to 2.6-3.0 - good quality and CE = 3.1-4.0 - high quality or usefulness of feed, diet, feed additives or their components, as well as feed mixtures formulated in industrial or experimental conditions.

In livestock practice, juicy, coarse, concentrated feeds, animal feed and microbial feed, mineral feed and animal feed are distinguished. Moreover, each type of feed includes a significant number of subspecies. For example, green fodder, which is part of succulent fodder, has more than 20 subspecies (alfalfa, clover, sainfoin, sweet clover, meadow oatmeal, beetroot boneless, hedgehog team, vetch, pea, vetch-oat mix, pea-oat mix, corn, sorghum , winter rye, etc.) A similar set of subspecies include coarse, concentrated, animal and microbial feed, mineral feed and animal feed. This makes it necessary to take into account the specific gravity of various food groups included in their composition when compiling feed rations. In this regard, the use in animal husbandry practice of an accelerated and objective assessment of the quality and usefulness of the proposed diet, depending on changes in the composition of its components, is an important task in improving the feeding system of agricultural workers. animals by developing complete diets.

For a comparative assessment of the effectiveness and usefulness of the proposed rations, feeds, feed mixtures and combined feeds, as a reference in the proposed test system, we use full-feed combined feeds for growing broilers of the 1st period - PCCT (testing of bird rations) and complete combined feeds for cattle PKKkrs (testing of feed for cattle).

The implementation of the method is illustrated by the following examples.

Example 1. The study of the influence of the reference diet (feed) - PKKkrs on the accumulation of biomass of the fungus Medusomyces Gisevii Lindau

The experiments were carried out in parallel in two 3-liter jars, the first of which served as experience, the second as control. At the bottom of the experimental can, a sample (4-5 g) of PKKkrs reference food was placed in a gauze bag tied with a thread, the free end of which was carried out and fixed on the neck of the can. The lower layers were separated from the mother fungus, placed in both cans, a solution of tea with sugar of 1.5-2.0 liters was poured. Both banks were placed in a warm place at 23-25 ° C and incubated for 12-14 days (time of maximum accumulation of fungal biomass). After the specified exposure, the obtained biomass was removed from the experimental and control cans, placed on the bottom of the sieve, kept for 3-5 minutes until the liquid drained completely, dried with filter paper, placed on a cup of laboratory balance and the fungus biomass was weighed from the control and experimental cans. The results of weighing the biomass of the fungus from the experimental and control cans and the determination of the efficiency coefficient showed that the multiplicity of the increase in the biomass of the fungus when growing it in the presence of the test food (PKKkrs), in comparison with the control (FE), was 3.58 times, with the values of the biomass of the fungus experience - 405.0 g and biomass of the control sample - 113.0 g

Example 2. The dependence of the growth of biomass of the fungus on the quantitative content of the sample of test food

While maintaining the conditions of example 1, biomass of Kombucha was obtained with placing in the culture medium a different number of test samples (from 1 to 10 g). The weighing results showed that the maximum increase in the fungal biomass in the experiment occurred when the test sample was added to the culture medium in an amount of 4-5 g. A further increase in the amount of the introduced sample did not lead to biomass growth, the excess amount of the introduced substrate was not assimilated by the microorganism and remained at the bottom of the can.

Example 3. The study of the dependence of the growth of biomass of the fungus on the time of cultivation

While maintaining the conditions of example 1, biomass of Kombucha was obtained by growing a microorganism for 3-14 days. The biomass weighing results showed that the maximum growth of the fungus in the experiment was recorded on the 12th day, the biomass was 401.9 g with a control value of 107.1 g; the multiplicity (FE) was 3.75 times. A further increase in the time of cultivation of the fungus in the experimental bank did not lead to a significant increase in the biomass of the fungus. On day 14, the biomass in the experimental bank was 404.9 g, while the control value was 113.1 g (CE = 3.58).

Example 4. The study of the effect on the growth of biomass of the fungus of one of the main components of the feed - soy

One of the most important components of a full-fledged combined feed for cattle breeding is soy, which is of exceptional importance for solving the protein problem in animal husbandry. In this regard, a study was conducted of the effect of this component on the growth of the biomass of the fungus when growing it in the presence of this culture. While maintaining the conditions of example 1, biomass of Kombucha was obtained by adding 4-5 g of soybean to the culture medium. The process of incubation of the microorganism lasted for 14 days. The result of the determination of biomass in the experimental and control banks showed that such in the experimental bank was 321.3 g, against 112.8 g in the control, CE = 2.85 times.

Example 5. The study of the effect on the growth of biomass of the fungus of one of the main components of the feed - powder

While maintaining the conditions of example 1, kombucha biomass was obtained when 4–5 g of powder of rhombus was introduced into the culture medium. It was found that on day 14 the fungus biomass in the experiment was 298.7 g versus 112.5 g in the control; CE was equal to - 2.65 times.

Example 6. The effect of soy-reverse mixture on the growth of biomass of the fungus

While maintaining the conditions of example 1, biomass of Kombucha was obtained by adding a soy-inverse mixture in the ratio of 1: 1 to the culture medium. It was found that this mixture had a stimulating effect on the growth of the fungus biomass. The weight of the fungus in the experiment was 341.3 g against 113.1 g in the control. Calculations showed that the EC of this mixture was 3.01 times.

Example 7. The effect of biologically active feed additives "Vita-Forte M" on the growth of biomass of the fungus

Considering that the usefulness of the PKKkrs reference feed is achieved by including feed additives (premixes, minerals, vitamins, etc.) in its composition, we studied the effect on the growth and development of the fungus of the biologically active feed additive “Vita-Forte M” containing products beekeeping (propolis, bee bread, salmon, bee killing, drone brood), flour (blood, coniferous and grassy) and bentonite, which has immunostimulating, anti-infectious, adaptogenic and radioprotective properties, increases the digestibility of feeds, and stimulates ro m and development of young animals, increases resistance under conditions exposing it to stressors - physical (irradiation), chemical (ecotoxicants) and biological (pathogenic and conditionally pathogenic microflora) and the chemical composition and the biological effects superior to known feed additives. When setting up the experiment, we took into account the fact that the high biological activity of the drug and the multicomponent chemical composition (the presence of up to 27 trace elements, vitamins A, B, C, D, E, PP, enzymes, amino acids, fatty acids, chitin, chitosan, etc.) can provide an effect even with a minimum concentration of biologically active substances. While maintaining the conditions of Example 1, kombucha biomass was obtained when samples of the Vita-Forte M preparation weighing from 1 to 10 g were added to the culture medium. It was established that the maximum effect was noted when 3-5 g of the drug was added to the culture fluid. The biomass increase for 14 days of cultivation was 438.1 g versus 115.3 g in the control (CE = 3.79). A further increase in the amount of the test drug did not lead to a significant increase in biomass, the excess feed additive was not absorbed by the microorganism, remained at the bottom of the can and did not enter the metabolic process.

Example 8. Evaluation of the effect of the mixture PKKkrs-"Vita-Forte M" on the growth of biomass of the fungus

While maintaining the conditions of Example 1, various quantitative combinations of the PKKkrs reference feed and the Vita-Forze M dietary supplement were introduced into the culture fluid. In this case, the following quantitative ratios of components were compiled: 9.5: 0.5; 9.0: 1.0; 8.5: 1.5; 8.0: 2.0; 7.5: 2.5; 7.0: 3.0; 6.5: 3.5; 6.0: 4.0 (g). It was found that the introduction of a mixture of feed and feed additive in a ratio of 8.5: 1.5 into the culture medium provided the most effective weight gain of the fungus - 465.3 g versus 113.5 g in the control, which corresponded to CE = 4.1 times.

Thus, the use of an in-vivo test system provided an objective accelerated rapid assessment of the quality and usefulness of feed, feed mixtures and dietary supplements.

The invention, in comparison with known methods, has the following advantages:

- there is no need to use chemical reagents, chemical glassware, expensive instruments and equipment;

- the method is safe for performers;

- no complex mathematical calculations and development of calibration equations are required;

- the studied indicator (biomass measurement) is an integral, most important production indicator that allows determining the economic and metabolic coefficients, obtaining information about the mass, volume, linear dimensions, weight of some biomass components, the mass of the substrate consumed and the resulting product (tea beverage), its taste metabolic rate and daily average weight gain;

- low cost, environmental friendliness, ease of technical performance and high reliability of the results.

- allows to reduce the time of research by 15-30 times, to simplify the selection, preparation and determination of feed quality.

The method has been tested in multiple laboratory tests using a significant assortment of feed, rations, food and feed additives, feed mixtures and their individual components (more than 150 items) with their full assessment.

Claims (1)

A method for assessing the quality of feed in an in-vivo test system is that for this purpose, samples of the studied feed are preliminarily taken, crushed, placed in gauze bags of 3-5 g and tied with threads; quality assessment is carried out by incubation for 12-14 days of the Kombucha microorganism strain Medusomyces Gisevii Lindau in the presence of a test sample of feed; then the biomass of the fungus is weighed and the efficiency coefficient (CE) is determined by the multiplicity of the biomass of the fungus in the experiment and control (g); on the coefficient of efficiency equal to 1.1-1.6, judge the low quality of feed; 1.7-2.5 - average quality; 2.6-3.0 - good quality and 3.1-4.0 - the highest quality feed.