RU2541676C1 - Feed additive for fish - Google Patents

Abstract

FIELD: agriculture.

SUBSTANCE: additive comprises Halobacterium halobium cells and salts of ethylenediaminetetraacetic acid, at that Halobacterium halobium cells are contained in the additive as a sterile dry biomass obtained when their cultivating and subsequent dehydration of the culture liquid containing these cells. The ingredients are contained in the feed mixture in the following concentration, wt %: Halobacterium halobium cells in the form of a dry sterilised biomass - 10-20, salts of ethylenediaminetetraacetic acid 5-10, feed mixture - the rest. At that in case, if the feed does not contain vitamin A, the ingredients in the feed mixture are taken in the following ratio, wt %: Halobacterium halobium cells in the form of a dry sterile biomass - 18-20, salts of ethylenediaminetetraacetic acid - 5-10 , feed mixture - the rest.

EFFECT: use of the invention enables to influence positively on biological indicators of grown fish, and to protect the fish body from the exposure to heavy metals.

2 cl, 3 tbl

Description

The invention is intended for use in fish farming, in particular, relates to feed for fish when grown in natural and artificial reservoirs.

Currently, a large number of fish feeds are known that are used for their cultivation in natural (ponds, rivers) and artificial (aquariums) water bodies [www.aquafeed.ru/ru.ask.com/; Feed + For + Fishes www.centromall.ru, etc.]. The choice of a particular food is determined by the breed of fish, the characteristics of the reservoir, the set effect.

Typically, fish food consists of feed mixtures and feed additives. The composition of feed mixtures for fish grown in ponds includes meal, meal, mixed feed, waste from the food and alcohol industries, some crops (barley, lupine, alfalfa), etc. The quality of feed depends on the content of nutrients in them (proteins, fats, carbohydrates), as well as vitamins. [http://www.cnshb.ru/AKDiL/0015/base/RK/000174.shtm]. At the same time, the needs of fish in nutrients are individual for each species, depending on several factors (age of the fish, its body weight and mobility).

The introduction of feed additives into the feed mixture allows optimizing the nutritional composition of the feed, improving its bioavailability and preserving the properties of the feed during storage, providing a therapeutic and preventive effect, affecting the speed of fish development, etc. Feed additives include, in particular, trace elements, vitamins B and C, lignins, carboxymethyl cellulose [http://aquariumfishfood.ru/; SU 1567140, 1990; GB 1466003, 1977].

Of the greatest interest as feed additives are substances that affect the biological activity of the fish organism. So, it is known the introduction of chitosan into the feed mixture, which has non-specific biological activity, but is used as a binding component [RU 2221437, 2004], however, its effect on the fish growth rate and their protection from the negative impact of the external environment is very small.

The closest in technical essence to the claimed invention is the use as an additive in the feed mixture of baxin and succinic acid, which are contained in the feed mixture in the amount, mg / kg of feed mixture: baxin - 100-1000, succinic acid and / or its compounds - 50- 500 [RU 2312517, 2007].

The baksin used in the additive is a dried biomass grown in an aqueous-mineral nutrient medium and inactivated cells of one of the species of extremophilic halobacteria (Halobacterium halobium 353P, VKPM v-1739) living in highly saline waters of some lakes in South Africa, the Dead Sea, and Kara Bay -Bogaz and others. At present, buxin is recommended as a biologically active human food supplement [RU 2109515, 1998]. The drug is a lyophilized dried powder of biomass halobacteria. Suitable as a biologically active food supplement, as a means of reducing the toxic effect of antitumor compounds. Suitable for the prevention and treatment of radiation sickness.

The disadvantage of a feed additive is the insufficient stimulating effect on the development of fish and the protection of their body from the effects of heavy metals.

The problem solved by the authors was to expand the range of feed additives used for fish in order to achieve more accelerated development of fish and to protect their body from the effects of heavy metals.

The technical task was to create a composition based on Halobacterium halobium, capable of providing the above task.

The technical result was achieved by creating a preparation containing a dry sterilized culture fluid obtained by culturing a strain of Halobacteriuin halobium 353P, VKPM B-1739 (KZhbaksin), and salts of ethylenediaminetetraacetic acid (EDTA) in the following ratio of ingredients (wt.%) From feed:

KZhbaksin - 10-20%,

EDTA - 5-10%,

Feed mixture - the rest.

The supplement contains a mixture of nutrient medium components with Halobacteriuin halobium (Baksin) cells and EDTA.

Freeze-dried Halobacterium halobium sterilized culture fluid contains from 0.05 to 0.20 mg / g of the total amount of carotenoids, which work as free radical traps, increase the immune status, improve skin condition, and have a beneficial effect on vision. The betaines and ectoins contained in it are able to protect biological molecules and living cells from extreme influences (freezing / thawing, drying, heating). In addition, it, in particular, due to metabolites produced by bacteria during their growth and during dehydration, has increased antioxidant activity.

The results of evaluating the antioxidant activity of a sterilized Halobacterium halobium culture fluid by its effect on the rubbed tissue of mouse liver [by the method of inhibiting the intermediate oxidation product — diene conjugates (DC) and by inhibiting the end oxidation product — malondialdehyde (MDA) in the in vitro system — in the liver homogenate mice and in vivo system in mice with toxicity induced by carbon tetrachloride] are shown in table 1.

Table 1 Evaluation of the antioxidant activity of dry culture fluid Halobacterium halobium Object name Antioxidant activity, cu / mg Dry culture liquid Halobacterium halobium 10 ± 1 Dry Halobacteriumhalobium cells 7 ± 1 Alpha tocopherol 2 ± 0.3

The optimal concentration in a mixture of dry sterilized culture fluid Halobacterium halobium (buxin) is 15 wt.%.

A decrease in its concentration of less than 10 wt.% Significantly reduces the antioxidant effect of the drug. Increasing the concentration of more than 15 wt.% Practically does not affect the properties of the drug, but increases the possibility of the destruction of metabolites under the action of their own hydrobiont enzymes. When using food containing no vitamin A as a feed mixture, the amount of KZhbaksin should be increased to 18-20 wt.%.

One component of the proposed supplement is EDTA. Currently, EDTA is usually used in the form of disodium salt dihydrate (complex III, Trilon B, Na ₂ -EDTA) or calcium-sodium salt as an antioxidant and color stabilizer in the production of margarine and mayonnaise in an amount of up to 100 mg / kg, fish and canned seafood (crustaceans and mollusks) - up to 75 mg / kg [am-am.su/210-antioksidante385-cana2-edta.html], as well as in the form of a 5% oral solution to neutralize heavy metals during oral chelation therapy [ http://www.nmiff.ru/paradeigma/edta.htm]. In the body, EDTA is practically not absorbed. The permissible daily norm for its introduction into the human body in the form of dietary supplements is about 2.5 mg / kg body weight per day.

A feature of the inventive feed additive is the use of a sterilized dry culture fluid obtained as a result of culturing the Halobacterium halobium cells as a preparation containing cells by increasing the concentration of this ingredient in the feed from 0.1 wt.% To 20-40 wt.% With simultaneous administration of feed significantly higher amounts of EDTA in the amount of 5-10 wt.% instead of 0.1-5 wt.%.

As the experiments showed, the new complex supplement significantly increases metabolic processes in the fish’s body, increasing the size and rate of weight gain and cleansing the fish’s body of negative elements, in particular heavy metals, which are enzyme poisons, affect the metal enzymes responsible for cleansing the body from xenobiotics and for cell reproduction. Feeding fish with food, including a Baksin-containing component and EDTA, allows you to normalize the metabolism of fish, detoxify the body, increase resistance to adverse environmental conditions and realize the reserves of their growth and development within the genotype, intensify tissue regeneration

The action of the drug is based on the introduction of biocarotenoids and EDTA into the intestinal cavity, which allows maintaining the activity of the biological components of the drug, diffusion of antioxidants and a detoxifying effect in the gastrointestinal tract, with sorption of toxic compounds, which is especially important for general detoxification of the body.

The use of dry KZhbaksin and EDTA allows for the preparation of feed by extrusion with short-term heating to 95 ° C, provides uniform distribution in the feed mixture due to fractional dilution. This technology is convenient and technologically advanced in the production process, including directly in the field of application, as it does not require special equipment.

As feed mixtures, as a rule, standard fish feeds are used, for example, those mentioned in GOST 10385-76 (Compound feeds for pond fish, Moscow, 1979, p. 76), such as formula 16-80 feed mixtures containing the following components: protein-vitamin hydrolysis yeast concentrate, fish meal, wheat flour, soybean meal, sunflower meal, methionine, premix; as well as production feed for the cultivation of pond carp "Carp pond 26/3 and 26/5" containing fish meal, meat and bone meal, yeast, sunflower meal, soybean meal, wheat, barley, bran, premix PKP. Feed may additionally contain a supplement of vitamin A (retinol) in a dose of 15,000 units.

Feed additive is prepared as follows. Initially receive the drug KZhbaksin as follows. Microorganisms Halobacterium halobium grown by deep cultivation in fermenters having a volume of 250 liters Maintaining the working pH value (7.2-7.4) is carried out using the control and regulation circuit. Lighting the growing medium in the fermenter is carried out by fluorescent lamps located in the light-transmitting channels of the fermenter. Air for aeration of the growing bacterial suspension is fed to the fermenter through the ejectors of the circulation pumps. When the optical density is not less than 1.0, measured in a 3 mm cuvette against distilled water at 490 nm, a solution of nutrient salts (sodium chlorine, magnesium sulfate, potassium) is fed into the fermenter with a metering pump from a measuring device at a speed of 1-10 l / h chloride, sodium citrate), and from the measuring element of micronutrients a metering pump begins to supply a solution of trace elements (phosphoric acid, manganese chloride, iron (II) sulfate, zinc sulfate) at a rate of 50-300 ml / hour. Upon reaching a working volume of 150 l, the slurry starts to ferment from the fermenter through a drain fitting, which is sent to the separation. The amount of spent culture fluid in the total volume of the medium varies in the range of 25-50%, choosing it so as to prevent the optical density of the system from falling below 0.7. A suspension of commercial halobacteria biomass is collected in a collection tank and poured into pressure vessels, from which it enters the separation. Separation is carried out on centrifugal separators. The flow rate of the separated suspension varies in the range of 8-40 l / h, selecting it so that the yield of the condensed paste of halobacteria biomass does not decrease below 8 g / l of the suspension. For drying, use the lyophilization method or drying under vacuum. When using lyophilization, the condensed product paste is frozen in a freezer at a temperature of -50 ° C. The amount of paste loaded into the tray is 400-600 g for a dryer type TG-5 or 500-1000 g for a dryer type LZ-9. The drying cycle lasts 48-72 hours. During the drying process, the following parameters are supported:

- temperature of heated plates, not more than - 15-20 ° С,

- residual pressure, not more than - 0.5 mm Hg

When drying under vacuum, the product is loaded into stainless steel trays in an amount of 400-600 g and placed in a vacuum drying oven. In the drying process, lasting 24-48 hours, maintain a temperature of 25-50 ° C.

The resulting dry preparation, together with a predetermined amount of EDTA, is added to the feed immediately before use or loaded into the feed hopper in the prescribed proportion.

The addition of KZhbaksin to various fish feeds in an amount of 100 to 200 g per kg of feed mixture activates the immune system of the fish, accelerates its growth and development. The introduction of EDTA in an amount of from 50 to 100 g / kg of feed mixture together with KZhbaksin increases the effectiveness of the latter, and the combination of buxins - EDTA exerts a synergistic effect on the fish organism.

The invention is illustrated by the following examples.

Example 1. Tests of feed additives were carried out on carp yearlings weighing 38-45 g in laboratory conditions in flowing aquariums. Carrot yearlings of 20 pcs. kept in 70 l flow aquariums at a water temperature during the entire experiment in the range of 18-21 ° C, and with an oxygen content of 6.8 to 9 mg / l. The total duration of the experiment was 45 days. Of these, 14 days were spent on the process of adaptation of fish to aquarium conditions and their transfer to a normalized feeding regimen - 4% per day. The feed feeding course was 24 days.

In parallel, the following experiments were conducted with the feed mixture “Carp pond 26/5”:

- option 1 - feed mixture without additives;

- option 2 - feed mixture with the addition of vitamin A (15000 units);

- option 3 - feed mixture with the addition of 0.5% baxin and 0.2% succinic acid (the closest analogue);

- option 4 - feed mixture with the addition of 8% EDTA;

- option 5 - feed mixture with the addition of 15% KZhbaksina;

- option 6 - feed mixture with the addition of 15% KZhbaksina and 8% EDTA;

- option 7 - feed mixture with the addition of 15% buxin and 8% EDTA;

- option 8 - feed mixture with the addition of 10% KZhbaksina and 5% EDTA;

- option 9 - feed mixture with the addition of 20% KZhbaksina and 10% EDTA;

- option 10 - feed mixture with vitamin A and the addition of 20% KZhbaksin and 8% EDTA.

, который характеризует интенсивность роста рыб (%) в сравнении с начальной массой или в среднем за период по формуле:At the end of the feeding course and at all stages of the experiment, a comparative assessment of the physiological state of the fish was carried out according to hematological parameters. Fish breeding calculations of the absolute, relative, and average daily gain in average weight were carried out according to generally accepted methods. In this case, the relative weight gain of the fish was estimated $$\Delta M_{\mathrm{about}tn}^{\mathrm{about}}$$

, which characterizes the intensity of fish growth (%) in comparison with the initial mass or on average for the period according to the formula:

The results are shown in table 2. The analysis of the data showed that the values of the main biochemical parameters of mucus in fish in the experimental and control groups practically do not differ and correspond to the physiological norm. In this case, normalization of the hemoglobin level in the mucus of fish is noted, because the claimed additive normalizes metabolism, while the relative increase almost doubled, and compared with the closest analogue increased by 20%. Comparison of the results of using the claimed formulation and its ingredients showed a significant increase in performance with the use of additives and the synergism achieved. At the same time, as the experiments showed, an increase is noted in the use of KZhbaksin in its composition compared with buksin, apparently due to the enrichment of the mixture with metabolites and nutrients included in the QOL.

Example 2. In the conditions of example 1, we studied the effect of feed additives on fish growing in the presence of heavy metals. For this purpose, salts of heavy metals were added to water to a cadmium content of 10.0 μg / L; lead - 120 μg / l, zinc 120 μg / l, copper 120 μg / l

The feed mixture used was the formulation 16-80, as feed additives used

- option 1 - feed mixture without additives;

- option 3 - feed mixture with the addition of 0.5% baxin and 0.2% succinic acid (the closest analogue);

- option 6 - feed mixture with the addition of 15% KZhbaksina and 8% EDTA;

- option 9 - feed mixture with the addition of 20% KZhbaksina and 10% EDTA;

- option 10 - feed mixture with vitamin A and the addition of 20% KZhbaksin and 10% EDTA.

The results showed that the use of the inventive additives significantly reduces the content of heavy metals in the body of fish. In the case of using a mixture of buxin with succinic acid, the protective effect is negligible.

The above examples illustrate that fish feed, which is a standard feed mixture with the addition of KZhbaksin and EDTA, has a positive effect on the fish-breeding and biological indicators of fish farming. The effect of growth stimulation, detoxification, immunocorrection and survival of artificially infected fish is most pronounced.

table 2 The effect of the nature of the feed additive on the content of vital signs Indicator Supplement option norm one 2 3 four 5 6 7 8 9 10 Relative increase *,% 6.79 6.81 10.7 8.89 9.34 12.71 11.71 10.66 12.76 12.82 Slime Hemoglobin, er./l 0-10 105.4 ± 26.1 97.1 ± 12.5 13.0 ± 2.0 12.0 ± 1.8 12.0 ± 1.8 9.2 ± 0.1 9.1 ± 0.1 9.3 ± 0.1 9.1 ± 0.1 8.5 ± 0.1 Protein, g / l 0.3-1.0 0.98 ± 0.09 0.86 ± 0.05 0.79 ± 0.1 0.89 ± 0.1 0.89 ± 0.1 0.68 ± 0.4 0.72 ± 0.4 0.55 ± 0.3 0.72 ± 0.4 0.66 ± 0.4 Blood Blood glucose, mcg / l 70-80 103.0 ± 7.6 101.0 ± 5.8 108 ± 13.8 87.0 ± 10.1 91.0 ± 8.4 98.1 ± 3.4 105.1 ± 3.4 98.0 ± 6.4 105.1 ± 3.4 98.3 ± 3.4 Hemoglobin, g / l 70-90 89.1 ± 3.8 89.6 ± 7.5 66.4 ± 3.3 76.4 ± 2.3 77.8 ± 3.3 80.3 ± 2.5 85.3 ± 2.5 80.9 ± 2.5 85.3 ± 2.5 85.2 ± 2.5 Red blood cells, mln / μl 1.0-1.2 1.19 ± 0.2 1.19 ± 0.2 1.03 ± 0.06 1.01 ± 0.01 1.0 ± 0.01 1.10 ± 0.01 1.15 ± 0.01 1.1 ± 0.01 1.15 ± 0.01 1.16 ± 0.01 White blood cells, thousand / μl 20-25 24.1 ± 3.4 24.7 ± 3.6 42.1 ± 6.0 24.2 ± 3.0 22.8 ± 2.1 23.8 ± 2.3 25.2 ± 2.3 22.0 ± 2.2 25.2 ± 2.3 25.4 ± 2.3

Table 3 The effect of the nature of the feed additive on the content of heavy metals in the body of fish. Metal mg / kg Norm * MPC one 3 6 9 10 Cadmium 0.2 2.2 ± 0.4 2.0 ± 0.4 0.8 ± 0.4 0.8 ± 0.4 0.7 ± 0.4 Lead 1,0 5.1 ± 1.2 4.8 ± 1.1 3.0 ± 1.2 2.8 ± 1.2 3.4 ± 1.2 Zinc 40 49.1 ± 5.9 52.2 ± 5.2 32.5 ± 3.8 30.2 ± 3.5 35.5 ± 3.2 Copper 10 35.0 ± 5.0 33.0 ± 5.0 22.4 ± 2.4 20.4 ± 3.2 22.0 ± 2.4 * http: //www.dietolog.org/components/food-toxins/heavy-metals/#requirements-the-content-heavy-metals-foods

Claims (2)

1. Additive to fish feed containing Halobacterium halobium cells, characterized in that it additionally contains salts of ethylenediaminetetraacetic acid, and Halobacterium halobium cells are contained in the additive in the form of dry sterilized biomass obtained during their cultivation and subsequent dehydration of the culture fluid containing these cells moreover, the ingredients are contained in the feed mixture in a concentration, wt.%:
dry culture fluid - 10-20,
salts of ethylenediaminetetraacetic acid 5-10,
feed mixture - the rest. 2. The feed additive for fish according to claim 1, characterized in that for feed that does not contain vitamin A, the content of ingredients in the feed mixture is, wt.%:
dry culture fluid 18-20,
salts of ethylenediaminetetraacetic acid 5-10,
feed mixture - the rest.