RU2264221C1 - Method for preparing complex immune modulator - Google Patents

Abstract

FIELD: veterinary science.

SUBSTANCE: the present innovation deals with cooling selected fertilized uninfected eggs at 2-4 C up to 7 d followed by homogenization, diluting with physiological solution of egg mass, bactericidal treatment, conservation with phenol solution up to its concentration being 0.3% at subsequent filtration of target product. Moreover, egg mass should be diluted with physiological solution at the ratio of 1:5, then comes thermal treatment upon water bath at 70-80 C for 1 h and before conservation one should cool the target product in bactericidal chamber under the lamp with ultraviolet irradiation for 30-40 min. The method provides increased total and specific body resistance, it enables to obtain cheaper and more acceptable biostimulant and simplified way for preparing.

EFFECT: higher efficiency.

4 ex, 3 tbl

Description

FIELD OF THE INVENTION

The invention relates to veterinary medicine, in particular to pharmaceutical preparations for increasing the general and specific resistance of an animal organism through the use of non-specific stimulating preparations having a regulatory effect on the immune system - immunomodulators.

State of the art

Known biostimulant-placenta, the active principle of the placenta of women. The preparation is made according to a new technology, without heat treatment using the cavitation effect and newly discovered chemicals when sterilizing (see. A new generation of tissue preparations - PAN // "Theoretical and practical aspects of the emergence and development of animal diseases and the protection of their health in modern conditions." International Conference. Voronezh, October 3-4, 2002. Conference proceedings. - Volume 1. - Voronezh, 2000. - 295 p.).

The disadvantage of this method of preparing a biostimulant is the high cost, complexity and multi-stage preparation.

There is also a method of preparing a biostimulator for feeding birds, which includes introducing into the main diet a biostimulator prepared by cooling at a temperature of 2-4 ° C for 4-5 days, diluting with water in a five-fold volume and settling for 24 hours, followed by autoclaving the resulting mass, with In this case, egg is used as a biostimulator and, after treatment, it is introduced into the diet in an amount of 0.2-0.3 ml per 1 kg of live weight with an interval of 6-7 days (see AS USSR No. 150163, class A 23 K 1/16).

The disadvantage of this method of preparing a biostimulant for feeding birds is the insufficiently high effect of increasing the general and specific resistance of the body.

The closest in technical essence and the achieved positive effect to the claimed method is the "Method of preparation of an embryonic biostimulator (STEMB)." The essence of the method is as follows. Fresh uninfected chicken eggs are selected, then they are incubated for 9 days, while on days 5, 6, 7, the eggs are irradiated for 30 seconds at a modulation frequency of 9 Hz and a radiation power of 9 mW, then the eggs are cooled for 6-7 days, homogenization is carried out, the egg mass is diluted 1:10 with physiological solution, mixed thoroughly, 5% phenol solution is added to the resulting mixture so that the phenol concentration is 0.3%, the resulting solution is filtered and poured into sterile vials, closed and stored in a refrigerator at a temperature 2-6 ° C (RF patent 2197251, MKI 7 A 61 K 35/12).

The disadvantage of this method is the high cost of materials, complexity and multi-stage, high energy consumption and the cost of the drug.

Disclosure of invention

The aim of the invention is the creation of a complex immune modulator (CMM), which helps to intensify the treatment and prevention of a number of diseases and pathological conditions of animals, accompanied by a decrease in the immunobiological properties of the body, and provides the preparation of a cheap and effective immunomodulator, in which a fertilized chicken egg is used as a substrate. In this case, rational methods were used, aimed both at preserving the initial properties of the substrate, and at obtaining a number of biologically active substances.

The technical result that can be achieved using the proposed method is to increase the overall and specific resistance of the body, to obtain a cheaper and more affordable biostimulator, to simplify the method of preparation. The technical result is achieved using the method of preparing a complex immune modulator, which includes cooling the selected fertilized uninfected eggs at a temperature of 2-4 ° C for up to 7 days, homogenization, dilution with physiological saline of the egg mass, bactericidal treatment, preservation with phenol solution to its concentration of 0.3%, followed by filtration of the target product, characterized in that the egg mass is diluted with saline in a ratio of 1: 5, then heat treatment is carried out in a water bath at a temperature of 70-80 ° C, for 1 hour and before preservation, the target product is cooled in a bactericidal chamber under a lamp with ultraviolet radiation for 30-40 minutes.

The essence of the proposed method

At the first stage, eggs are selected from a farm that is safe for infectious diseases. Use eggs of medium size, approximately the same in mass, without visible damage and contamination of the shell. At the second stage, eggs are placed in a dark refrigeration chamber at a temperature of 2-4 ° C for 5-7 days, which leads to slow death of the embryo with the production of nutrients in adverse conditions. Eggs are removed from the refrigerator, opened, the yolk is inspected for the presence of an embryo, mass volume is noted, homogenization is carried out. The resulting homogenizate is diluted with 0.9% saline in a ratio of 1: 5. Maintain the resulting mixture in a water bath at a temperature of 70-80 ° C for 1 hour. The homogenizate is placed in a bactericidal chamber and incubated in it for 30-40 minutes. Then filtered and poured into sterile vials. For preservation, use a 0.5% phenol solution to a concentration of 0.3%.

The implementation of the invention

Examples of specific performance of the proposed method

Example 1. At the first stage, eggs are selected from a farm that is successful in infectious diseases. Use eggs of medium size, approximately the same in mass, without visible damage and contamination of the shell. Cooling of selected fertilized uninfected eggs at a temperature of 2-4 ° C for 2-3 days, homogenization, dilution with physiological saline of egg mass in a ratio of 1: 3, heat treatment in an autoclave at a temperature of 120 ° C and a pressure of 1.5 atm. When holding in a refrigerator for 2-3 days after heat treatment of the mass in an autoclave, the mass curled up, turned out to be unsuitable for further work with it.

Example 2. At the first stage, eggs are selected from a farm that is successful in infectious diseases. Use eggs of medium size, approximately the same in mass, without visible damage and contamination of the shell. Cooling the selected fertilized uninfected eggs at a temperature of 2-4 ° C for 5-7 days, homogenization, dilution with physiological saline solution of the egg mass in a ratio of 1: 3, heat treatment in a water bath at a temperature of 70-80 ° C, for 1 hour, cooled in a bactericidal chamber under a lamp with ultraviolet radiation for 40 minutes. Preservation is carried out with a 0.5% solution of phenol to its concentration of 0.1% of the mass fraction of egg mass. The amount of biologically active substances in the resulting preparation was very low, because egg protein curled up and remained on the filter. The phenol concentration is low, and the sterility culture is positive.

Example 3. At the first stage, eggs are selected from a farm that is successful in infectious diseases. Use eggs of medium size, approximately the same in mass, without visible damage and contamination of the shell. This method involves cooling the selected fertilized uninfected eggs at a temperature of 2-4 ° C for 7 days, homogenization, dilution with physiological saline solution of the egg mass, bactericidal treatment, preservation with phenol solution to its concentration of 0.3%, followed by filtration of the target product, characterized in that the egg mass is diluted with saline in a ratio of 1: 5, then heat treatment is carried out in a water bath at a temperature of 70-80 ° C for 1 hour and before preservation the target product is cooled in a bacterium an acid chamber under a UV lamp for 30-40 minutes.

This method is the most effective, because the concentration of active substances is the highest, the drug is safe at a given concentration of preservative when used in animals.

Example 4. At the first stage, eggs are selected from a farm that is successful in infectious diseases. Use eggs of medium size, approximately the same in mass, without visible damage and contamination of the shell. The difference of this method is the cooling of selected fertilized uninfected eggs at a temperature of 2-4 ° C for up to 7 days, homogenization, dilution with physiological saline solution of egg mass in a ratio of 1: 7, thermal, bactericidal treatment, filtration. Heat treatment is carried out in a water bath at a temperature of 70-80 ° C, for one hour, cooled in a bactericidal chamber under a UV lamp for 30-40 minutes, preservation is carried out with a 0.5% phenol solution to a concentration of 1%, filtration .

With the introduction of this drug to animals, infiltrates appeared at the injection sites, the drug had an irritating effect on the skin of the animal, due to the high concentration of phenol. As a result of a decrease in the concentration of the drug itself, its effectiveness decreased markedly.

KIM Animal Test Results

In the manufacture of an experimental batch of the drug, a biological test was conducted for its harmlessness. The test was carried out on laboratory mice and rabbits.

KIM drug test results in laboratory animals

The development and use of biostimulant preparations used to increase the immunobiological reactivity in animals as a preventive measure and as part of various treatment regimens are a serious task.

Biological testing of the drug was carried out in a laboratory on white mice. Two groups of five mice were taken in the experiment: the first group was experimental, the second served as control. Animals were in the same conditions of keeping and feeding. Weighing of animals was carried out: before setting up the experiment and every two days. Observation of the physiological state of the animals was carried out daily. The mice of the experimental group were injected with the drug (CIM) at a dose of 0.5 ml / goal. According to the weighing results, the following data were obtained: in the experimental group received an increase in live weight by an average of 25% more than in animals of the control group. The results of using the CMM preparation on minks are given in table 1.

Table 1
The results of the use of the drug KIM on minks Age and gender groups Number of goals Dose, ml / kg Multiplicity of introduction Note Adult animals:
Males
twenty
0.1
3 times h / w 3 days - improvement of appetite; an increase in the number of coatings during the rut and a quick recovery after it; improved sperm quality. Females twenty 0.1 3 times w / w 3 days - reduction in cases of cannibalism; improved lactation; improvement of the safety of young animals before jigging; rapid recovery of females after lactation depletion Young growth:
Newborns
twenty
0.01 ml / goal
3 times w / w 3 days - good appetite is noted; increase in live weight; quick
accustoming to self-nutrition. 30-45 days twenty 0.05 ml / kg 3 times w / w 3 days - good appetite is noted; increase in live weight; fast accustoming to self-nutrition. 45-60 days twenty 0.05 ml / kg 3 times w / w 3 days - good appetite is noted; increase in live weight; fast accustoming to self-nutrition; reduction of cases of cannibalism; decrease in the body's response to vaccination.

Studies conducted at the Krasnogvardeiskoye breeding enterprise in the Stavropol Territory in 2003 convincingly show that the use of the KIM preparation at a dose of 0.05 ml per 1 kg of live weight three times in 7 days contributes to an increase in blood of boar-producing hemoglobin by 6.33 g / l, bactericidal activity of blood serum by 5.75%, lysozyme activity of blood serum by 3.97%. The activity of the transamination enzymes ALT and ACT increases by 0.07 and 0.08 µkat / l, reserve alkalinity by 4.4% vol. CO ² , the total protein in the blood by 10.2 g / l, the content of γ-globulins by 6, 27% compared with the control group.

A study of the biochemical parameters of the mink blood showed that the amount of hemoglobin in the males of the experimental group, where the complex immunomodulator KIM was used, was 4.7 g / l more than the analogues of the control group. In terms of the amount of total protein in the blood serum, there was also an advantage in males where a 3.5 g / L Kim drug was used. According to the albumin fraction, differences between the groups were in favor of the animals of the experimental group. In the gamma-globulin fraction, the animals of the experimental group exceeded the control by 1.8%.

All of the above indicates an increase in the resistance of the organism of the boars-producers and male minks, as well as the immunomodulating properties of the KIM preparation.

Biochemical analysis of the drug is important as an indicator of their quality. According to many scientists, a number of compounds have high biological potential and can determine the main biostimulating function of the drug. These substances are primarily amino acids. The amino acid composition of the drug is shown in table 2.

table 2
Amino acid composition of the drug KIM Amino acids Mg% Interchangeable Aspartic acid 5,4 Serine 3.4 Glutamic acid 10.0 Alanine 7.6 Tyrazine 3.2 Total 29.6 Irreplaceable Trianine 3.8 Glycine 4.9 Valine 6.2 Methian 1,5 Isoleucine 5.2 Leucine 9.2 Phenylalanine 5.2 Histidine 3,7 Lysine 3.3 Arginine 4.0 Total 47.0

To characterize the biological value of this drug, they use such a concept as the amino acid index, which determines the amino acid usefulness, it is equal to the ratio of essential amino acids to essential ones. In biologically complete substances, it is higher than 1, the amino acid index (KIM) = 47.0 / 29.6 = 1.59.

The following hormones are contained in the KIM preparation (Table 3)

Table 3
The hormone content in the KIM preparation Name of hormones number TTG (thyroid-stimulating hormone), mMed / l 0.09 FSH (follicle-stimulating hormone), mIU / ml 1,5 Free T4 (L-thyroxine), pmol / L 0.2 LH (luteinizing hormone), mIU / ml 2.2 Progesterone, nmol / L > 100.0 Testosterone, nmol / L 5.8 Cornisol, nmol / L 0.1 Prolactin, mIU / ml 75.0 STH (growth hormone), mMed / l 10.0 DHEAc (dehydroepiandrosterone), μg / ml 0.26 ACTH (adrenocorticotropic hormone), pg / mg 0.9

The quantitative composition of the drug

100 ml of the drug includes:

- 20 ml of biologically active egg mass;

- 79.7 ml of 0.9% sodium chloride solution;

- 0.3 ml of phenol as a preservative.

This drug is used in fur farming to reduce immunodeficiency conditions associated with the growth and shedding of puppies. Also, this drug has proven itself in the preparation of animals for rutting and whelping.

Thus, the immunomodulator has a high effect of increasing the general and specific resistance of the body.

The invention in comparison with the prototype and other known technical solutions has the following advantages:

- biostimulant obtained using the present invention has the effect of tissue therapy, protein hydrolyzate therapy, organotherapy, nonspecific protein therapy, as well as a bactericidal effect;

- the preparation method allows not only to obtain a drug with the effect of tissue and other types of therapy, but also allows you to save the beneficial properties of the egg: high amino acid composition, vitamins and hormones, high immune parameters, activation of the metabolic rate;

- cheaper manufacturing;

- The main methods, drugs, devices used in the preparation of the drug KIM are readily available.

Claims (1)

The method of preparation of the immune modulator, including cooling the selected fertilized uninfected eggs at a temperature of 2-4 ° C for 7 days, homogenization, dilution with physiological saline egg mass, bactericidal treatment, preservation with phenol solution to its 0.3% concentration, followed by filtration of the target product, characterized in that the egg mass is diluted with saline in a ratio of 1: 5, then heat treated in a water bath at a temperature of 70-80 ° C for 1 hour and before preservation her target product is cooled in a bactericidal chamber under a lamp with ultraviolet radiation for 30-40 minutes