Method of higher meat production of broilers and related preparation

FIELD: medicine; veterinary science.

SUBSTANCE: method of higher meat production of broilers provides single injection for day birds of liposomal forms of preparation containing chimeric protein with water insoluble enzyme-inactive chloramphenicol acetyltransferase without 10 S-terminal aminoacids, aminoacid spacer (Sp)n, where n=1, 2, 4, 8 and somatostatin-14 with aminoacid sequence AGCFWKTFTSC, with median size of liposomes 250±50 nm. And preparation is introduced in combination with Marek's disease factor vaccine.

EFFECT: invention allows for higher effective meat production of broilers using single injection of preparation during the whole fattening period.

2 cl, 1 tbl

 

The invention relates to a meat poultry, and particularly to a method of increasing meat efficiency of broilers and preparation for its implementation.

Modern poultry in several ways solves the problem of maximum utilization of the genetic potential of the birds while maintaining its productive health. Accelerated growth and increased daily liveweight gain of broilers at lower economic cost per 1 kg increase in live body weight is one of the main tasks in meat poultry. For many years, were used for growth promoting antibiotics, get the bird to feed at certain concentrations. However, the advent and widespread in the late 80-ies of XX century antibiotic-resistant strains of microorganisms, with cross-resistance to antibacterial compounds used in medicine, has resulted in a dramatic reduction in the frequency and volume of use of antibiotics in poultry production. At the present stage in many countries, in particular in the States of the EEC, have banned the use of antimicrobial drugs as growth-promoting additives in feed the bird.

There are other, indirect methods of raising meat efficiency of broilers. These include the introduction of trace elements (the particular selenium), vitamins to feed the bird, the application of humic substances. All of these methods provide for multiple use of feed additives and is aimed primarily at optimizing the composition of feed mixtures for a number of controllable parameters.

The direct method of raising meat efficiency of broilers aimed at increasing concentrations of growth hormone (somatotropin) in the body of the bird. This can be achieved by using chemically synthesized somatotropin. However, the drug's high cost makes this method of increasing productivity is not always cost-effective, and the use of hormonal drugs is a negative attitude in society.

There is another method of increasing the concentration of endogenous somatotropin, by reducing the concentration in the body of his antagonist - somatostatin.

Somatostatin is a biologically active tetradecapeptide. Produced in the hypothalamus and the gastrointestinal tract of animals and birds. The sequence of somatostatin-14 markedly conservative among vertebrates. Somatostatin exerts a strong inhibitory effect on a wide range of hormones and related body functions. A broad spectrum of action of somatostatin on the factors necessary for the disposal of food, opens up great prospects for use as PE is ulator poultry. In this regard, great interest is the autoimmune reaction of the body of the bird on the introduction somatostatinergic protein, leading to a decrease in the concentration of peptide in the blood and, as a consequence, the induction of anabolic factors and the growth of birds.

Research showed that immunized drugs somatostatin animals showed increase in daily gain of body weight by 10-20%, increase in the efficiency of digestion and utilization of food by 11%. While there is an improved absorption of nutrients and slow its passage through the GI tract. Immunized with somatostatin animals and their offspring have the correct proportions and mass distribution of the body. However, the practical distribution of livestock antisomatostatin immunotherapy has not received due to the high cost of chemically synthesized somatostatin. Because of the small size of the somatostatin-14 does not allow its direct microbial synthesis using recombinant DNA technology described several ways to carry out the synthesis in the form of a chimeric protein with the specific selection of the target product, not given, however, satisfactory results. The main disadvantage of the above methods is extremely low immunogenicity obtained drugs in related and somatostatin, due to its masking of the molecule in the chimeric protein, therefore, these methods of producing chimeric proteins did not find wide practical use (R. Itakura et al., 1977 Expression in E. Coli of a chemically synthesired gene of hormone somatostatin, Science, 1986, 1056-1063; A.A. Shishkin and other Synthesis fragment somatostatin genes. Chemistry of natural compounds, 1988, No. 6, s-615).

The known method of constructing chimeric somatostatinergic proteins using amino acid spacer containing arginine and Proline and contributing to the localization of somatostatin on the surface of carrier protein and, thereby, the high immunogenicity of the drug (RU C1 No. 2031121, IPC 6 C12N 15/12, 1995).

The design consists of a water-insoluble protein carrier (fragment of bacterial chloramphenicolchloramphenicol without the 10 C-terminal amino acids), tetramer spacer and C-terminal somatostatin-14. The molecular weight of the chimeric Beja is 28 kDa. This chimeric protein is expressed by a strain of E.coli In-6519 transformed by the plasmid pC(Sp)4S. Strain deposited in Russian national collection of industrial microorganisms (VKPM). Chimeric protein with the exposed somatostatin is a water-insoluble enzyme inactive chloramphenicolchloramphenicol without 10 terminal amino acid residues, which through the spacer elements of the sequence p is soedineni sequence of somatostatin-14.

Method antisomatostatin immunization of animals with the use of this chimeric somatostatinergic protein is used in the cattle industry (EN C1 No. 2034457, IPC 6 AC 67/02, 1995).

The result of the invention consists in developing a method of raising meat efficiency of broilers by applying a liposomal form of the drug with chimeric somatostatinomas protein.

This result is achieved by way of increasing meat efficiency of broilers provides a single injection at day old birds liposomal form of the drug containing chimeric protein with a water-insoluble enzyme inactive chloramphenicolchloramphenicol without the 10 C-terminal amino acids, amino acid spacer (Sp)n, where n=1, 2, 4, 8 and somatostatin-14 amino acid sequence AGCFWKTFTSC in liposomal form, with a median size of liposomes 250±50 nm. The introduction of the drug carried out in conjunction with a vaccine against the causative agent of Marek's disease.

Accordingly, a drug for improving meat efficiency of broilers contains liposomal form an effective amount of a chimeric protein with a water-insoluble enzyme inactive chloramphenicolchloramphenicol without the 10 C-terminal amino acids, amino acid spacer (Sp)n, where n=1, 2, 4, 8 and somatostatin-14 last is the sequence of amino acids AGCFWKTFTSC, with a median size of liposomes is 250±50 nm.

The mechanism of drug action based on the temporary blocking activity of endogenous somatostatin birds and the increased concentration of endogenous growth hormone. While there is an improved absorption of nutrients and slow its passage through the gastro-intestinal tract of the bird. Once (during the whole period of fattening poultry) the introduction of the drug can significantly minimize additional technological operations personnel.

A distinctive feature of the chimeric somatostatinergic protein is its extremely low solubility in aqueous media, as well as its aggregation and subsequent occulation when attempting to obtain a homogeneous dispersion in the case of the introduction in aqueous solutions of the true protein solution in other solvents, easily miscible with water. As a consequence, may be difficult to obtain a homogeneous dispersion with a low protein concentration (1·10-3g/ml) in the aquatic environment for the introduction of living organisms with body weight is 40-50,

This purpose was developed liposomal form of the drug, in which chimeric somatostatinergic protein is located within the highly dispersed phospholipid vesicles. When this protein inside each vesicles, not to communicate with the t protein, prisoners in other phospholipid education. This factor prevents the formation of protein aggregates. It is also important that the high stability of the obtained fine liposomal system (the median size of liposomes 200 nm, the stability of more than 1 year) allows to obtain a homogeneous concentration of protein in the whole volume of the system.

The feasibility of introducing the drug in conjunction with a vaccine against the causative agent of Marek's disease is caused only by the desire to reduce the degree of injury to birds by conducting several courses of vaccination. The drug can be used alone, without vaccines against the causative agent of Marek's disease, which on the effectiveness of the drug has no effect.

The possibility of carrying out the invention is illustrated by the example of getting a drug to increase meat productivity of broiler chickens.

Obtained and purified from the impurities of the drug is protein dissolved in buffer 0.2 M Tris - HCl pH 8.0, containing 6 M guanidine hydrochloride and 2M of META. Add 50-fold molar excess of β-mercaptoethanol in the calculation of the number of S-S groups chimeric protein and the solution is quickly diluted 10-fold volume of buffer without handinhand. The resulting hybrid protein precipitate is separated by centrifugation for 15 minutes at 12000g and a temperature of 4°and freeze-dried to posleduyuschaya.

Then lyophilized preparation of protein dissolved in an aprotic solvent at the desired concentration (1·10-5-1·10-3M). The resulting solution was transferred in liposomal form by using the processing apparatus LXIV (linear stepper induction rotator) for 10-20 minutes, together with calculated amounts of phospholipids (20% alcohol solution) and distilled water. Received liposomal preparation is characterized by the following parameters: diameter of liposomes 250±50 nm; the content of phospholipids 2,5±1,0%; dry residue 5-20%. Concentration somatostatinergic protein in the final recipe of the drug is 1.35-2,70 mg/ml. Liposomal drug Packed in consumer packaging according to the normative-technical documentation.

The efficacy of the drug to increase meat productivity of broiler chickens illustrated by the following.

It was formed three analog group broiler day old 1000 animals each.

Liposomal form somatostatinergic (PRS) of the drug in a volume of 1.5 ml with a concentration of protein of 1.35-2,70 mg/ml in compliance with the rules of asepsis immediately before vaccination birds add in a bottle (200 ml) with a vaccine preparation against Marek's disease. Carefully (by shaking) mix the contents of the vial is to obtain a homogeneous concentration of the suspension in the whole volume of the vial. Inject 0.2 ml/head in accordance with the advice of immunization.

Data on the effect of liposomal forms somatostatinergic chimeric protein to meat productivity of broiler chickens (growing period 41 days) are presented in the table.

As follows from the table, a single application at day old broilers liposomal forms somatostatinergic protein at a concentration of from 2 to 4 mcg/head in conjunction with a vaccine preparation against Marek's disease leads to an increase in daily liveweight gain of a bird is in the range from 3.1% to 7.3% at the same time increase the safety of the poultry population of 1.2 to 1.6%.

OptionsSafetyAverage daily gain in live weight per head
goals% of controlg% of control
Control95610038.3100
PRS968101,239,5103,1
2 mcg/1 head
PRS971101,641,1107,3
4 mcg/1 head

1. The way to increase meat productivity of broilers, providing a single injection at day old birds liposomal form of the drug containing chimeric protein with a water-insoluble enzyme inactive chloramphenicolchloramphenicol without the 10 C-terminal amino acids, amino acid spacer (Sp)n, where n=1, 2, 4, 8 and somatostatin-14 amino acid sequence AGCFWKTFTSC in liposomal form, with a median size of liposomes 250±50 nm, and the introduction of the drug carried out in conjunction with a vaccine against the causative agent of Marek's disease.

2. Drug to increase meat efficiency of broilers containing liposomal form an effective amount of a chimeric protein with a water-insoluble enzyme inactive chloramphenicolchloramphenicol without the 10 C-terminal amino acids, amino acid spacer (Sp)n, where n=1, 2, 4, 8 and somatostatin-14 amino acid sequence AGCFWKTFTSC, with a median size of liposomes is 250±50 nm.



 

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3 tbl, 3 ex

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