EA019956B1 - Pharmaceutical composition for treating diseases of bacterial and mycoplasmal aetiology - Google Patents

Abstract

The invention relates to veterinary medicine and is used for the treatment of bacterial infections. A pharmaceutical composition comprises two active substances relating to tetracycline and amfenikol groups in aprotic solvents. The composition can further comprise cations of bivalent metals. There can be also used doxycycline and florfenicol and their nearest analogies as pharmaceutically acceptable salts as the active substances relating to the to tetracycline and amfenikol groups.

Description

The invention relates to the field of veterinary medicine, is an antibacterial complex drug for the treatment of respiratory diseases and diseases of the gastrointestinal tract in farm animals.

Gastrointestinal and respiratory diseases significantly limit the development of livestock. Most often, infectious diseases occur in mixed form, which complicates the course of the disease and the choice of an effective drug. For the treatment of such associated infections, it is advisable to use combined drugs to overcome the problem of antibiotic resistance and significantly expand the spectrum of antimicrobial activity.

Known antibacterial composition (KN 1341765, IPC A61K 31/65, publ. 06.2001), including ascorbic acid, chloramphenicol and propylene glycol as solvent, in order to increase biological activity additionally containing sulfadimesin, oxytetracycline hydrochloride, novocaine and polyethylene glycol-400 at the ratio of components, wt.%:

Ascorbic acid 0.05-0.15

Chloramphenicol 2-6

Sulfadimesin 1-3;

Oxytetracycline hydrochloride 2-6 Polyethylene glycol-40010-30

Novocaine 0.5-1.5

Propylene glycol up to 100

Known antibacterial drug (KI 2089183, IPC A61K 31/00, publ. 09/10/1997), containing the following ratio of components, wt.%:

Novocaine or another anesthetic 0.5-1.5

Chloramphenicol 2-6

Oxytetracycline or another tetracycline antibiotic 1 3

Erythromycin

Dimexide

2-6

Before

one hundred

The lack of these analogues is associated with low efficiency in the presence of novocaine sulfonamides, which are used in these drugs along with old-generation antibiotics, to which many strains have resistance.

The closest analogue is a drug for the treatment of animals (KI 2277418, IPC A61K 31/65, publ. 10.06.2006), containing chloramphenicol, oxytetracycline hydrochloride, magnesium chloride, 1,2-propylene glycol, polyethylene oxide 400 in the following ratio, g per 100 3 cm of the drug:

Chloramphenicol

Oxytetracycline hydrochloride Magnesium chloride

1,2 Propylene glycol polyethylene oxide

5,0

5,0

0.9

75cm ³

Up to 100 cm ³

This analogue has the following disadvantages: it contains insufficiently effective antibiotics of the old generation, including chloramphenicol, which has a toxic effect on the hematopoietic and immune systems, is characterized by a narrower spectrum of action and is used only for the treatment of bronchopneumonia in one animal species (cattle calves).

The problem to which the invention is directed, is to create a composition based on doxycycline and florfenicol for the treatment of diseases of bacterial etiology with higher efficiency and a wide spectrum of action due to the introduction of newer antibiotics and having a synergistic effect.

The technical result of the invention is to increase the effectiveness of the treatment of diseases of bacterial and mycoplasma etiology in cattle, small cattle and pigs with lower doses with a decrease in toxic effect.

The technical result is achieved by the use of a complex preparation comprising the following components (in various combinations): doxycycline or its pharmaceutically acceptable salts, florfenicol, an organic solvent and additional excipients: divalent metal cations, water, antioxidant, solubilizer.

It is known that a wide range of antimicrobial activity of drugs is usually achieved by combining several drugs based on one or more groups of chemical compounds. Moreover, the combination of various chemical structures in the composition allows to achieve

- 1 019956 their synergistic effect and obtain drugs with a wider spectrum of action. This indicates the relevance of developing an easy-to-use and safe complex antibacterial drug based on doxycycline and florfenicol, the use of which will increase the effectiveness of the treatment of a wide range of mixed forms of infectious diseases in farm animals. Doxycycline and florfenicol are antibiotics of a wide spectrum of bacteriostatic action, covering microorganisms of various morphological groups, are characterized by long half-lives from the body, which reduces the frequency of use of the drug.

Doxycycline is a semi-synthetic antibiotic from the tetracycline group, it is a drug of a wide spectrum of bacteriostatic action. By its chemical structure, it is 6-dioxi5-oxytetracycline and has a mechanism of action common to all tetracyclines - suppression of protein synthesis in a microbial cell with a disruption in the connection of transport aminoacyl RNAs with 308 subunits of the ribosomal membrane. Doxycycline, being a representative of the tetracycline group, in comparison with other antibiotics of this series has a higher ability to penetrate into the microbial cell due to its lipophilic properties. In commonly used concentrations, it acts bacteriostatically on extracellularly and intracellularly located pathogens.

To doxycycline highly sensitive: gram-positive microorganisms 81 aryu1ossoss krr. (including 81 aryu1ossossik igeyik, 81aryu1osossik eryegtysh), 81er1osossik krr. (including 81er1ossossic ritual), S1ok1gIsht krr., L1k1eg1a krr .; and gram-negative microorganisms: No. kkepa dopoggyoeae, No. kkepa teshid SHLk, Naetoryyik tpien / ae, Kliebkel11a cr. (formerly Rak1eige11a krr.), Vas1ego1yek krr., Teropeta krr. (including strains resistant to other antibiotics, such as modern penicillins and cephalosporins). The most sensitive are the Nathorrhync nipple and intracellular pathogens. Doxycycline is active against most pathogens of dangerous infectious diseases: plague, tularemia, anthrax microbes, legionella, brucella, cholera vibrio, rickettsia, glanders, chlamydia. Does not affect most strains of Proteus, Pseudomonas aeruginosa and fungi. To a lesser extent than other tetracycline antibiotics, it inhibits the intestinal flora. Doxycycline is superior to natural tetracyclines in the degree of antibacterial activity. Unlike tetracycline and oxytetracycline, it has higher therapeutic efficacy.

Florfenicol belongs to the group of amphenicol. Features of the molecular structure of florfenicol determine its lower toxicity and higher antimicrobial activity compared to its analogue, chloramphenicol (chloramphenicol). First, the substitution of one of the hydroxyl groups with a fluorine atom increases the sensitivity of microorganisms resistant to chloramphenicol. Secondly, in the florfenicol molecule, unlike chloramphenicol, there is no nitro group that has a toxic effect on the hematopoietic system and the immune system, which increases the safety of its use. In addition, florfenicol was little used previously in veterinary practice on the territory of the Russian Federation, and at present no resistant microorganisms should exist for it.

The mechanism of action of florfenicol is associated with impaired protein synthesis in microbial cells. Florfenicol has a bacteriostatic effect on microorganisms sensitive to it. It is effective against many gram-positive and gram-negative bacteria, rickettsia, spirochetes, acts on strains of bacteria resistant to penicillin, streptomycin, sulfanilamides, effective against bacteria that produce acetyltransferase and are resistant to chloramphenicol. Florfenicol has a broad spectrum of antibacterial activity including against LsypoasShik r1eigorpeitop1ae, Rak1eige11a tiyosya, Rak1eige11a yaetoyysa, Vogye1e11a gopsYkerysa, Naetoryyik kr., Myuor1akta yuorpeitoshae, Myuor1akta yuogYshk, Otyoas1egsht gYpoyasyea1e, E. soi, 8a1tope11a kr., 81aryu1osossik aigeik, 81ger1osossik cr ., 81 idea cr., K1ebk1e11a kr., Prgoek kr. It is weakly active against acid-resistant bacteria, Pseudomonas aeruginosa, clostridia and protozoa. Drug resistance to the drug develops relatively slowly, while, as a rule, cross-resistance to other chemotherapeutic agents does not occur.

Preparation and use of the composition

A method of preparing a composition is to mix the components. At the same time, a weighed portion of doxycycline hyclate (1-10%) and antioxidant (0.1-0.5%) are added to the calculated amount of the organic solvent, then, with stirring, the calculated amount of water (2-17%) is added (the reaction is exothermic), the magnesium compound (0.1-5%) and after cooling to room temperature - a weighed portion of florfenicol (5-15%) and polyvinylpyrrolidone (collidone) (1-5%), stirred until completely homogeneous and filtered through a filter with a pore diameter of 0.45 microns.

- 2 019956

Examples of specific performance Example 1.

Component Name Component Action The content of the component mass% Doxycycline hyclate Active substance 1 6.00 Florfenicol Active substance 2 10.00 Dimethylacetamide (DMA) Solvent 1 69.88 Kollvdon 12 PP Solubilizer 2,50 Rongalit (sodium formaldehyde sulfoxylate) Antioxidant 0.41 Water Solvent 2 10.00 ΜβΟ Stabilizer 0.71 Lidocaine hydrochloride Local anesthetic 0.50

This composition has the maximum synergistic effect and minimal pain effect.

Example 2

Component Name Component Action The content of the component, mass% Doxycycline hyclate Active substance 1 6.00 Florfenicol Active substance 2 10.00 Dimethylacetamide (DMA) Solvent 1 70.38 Kollidon 12 PP Solubilizer 2,50 Rongalit Antioxidant 0.41 Water Solvent 2 10.00 M ₈ O Stabilizer 0.71

This example has the maximum synergistic effect.

Example 3

Component Name Component Action The content of the component, mass% Doxycycline hyclate Active substance 1 1.00 Florfenicol Active substance 2 15.00 Dimethylacetamide (DMA) Solvent 1 79.30 Kollidon 12 PP Solubilizer 2,50 Rongalit Antioxidant 0.40 Water Solvent 2 1.70 MDO Stabilizer 0.10

This composition has maximum efficiency in relation to gram-negative microorganisms.

Example 4

Component Name Component Action The content of the component, mass% Doxycycline hyclate Active substance I 10.00 Florfenicol Active substance 2 5.00 Dimethylacetamide (DMA) Solvent 1 60.10 Kollidon 12 PP Solubilizer 2,50 Rongalit Antioxidant 0.40 Water Solvent 2 17.00 M ^ O Stabilizer 5.00

The composition of this composition has maximum efficiency in relation to gram-positive microorganisms.

- 3 019956

Example 5

Component Name Component Action The content of the component, mass% Doxycycline hyclate Active substance 1 6.00 Florfenicol Active substance 2 10.00 Dimethylacetamide (DMA) Solvent 1 71.79 Kollidon 12 PP Solubilizer 1.00 Rongalit Antioxidant 0.50 Water Solvent 2 10.00 ΜβΟ Stabilizer 0.71

This composition has a minimum viscosity.

Example 6

Component Name Component Action The content of the component, mass% Doxycycline hyclate Active substance 1 6.00 Florfenicol Active substance 2 10.00 Dimethylacetamide (DMA) Solvent 1 68.19 Kollidon 12 PP Solubilizer 5.00 Rongalit Antioxidant 0.10 Water Solvent 2 10.00 M $ O Stabilizer 0.71

This composition has the maximum physico-chemical stability.

The following are data on therapeutic efficacy and titration of doses when using the composition according to example 1.

In total, 92 heads of cattle were used in the experiment, with a diagnosis of tracheitis 14 goals, mastitis, endometritis - 5 animals, dyspepsia - 3 heads, bronchopneumonia - 70 goals; small cattle with a diagnosis of bronchopneumonia - 15 goals; enteritis pigs - 20 animals.

The tolerance and therapeutic efficacy of the drug were determined on young cattle, small cattle and pigs on the basis of farms in the Saratov region. The study design is presented in table. 1-5.

Table 1. The study design of the drug in calves

Options The inventive drug Kind of animal Cattle, calves Number of goals, age 14 goals at the age of 4 months Dosage ml 1 ml / 15 kg m., 1p / day, 3-5 days in a row, intramuscularly Diagnosis Tracheitis

The clinical condition of animals diagnosed with tracheitis was assessed daily throughout the experiment. Blood samples were taken for morphological and biochemical studies before administration, 1 hour and 120 hours after drug administration. Treatment was prescribed according to the developed scheme presented in table. one.

After the first injection at the beginning of the second day in 30% of the animals, cough decreased and appetite appeared. On the third day, the animals disappeared by 64-79% of symptoms such as cough, soreness of the trachea, wheezing. On the fourth day, the percentage of recovery was 91%. By the end of the fifth day, the animals looked clinically healthy.

Table 2. The study design of the drug in cows

Options The inventive drug Kind of animal Cattle, cows Number of goals, age 5 goals at the age of 5 years Dosage ml 1 ml / 10 kg m., 1p / day, once, intramuscularly Diagnosis Mastitis, endometritis

The clinical condition of animals diagnosed with mastitis, endometritis and dyspepsia was observed for 4 days, i.e. until complete recovery. Treatment was prescribed according to the developed experimental design, presented in table. 3. After injection of the drug in a dose of 1 ml / 10 kg m. for 4 days in cows

- 4 019956 recorded a complete clinical recovery.

The determination of preliminary tolerance and therapeutic efficacy of the drug was also carried out on calves with a diagnosis of dyspepsia. The experiment was carried out according to the research scheme presented in table. 3, at a dose of 1 ml / 15 kg m., 1 r / day, for three days, intramuscularly. In animals, after the third injection, a complete clinical recovery occurred.

Table 3. The study design of the drug in calves

Options The inventive drug Kind of animal Cattle, calves Number of goals, age 3 heads at the age of 10 days Dosage ml 1 ml / 15 kg m.zh., 1 r / day, for 3 days, intramuscularly Diagnosis Dyspepsia

The scheme of the study of tolerability and therapeutic efficacy of the drug in MPC and pigs is presented in table. 4 and 5.

The clinical status of lambs diagnosed with bronchopneumonia and piglets with enteritis was evaluated daily throughout the experiment.

Table 4. Scheme of study of the drug in small cattle

Options Claimed drug Kind of animal Cattle, lambs Number of goals, age 15 goals at the age of 1.5 months Dosage ml 1 ml / 10 kg m. _L 1p / day, 3 days in a row, intramuscularly Diagnosis Bronchopneumonia

Table 5. The study design of the drug in pigs

Options Claimed drug Kind of animal Pigs Number of goals, age 20 goals at the age of 2 months Dosage ml 1 ml / 10 kg bw, 1 r / day, 3 days in a row, intramuscularly The diagnosis. Enteritis

The lambs were diagnosed on the basis of clinical signs and history. For therapeutic purposes, animals were prescribed treatment according to the developed scheme (Table 4). On the fourth day, the percentage of recovery was 90%. By the end of the fifth day, the animals looked clinically healthy.

From the above results it follows that intramuscular administration of the claimed drug to lambs with bronchopneumonia in a dose of 1 ml / 10 kg m., 1 time per day for 3 days, leads to a therapeutic effect.

Determination of tolerance and therapeutic efficacy of the claimed drug was also carried out on pigs. Based on the clinical picture and anamnestic data, animals were diagnosed with enteritis. For therapeutic purposes, the animals were used in the claimed preparation at a dose of 1 ml / 10 kg m., 1 time per day for 3 days.

After the injection of drugs, the improvement of the animals came at the end of the first day. Piglets have disappeared symptoms such as pain on palpation, weakness, depression; improved appetite. On the third day, feces assumed a physiologically normal consistency; 70% of the animals developed an appetite; body temperature returned to normal. On the fourth day, the pigs looked clinically healthy. The percentage of recovery of animals was: on the second day - 53%; on the third day - 87%; on the fourth day - 100%.

When titrating doses of the drug, groups of animals were formed according to the principle of analogues taking into account age, live weight, feeding conditions, content and diagnosis. The drug was used, guided by the dose titration scheme (Table 6), calves of 4 months of age with a diagnosis of bronchopneumonia were used.

The studies carried out are confirmed by the results of hematological and biochemical analysis of blood (tab. 7, 8).

- 5 019956

Table 6. The scheme of titration of doses of the drug

Multiplicity of introduction Group number Dose, ml / kg m. Dose, ml / animal Number of animals, goals The weight of the animal, kg. 1 time / day for 5 days one 1 / 6.7 9.0 10 60 2 1/10 6.0 10 3 1/15 4.0 10 1 time / day, with an interval of 48 hours 4 1 / 6.7 9.0 10 5 1/10 6.0 10 6 1/15 4.0 10 1 time / day, with an interval of 72 hours 7 1/5 12.0 10

We studied the morphological composition of peripheral blood and biochemical parameters of blood plasma.

According to the results of studies of the morphological composition of the peripheral blood of calves with intramuscular injection of the drug, there is a slight increase in hemoglobin and the number of red blood cells, as well as an insignificant increase in white blood cells within the physiological norm in this group of animals after drug administration.

Table 7. Hematological parameters

Index Doses of drug administration 1 ml / 7 _g 5 _kg daily 1ml / 10.0kg daily 1 ml / 15.0 kg daily 1ml / 5, dkg in 72 hours 2me / 7.5 "g after 48 hours 7 ml / 16.0 kg after 48 tea 1 ml / 15D kg after 48 hours White blood cells, 10¾ slight fluctuations slight fluctuations slight fluctuations 9.40 increase increase by 20.47 increase by 7.83 lowering by 5.9 Monocytes,% slight fluctuations slight fluctuations slight fluctuations slight fluctuations slight fluctuations slight fluctuations slight fluctuations Lymphocytes,% down 13.33 lower yes 15.17 slight fluctuations slight fluctuations lowering yes 15,23 slight fluctuations lowering at 27.15 Granulocytes,% an increase of 14.68 increase of 13.77 7.80 increase slight fluctuations increase by 7.8 slight fluctuations lowering at 27.33

From the presented results of studies of biochemical parameters of blood serum of calves with intramuscular injection of the drug, it follows that it does not have toxic properties.

The creatinine kinase, aspartate aminotransferase and alanine aminotransferase enzymes are reduced throughout the experiment, and creatinine kinase decreases to normal values, which indicates a pronounced positive dynamics of recovery.

The level of the enzyme alkaline phosphatase throughout the experiment does not change and remains within the initial values. An elevated level of this enzyme may be associated with partial hemolysis of red blood cells.

The concentrations of creatinine, urea, glucose and bilirubin in the blood serum of calves do not change and remain within normal values, therefore, the drug does not have hepato- and nephrotoxicity.

The indicators of total protein and albumin increase in the blood serum of calves, which once again proves the beneficial effect of the drug on the biochemical and metabolic processes of the animal's body.

- 6 019956

Table 8. The results of biochemical studies of blood serum of calves

Creatine kinase activity exceeds normal values before the start of the experiment, which is typical for acute myocarditis, developing against the background of bronchopneumonia. During the experiment, in animals of groups 6 and 7, the activity decreases, which, obviously, is associated with the correct selection of the administered dose of the drug and the achievement of the therapeutic effect.

Alkaline phosphatase is present in all tissues, especially a high concentration in the liver, bile duct, kidneys, bones and placenta. During the experiment decreases in all groups except the group

1. The therapeutic effect is achieved.

An increase in aspartate aminotransferase (AST) in excess of an increase in ALT is characteristic of damage to the heart muscle; if ALT is higher than AST, then this usually indicates the destruction of liver cells. The activity of aspartate aminotransferase increases in groups 1, 2, 3 and 5 compared with the starting point, in groups 4, 6 and 7, a decrease in activity occurs. All fluctuations in activity occur within the framework of physiological norms.

All fluctuations in the activity of alanine aminotransferase occur within physiological norms, with the exception of group 1.

In the claimed complex preparation, a synergistic effect of doxycycline and florfenicol was revealed for strains §1herChoseo8 ruodeyeki 163 and С1о81пбшш ποννί 198 - table. 9 (Report FGUZ Ros NIPCHI Microbe, 2011).

Table 9. Indicators of BMD (minimum inhibitory concentration) of antibacterial drugs for test strains

Rod, species microorganism Strain number MPC, mg / ml Doxycycline Florfenicol The claimed drug doxycycline + florfenicol 8 {ger {ossoss ruo ^ epese 163 0.195 1,56 0.097 + 0.195 SOZRiNit pooug 198 0,097 12.5 0.012 + 0.024

Similarly, the effectiveness of the treatment was confirmed for the other examples cited. Conclusions:

1. The drug is well tolerated by animals and shows high therapeutic efficacy with intramuscular injection at a therapeutic dose of 1 ml / 10 kg mw, both with daily use and with an interval of 48 hours; and at a dose of 1 ml / 5 kg bw, with an interval of 72 hours with fractional administration of the drug no more than 5 ml at one injection site.

2. Allergic reactions with the introduction of the drug in the studied doses are absent.

3. Side effects (impaired coordination of movements with signs of pain, twitching of the hind limb on the injection side, lateral position) were present in isolated cases when the drug was administered at a dose of 1 ml / 6.7 kg m., Both with daily use and with an interval of 48 hours, which may be associated with individual intolerance to the drug by animals.

4. The drug has a synergistic effect of doxycycline and florfenicol in relation to the strains of §1er Chocodes 8 rhoodeic 163 and C1o81pcp and UChU 198.

Thus, the inventive preparation is characterized by a wide spectrum of antimicrobial action against gram-positive and gram-negative microorganisms and has a synergistic effect. The drug has shown high efficiency in the treatment of diseases of bacterial and mycoplasma etiology in cattle, small cattle and piglets.

The drug does not adversely affect the body of animals, does not have side effects, with the exception of cases of individual intolerance.

The invention provides an increase in the effectiveness of treatment compared with drugs based on separately doxycycline and florfenicol and an analogue containing chloramphenicol and oxytetracycline hydrochloride.

Claims (6)

CLAIM 1. A pharmaceutical composition for treating diseases of bacterial and mycoplasma etiology, containing florfenicol and doxycycline in at least one aprotic polar solvent, characterized in that it additionally contains divalent metal cations in an amount, wt.%: Florfenicol - 5-15, doxycycline - 1-10, divalent metal cations - 0.1-5, water - 217, aprotic polar solvent - the rest. 2. The composition according to claim 1, characterized in that as the divalent metal cations it contains magnesium and calcium cations in the form of compounds such as oxides, chlorides, acetates, sulfates. 3. The composition according to claim 1, characterized in that, as an aprotic polar solvent, it contains a pyrrolidone solvent, or Ν, Ν-dimethylacetamide, or Ν, Ν-dimethylformamide, or dimethyl sulfoxide, or a combination thereof. 4. The composition according to claim 3, characterized in that as the aprotic polar pyrrolidone solvent contains a solvent that is I-methyl-2-pyrrolidone or 2-pyrrolidone. 5. The composition according to claim 1, characterized in that it further comprises sodium formaldehyde sulfoxylate as an antioxidant in an amount of 0.1-0.5%. 6. The composition according to claim 1, characterized in that it further comprises polyvinylpyrrolidone as a solubilizer in an amount of 1-5%. Eurasian Patent Organization, EAPO Russia, 109012, Moscow, Maly Cherkassky per., 2