RU2472500C1 - Method for normalising blood antithrombin iii activity in newborn calves with iron-deficiency anaemia - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: invention relates to veterinary science. A method involves the use of intramuscular injections of ferroglucinum 75 mg at 15 mg of iron per 1 kg of calf's body weight, once in a combination with feeding of glycopine 6.0 mg/day in the morning for 6 days starting simultaneously with injections of ferroglucinum 75.

EFFECT: method is high-effective in correction of blood antithrombin III activity.

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Description

The invention relates to veterinary medicine and biology, namely to hematology.

Analogues of the proposed method for correcting the activity of antithrombin III (AT III) in the blood of newborn calves with iron deficiency anemia does not exist.

In the literature, there is information about the use of ferroglukin in anemia in calves (Internal non-communicable diseases of farm animals. Under the editorship of V.M.Danilevsky. M: Agropromizdat, 1991. - 576 p.).

The literature also contains recommendations for the use in animals for the purpose of immunostimulation of glycopine (N-acetylglucosaminyl-N-acetyl-muramyl-alanyl-D-isoglutamine) (Andronova T.M., Pinegin B.V., Kozlov I.G., Ustinova GI The use of the glycopin immunomodulator for the prevention and treatment of animal diseases. Methodical recommendations. M., 2009. - 12 pp.) However, never before has glycopene, and especially its combination with ferroglucin, been used in newborn calves with iron deficiency anemia to correct existing they have attenuation of AT III activity the blood that can cause thrombogenic danger of animals, sharply worsening the anemia, weakening the reactivity of a living organism, often leading to his death.

The aim of the invention is the correction of the activity of AT III in the blood of newborn calves with iron deficiency anemia.

The essence of the proposed method is that for the correction of AT III in the blood of newborn calves with iron deficiency anemia, a solution of ferroglukin-75 (1 ml) containing 75 mg of iron in 1 ml of a solution of 15 mg of iron per 1 kg of live weight is used once intramuscularly the calf’s body and glycopine drinking at 6.0 mg / day in the morning (diluted in a small amount (100.0-150.0 ml) of water or milk or colostrum intended for calves to be fed) for 6 days, starting simultaneously with the injection of ferroglukin- 75.

The method allows to stably normalize the activity of AT III in the blood of newborn calves with iron deficiency anemia the day after the end of treatment, which will significantly improve the herd, reduce mortality, increase weight gain and receive healthy offspring from these animals.

The inventive method is as follows.

The determination of the activity of AT III in the blood of newborn calves with iron deficiency anemia is carried out as follows (Barkagan Z.S., Momot A.P. Basics of the diagnosis of hemostatic disorders. M., 1999. - 214 p.).

The principle of the method is based on the detection of a decrease in activity (in the absence of heparin) in AT III plasma, which is determined by the ability of the studied plasma to inactivate thrombin, for which it is pre-treated with a heparin sorbent, then subjected to thermal defibrination and mixed with a standard amount of thrombin. After incubation of the mixture, the residual coagulation activity of thrombin is determined in it, and the activity of AT III in the studied plasma is estimated by the level of decrease in thrombin activity as a percentage of the norm.

The following reagents are used. 1. Tris-HCl buffer 0.05 M, pH 7.4. 2. Thrombin solution in buffer: 0.2 ml of the solution should coagulate with 0.3 ml of 0.4% fibrinogen solution for 15-16 seconds. The thrombin working solution is prepared in a plastic or silicone tube, stored at room temperature and used in the test on the day of the study. 3. Sorbent of heparin Genasorb-1 (firm "Technology-Standard"). 4. Freshly prepared solution of fibrinogen at a concentration of 3.5-4.0 g / l, which is used to test the residual activity of thrombin. It is allowed to replace the fibrinogen solution with a normal platelet-poor nitrate plasma diluted in 2 times with buffer. 5. Fresh platelet-poor plasma obtained from 6-8 healthy calves, or proprietary lyophilized, with known activity of AT III, which is used to build a calibration graph.

The material for the study is nitrate-poor platelet plasma.

To prepare sorbed and defibrinated plasma in a test tube, 10 mg of heparin sorbent is mixed with 1.0 ml of the studied plasma. Constantly shaking the tube, mix its contents at room temperature for 8 minutes, excluding the formation of foam and sediment at its bottom. After that, the mixture is centrifuged for 2 minutes at 1000-1500 rpm. The plasma in the supernatant is transferred to a clean tube and defibrinated in a water bath at + 56 ° C for 6 min, after which it is again centrifuged at 3000 rpm for 10 min, the supernatant is removed and examined for the first 2 hours.

The course of determination. 0.4 ml of a thrombin working solution is added to the test tube and heated in a water bath at + 37 ° C for 2 minutes. Then add 0.1 ml of the studied adsorbed and defibrified plasma, include a stopwatch. After 2 minutes (exactly!), 0.2 ml of the mixture is transferred into a test tube with 0.3 ml of fibrinogen solution (or diluted normal plasma), preheated to + 37 ° C. Immediately turn on the stopwatch and record the clotting time.

The calibration curve is used to find the activity of AT III in percent.

The construction of the calibration curve is as follows. The plasma of healthy calves is defibrated as described above. Then, dilutions are prepared from it for constructing a calibration curve in accordance with Table 1.

Table 1 Scheme for preparing plasma dilutions for constructing a calibration curve in determining the activity of AT III Breeding number Plasma + buffer Activity AT III,% one Buffer free plasma one hundred 2 0.2 ml + 0.2 ml fifty 3 0.1 ml + 0.3 ml 25

With each of the dilutions, the clotting time is determined by the above procedure. All measurements are carried out three times, the average result is calculated. When constructing a calibration curve along the ordinate axis, the coagulation time in seconds is postponed, and the abscissa axis shows the activity of AT III in% in accordance with the above dilutions. An exemplary calibration curve is shown in figure 1.

A calibration curve for one type and one series of thrombin and buffer is built once. Only the activity of the thrombin working solution is periodically monitored.

Normally, the activity of AT III in plasma is 85-115%.

To correct a reduced AT III in the blood of newborn calves with iron deficiency anemia, ferroglukin-75 is prescribed once, containing in a 1 ml solution of 75 mg of iron at the rate of 15 mg of iron per 1 kg of calf body weight, and glycopine is taken out at 6.0 mg / day in the morning within 6 days, starting simultaneously with the injection of ferroglukin-75.

The method allows to normalize the activity of blood III AT in newborn calves with iron deficiency anemia on the next day after treatment, translating it to a level characteristic of healthy animals, which increases the reactivity of the animal’s body, treats the underlying disease, significantly reducing the risk of thrombotic complications and contributing to normal growth and animal development.

Example 1. Calf No. 74, the third day of life, body weight 27.0 kg, with iron deficiency anemia, examined in a calf. Blood was taken and examined from the calf with an assessment of the activity of AT III in it, which turned out to be reduced to 75%. The calf was prescribed 5.4 ml of a solution of ferroglukin-75 intramuscularly once and drinking the glycopine milk diluted in 100.0 ml at 6.0 mg / day in the morning for 6 days, starting simultaneously with the injection of ferroglukin-75. The use of this correction made it possible to raise the activity of AT III to the standard values of 110.0% the next day after its completion, eliminating the risk of further attenuation.

In the future, it was recommended to contain this calf on a rational diet for the prevention of a relapse of iron deficiency and weakening of AT III in the blood.

Example 2. Calf No. 82, the fourth day of life, body weight 31.5 kg, with iron deficiency anemia, examined in a calf. Blood was taken and examined from the calf with an assessment of the activity of AT III in it, which turned out to be reduced - 78%. The calf was prescribed 6.3 ml of a solution of ferroglukin-75 intramuscularly once and drinking the glycopine milk diluted in 150.0 ml at 6.0 mg / day in the morning for 6 days, starting simultaneously with the injection of ferroglukin-75. This allowed the next day after the final correction to raise the activity of AT III to standard values - 99%, eliminating the risk of its weakening in the future.

In the future, it was recommended to contain this calf on a rational diet to prevent it from recurring iron deficiency and weakening the activity of AT III in the blood.

Example 3. Calf No. 94, the second day of life, body weight 30.5 kg, with iron deficiency anemia, examined in a calf. Blood was taken and examined from the calf with an assessment of the activity of AT III in it, which turned out to be reduced to 73%. The calf was prescribed 6.1 ml of a solution of ferroglukin-75 intramuscularly once and drinking the glycopine milk diluted in 150.0 ml at 6.0 mg / day in the morning for 6 days, starting simultaneously with the injection of ferroglukin-75. This allowed the next day after the end of the correction to raise the activity of AT III to standard values - 107%, eliminating the risk of its weakening in the future.

In the future, it was recommended to contain this calf on a rational diet for the prevention of a relapse of iron deficiency and a decrease in the activity of AT III in the blood.

The claimed method was tested on 149 newborn calves with iron deficiency anemia and showed in 100% of cases the effectiveness of the correction of iron deficiency anemia and the level of activity of AT III in the blood.

Claims (1)

A method of normalizing the activity of antithrombin III in the blood of newborn calves with iron deficiency anemia, including the use of a solution of ferroglukin -75 at the rate of 15 mg of iron per 1 kg of calf body weight intramuscularly, once in combination with the release of glycopine at 6.0 mg / day in the morning, for 6 days, starting simultaneously with the injection of ferroglukin-75.

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