RU1814900C - Method for producing extract of saga horns - Google Patents

Abstract

The invention relates to medicine and the medical industry, and in particular to methods for producing an extract of saiga horn. The aim of the invention is to intensify the method. The goal is achieved in that the middle layer of horns is additionally used as raw material, the raw material is crushed on a disintegrator to a powder state, extraction is carried out with distilled water at 20-25 ° C for min. when the ratio of raw materials: solvent 1:50 with simultaneous exposure to ultrasound. 2 ill., 4 tab.

Description

The invention relates to biological and medical chemistry and relates to the preparation of biologically active substances having a pharmacological effect.

The aim of the present invention is to expand the raw material base used to produce biologically active compounds, increase the yield of the product and increase its activity. It is proposed to use saiga horns to expand the raw material base, and the preparation obtained in this way is called saigorin.

In experiments that made it possible to evaluate the reserve capacity of the organism for the time of swimming of animals, it was found that the extract from saiga horns had a stimulating effect (Table 1). The swimming method and the like are exceptionally laborious. When developing the method for producing saigorin, it was found that the preparation had donor properties, i.e. the presence of compounds with a mobile hydrogen atom. The high biological activity of compounds with a mobile hydrogen atom is due to their reducing properties. Glutathione, the free SH group of which determines its properties as an intramolecular reducing agent and in connection with, can be classified as known compounds possessing this ability. this his wide participation in metabolism. The biological role of NADH and NADPH is associated with the presence of mobile hydrogen. Hydrogen donors are a number of reduced forms of organic compounds, for example, ascorbic acid, which plays the role of a natural antioxidant, etc. It has been established that there is a linear relationship between the duration of swimming of animals and the donor properties of the administered drug (Fig. 1), and therefore in the future when practicing the method half. The students of saigorin monitored its activity by this indicator.

Figure 1 shows the dependence of the swimming time of mice on the amount of the drug administered, we estimate HR by the presence of donor properties; figure 2 - dependence of the yield on the extraction conditions, where

ate

with

oo

Ј

H)

1 - traditional extraction methods; 2 - shear disintegration method; 3 - disintegration with a shift + exposure to pre-hypersound.

There are various methods for determining donor properties, however, due to a number of inherent disadvantages, we have used our own method for determining this indicator. To do this, 250 µl and 0.1 M phosphate buffer, pH 8.0, are added to the luminometer cuvette; 50 μl of a 5-10 M solution of horseradish peroxidase and placed in the cuvette block of the device. The reaction is started by the introduction of 200 µl of a 5 M luminal solution containing M hydrogen peroxide and the reaction kinetics are recorded on the recorder tape until it is complete. The area under the kinetic curve is taken as 100%. the reaction kinetics are recorded in the presence of the test samples, the areas under the kinetic curves are determined, and the activity of the test samples is judged by the magnitude of their decrease. The unit of activity is the amount of the drug that causes 50% and inhibition of luminescence arising in the reaction of peroxidase oxidation of luminol.

The studies showed that only the cover and the middle layer of the horns have biological activity, in contrast to the core, where such properties were not found (Table 2). In this regard, in the future, only a cover and a middle layer are taken to obtain saigorin, which allows reducing the amount of impurities in the target product, reducing the amount of raw material processed in the future by approximately 30%, and increasing its specific activity.

For a sufficiently complete and efficient extraction, a combination of several factors is necessary: the correct choice of extractants, degree of grinding, extraction conditions (temperature, time, etc.).

In the prototype, when analyzing the preceding methods for preparing the preparation, it is noted that simple grinding of antlers with repeated emulsification does not provide either a high quality of the preparation or its satisfactory yield. To increase the yield of the target product in the prototype, it was proposed to grind antlers with direct steam before grinding. Such processing obviously reduces the quality requirements for grinding, but cannot contribute to improving the quality of the product. This is due to the fact that the high temperature during heat treatment causes the denaturation of thermolabile protein compounds and, first of all, having biological activity. The decrease in the biological activity of antlers during heat treatment with water at a temperature of 98-99 ° C for 100 + 40 min was noted during the development of the method of preservation of antlers, in connection with which the authors proposed to reduce the time of welding of antlers to 30 min + 30 min

due to this, increase their biological activity by 18.7%.

The analysis of the disadvantages inherent in analogues and prototype leads to the conclusion that to increase the yield

5 product and increase the activity of the drug (and therefore, increase production efficiency) it is necessary: 1) improvement of grinding technology, 2) the use of zones. possessing biological activity. 3) the use of gentle extraction modes .; In order to increase the yield of product I, a method was developed for disintegrating antler tissue into protoplasm. For this, raw materials i

5 are triturated to a powdery state in the gap between the surfaces of the female and male bodies, the latter of which is subjected to a precessional vibration providing specific pressure;

0 equal to the tensile strength of the cells of the treated syllables. The combination of compression with simultaneous shear creates the conditions for increasing the efficiency of subsequent extraction. In order to confirm the feasibility of such an approach, a comparative analysis of the effectiveness of traditional grinding methods (impact, pressure, planing, etc.) and the disintegration method used was carried out. In order to preserve

0 of the activity of the preparation, extraction was carried out at a temperature of 20-25 ° C using a magnetic stirrer for stirring.

To simplify and reduce the cost of the process, an attempt was made to replace ethanol with distilled water. The premise and justification for this was that, in the range of solvents for chromatography, distilled water is a more polar compound. The results of the comparative analysis are presented in Table 3. It can be seen that aqueous extracts with the same sample volume to a greater extent reduce the luminosum of the glow (i.e., contain a larger amount

5 active substance) for both the cover and the middle layer. We have calculated the amount of extract that is necessary to reduce the luminosity of the glow by 50% (Vso). As can be seen from the table of the aqueous extract from the cover and the middle layer, 1.6 and 2.3 are needed

times less than alcohol, i.e. by the same amount, respectively, extraction with distilled water under the given conditions is more efficient. In this regard, it can be considered that this extraction technique is much cheaper and economically viable.

- When the dependence of the product yield on the extraction time is emitted (Fig. 2), it can be seen that the use of the described method of raw material disintegration compared with the traditional one allows 1.8-2.0 times increase in the yield of the target product with a more than ten-fold reduction in time extraction, replacing ethanol with widely available distilled water, carrying out the extraction process at a temperature of 20-25 ° C.

Taking into account the data indicating an increase in the biological activity of canned antlers with a shorter heat treatment time, we studied the effect of a 60-minute incubation of the obtained extract at 80 ° C, which revealed a decrease in the activity of the drug by 30-50%.

In order to more fully extract the target product from the raw material, the raw material-solvent system was exposed to extraction in the process of extraction with ultrasonic waves in the MHz range (F 2.64 MHz, I 0.6-1.0 W / cm2, T 600-900 s) that allowed to increase the product yield by an additional 10-12% (figure 2, curve 3).

Extraction is carried out at a ratio of feed: solvent (NaO) of 1; 50 (weight of feed in g: volume of water in ml). During the processing of the method, this ratio was selected as optimal, which makes it possible to extract 90–96% of the target product. A further increase in the volume of extractant, without leading to a significant increase in the yield of the product, complicates further work, since in the subsequent stages of isolation it is necessary to deal with large volumes of liquid, reducing the amount of H20

below this ratio leads to a rapid increase in the loss of active substance in the extraction step. Stirring is carried out on a mixer of any type at room temperature for 10-15 minutes, after which the liquid is centrifuged at 4500 g for 10 minutes. The supernatant is drained, freeze-dried and the desired product is collected. Lyophilization did not lead to a decrease in activity, which was established by comparing the activity of the substance before and after the lyophilization process (Table 4).

The method is carried out as follows (specific example):

Core (core) is removed from the feedstock (saiga horn). The remainder is triturated to a powder state as described in the description on page 5. Ten grams of the powder is poured into 500 ml of distilled water, placed on a magnetic stirrer, the emitter of the ultrasonic vibration generator (, 64 MHz, 7 W / cm2) is placed on top so that the surface of the emitter touches the water, the extraction process is carried out for 10-15 min. centrifuged for 10 min at 4500 d, the supernatant is collected and subjected to lyophilization, get 2.5-3.5 g of the target product.

Claims (1)

The claims A method of producing an extract of saiga horns, including grinding the outer layer of the horns, extraction followed by separation of the precipitate, characterized in that, in order to intensify the method, the middle layer of horns is additionally used as a raw material, the raw material is ground on a disintegrator to a powder state, the extraction is distilled water at 20-25 ° C for 5-8 minutes with a ratio of raw materials: water 1:50 with simultaneous exposure to ultrasound. Table 1 The dependence of the swimming time of mice on the amount of drug administered The activity of the drug by the light sum of suppression of luminescence in% Dependence of the activity of the extract on the type of extractant The effect of freeze drying on the activity of the drug table 2 Table 3 Table 4 Kol-60 drug. Figure 1 ° / in Output No. Ш Swim time, min. 43 Fae. 2 6060 T extractions.