RU2472519C1 - Method for increasing degree of ecdysteroid extraction from herbal objects - Google Patents

Abstract

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to pharmaceutical industry, particularly a method for ecdysteroid extraction. The method for ecdysteroid extraction from an elevated part of Serratula cupuliformis involving raw material extraction in 70% ethanol simultaneously exposed to ultrasound under certain environment.

EFFECT: method provides higher ecdysteroid yield.

1 tbl, 4 ex

Description

The invention relates to medicine, in particular to methods for producing therapeutic agents from plant materials.

A known method of ultrasonic treatment with water and aqueous-alcoholic solutions of plant materials containing cardiac glycosides from digitalis grass. Processing time ranged from 5 to 30 minutes. The greatest number of compounds is extracted with water in 5 minutes, while alcohol solutions are much more and in 30 minutes (Golchanov G.I. Ultrasound in pharmacy. M: Medicine, 1980, 176 p.).

The disadvantages of this method: it is designed for unstable compounds - cardiac glycosides, for which more intense and prolonged exposure can lead to overheating of biological structures and their destruction, which does not allow to extract them completely under the influence of ultrasound.

A known method of exposure to ultrasound with a frequency of 19-44 kHz on the release of phenolic compounds (flavonoids, tannins, phenol glycosides, coumarins, anthocyanins, phenol carboxylic acids) from plants with a reduction in the extraction process (Molchanov G.I. Ultrasound in pharmacy. M .: Medicine, 1980, 176 pp .; VM Lysyansky, The process of sugar extraction from beet, Moscow: Food industry, 1973, 223 pp .; MM Brook et al. Obtaining medicines from plant and animal raw materials under the influence of ultrasound. Ultrasound in physiology and medicine. T.1, Rostov-on-Don, 1972, p. 115-116).

Disadvantages: the method is developed for phenolic compounds and is mainly applicable to fruit crops.

The closest (prototype) is the method proposed in patent RU 2321420 C1, 04/10/2008, which developed a method for producing a fraction from Serratula coronata containing 80% ecdysteroids, including extraction of the aerial parts of plants with 70-96% ethyl alcohol for 4.5-36 three times with simultaneous sonication for 5 minutes. The disadvantages of the method are

1. the preparation of raw materials for extraction with hot water at 100 ° C leads to the destruction of ecdysteroids, since they are thermolabile compounds at temperatures above 60 ° C, while the use of ultrasonic treatment leads to an increase in the temperature of the extract (raw material) (table 1, example 11) .

2. the use of a hydraulic module of the extraction process 1: 9-15 for plants that are super concentrators of ecdysteroids - more than 1%, such as types of sickles (1-2%), is not enough, especially when isolating the sum of ecdysteroids from 4 kg of plant material.

The objective of the claimed invention is to develop a method of processing plant materials using ultrasound in the process of extraction in 70% ethanol, which increases the degree of extraction of ecdysteroids and simplifies the preparation of extracts by reducing the time of the process.

The problem is solved in that the method of increasing the degree of extraction of ecdysteroids from plant objects involves the extraction of plant objects with an extractant and simultaneous sonication, but unlike the prototype, extraction is carried out in an ultrasonic extractor once for 30-60 minutes.

The ultrasonic extraction method is economically viable in laboratories and industry. The use of high-intensity ultrasound can increase the speed of the process, increase the yield of extractable substances. The regime of ultrasonic waves, the exposure time, the temperature regime is individual for each type of plant. The method of ultrasonic extraction of biologically active substances from plant materials can be widely applied at the enterprises of the pharmaceutical industry for the manufacture of drugs with a high yield of ecdysteroids at low energy costs.

The use of ultrasound in the inventive method for extracting substances from plant materials can reduce the extraction time. Under ultrasonic action, diffusion processes accelerate and the temperature rises due to cavitation. High frequencies can negatively affect biological structures and lead to their destruction; therefore, it is important to determine the conditions for the extraction not only for a specific plant species, but also for groups of biologically active compounds. Examples of specific uses of the invention are given below.

Example 1

0.5 g of dry ground ground mass of Serratula cupuliformis serpus is placed in a flask, filled with 70% ethyl alcohol at a ratio of 1:20 and placed in an Elmasonic S 60 N ultrasonic extractor, frequency 37 kHz. The extraction time is 0.5 hours. Extraction is carried out under the influence of both ultrasound and the accompanying heating.

Since ecdysteroids are chemically labile substances and are easily destroyed at high temperatures, a temperature level of 50-60 ° C was maintained during U3 extraction. The destructive effect of ultrasound on ecdysteroids was not observed, which is confirmed by UV spectra, HPLC and a high content of these substances.

Quantitative analysis of ecdysteroids was carried out by a chromatospectrophotometric method, which consisted of preliminary chromatographic separation of the concentrated extracts on 20 × 20 cm glass plates coated with a mixture of silica gels LS 5/40 and LL ₂₅₄ 5/40 and aluminum oxide. An insignificant addition of SiO ₂ LL ₂₅₄ , which contains an indicator with short-wavelength (254 nm) fluorescence, made it possible to identify the localization zones of ecdysteroids in the ultraviolet light of the ultrachemiscope and to exclude the procedure for detecting them with a solution of vanillin in sulfuric acid [4, 5]; In order to improve the separation of the aerial parts of plants with a high content of flavonoids and other associated pigments, the ratio of the components of the adsorption mixture was used by increasing the proportion of Al ₂ O ₃ (SiO ₂ LS-SiO ₂ LL ₂₅₄ -Al ₂ O ₃ = 7: 1: 7 ) and the solvent system chloroform - ethanol - acetone in a ratio of 5: 3: 1. Elution of ecdysteroids from sorbents was carried out with 95% ethanol. The absorption maximum due to the presence of a keto group conjugated to a double bond in the ecdysteroids molecule was fixed at λ = 242 nm. The optical density was measured on a Shimadzu 1800 spectrophotometer. The desorption completeness of the compounds was determined by chromatography of solutions of a reference sample of known concentrations. The level of 20-hydroxyedisone determined by the chromatospectrophotometric method amounted to 1.62% by weight of absolutely dry raw materials.

Example 2

0.5 g of dry ground ground mass of Serratula cupuliformis serpus is placed in a flask, filled with 70% ethyl alcohol at a ratio of 1:20 and placed in an Elmasonic S 60 N ultrasonic extractor, frequency 37 kHz. The extraction time is 1 hour. Extraction is carried out under the influence of both ultrasound and the accompanying heating.

The temperature during Y3 extraction was maintained at 50-60 ° C. The destructive effect of ultrasound on ecdysteroids was not observed, which is confirmed by UV spectra and a high content of these substances. The level of 20-hydroxyedisone determined by the chromatospectrophotometric method amounted to 1.85% by weight of absolutely dry raw materials.

Example 3

0.5 g of dry ground ground mass of serrata Serratula cupuliformis is placed in a flask, filled with 70% ethyl alcohol in a ratio of 1:20 and placed in an Elmasonic-S 60 N ultrasonic extractor, frequency 37 kHz. The extraction time is 1.5 hours. Extraction is carried out under the influence of both ultrasound and the accompanying heating.

The temperature during Y3 extraction was maintained at 50-60 ° C. The destructive effect of ultrasound on ecdysteroids was not observed, which is confirmed by UV spectra and a high content of these substances. The level of 20-hydroxyecdysone determined by the chromatospectrophotometric method amounted to 1.54% by weight of absolutely dry raw materials.

Example 4

The extraction of 0.5 g of Serratula cupuliformis without the use of ultrasonic treatment was carried out for 12 hours, with the expense of large volumes of ethanol extractant (about 4-5 times more). The content of 20-hydroxyecdysone was 1.87%.

As a result, it was found that the inventive tool reduces the extraction time by 9-36 times (based on the formula from 0.5 to 1 hour), accelerates the analysis and release of ecdysteroids, and therefore, more effective than the prototype ..

Table 1 shows the content of ecdysteroids from the time of extraction.

Table 1 The content of ecdysteroids from the time of extraction Example Extraction time, h The content of ecdysteroids,% calculated on absolutely dry raw materials Example 1 0.5 1.62 Example 2 1.0 1.85 Example 3 1.5 1.54 Example 4 12.0 1.87

Claims (1)

The method of extraction of ecdysteroids from the aerial part of Serratula cupuliformis, including the extraction of plant materials with 70% ethyl alcohol with simultaneous sonication, characterized in that the extraction is carried out in a ratio of plant materials and extractant 1:20, at a temperature of 50-60 ° C in an ultrasonic extractor at a frequency 37 kHz once for 0.5 to 1 hour