RU2720457C1 - Method for extracting hyperforin and adhyperforin from st_ john's wort herb - Google Patents

Abstract

FIELD: pharmaceutics.

SUBSTANCE: invention refers to chemical-pharmaceutical industry, namely to a method for extracting hyperforin and adhyperforin from St. John's wort herb. Method comprises grinding common St. John's wort herb, extraction of ground raw material with organic solvent, evaporation of solvent, organic liquid used is an easily boiling liquid – perfluorobutane methyl ester with raw material/solvent ratio of 1:4.5–9.0 wt/wt; extraction of the raw material is carried out by solvent circulation for 1–3 hours; the obtained extract is evaporated.

EFFECT: invention enables to isolate hyperforin and adhyperhorine from ground St_ John's wort herb with perfluorobutane methyl ester (Novec 7100) at minimum power inputs, normal pressure, using standard equipment specific for chemical-pharmaceutical and/or essential oil-producing enterprises using environmentally safe technology.

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Description

The invention relates to the pharmaceutical industry and can be used to isolate hyperforin and adgerforin from perforated hypericum (Hypericum perforatum L.), family of hypericum (Hypericaceae).

М., Hohmann J., Keve Т.,

В. (2011). Extraction of hyperforin and hypericin from st. John's wort (Hypericum perforatum L.) with different solvents. Journal of Food Process Engineering, 35(2), 222-235 (doi: 10.1111/j. 1745-4530.2010.00583.x).Currently, several different methods are described in the literature for the isolation of phloroglucin derivatives (hyperforin and adgerforin) from perforated St. John's wort - extraction using organic solvents and liquefied gases (Cossuta D., Vatai T.,

M., Hohmann J., Keve T.,

B. (2011). Extraction of hyperforin and hypericin from st. John's wort (Hypericum perforatum L.) with different solvents. Journal of Food Process Engineering, 35 (2), 222-235 (doi: 10.1111 / j. 1745-4530.2010.00583.x).

A known method of obtaining an extract from the herb of St. John's wort perforated using 227 ea freon (Dowdle P.A., Merseyside SSH, Corr S., Cheshire A., Harris H., Cheshire W. Solvent extraction process. Patent US 2004/0105899 Al. A61K 35/78 Pub. Date: Jun.3, 2004). This method involves the extraction of St. John's wort grass perforated in a special installation for 4 hours, using liquefied gas HFC 227 ea at a pressure of 15 atm., And a temperature of 50 ° C. At the same time, a quantitative yield of a yellow gum-like product of 2.4% is achieved.

However, this method requires equipment that must operate under an overpressure of more than 15 atm. Another disadvantage of this method is the need for energy consumption for the operation of evaporation of the extract and the operation of condensation of solvent vapor in the refrigeration unit. Another disadvantage of this method is the use of freon with a high global warming potential equal to GWP = 3300.

Н., Seger С, Kaiser C.S., Haslinger Е., Schmidt Р.С.(2004). Enrichment of hyperforin from St. John's Wort (Hypericum perforatum) by pilot-scale supercritical carbon dioxide extraction. European Journal of Pharmaceutical Sciences, 21(4), 443-451 (doi:10.1016/j.ejps.2003.10.026)). Данный способ включает экстракцию травы зверобоя продырявленного в специальной установке в течение 1 часа, с помощью сверхкритического диоксида углерода при давлении 120 бар, и температуре 40°С. При этом, достигается выход экстракта 2,4% масс. с содержанием гиперфорина в нем 33%.Closest to the claimed technical solution for the technical nature and the achieved technical result is the method described in (

N., Seger C, Kaiser CS, Haslinger E., Schmidt R.S. (2004). Enrichment of hyperforin from St. John's Wort (Hypericum perforatum) by pilot-scale supercritical carbon dioxide extraction. European Journal of Pharmaceutical Sciences, 21 (4), 443-451 (doi: 10.1016 / j.ejps.2003.10.026)). This method involves the extraction of St. John's wort grass perforated in a special installation for 1 hour using supercritical carbon dioxide at a pressure of 120 bar and a temperature of 40 ° C. In this case, the yield of the extract is 2.4% of the mass. with hyperforin content in it 33%.

The main disadvantages of this method include: the need to use special equipment working under pressure, which is one hundred and twenty times higher than atmospheric, as well as the need for energy consumption for the operation of evaporation of the extract and the operation of condensation of solvent vapor in the refrigeration unit.

An object of the invention is the development of a more efficient method for the isolation of hyperforin and adgerforin from perforated St. John's wort grass, simple in execution in which special and complex equipment is not required.

The problem is solved using the proposed method for the isolation of hyperforin and adgerforin from perforated St. John's wort grass, including grinding the raw materials, extracting the crushed raw materials with an organic solvent, evaporating the solvent, moreover, an easily boiling liquid is used as an organic solvent - perfluorobutane methyl ether (hereinafter Novec 7100) at a ratio raw material / solvent 1: (4.5-9.0) mass / mass, extraction of the raw material is carried out for 1-3 hours in the Soxhlet apparatus, the obtained extract is evaporated at a temperature of 65 ° C, to obtain the amount of hyperforin and adhyperforin, and the extractant is condensed. The raw material, without removing from the extractor, is heated to 65 ° C, purged with air until the extractant is completely removed and condensed. The regenerated extractant is combined with the bulk of the extractant and reused for extraction of a new batch of raw materials.

The range of the ratio of raw materials / solvent 1: (4.5-9.0) mass / mass, provides effective extraction of hyperforin and adgerforin from plant materials and is economically feasible. The lower limit value of 1: 4.5 mass / mass is due to the fact that this amount of extractant allows you to cover the bulk volume of plant material in the extractor with a small excess of extractant necessary to enable its circulation in the extraction apparatus. The increase in the ratio of raw materials / solvent more than 1: 9.0 wt./mass., Does not lead to an increase in the output of hyperforin and adperforin and requires unreasonably high energy consumption and extractant.

The claimed extraction time interval of 1-3 hours provides a given level of extraction of hyperforin and adgerforin from raw materials. Reducing the time of extraction leads to a significant under-extraction of hyperforin and adgerforin from the St. John's wort perforated. Increasing the extraction time is not economically feasible, as well as energy consumption, since it does not significantly increase the yield of hyperforin and adgerforin from the St. John's wort perforated.

The obtained extract of hyperforin and adgerforin from the herb St. John's wort perforated can be used in medicine and veterinary medicine.

Quantitative analysis of hyperforin and adgerforin in extracts was performed using reverse phase high performance liquid chromatography (RP HPLC) using an Agilent Technologies) chromatograph (Agilent 1200 Infinity series)), manufactured in the USA, under the following conditions: mobile phase (A) - 1% aqueous solution of formic acid, mobile phase (B) - ethanol in a linear gradient feed mode; chromatographic column - Supelco Ascentis express C18, column dimensions 100 mm × 4.6 mm, particle size 2.7 μm; the speed of the mobile phase is 0.5 ml / min; chromatographic column temperature + 35 ° C; sample volume - 1 μl. Mass spectrometric detection was carried out in the ESI mode (ionization by spraying in an electric field) with a positive scanning mode in the mass range 200-1000 Da, MM-APCI, 50 V. Before quantitative analysis, the exact weighed portion of the extract was dissolved in ethanol 96% vol, and centrifuged at 13000 rpm for 5 minutes The ethanol solution was drained and analyzed.

Description of drawings:

FIG. 1 - hyperforin yield depending on the time of circulation of the extractant.

FIG. 2 - chromatogram of the extract obtained using Novec 7100 at an analytical wavelength of 275 nm, designation: I - hyperforin; II - agiperforin.

FIG. 3 - characteristic of hyperforin in the extract obtained using Novec 7100.

FIG. 4 - characteristics of adgerforin in the extract obtained using Novec 7100.

Example 1

A portion of the ground grass of St. John's wort perforated with a mass of 5.00 g (particle fraction 0.1-0.5 mm) is placed in a Soxhlet circulation extractor. The raw material is poured with extractant 1: 7.5 wt./mass., I.e. 37.5 g (25.0 ml) and start the extraction process within 1 hour. The resulting extract was evaporated at a temperature of 65 ° C, to obtain a dry extract, and the extractant was condensed. The raw materials are not removed from the extractor, heated to 65 ° C, purged with air until complete removal and condensation of the extractant. The regenerated extractant is combined with the bulk of the extractant and reused for extraction of a new batch of raw materials.

Some physico-chemical, environmental and toxic indicators of supercritical carbon dioxide and Novec 7100 fluid for comparison are shown in table 1.

As can be seen from the data in Table 1, the Novec 7100 low-boiling liquid has a number of advantages, ceteris paribus, compared to similar indicators of carbon dioxide in the supercritical state, namely, the Novec 7100 boiling point is 61 ° С, i.e. under normal conditions, it is a liquid, the heat capacity is half as much (1183 <2961 J / (kg⋅K)), the heat of vaporization is 28% less than ck-CO2 (112 <155 kJ / kg).

The comparison results for the main features of the prototype and new technology are shown in table 2.

As can be seen from the data in table 2, the results of comparing the new technology over the technology of the prototype demonstrates its advantages, namely: the absence of special equipment working under pressure, the working pressure is 1 atm., The temperature of the condenser is 20-30 ° C, the absence of energy costs for condensing the extractant vapor , while in one hour of circulation of the solvent, the yield of hyperforin at the level of 87 ± 4% is achieved.

The loss of extractant amounted to 1.5 ± 0.2 g (1.0 ± 0.1 ml), which is equivalent to 4% of its initial amount.

Example 2

The kinetics of hyperforin yield versus Novec 7100 solvent circulation time is shown in FIG. 1. The conditions are the same as in example 1.

As can be seen from the data of FIG. 1, the yield of hyperforin by the first hour of extraction reaches 87 ± 4%, by the second hour 95 ± 5%, and by the third of its limit value 99 ± 5%. Therefore, the optimal extraction time can be considered a range from 1 to 2 hours of extraction.

Example 3

In FIG. 2 and FIG. 3 shows an RP-HPLC chromatogram of the extract obtained using Novec 7100 in Example 1.

As can be seen from FIG. 2, two basic substances (hyperforin and adiperforin) are present in the extract obtained using Novec 7100 solvent, the parameters of which are presented in FIG. 3 and FIG. 4.

Н., Sturm S., Haslinger Е., Schmidt Р.С., Hadacek F. (2004). Characterization of supercritical fluid extracts of St. John's Wort (Hypericum perforatum L.) by HPLC-MS and GC-MS. European Journal of Pharmaceutical Sciences, 21(4), 453-463 (doi: 10.1016/j.ejps.2003.10.021)).As seen in FIG. 3 and FIG. 4, the presence of two dominant substances was experimentally proved in the extract: hyperforin (70.3 ± 3.5%) and adderforin (5.1 ± 0.2%), while their experimentally found values of the absorption maximum in the UV spectrum and molecular weight coincide with those described in the literature (Seger C,

H., Sturm S., Haslinger E., Schmidt R.S., Hadacek F. (2004). Characterization of supercritical fluid extracts of St. John's Wort (Hypericum perforatum L.) by HPLC-MS and GC-MS. European Journal of Pharmaceutical Sciences, 21 (4), 453-463 (doi: 10.1016 / j.ejps.2003.10.021)).

The technical result is an effective and environmentally friendly method for the isolation of hyperforin and adgerforin from St. John's wort perforated simple to perform under ordinary conditions, without special and complex equipment.

Thus, the method of isolating hyperforin and adgerforin from perforated St. John's wort grass allows one to isolate hyperforin and adgerforin from ground herb hypericum perforated with methyl perfluorobutane (Novec 7100) at minimal energy consumption, normal pressure, using standard pharmaceutical equipment or chemical essential oil enterprises using environmentally friendly technology.

Claims (1)

A method of isolating hyperforin and adgerforin from perforated Hypericum herb, including crushing the Hypericum perforatum perforated, extracting the crushed material with an organic solvent, evaporating the solvent, characterized in that an easily boiling liquid is used as an organic solvent - perfluorobutane methyl ether at a ratio of raw material, solvent 1: 1 5-9.0 wt./mass., The extraction of raw materials is carried out by the solvent circulation method for 1-3 hours, the resulting extract is evaporated at temperatures 65 ° C to obtain the final product.

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