The method of obtaining inhibitor bromeline

 

(57) Abstract:

Usage: the invention relates to biotechnology and concerns a method for obtaining inhibitor bromeline. The essence of the method lies in the fact that proliferating the tops of the fruit of the pineapple is extracted with water in the ratio of raw materials - solvent 1:(2-4) and then hold clearing sepharose with immobilized cysteine proteinase type in acetate buffer pH of 4.4 to 4.6, - laundering of the sorbent was carried out in 0.1 M phosphate buffer pH of 7.6 to 7.8, containing 0.5 m sodium chloride, and the target product elute in 0.01 N sodium hydroxide solution containing 0.1 m sodium chloride. The technical result is to simplify the method.

The invention relates to biotechnology, namely to methods for enzyme inhibitors, in particular inhibitors of cysteine proteases from plant material inhibitor bromeline.

Inhibitors of cysteine proteins of plants find wide application in medicine and scientific research. These inhibitors found in the sources, both animal and vegetable origin. They were isolated from the epidermis, internal organs, from the blood plasma of animals and humans. Inhibitors found in the tubers of cards is hibitor bromeline (cysteine proteinase pineapple juice from the stems of pineapple), which consists in grinding the stems of pineapple, wringing from them juice with subsequent two-stage precipitation with acetone. Then the acetone powder of the stem of the pineapple is subjected to gel chromatography on Sephadex G-75 and ion-exchange chromatography on DEAE Sephadex at pH of 7.55 [1]

The known method is complex, involving two-stage precipitation with acetone and subsequent clearance of the drug inhibitor in two stages. In addition, the use of acetone increases the risk of production, especially in hot climates.

In connection with the above, the technical result will be to simplify the method.

The proposed method for inhibitor bromeline is the following. As a source of raw materials use waste from industrial processing of pineapples (proliferous the tops of the fruit of the pineapple), extraction of proteins is carried out by extraction with water. When this ratio of the raw material and the extractant is 1:(2-4) g/ml of Raw materials with the extractant is placed in a homogenizer, crushed and then extracted for one hour at room temperature, stirring with a mechanical stirrer. The obtained mass press, centrifuged and the clarified protein solution, podcastanswerman.com, balanced 0.1 M acetate buffer, pH 4,60. Nonspecific adsorbed proteins remove 0.1 m phosphate buffer, pH of 7.6 to 7.8, containing 0.5 M NaCl. The evolution of the associated column of inhibitor implement 0.01 n NaOH solution, containing 0.1 M NaCl, pH of 11.5 to 12.0. It should be noted the importance of compliance with all these conditions allocation of the inhibitor. Numerous attempts to allocate inhibitor under conditions different from those described above did not lead to the obtaining of the inhibitor. Thus, the binding of the inhibitor bromeline carried out exclusively at pH of 4.4 to 4.6, and the elution of the inhibitor from the medium can be performed only in a narrow range of high alkaline pH of 11.5 to 12.0.

P R I m e R 1. To 250 g of crushed mass of proliferating tops pineapple add 500 ml of water and conducting further ground in a homogenizer. The extraction continued with stirring with a mechanical stirrer for one hour at room temperature with the addition of 500 ml of water. The resulting mass is squeezed through several layers of gauze, center at 3000 xg for one hour. Get 920 ml of a solution containing 1500 mg of protein with specific activity 0,0086 I. IU/mg. Per unit of inhibitory activity (I. units) made this quantity of inhibitor that th and applied on the column with 50 ml sepharose with immobilized bromelain, balanced 0.1 M acetate buffer, pH 4,60. Nonspecific adsorbed proteins remove 0.1 M phosphate buffer, pH of 7.60, containing 0.5 M NaCl. The evolution of the bound inhibitor with immobilized bromeline exercise 0.01 n NaOH solution, containing 0.1 M NaCl. The output of the inhibitor reaches 94% increase in inhibitory activity compared to the original 389 times. Molecular weight inhibitor of about 6 kDa purified inhibitor effect on papain, fitsin, bromelain, trypsin.

P R I m m e R 2. The method is carried out according to example 1, but the weight ratio of raw material:solvent is 1:4, and the sorbent using immobilized on sepharose 6V papain. As a result, the output of the inhibitor is 90% increase in inhibitory activity compared to the original 388 times.

Summary information about the efficiency of the method is reflected in the table.

The METHOD of OBTAINING INHIBITOR BROMELINE by extraction of the crushed waste from industrial processing of pineapples with subsequent chromatographic isolation of target product, wherein the proliferating use the tops of the fruit of the pineapple, the extraction was carried out with water at a mass ratio of raw material: solvent 1:2-4, and highlighted steinbaugh type in acetate buffer pH 4,4-4,6, laundering sorbent is carried out in 0.1 M phosphate buffer pH of 7.6 to 7.8, containing 0.5 M sodium chloride, and the elution of the target product of 0.01 N. the solution of sodium hydroxide containing 0.1 M sodium chloride at a pH of 11.5 to 12.0.

 

Same patents:

The invention relates to biochemistry

The invention relates to methods of processing of medicinal raw materials, particularly to a method of processing plantain production pentaglycine - treatment for stomach ulcers

The invention relates to pharmaceutical industry and relates to a method of obtaining copper derivative of chlorophyll

The invention relates to chemical-pharmaceutical and cosmetic industries and concerns extractants or liquid solvents used for the extraction of bioactive components from plant materials and the use of extractants in cosmetic compositions

The invention relates to medicine and for the destruction of tumor tissue

The invention relates to the extraction equipment for the production of hydro-alcoholic extracts of plant material, in particular of aromatic and biologically active substances for alcohol balms
The invention relates to pharmaceutical industry and relates to the production of biologically active drug from green walnuts

FIELD: medicine, cardiology, phytotherapy, pharmacy.

SUBSTANCE: invention relates to the field in development of an agent of vegetable origin used for improvement of the functional state and prophylaxis of organic alterations in cardiovascular system, in particular, atherosclerosis. The biologically active supplement used for prophylaxis of cardiovascular diseases, in particular, atherosclerosis comprises clover dry extract, haw-thorn flowers powder, ascorbic and nicotinic acids, and accessory substances taken in the definite ratio. The biologically active supplement is made in the form of tablet or capsule. No adverse responses observed in intake of the supplement. The blood cholesterol level was reduced to the normal value in 68% of patients and reducing morbidity with acute respiratory-viral infections has been noticed. In patients with ischemic heart disease with cholesterolemia the improvement of the state is noted on the background of conventional therapy: the moderate reducing the arterial pressure value in its elevation, improved in the state of health, enhanced working capacity, reduced retrosternal pains, ear noise, vertigo, headache.

EFFECT: valuable medicinal properties of supplement.

1 ex

FIELD: medicine, hematology.

SUBSTANCE: invention relates to applying elm-leaf dropwort (Filipendula) above-ground part 40% alcoholic tincture as an agent stimulating erythroidal and granulocytic hemopoiesis stems in cytostatic myelorepressions. Agent promotes to effective stimulation of erythroidal and granulocytic hemopoiesis stems in cytostatic myelodepression. Invention is used for correction of disorders arising in blood system in administration of cytostatic preparations.

EFFECT: valuable medicinal properties of agent.

3 tbl

FIELD: organic chemistry, chemical technology, agriculture.

SUBSTANCE: invention relates to a method for preparing the preparation comprising triterpenic acid water-soluble salts and additionally added protein-containing product and vegetable raw, the source of triterpenic acids taken in the following ratio of components, wt.-%: protein-containing product, 10-17; triterpenic acid sodium salts, 4-5, and vegetable raw, the balance. Method involves mixing triterpenic acid-containing vegetable raw with the protein-containing product taken in the ratio = (9-11):(1-2), mechanical-chemical treatment of this mixture in activator device, mixing of prepared semi-finished product with sodium carbonate or sodium hydrocarbonate taken in the ratio = (92-97):(3.5-8.3) and repeated treatment in the activator device. Method involves applying flow-type ball vibration-centrifugal or ellipse-centrifugal mills as the activator device that provide the acceleration of milling bodies up to 170-250 m/c2 and time for treatment for 1.5-3 min. Invention provides simplifying the process and the complex processing waste in lumber industry.

EFFECT: improved preparing method.

6 cl, 1 tbl, 6 ex

FIELD: beauty products manufacturing.

SUBSTANCE: the present innovation deals with manufacturing cosmetic composition that contains solubilized cumic alcohol of a certain formula, glucose, ascorbic acid or compound which is split in skin up to glucose or ascorbic acid and a cosmetically acceptable carrier. Components should be taken at a certain quantitative ratio. The suggested composition improves expression of transglutaminase-1 and ceramides in skin cells and enhances absorption of glucose and ascorbic acid by cells.

EFFECT: higher efficiency.

1 cl, 5 ex, 12 tbl

FIELD: organic chemistry, natural compounds, medicine, oncology.

SUBSTANCE: invention represents new saponin mixtures used for inhibition of initiation and activation of mammalian epithelial cell in pre-malignant or malignant state, for stimulation of apoptosis of mammalian malignant cell, prophylaxis of anomalous proliferation of mammalian epithelial cell, for treatment of inflammatory and regulation of angiogenesis in mammal. These mixtures are isolated form plants of species Acacia victoriae. Also, invention relates to methods for their applying. These compounds can comprise triterpene component, such as acacic or oleanolic acid to which oligosaccharides and monoterpenoid components are joined. Mixtures and compounds elicit properties associated with regulation of apoptosis and cytotoxicity of cells and strong anti-tumor effect with respect to different tumor cells.

EFFECT: valuable medicinal properties of compositions.

43 cl, 53 tbl, 50 dwg, 44 ex

FIELD: pharmaceutical industry and technology, pharmacy.

SUBSTANCE: invention relates to a method for preparing extract based on walnuts at milkwax stage of ripeness. Method for preparing extract based on walnuts involves grinding walnuts at milkwax stage of ripeness taken with pericarps, drying to air-dry state, mixing with wild rose fruits ground and dried to air-dry state and taken in the definite ratio of components followed by treatment with aviation kerosene and stirring at room temperature. Method provides preparing extract with the higher content of biologically valuable substances and to increase the assortment of vegetable supplements.

EFFECT: improved preparing method.

2 tbl

FIELD: medicine.

SUBSTANCE: method involves using air-dried vegetable raw. Hollow horse-mint (Monarda) raw treated by steam is extracted with vegetable oil in the ratio raw : extract = 1:10 at heating for 5 h on water bath at temperature 60-70oC followed by filtration. Method provides the possibility for year-round manufacturing the hollow horse-mint (Monarda) oily extract eliciting with the high antibacterial and expressed anti-fungal activity.

EFFECT: improved preparing method.

1 tbl, 1 ex

FIELD: medicine, pharmaceutical industry.

SUBSTANCE: invention relates to agents used for treatment of chronic hepatitis. The hepatoprotective agent representing extract prepared from biomass of Maackia amurensis Rupr. et Maxim and obtained by the callus culture method comprises polyphenolic complex consisting of daidzein, retusin, genistein, formononetin, maackianin and medicarpin. The hepatoprotective agent promotes to effective treatment of chronic hepatitis.

EFFECT: valuable medicinal properties of agent.

2 tbl

FIELD: medicine, virology, pharmaceutical industry, pharmacy.

SUBSTANCE: invention proposes the preparation used for treatment of viral hepatitis C that comprises birch bark extract with the content of betulin above 70% and a pharmaceutically acceptable carrier. The preparation is administrated to patient by oral route. The preparation promotes to the effective treatment of viral hepatitis C. Invention can be used in treatment of viral hepatitis C.

EFFECT: valuable medicinal properties of agent.

4 cl, 2 tbl

FIELD: medicine, phytotherapy, pharmacy.

SUBSTANCE: invention relates to preparations of vegetable origin. Invention proposes the preparation eliciting immunomodulating effect and comprising aqueous-alcoholic extracts of the following plants, mas. p. p.: glabrous licorice roots, 5.0-15.0; fenestrate Saint-John's-wort herb 4.0-16.0; dandelion roots, 4.0-16.0; sandy immortelle flowers, 3.5-14.0; senna leaves, 3.5-14.0; thyme herb, 1.0-4.0; snowdon rose (rosewort) rhizomes and roots, 0.05-0.2. Also, invention proposes the preparation comprising condensed or an aqueous-alcoholic extract of above said plants and a filling agent. The preparation can be made as tablet or capsule. The preparation normalizes the blood free radicals content and provides elevating level of T-suppressors with the simultaneous reducing amount of T-helpers in blood of patients in treatment of acute and chronic inflammatory diseases of breathing organs.

EFFECT: valuable medicinal properties of preparation.

2 cl, 3 tbl, 1 ex

Up!