Chemistry: molecular biology and microbiology – Process of utilizing an enzyme or micro-organism to destroy... – Treating animal or plant material or micro-organism
Patent
1988-06-30
1990-12-11
Rosen, Sam
Chemistry: molecular biology and microbiology
Process of utilizing an enzyme or micro-organism to destroy...
Treating animal or plant material or micro-organism
260403, C11C 100, C07F 910
Patent
active
049770918
DESCRIPTION:
BRIEF SUMMARY
FIELD OF THE INVENTION
The present invention relates to the art of preparation of lipids and, more particularly, to a method for preparing phosphatidylinositol from a biological object.
Phosphatidylinositol pertains to the class of phospholipids and is useful as a biochemical reagent: for the preparation of membrane system simulating processes in live organisms; for the formation of liposomes serving for transferring foreign genetic information into cells of live organisms; for transportation of foreign proteins, vaccines and enzymes through membrane barriers; for delivering pharmaceutical preparations and other agents into the target tissues.
PRIOR ART
Known in the art are a number of methods for preparing phosphatidylinositol from various biological objects such as heart and brains of animals, from bacteria and seeds. Also known in the art is a method for preparing phosphatidylinositol from yeast (cf. Preparative Chemistry of Lipids, ed. by L. D. Bergelson and E. V. Dyatlovitskaya, 1981, Moscow, "Nauka" Publishers, p. 178).
The method for preparing phosphatidylinositol from yeast comprises autolysis of yeast by means of toluene for 5 hours, extraction of phospholipids with a mixture of chloroform:methanol (1:1) for 7 hours; filtration of the extract with washing of the filter cake with the abovementioned mixture of the organic solvents; stirring of the filtrate with distilled water for 20 hours; separation of the filtrate into chloroformic and aqueous phases; evaporation of the chloroformic phase; treatment of the evaporated chloroformic phase with isopropanol and its subsequent filtration through a glass filter. Then phospholipids are separated from the resulting filtrate in a column with silica gel by way of a successive, elution thereof first with 5-7 litres of a mixture chloroform-methanolammonia (80:20:2) and then with 1-2 liters of ethanol and in this manner ammonium salt of phosphatidylinositol is recorvered which is then transformed into sodium salt.
This method necessitates a great consumption rate of expensive organic solvents. Thus, for the preparation of 1 g of phosphatidylinositol it is necessary to employ more than 5 l of chloroform, more than 3 l of methanol, about 1 l of ethanol and other solvents. The method involves many labour-intensive operations taking a long time. All this considerably adds to the costs of the final product (according to the Calbiochem catalogue 1 g of phosphatidylinositol costs 6,950 U.S. dollars). Operation with great amounts of organic solvents creates harmful conditions of labour which hinders provision of normal environment for the operating personnel.
DISCLOSURE OF THE INVENTION
It is an object of the present invention to provide such a method for preparing phosphatidylinositol which would make it possible to substantially reduce the rate of consumtion of organic solvents, to lower labour-intensity and time consumption, improve labour conditions and considerably reduce costs of the product.
This object is accomplished by a method for preparing phosphatidylinositol from a biological object by way of its homogenization and extraction of the product which, according to the present invention, comprises:
homogenization of a biological object in an aqueo-saline solution with a molar concentration of from 0.005 to 1 M at a pH within the range of from 5.0 to 11.04;
separation of the resulting homogenizate into a cell-free water-salt phase containing particles of a protein-phosphatidylinositol complex and a precipitate containing non-broken cells and particles of other phospholipids by centrifugation at a gravity acceleration of from 3,000 to 30,000 g or by filtration on a filter with a pore size of from 3 to 50 .mu.m;
recovery of particles of the protein-phosphatidylinositol complex from the cell-free water-salt phase;
extraction of phosphatidylinositol from the recovered particles of the protein-phosphatidylinositol complex.
The recovery of the particles of the protein-phosphatidylinositol complex from the cell-free water-salt phase can be effected by gel-chro
REFERENCES:
patent: 2397874 (1946-04-01), Lloyd et al.
patent: 2508624 (1950-05-01), Singer et al.
patent: 2855416 (1958-10-01), Hennessy et al.
patent: 4169090 (1979-09-01), Murray
Preparative Chemistry of Lipids.
Dilbarkanova Rsai
Gilmanov Murat K.
Sultanbaev Beibyt E.
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