Process for preparing lipase

Details Process for preparing lipase Process for preparing lipase

1986-03-17

1988-12-13

Wiseman, Thomas G.

Chemistry: molecular biology and microbiology

Enzyme , proenzyme; compositions thereof; process for...

Hydrolase

435882, C12N 920

Patent

active

047910593

DESCRIPTION:

BRIEF SUMMARY
DESCRIPTION

1. Technical Field
The present invention is concerned with a process for preparing a novel lipase. More particularly, it relates to the process for preparing the novel lipase involving the use of strains of Staphylococcus capitis, which lipase provides free fatty acids and glycerin from triglyceride.
2. Background Art
For the preparation of lipase using microorganisms, particularly bacteria, the processes using bacteria belonging to the genera such as Pseudomonas, Chromo bacterium, Achromobacter, and Corynebacterium are known. However, when lipase derived from these microorganisms is used to hydrolyze fats or oils particularly triglycerides, substantial amounts of monoglycerides and diglycerides are formed in addition to fatty acids and glycerin. This is thought to arise from the recombination of fatty acids and glycerin by the action of lipase or from the termination of hydrolysis by lipase during the monoglyceride-forming stage or the di- glyceride-forming stage. Accordingly, the process for preparing fatty acids using such a known lipase by the hydrolysis of fats or oils has both technical and economical disadvantages.


DISCLOSURE OF THE INVENTION

Accordingly, the present invention provides a lipase for use in the preparation on an industrial scale of fatty acids by the enzymatic hydrolysis of fats or oils in which triglyceride is hydrolyzed so completely that fatty acids and glycerin are formed but virtually no monoglyceride and diglyceride are formed.
The present inventors found that such lipase is produced by Staphylococcus capitis, and they subsequently isolated and purified the same in a pure form.
Consequently, this invention provides a process for preparing a novel lipase characterized by the steps of: culturing a microorganism belonging to Staphylococcus capitis and capable of producing the novel lipase in a culture medium to accumulate the lipase in the fermentation broth and recovering the lipase from the broth. This lipase hydrolyzes triglyceride virtually completely to fatty acids and glycerin so that no monoglyceride and diglyceride are produced.


BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows the activities of the lipase of the present invention for different substrates relative to its activity for olive oil as a substrate, expressed as 100%;
FIG. 2 shows the activities of the lipase of the present invention for different substrates relative to its activity for triolein as a substrate, expressed as 100%;
FIG. 3 shows the effects of pH on the lipase of the present invention, wherein -.cndot.- represents the activity determined by using an 0.1M acetate buffer, -o- represents the activity determined by using an 0.1M phosphate buffer, and -.quadrature.- represents the activity determined by using an 0.1M glycine-NaOH buffer;
FIG. 4 shows the stability of the lipase of the present invention to pH wherein the signs used are as the same as those used in FIG. 3;
FIG. 5 shows the effects of temperatures on the activity of the lipase of the present invention;
FIG. 6 shows the stabilities of the lipase of the present invention to temperatures;
FIG. 7 shows thin-layer chromatograms comparing the lipase of the present invention and that of the prior art with respect to the reaction product;
FIG. 8 is a thin-layer chromatogram showing activities of the lipase of the present invention to different glyceryl oleates; and,
FIGS. 9 and 10 are thin-layer chromatograms showing the products resulting from hydrolysis of triolein with different enzyme solutions (from different strains) of the present invention.


THE BEST MODE FOR CARRYING OUT THE INVENTION



Lipase-producing Microorganisms

The lipase-producing microorganisms used in the process of the present invention are of the species Staphylococcus capitis having the ability to produce lipase. Any bacteria having such a property may be used in the process of the invention. For example, strains of Staphylococcus capitis which are preserved at the American Type Culture Collection (12301 Parklawn Drive, Rockville, Maryland 20852, U.S.A.) and

REFERENCES:
patent: 4565698 (1986-01-01), Yoshizumi et al.
patent: 4670255 (1987-06-01), Yoshizumi et al.
Chemical Abstracts vol. 81, No. 15, p. 112, right column, No. 87058u.

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