Investigation on the enantioselectivity of a partially purified lipase from Candida claussenii towards hydrolysis of a racemic ester

Date

10-1999

Degree

Bachelor of Science in Chemistry

College

College of Arts and Sciences (CAS)

Adviser/Committee Chair

Jamela R. Revilleza

Co-adviser

Milagros M Peralta

Abstract

A lipase from a Candida strain was isolated, purified and then investigated for its enantioselective hydrolysis of a racemic ester of an arylpropionic acid Four strains of Candida species were screened for their extracellular lipase and protease activities Candida claussenii showed the highest specific lipase activity of 312 00 U/mg and was then chosen for the study Production of the crude enzyme was optimized at pH 6 5 and 40°C. Two enzyme purification methods were used ion exchange chromatography (IEC) using DEAE-Sephadex and hydrophobic interaction chromatography (HIC) using phenyl sepharose. The fraction eluted from DEAE-Sephadex with 0.2 M NaCI in phosphate buffer at pH 6.5 showed the highest specific activity resulting in an 11-fold purification On the other hand, the fraction eluted with 0.4 M NaCI in phosphate buffer at pH 6 5 showed a purification factor of 0.27. Analysis of the partially purified enzymes from the two fractions using SDS-PAGE both showed two bands of 59 kD and 67 kD. The HIC was not as efficient as IEC as indicated by the low activities obtained for the HIC fractions. Fractions with high lipase activities eluted from DEAE-Sephadex were used in the hydrolysis of methyl arylpropionate and butyl arylpropionate Hydrolysates were analyzed via high pressure liquid chromatography (HPLC) using a S,S-Whelk-01, a chiral column. Results showed that hydrolysates of the butyl ester using fractions eluted with 0.2 M NaCI and 0 4 M NaCI in phosphate buffer at pH 6.5 exhibited apparent S(+) enantioselectivity Hydrolysis of the methyl ester using the partially purified lipases as catalyst produced no acid

Language

Filipino

Location

UPLB Main Library Special Collections Section (USCS)

Call Number

LG 993.5 1999 C4 A54

Document Type

Thesis

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