Partial purification and immobilization of bromelain from pineapple (Ananas comosus) wastes for use in the hydrolysis of racemic arylpropionic esters

Date

10-2000

Degree

Bachelor of Science in Agricultural Chemistry

College

College of Arts and Sciences (CAS)

Adviser/Committee Chair

Milagros M. Peralta

Co-adviser

Marivic S. Lacsamana

Abstract

RECUENCO, MARIAM CAGAYAT. University of the Philippines Los Banos, October 2000. " PARTIAL PURIFICATION AND IMMOBILIZATION OF BROMELAIN FROM PINEAPPLE (Ananas comosus) WASTES FOR USE IN THE HYDROLYSIS OF RACEMIC ARYLPROPIONIC ESTERS."

Adviser: Dr. Milagros M. Peralta

Co-adviser: Dr. Marivic S. Lacsamana

Proteases from the pineapple crown were extracted by 0.05 M phosphate buffer pH 6.5 with 1 mM EDTA. The crude extract was concentrated by acetone precipitation to obtain the acetone precipitate, 80 P. prior to gel filtration chromatography. Fractions within the peak in the elution profile were individually analyzed by denaturing gel electrophoresis to see which fractions have similar profiles. The gel filtration fractions were reduced to four upon pooling namely A (50-56), B (57-63), C (64-67), and D (68-85). Fraction A was 1.50 times more active than the crude enzyme while fractions B, C, and D had purification folds between 4.20-4.60.

SDS-PAGE of the crude enzyme, acetone precipitate (80P) and the gel fractions revealed four major bands with estimated molecular weights of 27.7 kD, 22.2 kD, 17 kD and 12 kD. respectively. confirming molecular weights of the components of bromelain reported in various literature.

In all stages of purification, enzyme aliquots were used to hydrolyze the methyl ester of racemic ibuprofen. The reaction using the crude enzyme was found to produce the highest amount of the product at three days of incubation. All fractions acted on both the R- and S-enantiomers in equal amounts. Immobilization of the enzyme by glutaraldehyde cross-linking increased the reactivity of the enzyme as indicated by the increase in the racemic product. The immobilized enzyme was also used for the hydrolysis of ethyl and butyl esters; however, no acid was detected during HPLC analysis of the hydrolysates

Language

English

Location

UPLB Main Library Special Collections Section (USCS)

Call Number

LG 993.5 2000 A13 R43

Document Type

Thesis

This document is currently not available here.

Share

COinS