Studies on lipase-affinity adsorption using response-surface analysis

doi:10.1042/BA20000081

2010-12-23

PRE

Eliana Setsuko Kamimura

Oscar Medieta

Maria Isabel Rodrigues

Francisco Maugeri

DOI：10.1042/BA20000081

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摘要

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Lipases are widely distributed enzymes that can be obtained from animals, plants and micro-organisms. Coupling lipases with a wide range of substrates allows the opportunity for synthesis of optically pure pharmaceutical preparations, flavour compounds and other food additives. Affinity chromatography owes its power as a purification method to specific biological interactions. Response-surface analysis was chosen to study column efficiency. This method allows the understanding of interactions between variables with advantages over conventional methods, which involve changing one variable while fixing others at certain levels. The aim of this work was to study the influence of the ratio bed height/column diameter (L/D) and superficial velocity (V) on the column efficiency. The experimental design involved the two variables, Un (2-10) and v (1-2 cm/min), at five levels. Lipase was obtained from Geotrichum sp, culture in a complex medium composed of 5% corn-steep liquor, 0.5% NH4NO3 and 1% alive oil at 30 degreesC, with I VVM (air volume/medium volume per min) aeration and 400 rev,/min agitation. Maximum lipase activity was 19 units/ml after almost 9h of fermentation. This lipase could potentially be used in esterification reactions to increase the content of gamma -linolenic acid and to produce bioaromas for food industries. The adsorption assays were carried out in a fixed-bed column with an affinity adsorbent, which was obtained by reaction of a gel with oleic acid as ligand, Breakthrough curves were obtained for all experiments. It has been shown that the lower the values of both Un and v, the higher the column efficiency (maximum 65.43%), Also, it was observed from the response surface that the efficiency reached a minimum at an L/D of around 8.

Keyword:

affinity chromatography

breakthrough curves

factorial design

fixed bed column

Geotrichum sp.