Novel reactive perstraction system applied to the hydrolysis of penicillin G.
The activity of penicillin acylase has been studied in aqueous and organic solvents, as free enzyme as well as immobilized within the membrane of liquid-core capsules. The activity of the enzyme is inhibited by the accumulation of the products of the hydrolysis reaction, namely phenyl acetic acid (PAA). In order to overcome this inhibition a range of organic solvents were tested for use in in situ product recovery. Of these solvents dibutyl sebacate (DBS) was chosen due to the rapid extraction rate, the high logP and to facilitate capsule production. The extraction efficiency at pH 3.5 for PAA was >80% for phase ratios of >50% free solvent with partition coefficients of 8 and 0.7 for PAA and penicillin G (PenG), respectively, thereby showing that PAA could be selectively extracted at pH 3.5 and 25 degrees C. Liquid-core capsules containing DBS were shown to efficiently remove PAA selectively and the PAA could be effectively back-extracted and the capsules re-used in a three-stage process resulting in high product separation. Immobilization of penicillin acylase onto the capsule membranes resulted in increased operational stability of the enzyme and a very high enzyme activity. Over 53.3% of the PAA formed could be recovered in the capsule core with a concentration over sevenfold higher than in the aqueous phase. Higher extraction efficiencies could be obtained by varying the substrate concentration and number of capsules. The enzyme immobilized on capsules could be stored for over 4 months at pH 8 and 4 degrees C with no loss of activity. Over 80% of the initial activity could be recovered over five repeated batch cycles of the bioconversion process. The importance of capsular perstraction and reactive capsular perstraction has been clearly demonstrated.
Wyss
A
A
Laboratory of Chemical and Biochemical Engineering, Swiss Federal Institute of Technology (EPFL), CH-1015 Lausanne, Switzerland.
Seitert
H
H
von Stockar
U
U
Marison
I W
IW
D003998:Dicarboxylic Acids / Q000737:chemistry*; D004789:Enzyme Activation; D004800:Enzymes, Immobilized / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D009930:Organic Chemicals / Q000737:chemistry*; D010399:Penicillin Amidase / Q000737:chemistry*; D010400:Penicillin G / Q000737:chemistry*; D010648:Phenylacetates / Q000737:chemistry* / Q000302:isolation & purification; D012997:Solvents / Q000737:chemistry*
D003998:Dicarboxylic Acids; D004800:Enzymes, Immobilized; D009930:Organic Chemicals; D010648:Phenylacetates; D012997:Solvents; C055481:dibutyl sebacate; D010399:Penicillin Amidase; C025136:phenylacetic acid; D010400:Penicillin G