A method of fast separation of lignin peroxidases using convective interaction media disks.
The HPLC separation of lignin peroxidase isoenzymes using Convective Interaction Media disks containing quaternary amine and diethylaminoethyl ion-exchange active groups is proposed. In contrast to standard HPLC procedures the separation can be performed within a few minutes without considerably affecting the separation resolution. The method is reproducible and gives a linear response of integrated peak area to protein concentration for all measured isoenzymes. The separation resolution is retained unchanged by applying crude culture filtrate instead of a sample previously frozen and dialyzed. The optimized method might therefore be used for on-line monitoring of lignin peroxidase isoenzyme composition during fermentation. On the other hand, the proposed method is comparable in time to the original method of lignin peroxidase activity measurement (proposed by Tien and Kirk), providing additionally the isoenzyme composition.
Faculty of Chemistry and Chemical Technology, University of Ljubljana, Ljubljana, Askerceva 5, 1001, Slovenia. helena.podgornik@uni-lj.si
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D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods* / Q000706:statistics & numerical data; D005069:Evaluation Studies as Topic; D007525:Isoelectric Focusing; D007526:Isoelectric Point; D007527:Isoenzymes / Q000737:chemistry / Q000302:isolation & purification*; D010544:Peroxidases / Q000737:chemistry / Q000302:isolation & purification*; D020075:Phanerochaete / Q000201:enzymology; D015203:Reproducibility of Results
D007527:Isoenzymes; D010544:Peroxidases; C042858:lignin peroxidase