PURIFICATION OF LACTIC-ACID DEHYDROGENASE FROM BOVINE HEART CRUDE EXTRACT BY COUNTERCURRENT CHROMATOGRAPHY

To test the utility of counter-current chromatography in purifying pro teins, lactic acid dehydrogenase (LDH) was extracted from a crude bovi ne heart filtrate using a cross-axis coil planet centrifuge. The purif ication was performed with several polymer phase systems composed of 1 6% (w/w) poly(ethylene glycol) (PEG) 1000-12.5% (w/w) potassium phosph ate buffers and 4.4% (w/w) PEG 8000-7.0 (w/w) dextran T500 at pH value s ranging from 6.5 to 11.0. The best purification was achieved using P EG 1000-potassium phosphate buffer system at pH 7.3 by eluting the upp er phase at 1.0 ml/min. Fractions were analyzed by LDH enzymatic activ ity and sodium dodecyl sulfate slab gel electrophoresis (SDS-PAGE), Th e LDH was purified directly from bovine heart crude extract within 3 h . (C) 1997 Elsevier Science B.V.