Purification of lactic acid dehydrogenase from bovine heart crude extract by counter-current chromatography.

To test the utility of counter-current chromatography in purifying proteins, lactic acid dehydrogenase (LDH) was extracted from a crude bovine heart filtrate using a cross-axis coil planet centrifuge. The purification was performed with several polymer phase systems composed of 16% (w/w) poly(ethylene glycol) (PEG) 1000-12.5% (w/w) potassium phosphate buffers and 4.4% (w/w) PEG 8000-7.0 (w/w) dextran T500 at pH values ranging from 6.5 to 11.0. The best purification was achieved using PEG 1000-potassium phosphate buffer system at pH 7.3 by eluting the upper phase at 1.0 ml/min. Fractions were analyzed by LDH enzymatic activity and sodium dodecyl sulfate slab gel electrophoresis (SDS-PAGE). The LDH was purified directly from bovine heart crude extract within 3 h.