Purification and characterization of hydroperoxide lyase from amaranth tricolor (Amaranthus mangostanus L.) leaves

Hydroperoxide lyase (HPL) was extracted from amaranth tricolor leaves using Triton X-100, and purified to electrophoretic homogeneity by ammonium sulfate precipitation, ion-exchange chromatography, hydrophobic interaction chromatography and hydroxyapatite chromatography. The purified HPL preparation consisted of a single band and spot with a molecular mass of about 55 kDa as shown in SDS-PAGE and 2-D PAGE, respectively; the isoelectric point was found to be about 5.4. The maximum activity of the enzyme was observed at pH 6.0 and 25 A degrees C, respectively. The HPL showed higher activity against 13-hydroperoxy-linolenic acid compared to 13-hydroperoxy-linoleic acid. K (m) value for 13-hydroperoxy-linolenic acid was 62.7 mu M, and the corresponding V (max) was 178.5 mu M min(-1). The activity of HPL was significantly inhibited by nordihydroguaiaretic acid, HgCl(2) and 2(E)-hexenal but not by EDTA and hexanal.