Detection of an enol intermediate in the hydroperoxide lyase chain cleavage reaction.
Guava (Psidium guajava) hydroperoxide lyase (HPL) preparations were incubated with [1-(14)C](9Z,11E,13S,15Z)-13-hydroperoxy-9,11,15-octadecatrienoic acid for 1 min at 0 degrees C, followed by rapid extraction/trimethylsilylation. Analysis of the trimethylsilylated products by gas chromatography-mass spectrometry and radio-high-performance liquid chromatography revealed a single predominant (14)C-labelled compound, identified by its (1)H-nuclear magnetic resonance, ultraviolet and mass spectra as the trimethylsilyl ether/ester of (9Z,11E)-12-hydroxy-9,11-dodecadienoic acid. Longer time incubations afford smaller yield of this enol due to its partial tautomerization into (9Z)-12-oxo-9-dodecenoic acid. The data obtained demonstrate that formation of (9Z)-12-oxo-9-dodecenoic acid in the HPL reaction is preceded by unstable enol oxylipin, and further suggest that hemiacetals are the true products of HPL catalysis.
Grechkin
Alexander N
AN
Kazan Institute of Biochemistry and Biophysics, Russian Academy of Sciences, Kazan, Russia. grechkin@sci.kcn.ru
Mukhtarova
Lucia S
LS
Hamberg
Mats
M
D000446:Aldehyde-Lyases / Q000378:metabolism*; D002250:Carbon Radioisotopes / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D005233:Fatty Alcohols / Q000032:analysis* / Q000378:metabolism; D008401:Gas Chromatography-Mass Spectrometry; D007850:Lauric Acids; D008042:Linolenic Acids / Q000378:metabolism; D008054:Lipid Peroxides / Q000378:metabolism; D010940:Plant Proteins / Q000378:metabolism; D031647:Psidium / Q000201:enzymology
D002250:Carbon Radioisotopes; D005233:Fatty Alcohols; D007850:Lauric Acids; D008042:Linolenic Acids; D008054:Lipid Peroxides; D010940:Plant Proteins; C057375:13-hydroperoxy-9,11,15-octadecatrienoic acid; C031263:12-hydroxydodecanoic acid; D003577:Cytochrome P-450 Enzyme System; D000446:Aldehyde-Lyases; C054540:hydroperoxide lyase