Comparison of fatty acid alpha-oxidation by rat hepatocytes and by liver microsomes fortified with NADPH, Fe3+ and phosphate.
Rat liver microsomes, when fortified with NADPH, Fe3+ and phosphate, can catalyze the oxidative decarboxylation (alpha-oxidation) of 3-methyl-substituted fatty acids (phytanic and 3-methylheptadecanoic acids) at rates that equal 60-70% of those observed in isolated hepatocytes (Huang, S., Van Veldhoven, P.P., Vanhoutte, F., Parmentier, G., Eyssen, H.J., and Mannaerts, G.P., 1992, Arch. Biochem. Biophys. 296, 214-223). In the present study we set out to identify and compare the products and possible intermediates of alpha-oxidation formed in rat hepatocytes and by rat liver microsomes. In the presence of NADPH, Fe3+ and phosphate, microsomes decarboxylated not only 3-methyl fatty acids but also 2-methyl fatty acids and even straight chain fatty acids. The decarboxylation products of 3-methylheptadecanoic and palmitic acids were purified by high-performance liquid chromatography and identified by gas chromatography/mass spectrometry as 2-methylhexadecanoic and pentadecanoic acids, respectively. Inclusion in the incubation mixtures of glutathione plus glutathione peroxidase inhibited decarboxylation by more than 90%, suggesting that a 2-hydroperoxy fatty acid is formed as a possible intermediate. However, we have not yet been able to unequivocally identify this intermediate. Instead, several possible rearrangement metabolites were identified. In isolated rat hepatocytes incubated with 3-methylheptadecanoic acid, the formation of the decarboxylation product, 2-methylhexadecanoic acid, was demonstrated, but no accumulation of putative intermediates or rearrangement products was observed. Our data do not allow us to draw conclusions on whether the reconstituted microsomal system is representative of the cellular alpha-oxidation system.(ABSTRACT TRUNCATED AT 250 WORDS)
Huang
S
S
Katholieke Universiteit Leuven, Afdeling Farmacologie, Belgium.
Van Veldhoven
P P
PP
Asselberghs
S
S
Eyssen
H J
HJ
de Hoffmann
E
E
Mannaerts
G P
GP
D000818:Animals; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D005227:Fatty Acids / Q000378:metabolism*; D005290:Ferric Compounds / Q000378:metabolism / Q000494:pharmacology*; D008401:Gas Chromatography-Mass Spectrometry; D005978:Glutathione / Q000494:pharmacology; D005979:Glutathione Peroxidase / Q000494:pharmacology; D008099:Liver / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects* / Q000378:metabolism*; D009249:NADP / Q000378:metabolism / Q000494:pharmacology*; D010084:Oxidation-Reduction; D010710:Phosphates / Q000378:metabolism / Q000494:pharmacology*; D051381:Rats; D017208:Rats, Wistar; D013379:Substrate Specificity
D005227:Fatty Acids; D005290:Ferric Compounds; D010710:Phosphates; D009249:NADP; D005979:Glutathione Peroxidase; D005978:Glutathione