High-performance liquid chromatography-thermospray mass spectrometry of omega-carboxyleukotriene B4 and omega-hydroxyleukotriene B4 from an incubation mixture of human colonic well-differentiated adenocarcinoma homogenate.
A method for the analysis of omega-carboxyleukotriene B4 and omega-hydroxyleukotriene B4 in human colonic carcinoma homogenate is described. The hydroxy groups of the leukotriene metabolite were acetylated by acetic anhydride, and the mixture was partially purified on a Sep-Pak C18 cartridge and analysed by reversed-phase HPLC-thermospray MS. Generally, the base ion, [MH-2(60)]+, is produced through elimination of two acetic acid (60 mass units) molecules from the protonated molecular ion. On selected-ion monitoring, standard curves for omega-carboxy- or omega-hydroxyleukotriene B4 showed a linear relationship over the range 72-1500 pmol. The assay based on selected-ion monitoring was applied to an extract from human colonic carcinoma homogenate. When a homogenate of human colonic well-differentiated adenocarcinoma was incubated with NADPH and leukotriene B4 (60.6 nmol) as a substrate, the conversion of precursor leukotriene B4 to omega-carboxyleukotriene B4 or omega-hydroxyleukotriene B4 was 0.33 or 3.17%, respectively. Based on these results, it is suggested that carcinoma cells themselves or leukocytes at the hostsite in a region of human colonic well-differentiated adenocarcinoma are performing omega-oxidation through NADPH-dependent omega-hydroxylation of leukotriene B4.
Yamane
M
M
Department of Biochemistry, Tokyo Medical College, Japan.
Shimizu
S
S
Abe
A
A
Sugiura
H
H
Miyaoka
M
M
Saitoh
T
T
D000230:Adenocarcinoma / Q000737:chemistry* / Q000473:pathology; D002454:Cell Differentiation; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003110:Colonic Neoplasms / Q000737:chemistry* / Q000473:pathology; D006801:Humans; D006893:Hydroxyeicosatetraenoic Acids / Q000032:analysis*; D007975:Leukotriene B4 / Q000031:analogs & derivatives* / Q000032:analysis; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*
D006893:Hydroxyeicosatetraenoic Acids; D007975:Leukotriene B4; C031571:5,12,20-trihydroxy-6,8,10,14-eicosatetraenoic acid; C031911:20-carboxyleukotriene B4