IPF_2α-I: An index of lipid peroxidation in humans

Isoprostanes are prostaglandin isomers produced from arachidonic acid by a free radical-catalyzed mechanism. Urinary excretion of 8-iso-prostaglandin F_2α, an isomer of the PGG/H synthase (cyclooxygenase or COX) enzyme product, prostaglandin F_2α (PGF_2α), has exhibited promise as an index of oxidant stress in vivo. We have developed a quantitative method to measure isoprostane F_2α-I, (IPF_2α-I) a class I isomer (8-iso-PGF_2α is class IV), using gas chromatography/mass spectrometry. IPF_2α-I is severalfold as abundant in human urine as 8-iso-PGF_2α, with mean values of 737 ± 20.6 pg/mg creatinine. Both isoprostanes are formed in a free radical-dependent manner in low density lipoprotein oxidized by copper in vitro. However, IPF_2α-I, unlike 8-iso-PGF_2α, is not formed in a COX-dependent manner by platelets activated by thrombin or collagen in vitro. Similarly, COX inhibition in vivo has no effect on IPF_2α-I. Neither serum IPF_2α-I, an index of cellular capacity to generate the isoprostane, nor urinary excretion of IPF_2α-I, an index of actual generation in vivo, is depressed by aspirin or indomethacin. In contrast, both serum thromboxane B₂ and urinary excretion of its 11-dehydro metabolite are depressed by the COX inhibitors. Although serum S-iso-PGF_2α formation is substantially depressed by COX inhibitors, urinary excretion of the compound is unaffected. Urinary IPF_2α-I is elevated in cigarette smokers compared with controls (1525 ± 180 versus 740 ± 40 pg/mg creatinine; P < 0.01) and is highly correlated with urinary 8-iso-PGF_2α (r = 0.9; P < 0.001). Urinary IPF_2α-I is a novel index of lipid peroxidation in vivo, which can be measured with precision and sensitivity. It is an abundant F₂-isoprostane formed in a free radical- but not COX-dependent manner. Although 8-iso-PGF_2α may be formed as a minor product of COX, this pathway contributes trivially, if at all, to levels in urine. Urinary excretion of both isoprostanes is elevated in cigarette smokers.