Identification of Novel Prostaglandin-Containing Glycerophospholipids in vivo

Abstract

We recently described the formation of a series of prostaglandin (PG)F₂-like compounds in vivo by a non-cyclooxygenase mechanism involving free radical catalyzed peroxidation of arachidonic acid. We now examined the possibility that these prostanoids may be formed from acylated arachidonic acid on phospholipids (PL). An initial suggestion that PGF₂-like compounds may exist acylated to PL emerged from the finding that levels of free PGF₂-like compounds detected following alkaline hydrolysis of liver PL from control rats and rats treated with CCI₄ to induce lipid peroxidation were over 100-fold higher in the treated rats. These increased levels were unaffected by treating the PL with reducing agents or butylated hydroxytoluene (BHT), suggesting that they did not arise from autoxidation of precursors ex vivo. In addition, the appearance of free PGF₂-like compounds in the circulation lagged behind the increase in apparent PL-associated PGF₂-like compounds in the liver, suggesting that the free PGF₂-like compounds arose secondarily from hydrolysis of PL. Further, free PGF₂-like compounds were released following incubation of PL with bee venom but, interestingly, not with snake venom phospholipase A₂. Normal phase HPLC of liver PL from CCI₄ treated rats resulted in the elution of PL which yielded PGF₂ compounds upon hydrolysis at a much more polar retention time than non-oxidized phosphatidylcholine (PC). Liquid secondary ionization mass spectrometric analysis of these more polar PL confirmed that they were PGF₂-containing species of PC. Thus we report the novel finding that PGF2-like compounds are actually formed in Situ, on PL and then subsequently released in free form, presumably by phospholipases.