SummaryAn efficient procedure for the synthesis of 3-hydroxyoxylipins labelled with tritium on all double bond positions is reported. The synthetic scheme involves a joint route for the formation of tetraacetylenic precursors followed by stereoselective reduction of the triple bonds either with hydrogen or tritium. The final tritiated products were obtained with specific activities ranging from 1.65 to 1.80 Ci/mmol. Copyright # 2003 John Wiley & Sons, Ltd.Key Words: tritium labelled fatty acids; 3(R)-hydroxy-oxylipins; 3(R)-HETE; 3(R),18(R/S)-DiHETE Introduction 3(R)-Hydroxy fatty acid derivatives of AA, a group of biologically active commonly named 3(R)-hydroxy-oxylipins, were identified in some fungal species (Dipodascopsis uninucleata and Mucor genevensis) AA was utilized as a substrate. 3(R)-hydroxy-oxylipins have been shown to be potent microbial growth factors 4,5 and to play a crucial role in the morphogenesis of Candida albicans.4 Moreover, 3-HETE was found to be a strong proinflammatory lipid mediator, which modulates several human neutrophil functions, such as chemotaxis and phagocytosis. Recently, a novel AA metabolite -3,18-DiHETE -has been identified in Candida albicans. 4Moreover, its formation was strongly suppresed by salicylic acid and its acetylated derivative. Despite progress in the research of 3(R)-hydroxyoxylipins, a number of open issues on their role in intracellular signal transduction pathways needs to be clarified. The site of the formation of 3,18-DiHETE still remains to be established, and whether its formation affects the pattern of endogenous 3(R)-hydroxy-oxylipins, and if the various oxylipins reveal different biological activities. The utilization of radiolabelled analogues will facilitate the detection of small changes in metabolic products. Hence, efficient synthetic methods for their preparation are required to understand the extensive biological role of 3(R)-hydroxy-oxylipins in mammalian cells. In the present study we describe simple and effective method for the synthesis of 3(R)-HETE, and 3(R),18(R/S)-DiHETE, selectively labelled with tritium on all double bond positions.Experimental 1 H and 13 C NMR spectra for compounds 5-7 were recorded either on a Bruker MSL 200 or Bruker MSL 300 spectrometers in CDCl 3 as solvent. Chemical shifts are referenced to tetramethylsilane as an internal standard for 1 H NMR or to the deuterium lock signal of CDCl 3 (d 13 C=77.19 ppm). HPLC analysis was carried out on a Shimadzu LC-10Avp liquid chromatograph connected to a SPD-10Advp UV detector. RP-HPLC analysis was performed on a Nucleosil C18-column; 250 Â 4 mm, 5 mm particle size (Machery-Nagel, D .u uren, Germany) with different solvent systems: MeOH/H 2 O (95:5, by vol.) and a flow rate of 1 ml/min were used for analysis of compounds 6, 8 and MeOH/H 2 O/Ac (85:15:0.1, by vol.) for acid 2 and MeOH/H 2 O/Ac (75:25:0.1, by vol.) for dihydroxy acids 7 and 9. Preparative HPLC was carried out on a Lichrospher 100 RP18 column; 250 Â 22.5 mm, 10 mm particle size (Knauer, Berlin, Germany) with MeO...