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family 1 to 3 are mainly epoxygenases, whereas CYP isoforms from family 4 are mainly -hydroxylases ( 4, 5 ).CYP epoxygenases convert arachidonic acid (AA) to four epoxyeicosatrienoic acid regioisomers (5,8,11,and 14, that function as lipid mediators. EETs produce vascular relaxation, have antiinfl ammatory effects on blood vessels and in the kidney, promote angiogenesis, and protect ischemic myocardium and brain [for review, see ( 6-8 )]. CYP epoxygenases also convert the 3-PUFA eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) to epoxy-derivatives ( 9 ), which are potent dilators of coronary arterioles ( 10-12 ) or pulmonary artery ( 13 ) and inhibit platelet aggregation ( 14 ). The EPA-derived epoxides account for fi ve epoxyeicosatetraenoic acids (5,8,11,14,and 17,, whereas the DHA-derived epoxides account for six epoxydocosapentaenoic acids (4,7,10,13,16,and 19,. A high regio-and stereoselectivity has been observed when testing the effects of chemically synthesized epoxides from AA and EPA. For example, in the rat, renal arteries were dilated by 11(R),12(S)-EET but not by 11(S),12(R)-EET or 14,15-EET enantiomers ( 15 ). Also in rats, among all EETeTr enantiomers, only the 17(R),18(S)-enantiomer, not 17(S),18(R), was effective on calcium-activated potassium (BK) channels in isolated cerebral arteries ( 11 ). However, in porcine coronary microvessels, all regioisomeric EETeTrs had similar Cytochromes P450 (CYPs) belong to a protein superfamily among which some members metabolize polyunsaturated long-chain fatty acids (PUFA-LC) to several classes of oxygenated metabolites. The product profi les depend on the involved CYP isoforms and may consist of a series of regio-and stereo-isomeric epoxides and hydroxylated compounds ( 1-3 ). In humans, CYP isoforms from

Human CYP450 v-hydroxylases of the CYP4 family are known to convert arachidonic acid (AA) to its metabolite . This study deals with hydroxylations of four PUFAs, eicosatrienoic acid (ETA), AA, eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA) by either human recombinant CYP4s enzymes or human liver microsomal preparations. CYP4F3A and CYP4F3B were the most efficient v-hydroxylases of these PUFAs. Moreover, the differences in the number of unsaturations of ETA, AA, and EPA allowed us to demonstrate a rise in the metabolic rate of hydroxylation when the double bond in 14-15 or 17-18 was missing. With the CYP4F enzymes, the main pathway was always the v-hydroxylation of PUFAs, whereas it was the (v-1)-hydroxylation with CYP1A1, CYP2C19, and CYP2E1. Finally, we demonstrated that the v9 and v3 PUFAs (ETA, EPA, and DHA) could all be used as alternative substrates in AA metabolism by human CYP4F2 and -4F3B. Thus, they decreased the ability of these enzymes to convert AA to 20-HETE. However, although ETA was the most hydroxylated substrate, EPA and DHA were the most potent inhibitors of the conversion of AA to 20-HETE. These findings suggest that some physiological effects of v3 FAs could partly result from a shift in the generation of active hydroxylated metabolites of AA through a CYP-mediated catalysis.-Fer, M., L. Corcos, Y. Dréano, E. Plée-Gautier, J-P. Salaün, F. Berthou, and Y. Amet. Cytochromes P450 from family 4 are the main omega hydroxylating enzymes in humans: CYP4F3B is the prominent player in PUFA metabolism. J. Lipid Res. 2008Res. . 49: 2379Res. -2389

Compared to other analytical methods, matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS) presents several unique advantages for the structural characterization of degradation products of carbohydrates. Our final goal is to implement this technique as a high-throughput platform, with the aim of exploring natural bio-diversity to discover new carbohydrate depolymerizing enzymes. In this approach, a variety of carbohydrates will be used as enzymes substrates and MALDI-MS will be employed to monitor the oligosaccharides produced. One drawback of MALDI, however, is that the choice of the matrix is largely dependent on the chemical properties of the analyte. In this context, our objective in the present work was to find the smallest set of MALDI matrices able to detect chemically heterogeneous oligosaccharides. This was done through the performance evaluation of more than 40 MALDI matrices preparations. Homogeneity of analyte-matrix deposits was considered as a critical feature, especially since the final objective is to fully automate the analyses. Evaluation of the matrices was done by means of a rigorous statistical approach. Amongst all tested compounds, our work proposes the use of the DHB/DMA ionic matrix as the most generic matrix, for rapid detection of a variety of polysaccharides including neutral, anionic, methylated, sulfated, and acetylated compounds. The selected matrices were then used to screen crude bacterial incubation media for the detection of enzymatic degradation products.