Substitution of vanadium into the framework of mesoporous MCM-41 molecular sieves has been achieved; these materials were found to be very efficient catalysts for the selective oxidation of large organic molecules using hydrogen peroxide.Vanadium silicate molecular sieves are a new class of catalysts with remarkable catalytic properties in the selective oxidation of various organic molecules.' All vanadium-modified molecular sieves reported previously are microporous solids with severe restrictions to oxidise large organic molecules.'Recently, Kresge et af.2 discovered a new family of mesoporous molecular sieves designated as M41S. MCM-41, one of the members of this extensive family of materials, possesses a hexagonal array of uniform mesopores.2J MCM-41 has been synthesized with uniform channels varying from approximately 15 A to more than 100 A in size. The pore size was found to be dependent on the chain length of the organic template (surfactant) used in the hydrothermal synthesis. In this communication, for the first time, we report the preparation and characterization of mesoporous vanadium silicate molecular sieve, V-MCM-41. The activity of V-MCM-41 in the oxidation of 1-naphthol and cyclododecane is also reported.The hydrothermal synthesis of V-MCM-41 was carried out using gels with the following molar composition: Si02-x-VO2-0. 17Na20.0.5RBr.30H20, where x S 0.02 and RBr designates the organic template.In a typical synthesis, fumed silica (Cab-0-Sil M5) was dissolved in a NaOH solution and stirred for 30 min. Then an aqueous solution of vanadyl sulfate was added to the mixture which was stirred for an additional 3 h before adding the organic template, dodecyltrimethylammonium bromide. After one more hour of stirring, the resulting mixture was poured into a 100 ml Teflon lined stainless-steel autoclave which was tumbled at 25 rpm at 373 K in an oven for 6 d. After crystallization, the product was filtered, washed with deionised water, dried at 373 K and calcined in air at 823 K for 6 h. The product yield was about 90%. The pure-silica polymorph of MCM-41 was synthesized using the same procedure, except that no vanadium was added.The X-ray diffraction pattern of V-MCM-41 ( Fig. 1) matches well that of the vanadium-free silica polymorph MCM-41 and also the patterns reported by Kresge et aZ.2 One major peak along with three small peaks were observed. Beck et aZ.3 indexed these peaks for a hexagonal unit cell the parameter of which was calculated using a0 = 2dl&h. It was also observed that the dlW d-spacing, which indicates approximate pore size, increases with the chain length of the organic template. Table 1 shows the d-spacing and unit cell parameter of V-MCM-41 and its silica polymorph. The increase in the unit cell parameter of V-MCM-41 compared to its pure silica analogue (Table 1) confirms the presence of vanadium in the silicate framework. Insertion of metal ions larger than silicon brings about an increase in the unit cell parameters because of the longer M -0 bond distance. Similar findings were rep...

20-Hydroxyeicosatetraenoic acid (HETE), the cytochrome P-450 (CYP) 4A ω-hydroxylation product of arachidonic acid, has potent biological effects on renal tubular and vascular functions and on the control of arterial pressure. We have expressed high levels of the rat CYP4A1, -4A2, -4A3, and -4A8 cDNAs, using baculovirus and Sf 9 insect cells. Arachidonic acid ω- and ω-1-hydroxylations were catalyzed by three of the CYP4A isoforms; the highest catalytic efficiency of 947 nM−1 ⋅ min−1for CYP4A1 was followed by 72 and 22 nM−1 ⋅ min−1for CYP4A2 and CYP4A3, respectively. CYP4A2 and CYP4A3 exhibited an additional arachidonate 11,12-epoxidation activity, whereas CYP4A1 operated solely as an ω-hydroxylase. CYP4A8 did not catalyze arachidonic or linoleic acid but did have a detectable lauric acid ω-hydroxylation activity. The inhibitory activity of various acetylenic and olefinic fatty acid analogs revealed differences and indicated isoform-specific inhibition. These studies suggest that CYP4A1, despite its low expression in extrahepatic tissues, may constitute the major source of 20-HETE synthesis. Moreover, the ability of CYP4A2 and -4A3 to catalyze the formation of two opposing biologically active metabolites, 20-HETE and 11,12-epoxyeicosatrienoic acid, may be of great significance to the regulation of vascular tone.

In our present studies utilizing a well characterized proximal tubule cell line, LLCPKcl4, we determined that all four EET regioisomers (5,6-EET, 8,9-EET, 11,12-EET, and 14,15-EET) stimulated [ 3 H]thymidine incorporation, with 14,15-EET being the most potent. In contrast, no mitogenic effects were seen with arachidonic acid, other cP450 arachidonate metabolites (12R-hydroxyeicosatetraenoic acid (12R-HETE), 14,15-dihydroxyeicosatrienoic acid (14,15-DHET), or 20-HETE), or lipoxygenase metabolites (5S-HETE, leukotriene B 4 , or lipoxin A4). We found that their metabolically more stable sulfonimide (SI) analogs (11,12-EET-SI and 14,15-EET-SI) were also potent mitogens. In addition 14,15-EET-SI also increased cell proliferation as well as expression of both c-fos and egr-1 mRNA. The protein kinase C and A inhibitors, H-7 and H-8, or the cyclooxygenase inhibitor, indomethacin, had no effect upon 14,15-EET-induced Cytochrome P450 epoxygenase catalyzes the NADPHdependent epoxidation of arachidonic acid to 5,6-, 8,9-, 11,12-, and 14,15-epoxyeicosatrienoic acids (EETs) 1 in a regio-and stereo-selective manner. The EETs are produced predominantly by epoxygenases of the 2C family of cP450s, which have been localized in the kidney to the mammalian proximal tubule. The proximal tubule contains the highest concentration of cP450 within the mammalian kidney (1) and expresses minimal cyclooxygenase or lipoxygenase activity (2). The EETs or their hydration products have been implicated in modulation of vascular tone and renal glomerular hemodynamics (3), renal proximal tubule function, and regulation of mitogenesis (4, 5). Recently, EETs have been suggested to be an endothelialderived hyperpolarizing factor (6), although there is controversy about this issue (7,8). Direct administration of EETs inhibits amiloride-sensitive sodium transport (9) and 86 Rb uptake in LLC-PK1, a nontransformed, immortalized cell line from pig kidney with certain proximal tubule characteristics (10).EETs have also been proposed as second messengers for hormones and growth factors in the proximal tubule. We have shown that epidermal growth factor (EGF) stimulates EET production in rat proximal tubule suspensions and primary cultured rabbit proximal tubule cells, and EETs may mediate both EGF-stimulated calcium influx and mitogenesis in proximal tubules (11). Omata et al. also demonstrated EET production upon stimulation with angiotensin II (12). EETs may mediate the effect of high (Ͼ10 Ϫ7 M) angiotensin II to increase cytosolic calcium ([Ca 2ϩ ] i ) and decrease Na ϩ /H ϩ exchange activity in cultured rabbit proximal tubule cells and isolated rat proximal tubules (13)(14)(15). In the present studies, we examined mitogenic signaling mechanisms of EETs in renal epithelial cells. We determined the mitogenic effects of all four EET regioisomers and utilized metabolically more stable sulfonimide analogs to determine potential intracellular signaling mechanisms mediating mitogenic effects. We demonstrate here that EETs activate pp60 c-src and initiate a ty...

The present study examined the effects of a series of 20-hydroxyeicosatetraenoic acid (20-HETE) derivatives on the diameter of renal arterioles to determine the structural requirements of the vasoconstrictor response to 20-HETE. The vascular responses to 5-, 8-, 12-, 15-, 19-, 20-, 21-HETEs, arachidonic acid (AA), and saturated, partially saturated, dimethyl, carboxyl, and 19-carbon derivatives of 20-HETE (10(-8) to 10(-6) M) were assessed in rat renal interlobular arteries (65-125 micrometer). 20-HETE, 21-HETE, dimethyl-20-HETE, and a partially saturated derivative of 20-HETE, 20-hydroxyeicosa-5(Z),14(Z)-dienoic acid, reduced vessel diameter by 19 +/- 3, 17 +/- 3, 16 +/- 2, and 28 +/- 2%, respectively. In contrast, 5-, 8-, 12-, 15-, and 19-HETE, AA, saturated, partially saturated, carboxyl, and the 19-carbon derivatives of 20-HETE had no effect on vessel diameter. Pretreatment with 5-, 15-, and 19-HETE, the 19-carbon derivative or 20-hydroxyeicosa-6(Z),15(Z)-dienoic acid (1 microM) completely blocked the vasoconstrictor response to 20-HETE in renal arterioles. Pretreatment with AA, carboxyl, saturated 19-carbon, and saturated 20-HETE derivatives (1 microM) partially blocked the response, whereas 8- and 12-HETE (1 microM) had no effect on the vasoconstrictor response to 20-HETE. These findings suggest that 20-HETE agonists and antagonists require a carboxyl or an ionizable group on carbon 1 and a double bond near the 14 or 15 carbon. 20-HETE agonists also require a functional group capable of hydrogen bonding on carbon 20 or 21, whereas antagonists lack this reactive group.

Epoxyeicosatrienoic acids (EETs) are endothelium-derived eicosanoids that activate potassium channels, hyperpolarize the membrane, and cause relaxation. We tested 19 analogs of 14,15-EET on vascular tone to determine the structural features required for activity. 14,15-EET relaxed bovine coronary arterial rings in a concentration-related manner (ED(50) = 10(-6) M). Changing the carboxyl to an alcohol eliminated dilator activity, whereas 14,15-EET-methyl ester and 14,15-EET-methylsulfonimide retained full activity. Shortening the distance between the carboxyl and epoxy groups reduced the agonist potency and activity. Removal of all three double bonds decreased potency. An analog with a Delta8 double bond had full activity and potency. However, the analogs with only a Delta5 or Delta11 double bond had reduced potency. Conversion of the epoxy oxygen to a sulfur or nitrogen resulted in loss of activity. 14(S),15(R)-EET was more potent than 14(R),15(S)-EET, and 14,15-(cis)-EET was more potent than 14,15-(trans)-EET. These studies indicate that the structural features of 14,15-EET required for relaxation of the bovine coronary artery include a carbon-1 acidic group, a Delta8 double bond, and a 14(S),15(R)-(cis)-epoxy group.

Previous structural analyses of diphosphoinositol polyphosphates in biological systems have relied largely on NMR analysis. For example, in Dictyostelium discoideum, diphosphoinositol pentakisphosphate was determined by NMR to be 4- and/or 6-PPInsP5, and the bisdiphosphoinositol tetrakisphosphate was found to be 4, 5-bisPPInsP4 and/or 5,6-bisPPInsP4 [Laussmann, Eujen, Weisshuhn, Thiel and Vogel (1996) Biochem. J. 315, 715-720]. We now describe three recent technical developments to aid the analysis of these compounds, not just in Dictyostelium, but also in a wider range of biological systems: (i) improved resolution and sensitivity of detection of PPInsP5 isomers by microbore metal-dye-detection HPLC; (ii) the use of the enantiomerically specific properties of a rat hepatic diphosphatase; (iii) chemical synthesis of enantiomerically pure reference standards of all six possible PPInsP5 isomers. Thus we now demonstrate that the major PPInsP5 isomer in Dictyostelium is 6-PPInsP5. Similar findings obtained using the same synthetic standards have been published [Laussmann, Reddy, Reddy, Falck and Vogel (1997) Biochem. J. 322, 31-33]. In addition, we show that 10-25% of the Dictyostelium PPInsP5 pool is comprised of 5-PPInsP5. The biological significance of this new observation was reinforced by our demonstration that 5-PPInsP5 is the predominant PPInsP5 isomer in four different mammalian cell lines (FTC human thyroid cancer cells, Swiss 3T3 fibroblasts, Jurkat T-cells and Chinese hamster ovary cells). The fact that the cellular spectrum of diphosphoinositol polyphosphates varies across phylogenetic boundaries underscores the value of our technological developments for future determinations of the structures of this class of compounds in other systems.

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