Formation of conjugated Delta11Delta13-double bonds by Delta12-linoleic acid (1,4)-acyl-lipid-desaturase in pomegranate seeds. (1/83)

For the biosynthesis of punicic acid (18:3Delta9Z,11E,13Z) a (11,14)-linoleoyl desaturase activity has been proposed. To isolate this acyl-lipid-desaturase, PCR-based cloning was used. This approach resulted in the isolation of two complete cDNAs. The first isolated full-length cDNA harbors a sequence of 1350 bp encoding a protein of 395 amino acids. The second cDNA was 1415 bp long encoding a protein of 387 amino acids. For functional identification proteins encoded by the cDNAs were expressed in Saccharomyces cerevisiae, and formation of newly formed fatty acids was analyzed by gas chromatography-free induction decay (GC-FID) and GC/MS. The expression of the heterologous enzymes resulted in the first case in a significant amount of linoleic acid and in the second case, after linoleic acid supplementation, in formation of punicic acid. The results presented here identify one cDNA coding for a classical Delta12-acyl-lipid-desaturase. The other one codes for a new type of (1,4)-acyl-lipid-desaturase that converts a cis double bond located in the Delta12-position of linoleic acid or gamma-linolenic acid, but not in alpha-linolenic acid, into a conjugated cis-trans double bond system.  (+info)

A novel mechanism for the inhibition of NF-kappaB activation in vascular endothelial cells by natural antioxidants. (2/83)

The activation of Nuclear Factor kappa B (NF-kappaB) in vascular endothelial cells, in response to biochemical or biomechanical stimuli, is associated with vascular pathologies such as atherosclerosis. The present manuscript studies the ability of the natural antioxidant-pomegranate wine (PW), to inhibit tumor necrosis factor alpha (TNF-alpha) or shear stress-mediated-NF-kappaB activation in vascular endothelial cells and compares it to that of red wine (RW) and N-acetyl cysteine (NAC). PW and RW act as potent antioxidants in vascular endothelial cells, inhibiting the oxidation of 2',7'-dichloroflurescin diacetate in TNF-alpha treated cells. PW (as well as RW and NAC) acted as potent inhibitors of NF-kappaB activation (migration into the nucleus and DNA binding activity) in vascular endothelial cells. Nevertheless, PW and NAC failed to inhibit TNF-a induced serine 32/36 phosphorylation and IkappaBalpha degradation. Surprisingly, these antioxidants alone induced enhanced IkappaB serine phosphorylation, which was not accompanied by IkappaBalpha degradation, or NF-kappaB nuclear translocation. This phosphorylation did not involve serine 32/36. Furthermore, we show for the first time that NAC inhibited TNF-alpha mediated phosphorylation of p65 (ser536), whereas PW had no effect on this phosphorylation. Thus, natural antioxidants may serve as potent NF-kappaB inhibitors in vascular endothelial cells, yet act through unique and divergent pathways.  (+info)

Delta 12-oleate desaturase-related enzymes associated with formation of conjugated trans-delta 11, cis-delta 13 double bonds. (3/83)

Conjugated linolenic acids are present as major seed oils in several plant species. Punicic acid (or trichosanic acid) is a conjugated linolenic acid isomer containing cis-delta9, trans-delta11, cis-delta13 double bonds in the C(18) carbon chain. Here we report cDNAs, TkFac and PgFac, isolated from Trichosanthes kirilowii and Punica granatum, that encode a class of conjugases associated with the formation of trans-delta11, cis-delta13 double bonds. Expression of TkFac and PgFac in Arabidopsis seeds under transcriptional control of the seed-specific napin promoter resulted in accumulation of punicic acid up to approximately 10% (w/w) of the total seed oils. In contrast, no punicic acid was found in lipids from leaves even when the conjugases were driven under control of the cauliflower mosaic virus 35S promoter. In yeast cells grown without exogenous fatty acids in the culture medium, TkFac and PgFac expression resulted in punicic acid accumulation accompanied by 16:2delta(9cis, 12cis) and 18:2delta(9cis, 12cis) production. Thus, TkFac and PgFac are defined as bifunctional enzymes having both conjugase and delta12-oleate desaturase activity. Furthermore, we demonstrate that 16:2delta(9cis, 12cis) and 18:3delta(9cis, 12cis, 15cis) as well as 18:2delta(9cis, 12cis) are potential substrates for the conjugases to form trans-delta11 and cis-delta13 double bonds.  (+info)

Potent inhibition by star fruit of human cytochrome P450 3A (CYP3A) activity. (4/83)

There has been very limited information on the capacities of tropical fruits to inhibit human cytochrome P450 3A (CYP3A) activity. Thus, the inhibitory effects of tropical fruits on midazolam 1'-hydroxylase activity of CYP3A in human liver microsomes were evaluated. Eight tropical fruits such as common papaw, dragon fruit, kiwi fruit, mango, passion fruit, pomegranate, rambutan, and star fruit were tested. We also examined the inhibition of CYP3A activity by grapefruit (white) and Valencia orange as controls. The juice of star fruit showed the most potent inhibition of CYP3A. The addition of a star fruit juice (5.0%, v/v) resulted in the almost complete inhibition of midazolam 1'-hydroxylase activity (residual activity of 0.1%). In the case of grape-fruit, the residual activity was 14.7%. The inhibition depended on the amount of fruit juice added to the incubation mixture (0.2-6.0%, v/v). The elongation of the preincubation period of a juice from star fruit (1.25 or 2.5%, v/v) with the microsomal fraction did not alter the CYP3A inhibition, suggesting that the star fruit did not contain a mechanism-based inhibitor. Thus, we discovered filtered extracts of star fruit juice to be inhibitors of human CYP3A activity in vitro.  (+info)

Punica granatum (Pomegranate) juice provides an HIV-1 entry inhibitor and candidate topical microbicide. (5/83)

BACKGROUND: For approximately 24 years the AIDS pandemic has claimed approximately 30 million lives, causing approximately 14,000 new HIV-1 infections daily worldwide in 2003. About 80% of infections occur by heterosexual transmission. In the absence of vaccines, topical microbicides, expected to block virus transmission, offer hope for controlling the pandemic. Antiretroviral chemotherapeutics have decreased AIDS mortality in industrialized countries, but only minimally in developing countries. To prevent an analogous dichotomy, microbicides should be: acceptable; accessible; affordable; and accelerative in transition from development to marketing. Already marketed pharmaceutical excipients or foods, with established safety records and adequate anti-HIV-1 activity, may provide this option. METHODS: Fruit juices were screened for inhibitory activity against HIV-1 IIIB using CD4 and CXCR4 as cell receptors. The best juice was tested for inhibition of: (1) infection by HIV-1 BaL, utilizing CCR5 as the cellular coreceptor; and (2) binding of gp120 IIIB and gp120 BaL, respectively, to CXCR4 and CCR5. To remove most colored juice components, the adsorption of the effective ingredient(s) to dispersible excipients and other foods was investigated. A selected complex was assayed for inhibition of infection by primary HIV-1 isolates. RESULTS: HIV-1 entry inhibitors from pomegranate juice adsorb onto corn starch. The resulting complex blocks virus binding to CD4 and CXCR4/CCR5 and inhibits infection by primary virus clades A to G and group O. CONCLUSION: These results suggest the possibility of producing an anti-HIV-1 microbicide from inexpensive, widely available sources, whose safety has been established throughout centuries, provided that its quality is adequately standardized and monitored.  (+info)

Effects of pomegranate juice on human cytochrome p450 3A (CYP3A) and carbamazepine pharmacokinetics in rats. (6/83)

In this study, we investigated whether components of pomegranate could inhibit CYP3A-mediated drug metabolism. The ability of pomegranate to inhibit the carbamazepine 10,11-epoxidase activity of CYP3A was examined using human liver microsomes, and pomegranate juice was shown to be a potent inhibitor of human CYP3A. Addition of 25 microl (5.0% v/v) of pomegranate juice resulted in almost complete inhibition of the carbamazepine 10,11-epoxidase activity of human CYP3A (1.8%). The inhibition potency of pomegranate juice was similar to that of grapefruit juice. In addition, we investigated the in vivo interaction between pomegranate juice and carbamazepine pharmacokinetics using rats. In comparison with water, the area under the concentration-time curve (AUC) of carbamazepine was approximately 1.5-fold higher when pomegranate juice (2 ml) was orally injected 1 h before the oral administration of the carbamazepine (50 mg/kg). On the other hand, the elimination half-life of carbamazepine and the AUC ratio of carbamazepine 10,11-epoxide to carbamazepine were not altered by the injection of pomegranate juice. These data suggest that pomegranate juice component(s) impairs the function of enteric but not hepatic CYP3A. Thus, we discovered that a component(s) of pomegranate inhibits the human CYP3A-mediated metabolism of carbamazepine. Furthermore, pomegranate juice alters the carbamazepine pharmacokinetics in rats.  (+info)

Beneficial effects of pomegranate juice on oxidation-sensitive genes and endothelial nitric oxide synthase activity at sites of perturbed shear stress. (7/83)

Atherosclerosis is enhanced in arterial segments exposed to disturbed flow. Perturbed shear stress increases the expression of oxidation-sensitive responsive genes (such as ELK-1 and p-JUN) in the endothelium. Evidence suggests that polyphenolic antioxidants contained in the juice derived from the pomegranate can contribute to the reduction of oxidative stress and atherogenesis. The aim of the present study was to evaluate the effects of intervention with pomegranate juice (PJ) on oxidation-sensitive genes and endothelial NO synthase (eNOS) expression induced by high shear stress in vitro and in vivo. Cultured human coronary artery endothelial cells (EC) exposed to high shear stress in vitro and hypercholesterolemic mice were used in this study. PJ concentrate reduced the activation of redox-sensitive genes (ELK-1 and p-JUN) and increased eNOS expression (which was decreased by perturbed shear stress) in cultured EC and in atherosclerosis-prone areas of hypercholesterolemic mice. Moreover, oral administration of PJ to hypercholesterolemic mice at various stages of disease reduced significantly the progression of atherosclerosis. This experimental study indicates that the proatherogenic effects induced by perturbed shear stress can be reversed by chronic administration of PJ. This approach may have implications for the prevention or treatment of atherosclerosis and its clinical manifestations.  (+info)

Pomegranate flower improves cardiac lipid metabolism in a diabetic rat model: role of lowering circulating lipids. (8/83)

Excess triglyceride (TG) accumulation and increased fatty acid (FA) oxidation in the diabetic heart contribute to cardiac dysfunction. Punica granatum flower (PGF) is a traditional antidiabetic medicine. Here, we investigated the effects and mechanisms of action of PGF extract on abnormal cardiac lipid metabolism both in vivo and in vitro. Long-term oral administration of PGF extract (500 mg kg(-1)) reduced cardiac TG content, accompanied by a decrease in plasma levels of TG and total cholesterol in Zucker diabetic fatty (ZDF) rats, indicating improvement by PGF extract of abnormal cardiac TG accumulation and hyperlipidemia in this diabetic model. Treatment of ZDF rats with PGF extract lowered plasma FA levels. Furthermore, the treatment suppressed cardiac overexpression of mRNAs encoding for FA transport protein, peroxisome proliferator-activated receptor (PPAR)-alpha, carnitine palmitoyltransferase-1, acyl-CoA oxidase and 5'-AMP-activated protein kinase alpha2, and restored downregulated cardiac acetyl-CoA carboxylase mRNA expression in ZDF rats, whereas it showed little effect in Zucker lean rats. The results suggest that PGF extract inhibits increased cardiac FA uptake and oxidation in the diabetic condition. PGF extract and its component oleanolic acid enhanced PPAR-alpha luciferase reporter gene activity in human embryonic kidney 293 cells, and this effect was completely suppressed by a selective PPAR-alpha antagonist MK-886, consistent with the presence of PPAR-alpha activator activity in the extract and this component. Our findings suggest that PGF extract improves abnormal cardiac lipid metabolism in ZDF rats by activating PPAR-alpha and thereby lowering circulating lipid and inhibiting its cardiac uptake.  (+info)