NADPH-dependent Microsomal Lipid Peroxidation Research Articles

Synthesis and Evaluation of Antioxidant Properties of Novel 2‐[2‐(4‐chlorophenyl) benzimidazole‐1‐yl]‐N‐(2‐arylmethylene amino) acetamides and 2‐[2‐(4‐chlorophenyl) benzimidazole‐1‐yl]‐N‐(4‐oxo‐2‐aryl‐thiazolidine‐3‐yl) acetamides‐I

Our approach was to synthesize and examine the antioxidant properties of some new 2-[2-(4-chlorophenyl)benzimidazole-1-yl]-N-(2-arylmethyleneamino) acetamide (1-18) and 2-[2-(4-chlorophenyl)benzimidazole-1-yl]-N-(4-oxo-2-aryl-thiazolidine-3-yl)acetamide (1t-18t) derivatives. Their in vitro effects on rat liver microsomal NADPH-dependent lipid peroxidation levels (LP assay) and microsomal ethoxyresorufin O-deethylase activities (EROD assay) were determined. The free radical scavenging properties of the compounds were also examined in vitro determining the interaction of 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical. The compounds showed significant effects in the above tests.

ChemInform Abstract: Synthesis and Antioxidant Properties of Some New Flavone Derivatives on Lipid Peroxidation in the Rat Liver.

A series of 2'-, 3'-, 4'- and 6-substituted flavone derivatives was synthesized and their in vitro effects on rat liver microsomal NADPH-dependent lipid peroxidation (LP) levels were determined. Retinoic acid, at 10(-4) M concentration, decreased the LP level by about 34%. A significant decrease in male liver microsomal LP level was noted for the compounds 3d, 1a, 3b and 4b at a concentration of 10(-4) M (100%, 95%, 75% and 62%, respectively).

Synthesis and evaluation of N-substituted indole-3-carboxamide derivatives as inhibitors of lipid peroxidation and superoxide anion formation

The in vitro antioxidant effects of novel N-substituted indole-3-carboxamides (I3CDs) 1-10 on rat liver microsomal NADPH-dependent lipid peroxidation (LP) levels and their free radicals scavenging properties were determined by the inhibition of superoxide anion formation (SOD). Among the synthesized compounds, 4, 5, 8 and 9 significantly inhibited SOD with an inhibition range at 84–100% at 10− 3 M concentration. The presence of halo substituents both ortho- and para- positions of these compounds resulted 100% inhibition of SOD. Comparison the activity results of halogenated and non-halogenated derivatives suggested that the halogenated compounds are more active than the non-halogenated compounds. On the other hand, the introduction of a para fluoro benzyl in the 1-position of indole (compounds 7, 8) has more impact on the SOD inhibition when the benzamide ring was mono halogenated. However, none of other compounds had a significant inhibitory effects on the level of lipid peroxidation.

Synthesis, antifungal and antioxidant screening of some novel benzimidazole derivatives

Some novel benzimidazole derivatives were synthesized and their in vitro effects on rat liver microsomal NADPH-dependent lipid peroxidation (LP) level, ethoxyresorufin O-deethylase (EROD) and antifungal activities were determined. A significant decrease in male rat liver microsomal LP level was noted by compounds 4c (52%), 4e (58%) and 4h (43%) at 10− 3 M concentration. Compounds 4c (100.0%), 4h (100.0%), 5c (98.0%) and 5h (100.0%) inhibited the microsomal ethoxyresorufin O-deethylase (EROD) enzyme activity better than that of the specific inhibitor caffeine (85%). Among these compounds, only compounds 4b and 4h exhibited moderate activity against C.albicans whereas the others had weak effects.

A study on the antioxidant activities of some new benzazole derivatives

The in vitro antioxidant properties of some new benzazole derivatives (1-10) such as benzoxazoles, benzimidazoles, and benzothiazoles were determined by their effects on the rat liver microsomal NADPH-dependent lipid peroxidation (LP) level, the scavenging of superoxide anion and the stable radical 2,2-diphenyl-1-picrylhydrazyl (DPPH). Compounds 1, 2, 4 and 6, showed potent scavenging effect on superoxide radical at 10(-3) M. Compound 8, 5-nitro-2-(phenoxymethyl)benzimidazole, strongly inhibited lipid peroxidation at 10(-3) M concentration.

Expression and activity of CYP2E1 in circulating lymphocytes are not altered in diabetic individuals

Cytochrome P4502E1 (CYP2E1) plays an important role in ROS production thus favouring accelerated membrane lipid peroxidation. This isoform is strongly expressed in the liver but it can be also found in lymphocytes. As such, lymphocyte may provide a non-invasive accessible pool for screening CYP2E1 expression in man. We have, therefore, analysed CYP2E1 expression and activity in lymphocyte microsomes from 12 healthy controls, 11 type 1 and 12 type 2 diabetic subjects by using Western blot and enzymatic activities. Immunoblotting did not show difference among CYP2E1 protein bands in controls, type 1 and type 2 diabetics. To assess CYP2E1 activity we used the 7-ethoxy-4-trifluoromethylcoumarin (7-EFC), as a fluorescent substrate. The rate of deethylation of 7-EFC from controls did not differ from type 1 and type 2 diabetic subjects. The lack of any difference in CYP2E1 activity also was confirmed by the NADPH-dependent microsomal lipid peroxidation CCL 4-induced assay showing similar peroxidation rates among controls and diabetic subjects. The results show that CYP2E1 expression/activity in lymphocytes is not enhanced in diabetes.

Antioxidant properties of flavone-6(4')-carboxaldehyde oxime ether derivatives.

The in vitro antioxidant properties of some flavone-6(4)-carboxaldehyde oxime ether derivatives (Ia-f, IIa-f) were determined by their effects on the rat liver microsomal NADPH-dependent lipid peroxidation (LP) levels by measuring the formation of 2-thiobarbituric acid reactive substances. The free radical scavenging properties of the compounds were also examined in vitro by determining their capacity to scavenge superoxide anions and interact with the stable free radical 2, 2-diphenyl-1-picrylhydrazyl (DPPH). The most active compounds, IIb (Flavone-4'-carboxaldehyde-O-ethyl oxime) and Id (Flavone-6-carboxaldehyde-O-[2-(1-pyrolidino) ethyl] oxime), caused 98 and 79% inhibition of superoxide anion production and DPPH stable free radical at 10(-3) M, respectively.

Synthesis and Antioxidant Properties of Novel Benzimidazole Derivatives

Some novel benzimidazole derivatives carrying thiosemicarbazide and triazole moieties at the N1 position were synthesized and their in vitro effects on rat liver microsomal NADPH-dependent lipid peroxidation (LP) levels determined by measuring the formation of 2-thiobarbituric acid reactive substance. The free radical scavenging properties of the compounds were also examined in vitro by determining the capacity to scavenge superoxide anion formation and the interaction with the stable free radical 2,2-diphenyl-1-picrylhydrazyl (DPPH). The compounds showed a significant effect in the above tests except to scavenge superoxide anion formation.

Synthesis and antioxidant properties of some novel benzimidazole derivatives on lipid peroxidation in the rat liver.

Some benzimidazole derivatives namely 1-[(substituted thiocarbamoylhydrazine carbonyl) methyl]-2-phenyl-1H-benzimidazoles (1a-13a), N-[(2-phenylbenzimidazol-1-yl methyl)-[1,3,4]-thiadiazole-2-yl]-substituted phenyl amines (1b-13b) and 5-(2-phenyl benzimidazol-1-yl-methyl)-4-substituted phenyl-4H-1,2,4-triazole-3-thiones (1c-13c) were synthesized, and their in vitro effects on the rat liver microsomal NADPH-dependent lipid peroxidation (LP) levels were determined. The most active compound 10a caused an 84% inhibition of LP at 10(-3) M, which is better than that of butylated hydroxytoluene (BHT) (65%).

Evaluation of the antioxidant effects of some flavonylthiazolidinediones by determining their effects on lipid peroxidation, superoxide anion formation, and 2,2-diphenyl-1-picrylhydrazyl stable free radical.

The in vitro antioxidant effects of some flavonylthiazolidinediones (Ia-d, IIa-d) on rat liver microsomal NADPH-dependent lipid peroxidation (LP) levels were determined by measuring the formation of 2-thiobarbituric acid reactive substance. The free radical-scavenging properties of the compounds were examined in vitro by determining the capacity to scavenge superoxide anion formation and of the interaction with the stable free radical 2,2-diphenyl-1-picrylhydrazyl (DPPH). The compounds had no inhibitory effects on LP. However, they had variable inhibitory influence on superoxide anion production and DPPH radical in a concentration-dependent manner. The most active compound, 3-(2,5-dimethoxyphenacyl)-5-[2-phenyl-4H-4-oxo-1-benzopyran-6-yl)methylenyl]-thiazolidine-2,4-dione, Id showed 98% inhibition of superoxide anion production and 95% inhibition of DPPH stable free radical at 10(-3) M.

ANTIOXIDANT ACTIVITY OF N-SUBSTITUTED İNDOLE 2- AND 3-CARBOXAMIDES : N-SÜBSTİTÜE İNDOL-2 VE 3-KARBOKSAMİTLERİN ANTİOKSİDAN AKTİVİTESİ

Bir seri N-sübstitüe indol amid türevinin, sıçan karaciğerinden elde edilen mikrozomal NADP'ye bağımlı lipid peroksidasyon (LP) ve süperoksit anyon şekillenmesi üzerinde olan invitro etkileri test edildi. Vitamin E (a-tokoferol), LP seviyesini 10'4 M konsantrasyonda yaklaşık % 63 kadar azalttı. Bileşiklerden 1,8 ve 10'ün 10'4 M konsantrasyonda, a-tokoferol ile karşılaştırıldıklarında erkek karaciğer mikrozomal LP düzeylerinde belirgin bir azalmaya neden olduğu bulundu. Bileşiklerden 8 ve 10'ün, ö2(~ anyonunu belirgin bir biçimde inhibe ettiği bulundu. Her iki aktivite sonucu, bu bileşiklerin ümit verici antioksidanlar olabileceklerini gösterdi

Synthesis and Antioxidative Properties of Novel Thiazolidinedione/Imidazolidinedione Compounds as Retinoids.

AbstractFor Abstract see ChemInform Abstract in Full Text.

Cinnamophilin as a novel antiperoxidative cytoprotectant and free radical scavenger

The antioxidant properties of cinnamophilin were evaluated by studying its ability to react with relevant reactive oxygen species, and its protective effect on cultured cells and biomacromolecules under oxidative stress. Cinnamophilin concentration-dependently suppressed non-enzymatic iron-induced lipid peroxidation in rat brain homogenates with an IC 50 value of 8.0±0.7 μM and iron ion/ADP/ascorbate-initiated rat liver mitochondrial lipid peroxidation with an IC 50 value of 17.7±0.2 μM. It also exerted an inhibitory activity on NADPH-dependent microsomal lipid peroxidation with an IC 50 value of 3.4±0.1 μM without affecting microsomal electron transport of NADPH-cytochrome P-450 reductase. Both 1,1-diphenyl-2-picrylhydrazyl and 2,2′-azo-bis(2-amidinopropane) dihydrochloride-derived peroxyl radical tests demonstrated that cinnamophilin possessed marked free radical scavenging capacity. Cinnamophilin significantly protected cultured rat aortic smooth muscle cells (A7r5) against alloxan/iron ion/H 2O 2-induced damage resulting in cytoplasmic membranous disturbance and mitochondrial potential decay. By the way, cinnamophilin inhibited copper-catalyzed oxidation of human low-density lipoprotein, as measured by fluorescence intensity and thiobarbituric acid-reactive substance formation in a concentration-dependent manner. On the other hand, it was reactive toward superoxide anions generated by the xanthine/xanthine oxidase system and the aortic segment from aged spontaneously hypertensive rat. Furthermore, cinnamophilin exerted a divergent effect on the respiratory burst of human neutrophil by different stimulators. Our results show that cinnamophilin acts as a novel antioxidant and cytoprotectant against oxidative damage.

CYP2E1 and CYP4A as microsomal catalysts of lipid peroxides in murine nonalcoholic steatohepatitis.

Nonalcoholic steatohepatitis (NASH) and alcoholic liver disease have similar pathological features. Because CYP2E1 plays a key role in alcoholic liver disease with its ability to stimulate lipid peroxidation, we tested the proposal that CYP2E1 could also be a factor in the development of NASH. In a dietary model - mice fed a methionine- and choline-deficient (MCD) diet - liver injury was associated with both induction of CYP2E1 and a 100-fold increase in hepatic content of lipid peroxides. Microsomal NADPH-dependent lipid oxidases contributed to the formation of these lipid peroxides, and in vitro inhibition studies demonstrated that CYP2E1 was the major catalyst. To further define the role of CYP2E1 as an initiator of oxidative stress in NASH, Cyp2e1(-/-)mice were administered the MCD diet. CYP2E1 deficiency neither prevented the development of NASH nor abrogated the increased microsomal NADPH-dependent lipid peroxidation, indicating the operation of a non-CYP2E1 peroxidase pathway. In Cyp2e1(-/-) mice with NASH (but not in wild-type mice), CYP4A10 and CYP4A14 were upregulated. Furthermore, hepatic microsomal lipid peroxidation was substantially inhibited by anti-mouse CYP4A10 antibody in vitro. These results show that experimental NASH is strongly associated with hepatic microsomal lipid peroxidation. CYP2E1, the main enzyme associated with that process in wild-type mice, is not unique among P450 proteins in catalyzing peroxidation of endogenous lipids. We have now identified CYP4A enzymes as alternative initiators of oxidative stress in the liver.

Pirfenidone inhibits NADPH-dependent microsomal lipid peroxidation and scavenges hydroxyl radicals.

Pirfenidone (Pf), a new broad-spectrum anti-fibrotic agent, is known to offer protection against lung fibrosis in vivo in laboratory animals, and against mitogenesis and collagen formation by human lung fibroblasts in vitro. Because reactive oxygen species are thought to be involved in these events, we investigated the mechanism(s) by which Pf ameliorates oxidative stress and its effects on NADPH-dependent lipid peroxidation. Pf has been shown to cause inhibit NADPH-dependent lipid peroxidation in sheep liver microsomes in a dose-dependent manner. The concentration of Pf required to cause 50% inhibition of lipid peroxidation was approximately 6 mM. Pf was found to be ineffective as a superoxide radical scavenger. Pf was also ineffective in decomposing H2O2 and chelating iron. In deoxyribose degradation assays, Pf was a potent scavenger of hydroxyl radicals with a rate constant of 5.4 x 10(9) M(-1) sec(-1). EPR spectroscopy in combination with spin trapping techniques, using a Fenton type reaction and DMPO as a spin-trapping agent, Pf scavenged hydroxyl radicals in a dose-dependent manner. The concentration of Pf required to inhibit 50% signal height was approximately 2.5 mM. Because iron was used in the Fenton reaction, the ability of Pf in chelating iron was verified in a fluorescent competitive assay using calcein as the fluorescent probe. Pf up to 10 mM concentration was ineffective in chelating either Fe2+ or Fe3+ in this system. We propose that Pf exerts its beneficial effects, at least in part, through its ability to scavenge toxic hydroxyl radicals.

Ａｎｔｈｒａｃｙｃｌｉｎｅ新規制癌剤ＭＸ２誘導マウス組織中過酸化脂質上昇に対するリポソーム化の効果

We examined the effect of liposomalization on the changes of lipid peroxide level and glutathione peroxidase (GSHpx) activity in mice tissues after 3'-deamino-3'-morpholino-13-deoxo-10-hydroxycarminomycin (MX2) administration. The agreement of the decrease in body weight on the 4th day after double MX2 administration (3.0mg/kg/day×2days, i.p.) was similar to those after adriamycin (ADR) administration (15mg/kg, i.p.). The lipid peroxide level in the heart and liver significantly increased after MX2 double treatment. GSHpx activity in the heart decreased to 80% on the 2nd day after MX2 injection, compared to normal activity. However, these changes are smaller than those after ADR administration (15mg/kg). NADPH-dependent microsomal lipid peroxidation in mouse liver increased, induced more by MX2 than by ADR. The combination of α-tocopherol with MX2 suppressed the increase in the lipid peroxide level by MX2 whereas did not suppress the decrease in GSHpx activity. Thus, it is considered that two mechanisms are involved, i.e., the increase in the lipid peroxide level occurs due to the production of active oxygen from MX2 and due to MX2-induced inhibition of GSHpx. The liposomalization of MX2 suppressed both MX2 induced changes in lipid peroxide level and GSHpx activity. Namely, it is considered that MX2 induced toxicity by these changes were reduced by α-tocopherol combination or liposomalization of MX2.

A study on the antioxidant capacities of some benzimidazoles in rat tissues

Seven benzimidazole compounds were synthesized and their in vitro effects on rat liver, lung and kidney microsomal NADPH-dependent lipid peroxidation (LP) levels were determined. The significant decrease in male rat liver microsomal LP level was noted only by the compound 4 at 10 −4 M (20%) and 10 −3 M (40%) concentrations whereas the other compounds were ineffective. In lung, only the compound 6 at 10 −4 M concentration exhibited significant alteration, i.e. 56% increase, in LP level. In kidney, however, apart from the compound 4, all the compounds increased LP level (35–52%) significantly. The classical antioxidant, butylated hydroxy toluene (BHT), at 10 −4 M concentration, significantly decreased LP level about 70%, in all the tissues studied. To clarify the effects of compounds 4 and 6 on LP, the responses of some CYPs, which are active in producing reactive oxygen species, to these compounds were also investigated. The compound 4 at 10 −4 and 10 −3 M concentrations inhibited the hepatic microsomal ethoxyresorufin O-deethylase (EROD) (37 and 65%) and pentoxyresorufin O-depenthylase (PROD) (14 and 62%) enzyme activities significantly. However, it did not alter the hepatic microsomal NADPH-cytochrome P450-reductase activity. BHT, at 10 −3 M concentration, significantly inhibited hepatic microsomal EROD (73%), PROD (62%) and NADPH-cytochrome P450 reductase (17%) enzyme activities. Caffeine (10 −3 M) and SKF 525A (10 −3 M), which are specific inhibitors of EROD and PROD enzyme activities, significantly decreased the enzyme activities 33 and 77%, respectively. Caffeine was unable to alter hepatic microsomal NADPH-cytochrome P450 reductase enzyme activity whereas SKF 525A significantly inhibited (80%) it. In lung and kidney, the compound 6 at 10 −4 M concentration significantly increased EROD (44 and 19%) and PROD (103 and 86%) enzyme activities. However, the elevation of PROD enzyme activity in both tissues was observed to be more pronounced than that of EROD enzyme activity. This compound was ineffective on lung and kidney microsomal P450-reductase enzyme activity. These results reveal that the synthesized benzimidazoles have variable tissue dependent in vitro effects on LP due to their distinct effects on CYP activities but not on NADPH-cytochrome P450 reductase activity in rats.

Tissue Specific Expression of Glutathione S-transferases, Glutathione Content and Lipid Peroxidation in Camel Tissues

Differential expression of glutathione S-transferase (GST) enzyme activity in various tissues of the camel was observed with a maximum activity in the liver. Compared with the rat and human livers, GST activity in camel liver was 50% lower than that of rat liver and similar to that of human liver. Extrahepatic tissues in camel have a comparable GST activity with those of similar tissues in the rat. Assay of GST activity using ethacrynic acid as substrate demonstrated maximum activity in the camel brain followed by intestine, liver and kidney. Microsomal GST activity in camel tissues was expressed in the order of liver > testis > intestine ≈ kidney ≈ brain. Phenotyping of GST was performed in camel hepatic and extrahepatic tissues using human specific antibodies to class α, μ, and π cytosolic GST isoenzymes and rat specific antibody to the microsomal GST. Western immunoblot and immunohistochemical analyses showed an abundant expression of GST α and μ in the camel liver, while π was very poorly expressed. Camel extrahepatic tissues however, had a significant expression of GST π. The camel GST isoenzymes were found to be predominantly expressed in the hepatocytes around the central vein with a gradual decrease in expression in the hepatocytes located toward the periphery. Kidney cortex exhibited a greater expression of the enzyme protein in the proximal tubules as compared to the glomeruli. Glutathione (GSH) concentration in rat tissues, except in the brain, was about 2-fold higher than that of camel tissues. Rate of NADPH-dependent microsomal lipid peroxidation was comparable both in the rat and camel tissues with the highest activity in the brain and lowest activity in the intestine. The differential expression of GST isoenzymes in different organs of the camel, GSH concentration and the rate of lipid peroxidation in different tissues may be important factors in determining the differential susceptibility of camel tissues to the toxic effects of xenobiotics.

Potentiation of iron-induced lipid peroxidation by a series of bipyridyls in relation to their ability to reduce iron

We determine the relative abilities of three bipyridyls (Paraquat P ++, Benzylviologen B ++ and Diquat D ++) to stimulate iron-induced lipid peroxidation in relation to their ability to redox cycle and to reduce iron. The ability to redox cycle increases with the redox potential, in the order: P ++ < B ++ < D ++. The initial rates of Fe 3+reduction increased from P ++ to D ++ and B ++. NADPH-dependent microsomal lipid peroxidation was measured in different conditions (1) increasing incubation time, 100 μM bipyridyl, 10 μM FeCl 3; (2) increasing concentrations of bipyridyls, 10 μM FeCl 3, 10 min incubation time; (3) increasing concentrations of FeCl 3, 100μM bipyridyl, 10 min incubation time. Bipyridyls potentiated the production of malondialdehyde (MDA), particularly with the shortest incubation times and the lowest concentrations of FeCl 3, in the order of activity P ++ < D ++ < B ++. The strong correlation between the initial rate of lipid peroxidation and Fe 3+ reduction indicates that the rate of Fe 3+ reduction determines the intensity of the potentiation effect of bipyridyls on lipid peroxidation—on the condition that the amount of peroxidizable substrate is not the limiting factor.

Marchantin H as a Natural Antioxidant and Free Radical Scavenger

The antioxidant activity of marchantin H was investigated using various experimental models. Marchantin H inhibited nonenzymatic iron-induced lipid peroxidation in rat brain homogenates with an IC50value of 0.51 ± 0.03 μm. It was more potent than desferrioxamine or other classical antioxidants. Marchantin H also suppressed NADPH-dependent microsomal lipid peroxidation with an IC50value of 0.32 ± 0.01 μmwithout affecting microsomal electron transport of NADPH–cytochrome P450 reductase. Marchantin H could scavenge the stable free radical 1,1-diphenyl-2-picrylhydrazyl and peroxyl radical derived from 2,2′-azobis(2-amidinopropane) dihydrochloride in aqueous phase, but not the peroxyl radical derived from 2,2′-azobis(2,4-dimethylvaleronitrile) in hexane. The oxygen consumption during peroxyl radical-induced human erythrocyte ghost oxidation was decreased in a concentration-dependent manner by marchantin H. Furthermore, it was reactive toward superoxide anion generated by the xanthine/xanthine oxidase system. On the other hand, marchantin H inhibited copper-catalyzed oxidation of human low-density lipoprotein, as measured by fluorescence intensity, thiobarbituric acid-reactive substance formation, and electrophoretic mobility in a concentration-dependent manner. Our results indicate that marchantin H is a potentially effective and versatile antioxidant and can be used as a chaperone protecting biomacromolecules against peroxidative damage.