Interaction of α-Phenyl-N-tert-Butyl Nitrone and Alternative Electron Acceptors with Complex I Indicates a Substrate Reduction Site Upstream from the Rotenone Binding Site

Abstract: Mitochondrial complexes I, II, and III were studied in isolated brain mitochondrial preparations with the goal of determining their relative abilities to reduce O₂ to hydrogen peroxide (H₂O₂) or to reduce the alternative electron acceptors nitroblue tetrazolium (NBT) and diphenyliodonium (DPI). Complex I and II stimulation caused H₂O₂ formation and reduced NBT and DPI as indicated by dichlorodihydrofluorescein oxidation, nitroformazan precipitation, and DPI-mediated enzyme inactivation. The O₂ consumption rate was more rapid under complex II (succinate) stimulation than under complex I (NADH) stimulation. In contrast, H₂O₂ generation and NBT and DPI reduction kinetics were favored by NADH addition but were virtually unobservable during succinate-linked respiration. NADH oxidation was strongly suppressed by rotenone, but NADH-coupled H₂O₂ flux was accelerated by rotenone. α-Phenyl- N-tert -butyl nitrone (PBN), a compound documented to inhibit oxidative stress in models of stroke, sepsis, and parkinsonism, partially inhibited complex I-stimulated H₂O₂ flux and NBT reduction and also protected complex I from DPI-mediated inactivation while trapping the phenyl radical product of DPI reduction. The results suggest that complex I may be the principal source of brain mitochondrial H₂O₂ synthesis, possessing an "electron leak" site upstream from the rotenone binding site (i.e., on the NADH side of the enzyme). The inhibition of H₂O₂ production by PBN suggests a novel explanation for the broad-spectrum antioxidant and antiinflammatory activity of this nitrone spin trap.     

Characterization of partially folded intermediates of papain in presence of cationic, anionic, and nonionic detergents at low pH

A systematic investigation of the effects of detergents [Sodium dodecyl sulphate (SDS), hexa decyltrimethyl ammonium bromide (CTAB) and Tween-20] on the structure of acid-unfolded papain (EC.3.4.22.2) was made using circular dichroism (CD), intrinsic tryptophan fluorescence, and 1-anilino 8-sulfonic acid (ANS) binding. At pH 2, papain exhibits a substantial amount of secondary structure and is relatively less denatured compared with 6 M GdnHCl (guanidine hydrochloride) but loses the persistent tertiary contacts of the native state. Addition of detergents caused an induction of alpha-helical structure as evident from the increase in the mean residue ellipticity value at 208 and 222 nm. Near-UV CD spectra also showed the regain of native-like spectral features in the presence of 8 mM SDS and 3.5 mM CTAB. Induction of structure in acid-unfolded papain was greater in the presence SDS followed by CTAB and Tween-20. Intrinsic tryptophan fluorescence studies indicate the change in the environment of tryptophan residues upon addition of detergents to acid-unfolded papain. Addition of 8 mM SDS resulted in the loss of ANS binding sites exhibited by a decrease in ANS fluorescence intensity, suggesting the burial of hydrophobic patches. Maximum ANS binding was obtained in the presence of 0.1 mM Tween-20 followed by CTAB, indicating a compact "molten-globule"-like conformation with enhanced exposure of hydrophobic surface area. Acid-unfolded papain in the presence of detergents showed the partial recovery of enzymatic activity. These results suggest that papain at low pH and in the presence of SDS exists in a partially folded state characterized by native-like secondary structure and tertiary folds. While in the presence of Tween, acid-unfolded papain exists as a compact intermediate with molten-globule-like characteristics, viz. enhanced hydrophobic surface area and retention of secondary structure. While in the presence of CTAB it exists as a compact intermediate with regain of native-like secondary and partial tertiary structure as well as high ANS binding with the partially recovered enzymatic activity, i.e., a molten globule state with tertiary folds.     

Omega 3 Fatty Acids Promote Macrophage Reverse Cholesterol Transport in Hamster Fed High Fat Diet

The aim of this study was to investigate macrophage reverse cholesterol transport (RCT) in hamster, a CETP-expressing species, fed omega 3 fatty acids (ω3PUFA) supplemented high fat diet (HFD). Three groups of hamsters (n = 6/group) were studied for 20 weeks: 1) control diet: Control, 2) HFD group: HF and 3) HFD group supplemented with ω3PUFA (EPA and DHA): HFω3. In vivo macrophage-to-feces RCT was assessed after an intraperitoneal injection of ³H-cholesterol-labelled hamster primary macrophages.Compared to Control, HF presented significant (p<0.05) increase in body weight, plasma TG (p<0.01) and cholesterol (p<0.001) with an increase in VLDL TG and in VLDL and LDL cholesterol (p<0.001).Compared to HF, HFω3 presented significant decrease in body weight. HFω3 showed less plasma TG (p<0.001) and cholesterol (p<0.001) related to a decrease in VLDL TG and HDL cholesterol respectively and higher LCAT activity (p<0.05) compared to HF. HFω3 showed a higher fecal bile acid excretion (p<0.05) compared to Control and HF groups and higher fecal cholesterol excretion (p<0.05) compared to HF. This increase was related to higher gene expression of ABCG5, ABCA1 and SR-B1 in HFω3 compared to Control and HF groups (<0.05) and in ABCG1 and CYP7A1 compared to HF group (p<0.05). A higher plasma efflux capacity was also measured in HFω3 using ³H- cholesterol labeled Fu5AH cells.In conclusion, EPA and DHA supplementation improved macrophage to feces reverse cholesterol transport in hamster fed HFD. This change was related to the higher cholesterol and fecal bile acids excretion and to the activation of major genes involved in RCT.     

Bioremediation potential and lead removal capacity of heavy metal-tolerant yeasts isolated from Dayet Oum Ghellaz Lake water (northwest of Algeria)

International Microbiology - Seven metal-resistant yeast strains were isolated and selected from Dayet Oum Ghellaz Lake water (northwest of Algeria) known as a highly polluted area by lead and...     

Feeding ecology of the vulnerable aoudad (Ammotragus lervia) in north-western Sahara

The feeding ecology of the aoudad (Ammotragus lervia) was investigated for the first time in north-western Sahara, Djebel Antar (Bechar province, Algeria), from autumn 2015 to summer 2016. Microhistological analyses of faeces revealed an annual diet composed of 23 identified taxa belonging to 16 plant families. The highest species diversity was recorded in spring and summer (23 species), despite a marked consumption of two species: Vachellia tortilis (17.7%) and Avena sterilis (14.0%); diet diversity was lower in autumn and winter (16 species), including mainly Teucrium polium (14.7%, 21.0%) and Gymnocarpos decander (19.7%, 10.0%). The main plant parts consumed during these seasons were stems (77.7%, 65.3%), while leaves and inflorescences were mostly consumed during spring and summer (54.7%, 52.3%). Forbs dominated the aoudad's diet, with 46.3% average relative abundance, including mainly T. polium, Limoniastrum feei, and Chrysanthemum macrocarpum. Woody plants including mainly V. tortilis and G. decander accounted for 33.3% (50.0% in autumn), and grasses including A. sterilis and H. murinum for 20.4% (32.0% in summer). Based on this diet, A. lervia can be classified as a “generalist mixed-feeder.”     

Prebiotics: tools to manipulate the gut microbiome and metabolome

Journal of Industrial Microbiology & Biotechnology - The human gut is an ecosystem comprising trillions of microbes interacting with the host. The composition of the microbiota and their...     

Development of functional foods using psyllium husk and wheat bran fractions: Phytic acid contents

Wheat grain is a rich source of phosphorus which is present mostly as phytic acid and is distributed mainly in the bran and germ fractions. Phytic acid has now been recognized as an important phytochemical having antioxidant properties. This study deals with the determination of total as well as phytic phosphorus contents of psyllium (PS), course (CB) and fine wheat bran (FB) enriched pan bread and Arabic flat bread. The concentration of phytic acid in CB, FB, wheat germ, wholegrain wheat flour (WGF), white wheat flour (WWF), and psyllium were found to be 8.86 mg/g, 8.52 mg/g, 6.05 mg/g, 1.74 mg/g, 0.46 mg/g and 0.02 mg/g, respectively. Most of the phosphorus existed as phytic phosphorus (74.7–90.8%) in FB, CB, germ, and WGF as compared to only 42.6% in WWF. The level of phytic phosphorus in pan bread containing 10% CB, 20% FB (both containing with 5% PS) was found to be 0.63 mg/g and 1.53 mg/g respectively, as compared to only 0.34 mg/g in WWF pan bread, and 0.90 mg/g in WGF pan bread. The phytic phosphorus content in Arabic bread made with WGF and 3% psyllium was 1.32 mg/g as compared to only 0.48 mg/g in WWF Arabic flat bread. The results obtained indicate that the level of phytic phosphorus significantly increased in bread formulations containing CB, FB, and WGF, but no change with psyllium addition was observed. Adding these wheat mill fractions, and psyllium will enable bakeries not only to produce fiber-enriched pan bread and Arabic bread but would also benefit consumers to increase their dietary fiber intakes, and health-promoting phytochemicals coming from wheat bran and germ fractions.