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In this study, the relationship between the inhibitory effects of extracts from rye-buckwheat ginger cakes supplemented with low and high rutin dosage baked without or with dough fermentation step on the formation of fluorescent advanced glycation end-products (AGEs), and phenolic compounds, rutin, D-chiro-inositol and antioxidant capacity were addressed. The cakes were based on rye flour substituted by light buckwheat flour or flour from roasted buckwheat groats at 30% level, and were produced with or without dough fermentation step. The inhibitory effect against AGEs formation was studied in bovine serum albumin (BSA)-glucose and BSA-methylglyoxal (MGO) systems. The antioxidant capacity was measured by 2,2-diphenyl--1-picrylhydrazyl (DPPH) and cyclic voltammetry (CV), rutin and D-chiro-inositol contents by HPLC and total phenolics (TPC) by spectrophotometry assays. The study showed the inhibitory effects of extracts from rye-buckwheat ginger cakes supplemented with low and high rutin dosage. The results of the inhibitory activity were highly correlated in two applied model systems. Enrichment of rye-buckwheat ginger cakes with rutin improved their antioxidant properties. The correlation studies showed that the inhibitory effects of rye-buckwheat ginger cakes produced with dough fermentation step and enhanced with rutin against formation of AGEs were highly correlated with TPC, rutin and D-chiroinositol contents, and antioxidant capacity. Moreover, the effect of rutin enrichment was clearly seen in cakes obtained with dough fermentation step, even the inhibitory activity was slightly lower as compared to the cakes produced without dough fermentation.
 Exposure to environmental pollutants often leads to an upsurge in the production of reactive oxygen species (ROS). ROS oxidize cellular fatty acids to produce lipid peroxyl radicals, subsequently transformed into lipid peroxides, which decrease membrane fluidity and increase the activity of various enzymes implicated in degenerative diseases and cancer formation. Edible plants that contain exogenous compounds like curcumeroid, β-carotene, turmeric, and so on, protect the aerobic cells from oxidation of free radicals. This study thus evaluates antioxidant and antimutagenic activities of ethyl acetate, aqueous and methanolic fractions of Holarrhena floribunda leaves. Inhibitory activities of the ethyl acetate fraction on Fe2+-induced lipid peroxidation in hen egg yolk; rat liver and brain tissues were also evaluated. The Allium cepa root assay was used to evaluate antimutagenic activity. Results showed that the ethyl acetate scavenged DPPH, OH•, and •O2- much stronger than other fractions, as evidenced by its lowest respective IC50 values. All the fractions displayed antimutagenic activities against cyclophosphamide-induced chromosomal aberrations. Likewise, all the fractions induced a reduction in mitotic index, a hallmark of cytotoxicity in the root meristem of Allium cepa. The decrease in mitotic index was most profound for the ethyl acetate fraction, which also demonstrated a significant lipid peroxidation inhibitory activity in the liver and brain homogenates, but not in egg yolk, compared with the ascorbic acid standard. In general, the results suggest that the ethyl acetate fraction might contain beneficial phytochemicals that should be explored as novel candidates for preclinical drug development.
The aim of this work was to determine the activity of proteases and protease inhibitors sampled from the body surface of tau-fluvalinate-sensitive and resistant V. destructor. Proteins were isolated from the tau-fluvalinate-sensitive and resistant mites, while mites untreated with tau-fluvalinate constituted the control. Subsequently, the following methodology was applied: protein concentration assay by the Lowry method — as modified by Schacterle and Pollack; assay of proteolytic activity in relation to various substrates (gelatine, haemoglobin, ovoalbumin, albumin, cytochrome C, casein) by the modified Anson method; identification of proteolytic activity in relation to diagnostic inhibitors of proteolytic enzymes (pepstatin A, PMSF, iodoacetamide, o-phenantrolin), using the Lee and Lin method; identification of acidic, neutral and basic protease activities by means of the modified Anson method; electrophoretic analysis of proteins in a polyacrylamide gel for protease detection with the Laemmli method and for protease inhibitor detection with the Felicioli method. The highest value of protein concentration was found in the tau-fluvalinate-sensitive V. destructor, while the highest activity levels of acidic, neutral and alkaline proteases were observed in the tau-fluvalinate-resistant mites. Aspartic, serine, thiolic and metallic proteases were found in the drug-resistant and drug-sensitive Varroa mites. The control samples were found to contain aspartic and serine proteases. In an acidic and alkaline environment, the results revealed a complete loss of inhibitor activities in the in vitro analyses and electrophoresis. Serine protease inhibitor activities (at pH 7.0) were high, especially in the group of tau-fluvalinate-resistant mites.
Increasing the potency of antihypertensive food-derived peptides is a critical and important step in the development of natural drugs for cardiovascular diseases prevention. We have proposed the egg-white protein precipitate (EWPP) obtained as a byproduct of cystatin and lysozyme isolation as a potential source of ACE-inhibitory peptides derived by pepsin digestion. The results indicated that hydrolysis of EWPP with pepsin produced the ACE inhibitory activity. During 3-h hydrolysis (DH: 38.3%), the IC50 value of EWPP hydrolysate was signifi cantly increased and fi nally reached IC50=643.1 μg/mL. This hydrolysate was further fractionated by RP-HPLC. The peptide fraction exhibiting the highest ACE inhibitory activity was rechromatographed. Three peptide subfractions exhibiting ACE-inhibitory activities of 69.0, 25.0, and 37.6 μg/mL were further characterised. In each of them, mixtures of peptides with different molecular masses were observed.
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