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Interactions of the gut microflora and the host in toxicology.

Food components and ingested chemicals may be metabolized in the gut, not only by digestive and intestinal mucosal enzymes, but also by the resident bacteria, which are found in greatest numbers in the large intestine. The gut microflora is a large (about 10(11) organisms per g colon contents) and diverse (over 400 species) population of organisms and possesses a correspondingly diverse range of metabolic activities, including reductions, hydrolyses and degradations. In many cases, these reactions both complement and antagonize those of the liver, which are mainly oxidative and synthetic. The metabolism, by the gut flora, of chemicals ingested in food or secreted in bile can have numerous toxicological sequelae, including activation to more toxic, mutagenic or carcinogenic derivatives, detoxication and enterohepatic circulation. These toxicological consequences may be modified by changes in the flora due to diet, drugs and interindividual differences. Examples presented of the interaction between the gut flora and its host are the influence of intestinal bacteria on hepatic tumor incidence in mice, the effect of bacterial hydrolysis of rutin on the activity of hepatic enzymes which activate dietary carcinogens, and the role of the gut flora in demethylation and detoxication of methylmercury.

Animals↗

Antioxidant properties of complexes of flavonoids with metal ions.

The formation of complexes of metal ions with the flavonoids quercetin (L1), rutin (L2), galangin (L3) and catechin (L4) has been investigated by UV-visible spectroscopy. The antioxidant activities of the compounds were evaluated by using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicalscavenging method. In this work, we have shown that the complexed flavonoids are much more effective free radical scavengers than the free flavonoids. We suggest that the higher antioxidant activity of the complexes is due to the acquisition of additional superoxide dismutating centers. Radical scavenging activities of the compounds were also investigated from an electrochemical point of view. There is a relationship between the logarithm of the antioxidant activity (represented by EC50) and the oxidation potential. The synergic effect of the complexes and ascorbic acid were studied by [13C]-NMR analyses. The results show that ascorbic acid can protect flavonoids from oxidative degradation, and reveal antioxidant synergies between ascorbic acid and the compounds.

Antioxidants↗

Field survey of Glycyrrhiza plants in Central Asia (3). Chemical characterization of G. glabra collected in Uzbekistan.

The chemical characteristics of Glycyrrhiza glabra L. were investigated at a habitat in Uzbekistan. HPLC analysis of the underground parts indicated that glycyrrhizin contents varied from 3.3 to 6.1% of dry weight, and that glabridin, a species-specific flavonoid for G. glabra, was detected in all underground samples (0.08-0.35% of dry weight). HPLC analysis of the leaves indicated that G. glabra plants collected in the present study could be divided into two types, RT-type and IQ-type, according to their major flavonol glycosides, rutin or isoquercitrin, respectively.

Asia, Central↗

In vivo modulation of signaling factors involved in cell survival.

In vivo expression of cell survival factors protein kinase C (PKC), nuclear factor kappaB (NFkappaB), and extracellular signal-regulated kinase (Erk), which may contribute to the development of radioresistance following radiotherapy, was looked for. Their modulation with natural compounds (curcumin, rutin or nicotinamide) was attempted in mice bearing a serially transplanted fibrosarcoma. Expression of protein kinase C was isoform specific. No translocation of any of the isozymes was noticed following gamma-irradiation as has been reported elsewhere. None of the isoforms could be significantly inhibited by the modulators. However, significant inhibition of radiation-induced ERK and NFkappaB was observed with both curcumin and nicotinamide. Therefore we conclude that use of inhibitors of MAP kinases or NFkappaB may be a more promising strategy to enhance tumour cell killing or to prevent the development of radioresistance during radiotherapy.

Animals↗

Isolation and measurement of quercetin glucosides in flower buds of Japanese butterbur (Petasites japonicus subsp. gigantea Kitam.).

Three quercetin glucosides were isolated from flower buds of Japanese butterbur (Petasites japonicus subsp. gigantea Kitam.) together with caffeic acid as the ingredients that had DPPH radical scavenging activity, using the DPPH-HPLC method for measuring the radical scavenging activity. These quercetin glucosides were identified as quercetin 3-O-beta-D-glucoside, quercetin 3-O-beta-D-6''-O-acetylglucoside, and rutin, and the amounts of the glucosides in flower buds were also examined by HPLC. The flower buds were harvested from four different sites, the total amount of quercetin glucosides in each site was 100-170 mg/100 g fr. wt., and there were no great differences of the amounts between growing fields.

Antioxidants↗

Quercetin cumulatively enhances copper induction of metallothionein in intestinal cells.

Wilson's disease, a genetic copper-overload condition, is currently treated with zinc because of the ability of zinc to induce metallothionein. We are interested in nonmetal chemicals that may alter intestinal copper metabolism and thus help to alleviate copper toxicity. Previously, we have shown that quercetin, a dietary flavonoid, can chelate copper. This study further examined the interaction of quercetin and copper in intestinal epithelial cells. We found that quercetin enhanced metallothoinein induction by copper and the effect was dose dependent. Quercetin also exerted a cumulative effect after repeated exposure. Repeated low-dose treatment (3-10 microM) of cells with quercetin can lead to the same effect on metallothoinein as one higher concentration treatment (100 microM). This property of quercetin is distinct from its chemical interaction with copper, but both can contribute to a reduction of copper toxicity. Among other flavonoids tested, two other copper chelators, catechin and rutin, did not increase copper induction of metallothionein, whereas genistein, an isoflavone that does not interact with copper chemically, increased copper induction of metallothionein. The effect of quercetin on copper metabolism is unique. Quercetin decreased zinc-stimulated metallothionein expression and had no effect on the cadmium induction of metallothionein. The clinical application of our observation needs to be explored.

Cadmium↗

Evaluation of radioprotective action of compounds using Saccharomyces cerevisiae.

The yeast Saccharomyces cerevisiae serves as a model eukaryotic system to screen radioprotectors that can be used primarily in radiotherapy as well as in occupational workers in nuclear and allied industries. A number of antioxidants are suggested to be radioprotectors by virtue of their ability to quench reactive oxygen species, but their radioprotective action has not been investigated so far. In this study, a number of antioxidants were tested for their efficacy in radioprotection using yeast cells. Sulfhydryl compounds (disulfiram at 100 and 200 microM) and reduced glutathione (10 and 100 mM), purified compounds of plant origin, such as curcumin (1 mM, 10 mM, and 100 mM), quercetin (100 and 500 microM), rutin (100 and 200 microM), ellagic acid (100,200, and 500 microM) and gallic acid (100 and 500 microM)-were studied. The results revealed that all compounds, except gallic acid, specifically protected normal yeast cells from gamma-radiation damage. Using rad 52 mutants, which lack recombinational DNA repair pathway, it has been found that protection was solely brought about by reducing DNA damage rather than by interfering with DNA repair. Results with DNA repair polymerase further substantiated this contention. We conclude that simple eukaryotic yeast cells can serve as a test system not only for rapid screening of radiomodifiers but also to study their mode of action.

Curcumin↗

Conflicting demands on detoxification pathways influence how common brushtail possums choose their diets.

Most herbivores eat more and survive better when they have access to a variety of foods. One explanation involves the detoxification of plant secondary metabolites (PSMs). By feeding from a variety of plants that contain different classes of PSMs, animals can use multiple detoxification pathways and presumably consume more food. Although popular, this theory is difficult to test because it requires knowledge of the detoxification pathways of each PSM in the diet. We established that common brushtail possums (Trichosurus vulpecula) use various combinations of oxidation, hydrolysis, and conjugation with glucuronic acid (GA) or glycine to detoxify six PSMs. Compared to their ingestion of a single PSM, possums ate more when offered a choice between two diets containing PSMs that require apparently independent detoxification pathways (benzoate and 1,8-cineole, benzoate and p-cymene, benzoate and orcinol, benzoate and salicin, or orcinol and 1,8-cineole). However, possums still did not eat as much of these diets as they did of a basal diet free of PSMs. This suggests that detoxification pathways are never independent, but are separated instead by degrees. In contrast, possums offered a choice of two PSMs that require competing detoxification pathways (1,8-cineole and p-cymene, 1,8-cineole and salicin, or orcinol and salicin) ate no more than when offered diets containing one of the compounds. There was an exception: even though both rutin and orcinol are detoxified via conjugation with GA, the feeding behavior of possums did not suggest competition for detoxification pathways. This implies that the supply of GA is not limiting. This study provides the first convincing evidence that herbivorous mammals can eat more by selecting mixed diets with a diversity of PSMs that make full use of their detoxification potential. It also emphasizes that other behavioral and physiological factors, such as transient food aversions, influence feeding behavior.

Animals↗

Plant polyphenolic-protein conjugates activate murine spleen cells and bind to multiple cell surface components.

Previous studies have shown that a flavonoid-phenolic-rich glycoprotein (TGP) is a mitogen to murine B cells. To test the hypothesis that this could be duplicated by a protein conjugate of the phenolic moiety, the response to rutin (R)-BSA was studied. It was demonstrated that: (I) R-BSA, like TGP, activates mouse B cells to proliferate; (ii) the kinetics of proliferation induced by R-BSA and TGP are similar; (iii) the mitogenic effects of neither R-BSA nor TGP are inhibitable by free flavonoids or phenolics; (iv) there is no age-dependent decrease in the proliferative response to either R-BSA or TGP; (v) both R-BSA and TGP induce spleen cells to secrete IL-6 by 2 hr of culture. They differ in that the proliferative response of congenitally athymic mice to R-BSA, but not to TGP, is significantly lower than that of euthymic mice, and in that TGP seems to stimulate a small subpopulation of T cells to enter cell cycle. The importance of the phenolic moiety in the response to TGP is supported by the observation that R-BSA immunoprecipitates cell surface components that seem to be identical to those precipitated by TGP. The apparent molecular sizes of the bands are approximately 110, 70, 55, 43, 34, 29, and 25-23 kDa. The 2-D analyses of the TGP and R-BSA precipitates are also striking in their similarity. The isoelectric point (pI) of the 28-kDa band is between pH 6.3 to 6.6. A band at approximately 23 kDa has a pI of 6.0, one band at approximately 25 kDa has a pI of approximately 5.4, and five bands ranging in size from approximately 26 to approximately 110 kDa all have a pI in the pH range of 4.6 to 5.4. The presence of multiple binding sites suggests that these compounds might activate cells via multiple distinct pathways.

Age Factors↗

Theoretical elucidation of activity differences of five phenolic antioxidants.

AIM: To verify the effectiveness of structure-activity relationship (SAR) and theoretical calculation methods for antioxidants. METHODS: Preliminary elucidation on the differences of activities of 5 antioxidants was performed by SAR. Then semiempirical quantum chemistry method AM1 was employed to calculate the delta HOF value, the difference between the heat of formation of antioxidant and its free radical, which was used as a theoretical parameter to elucidate the differences of activities of the antioxidants thoroughly. RESULTS: delta HOF values of antioxidants were obtained as follows: ferulic acid, 150.58 kJ.mol-1; anion of ferulic acid, 122.64 kJ.mol-1; modified ferulic acid, 137.70 kJ.mol-1; anion of modified ferulic acid, 118.99 kJ.mol-1; salvianic acid, 134.17 kJ.mol-1; rutin, 137.83 kJ.mol-1, L-EGCG, 124.39 kJ.mol-1; paeonol, 176.79 kJ.mol-1. The differences of the antioxidant activities were elucidated, and how to further enhance the antioxidant activity was investigated as well. CONCLUSION: The SAR and calculation methods are rather effective to elucidate the differences of antioxidant activities, and present some new clues for structural modification of antioxidants to increase their activities.

Acetophenones↗

[Antioxidative activity of histochrome and some other drugs used in ophthalmology].

The antioxidative activity (AOA) of histochrome (2,3,5,6,8-pentahydroxy-7-ethyl-1,4-naphthoquinone) and some other drugs with antioxidant properties used in ophthalmology has been studied in the hemoglobin--hydrogen peroxide--luminol system. The AOA of histochrome, ascorbate, dicinon, rutin, quercetin, and dihydroquercetin is comparable to that of trolox, the reference antioxidant in our studies. The drugs (in physiological concentrations) listed above are characterized by a high AOA. Cerebrolysin was 300 times less active than trolox, and emoxipin showed no antioxidant properties. Lacrimal AOA increased after a parabulbar injection of histochrome. Histochrome is believed to be a perspective antioxidant for the treatment of ocular diseases.

Amino Acids↗

Effects of citrus phytochemicals on liver and lung cytochrome P450 activity and on the in vitro metabolism of the tobacco-specific nitrosamine NNK.

NNK is a potent environmental carcinogen to which both smokers and nonsmokers are exposed. The response to NNK may be affected by factors including nutrition. We investigated the effects of five citrus phytochemicals on the in vitro metabolism of the tobacco-specific nitrosamine NNK and on the dealkylation of methoxyresorufin (MROD) and pentoxyresorufin (PROD) in liver and lung microsomes of the Syrian golden hamster. In the NNK metabolism experiments in vitro incubations contained 3 microCi [5-H3] NNK, 0.5 mg microsomal protein and 0.5 mumole of the citrus phytochemical diosmin, naringin, naringenin, quercetin or rutin. In the dealkylation studies incubations contained 0.5 microM methoxyresorufin or pentoxyresorufin, 0.5 mg microsomal protein and 0.5 mumole of citrus phytochemical. The major NNK metabolism pathway in hamster liver microsomes was NNK-reduction while in lung microsomes it was alpha-hydroxylation. The alpha-hydroxylation pathway produces metabolic products that methylate and pyridyloxobutylate DNA. Naringenin, a metabolite of naringin, and quercetin were the most potent inhibitors of alpha-hydroxylation of NNK in both liver and lung microsomes. This inhibition correlated with a potent inhibition of MROD and PROD activity in liver but not in lung microsomes. The metabolic activation of NNK is associated with cytochrome P450 isoforms 1A1, 1A2, 2B1, 2D6 and 2E1. Our results suggest that naringenin and quercetin from citrus fruits inhibit the activity of cytochrome P450 (CYP) isoforms that activate NNK and may afford protection against NNK-induced carcinogenesis.

Animals↗

[Separation and determination of flavonols in the flowers of Carthamus tinctorius by RP-HPLC].

Carthamus tinctorius L. is a kind of traditional Chinese medicine used in the treatment of uterine congestion, cardio-vascular disease, thrombosis and so on. Eight flavonoids have been isolated from its flowers. A reversed phase HPLC method for the separation and determination of the three flavonols: rutin (I), quercetin(II) and kaempferol(III) in the flowers of Carthamus tinctorius L. is described. The column was 25 cm x 4.6 mm ID, packed with YWG-C18(10 microns). The mobile phase was a mixture of MeOH-H2O-H3PO4(48.5:51.5:0.25, pH 3.5). The flow rate was 1.0 ml.min-1. Daidzein was used as an internal standard. UV detection was performed at 360 nm. The relative standard deviation of the method was 1.1-2.6%. The calibration curves showed good linearity over the range of 0.11-0.80 microgram, gamma = 0.9995-0.9998 and detection limits were 2.0-20.0 ng. The recoveries were 97.8%-98.9% for the three flavonols. The method is simple, fast, sensitive and accurate and has been applied to the analysis of Carthamus tinctorius L. of different sources. It can also be used for the quality control of safflower preparations for clinical uses.

Asteraceae↗

[Principal constituents from flowering aerial parts of wild pansy].

Dried flowering aerial parts of 12 harvested batches and 9 batches of commercial origin from Viola arvensis Murray were examined. The levels of principal compounds averaged respectively: total flavonoids 2.86 and 1.63%, rutin 1.15 and 0.57%, violanthin 0.80 and 0.82%, violarvensin 0.75 and 0.20%, mucilage 21.5 and 16.5%, total ashes 10.6 and 14.8%, potassium 2.75 and 2.85% and also salicy lic acid 0.11 and 0.09%. Saponins were not detected. Specifications were discussed for an European Pharmacopoeial monography.

Europe↗

Inhibitory effect of enzyme therapy and combination therapy with cyclosporin A on collagen-induced arthritis.

OBJECTIVE: There is increasing interest in the use of combination therapy for rheumatoid arthritis and in the possibility of combining the conventional drug approach with newer antirheumatic therapy. The present study investigates the efficacy of long-term prophylactic enzyme therapy and combination therapy with cyclosporin A in rats with collagen-induced arthritis. METHODS: Rats with collagen-induced arthritis were administered the following drugs: cyclosporin A (5 mg/kg/day and 10 mg/kg/day orally); a mixture of enzymes containing pure substances (bromelain, trypsin, rutin) in the same ratio as in Phlogenzym (PHL, 150 mg/kg, twice daily intrarectally); and a combination of 5 mg/kg/day cyclosporin A plus 300 mg/kg/day PHL for a period of 50 days from the immunization. Levels of serum albumin, serum nitrite/nitrate concentrations, changes in hind paw swelling and bone erosions were measured in the rats as variables of inflammation and destructive arthritis-associated changes. RESULTS: Treatment with 10 mg/kg cyclosporin A, as well as combination therapy with half dosages of cyclosporin A (5 mg/kg) plus PHL significantly inhibited both inflammation and destructive arthritis-associated changes. Significant differences in favor of combination therapy with 5 mg/kg CsA + 300 mg/kg PHL as compared to 5 mg/kg CsA alone were seen in hind paw swelling. Also, reduction of the radiographic scores was more significant in the combination therapy group. Five mg cyclosporin A or PHL alone reduced the disease markers studied to a lesser extent, and in the case of enzyme therapy this occurred at a later stage of arthritis development. CONCLUSION: Our results show the inhibitory effect of enzyme therapy on collagen-induced arthritis in rats, as well as the efficacy of cyclosporin A given in low doses in combination with enzyme therapy, which may be useful in the treatment of rheumatoid arthritis.

Animals↗

[Chemical constituents of the leaves of Crataegus scabrifolia (Franch.) Rehd].

OBJECTIVE: To explore the medicinal resources of Crataegus and seek compounds with pharmaceutical effects. METHOD: Column chromatography was employed for the isolation and purification of constituents and the structures were elucidated by spectral analysis and chemical evidence. RESULT: Six compounds were obtained and identified as rutin, hyperoside, vitexin, ursolic acid, daucosterol, and nonacosanol. CONCLUSION: The main constituents of the leaves of Crataegus scabrifolia are flavonoids.

Crataegus↗

Use of the SOS-chromotest spot assay as a screening system for detecting genotoxic compounds in crude plant extracts.

An SOS-chromotest spot assay was used to detect genotoxic compounds in crude plant extracts. The method allows simultaneous testing of extracts from different species in either a liquid or a solid crystalline form. Extracts from two species of the genus Senna, native to the state of Morelos, Mexico, were assayed. Four genotoxic compounds were isolated, and were identified as quercetin and rutin from S. wislizeni, and 5,7-di- O-methylrutin and 5,7-di-O-methylquercetin from S. skinneri. The SOS-chromotest spot assay proved to be useful for activity- guided fractionation at the beginning of screening for genotoxic compounds in crude plant extracts.

Chemical Fractionation↗

Efficacy and safety of phlogenzym--a protease formulation, in sepsis in children.

BACKGROUND: Infections are a major cause of hospitalisation wherein the host mounts an inflammatory response against the infecting agent. Administration of proteolytic enzymes could regulate the host's immune system and help early recovery from sepsis. OBJECTIVE: To test the efficacy and safety of an oral enzyme formulation, Phlogenzym (Mucos Pharma GmbH, Geretsried, Germany; constituents of each enteric-coated tablet were bromelain 90 mg, trypsin 48 mg, rutin 100 mg) as adjuvant therapy in treatment of sepsis in children. SUBJECTS AND METHODS: Double-blind, randomised, controlled phase III study at a tertiary care centre wherein 60 eligible children aged one month to 12 years with sepsis were randomised to receive either phlogenzym (n=30; 17 boys) or placebo (n=30; 22 boys) tablets (1 tablet/10 kg body weight up to maximum six tablets a day in two or three divided doses for 14-21 days) along with appropriate antibiotics and supportive treatment. RESULTS: Median time taken for fever to subside was three days (range 1-12; 95% CI--1.14 to 7.14) in the phlogenzym group vs four days (range 1-18; 95% CI--3.52 to 11.52) in the placebo group (p < 0.05); haemodynamic support was needed for two days (range 1-3; 95% CI--0.84 to 3.16) in the phlogenzym group but three days (range 1-8; 95% CI--0.76 to 5.24) in the placebo group (p < 0.05). The modified Glasgow coma scale score normalized in three days (range 1-14; 95% CI--4.62 to 9.62) in the phlogenzym group vs 5.5 days (range 1-18; 95% CI--2.52 to 13.52) in the placebo group (p > 0.05). Oral feeds could be started in four days (range 1-15; 95% CI--1.74 to 9.74) in the phlogenzym group vs five days (range 1-11; 95% CI--1.26 to 11.26) in the placebo group (p > 0.05). Two patients died in the placebo group. CONCLUSION: Phlogenzym is effective as an adjuvant with antibiotics and supportive treatment for early improvement of pediatric patients with sepsis.

Adjuvants, Immunologic↗