Kinetics of nitrobenzene degradation coupled to indigenous microorganism dissimilatory iron reduction stimulated by emulsified vegetable oil.

Widespread contamination by nitrobenzene (NB) in sediments and groundwater requires better understanding of the biogeochemical removal process of the pollutant. NB degradation, coupled with dissimilatory iron reduction, is one of the most efficient pollutant removal methods. However, research on NB degradation coupled to indigenous microorganism dissimilatory iron reduction stimulated by electron donors is still experimental. A model for remediation in an actual polluted site does not currently exist. Therefore, in this study, the dynamics was derived from the Michaelis-Menten model (when the mass ratio of emulsified vegetable oil and NB reached the critical value 91:1). The effect of SO42-, NO3-, Ca2+/Mg2+, and the grain size of aquifer media on the dynamics were studied, and the NB degradation dynamic model was then modified based on the most significant factors. Utilizing the model, the remediation time could be calculated in a contaminated site.

Selective Enrichment of Omega-3 Fatty Acids in Oils by Phospholipase A1.

Omega fatty acids are recognized as key nutrients for healthier ageing. Lipases are used to release ω-3 fatty acids from oils for preparing enriched ω-3 fatty acid supplements. However, use of lipases in enrichment of ω-3 fatty acids is limited due to their insufficient specificity for ω-3 fatty acids. In this study use of phospholipase A1 (PLA1), which possesses both sn-1 specific activity on phospholipids and lipase activity, was explored for hydrolysis of ω-3 fatty acids from anchovy oil. Substrate specificity of PLA1 from Thermomyces lenuginosus was initially tested with synthetic p-nitrophenyl esters along with a lipase from Bacillus subtilis (BSL), as a lipase control. Gas chromatographic characterization of the hydrolysate obtained upon treatment of anchovy oil with these enzymes indicated a selective retention of ω-3 fatty acids in the triglyceride fraction by PLA1 and not by BSL. 13C NMR spectroscopy based position analysis of fatty acids in enzyme treated and untreated samples indicated that PLA1 preferably retained ω-3 fatty acids in oil, while saturated fatty acids were hydrolysed irrespective of their position. Hydrolysis of structured triglyceride,1,3-dioleoyl-2-palmitoylglycerol, suggested that both the enzymes hydrolyse the fatty acids at both the positions. The observed discrimination against ω-3 fatty acids by PLA1 appears to be due to its fatty acid selectivity rather than positional specificity. These studies suggest that PLA1 could be used as a potential enzyme for selective concentrationof ω-3 fatty acids.

A new arginase enzymatic reactor: development and application for the research of plant-derived inhibitors.

This work was dedicated to the development of a new micro immobilized enzyme reactor (IMER) by using an in situ procedure. Arginase was covalently immobilized on an ethylenediamine (EDA) monolithic convective interaction media (CIM) disk (12mm × 3mm i.d.) previously derivatized with glutaraldehyde. The activity of this IMER was investigated by inserting this micro-IMER in a HPLC system. The effect of the arginase inhibitors was evaluated by the simultaneous injection of each inhibitor with the nitro guanidino benzene (NGB) substrate. The relative IC50 values were found in agreement with those derived by the conventional spectrometric method. This arginase micro-IMER system was also used to study the effects of plant-derived products on the arginase activity. The pet ether extract from the stem bark of the plant Ficus glomerata Roxob. and the procyanidin oligomers of cocoa and chocolate inhibit the arginase activity. Our results confirmed the direct effect of some plant extracts on the arginase activity and their interest in therapies for treating several NO-dependent smooth disorders.

Omega-3 Fatty acids and inflammation: novel interactions reveal a new step in neutrophil recruitment.

Inflammation is a physiological response to tissue trauma or infection, but leukocytes, which are the effector cells of the inflammatory process, have powerful tissue remodelling capabilities. Thus, to ensure their precise localisation, passage of leukocytes from the blood into inflamed tissue is tightly regulated. Recruitment of blood borne neutrophils to the tissue stroma occurs during early inflammation. In this process, peptide agonists of the chemokine family are assumed to provide a chemotactic stimulus capable of supporting the migration of neutrophils across vascular endothelial cells, through the basement membrane of the vessel wall, and out into the tissue stroma. Here, we show that, although an initial chemokine stimulus is essential for the recruitment of flowing neutrophils by endothelial cells stimulated with the inflammatory cytokine tumour necrosis factor-alpha, transit of the endothelial monolayer is regulated by an additional and downstream stimulus. This signal is supplied by the metabolism of the omega-6-polyunsaturated fatty acid (n-6-PUFA), arachidonic acid, into the eicosanoid prostaglandin-D(2) (PGD(2)) by cyclooxygenase (COX) enzymes. This new step in the neutrophil recruitment process was revealed when the dietary n-3-PUFA, eicosapentaenoic acid (EPA), was utilised as an alternative substrate for COX enzymes, leading to the generation of PGD(3). This alternative series eicosanoid inhibited the migration of neutrophils across endothelial cells by antagonising the PGD(2) receptor. Here, we describe a new step in the neutrophil recruitment process that relies upon a lipid-mediated signal to regulate the migration of neutrophils across endothelial cells. PGD(2) signalling is subordinate to the chemokine-mediated activation of neutrophils, but without the sequential delivery of this signal, neutrophils fail to penetrate the endothelial cell monolayer. Importantly, the ability of the dietary n-3-PUFA, EPA, to inhibit this process not only revealed an unsuspected level of regulation in the migration of inflammatory leukocytes, it also contributes to our understanding of the interactions of this bioactive lipid with the inflammatory system. Moreover, it indicates the potential for novel therapeutics that target the inflammatory system with greater affinity and/or specificity than supplementing the diet with n-3-PUFAs.

Short-term black tea intake modulates the excretion of urinary mutagens in rats treated with 2-amino-3-methylimidazo-[4,5-f]quinoline (IQ): role of CYP1A2 upregulation.

Rats were exposed to black tea (2.5% w/v) as their sole drinking liquid for either 1 day or 1 month, while controls were maintained on water. After this treatment period, all animals received a single oral dose IQ (2-amino-3-methylimidazo-[4,5-f]quinoline), and urine was collected for 48 h. Mutagenic activity of the urine was determined in the Ames test in the presence and absence of an activation system. The excretion of direct-acting mutagens was markedly reduced following tea intake, and was more pronounced after the 1-day treatment. Similarly, both tea treatments suppressed the excretion of indirect-acting mutagens. Furthermore, both tea treatments induced hepatic CYP1A2 activity and expression, but cytosolic glutathione S-transferase activity was only modestly induced in the group of animals receiving tea for 1 day, and only when DCNB (1,2-dichloro-4-nitrobenzene) was used as substrate; glucuronosyl activity was elevated modestly only in the animals receiving the tea for a month. It is concluded that even short-term exposure to black tea is capable of influencing the metabolic fate of IQ, and this is most likely related to the upregulation of CYP1A2.

The carry-through of residues of thiabendazole, tecnazene and chlorpropham from potatoes following manufacture into potato crisps and jacket potato crisps.

Potatoes, commercially treated with thiabendazole, tecnazene and chlorpropham, were processed into potato crisps and jacket potato crisps at a crisp factory using standard manufacturing conditions. A multi-residue method based on gas chromatography with mass spectrometric detection was developed and used to determine pesticide residue levels in the potatoes and potato crisps. Results showed that the residues of all three pesticides were significantly reduced to less than 2% and less than 10% of the maximum theoretical residue carry-through level for potato crisps and jacket potato crisps respectively.

Influence of dietary pectin on intestinal microfloral metabolism and toxicity of nitrobenzene.

Intestinal microfloral metabolism of nitrobenzene is essential for the production of methemoglobin. Since dietary pectin alters intestinal microflora, these studies were designed to examine the effects of dietary pectin on nitrobenzene-induced methemoglobinemia. Male Fischer-344 rats were fed either AIN-76A (purified diet containing 5% cellulose), AIN-76A with 5% pectin replacing the cellulose, or NIH-07 (cereal-based diet containing 8.4% pectin) for 28 days. Following this period, nitrobenzene (200 mg/kg) was administered by gastric intubation, and methemoglobin concentrations were determined after 1, 2, 4, 8, and 24 hr. Nitrobenzene-induced methemoglobinemia was evident as early as 1 hr, peaked at 4 hr, and diminished thereafter in rats fed NIH-07 diet. In contrast, nitrobenzene-induced methemoglobinemia was not detectable in rats fed AIN-76A; however, inclusion of 5% pectin in this diet resulted in methemoglobinemia comparable to that of NIH-07-fed animals at 4, 8, and 24 hr. Administration of 400 or 600 mg/kg nitrobenzene resulted in significant diet-related differences in methemoglobinemia. Administration of 600 mg/kg nitrobenzene to animals fed NIH-07 resulted in the highest methemoglobin concentrations (64 +/- 1%); those fed AIN-76A had the lowest (20 +/- 5%), and those fed AIN-76A containing pectin had intermediate methemoglobin concentrations (44 +/- 6%). No diet-related differences in the microbial population of the stomach or small intestine were observed. However, the number of anaerobes present in the ceca of rats fed AIN-76A containing pectin was 2 to 2.5 times greater than that of rats fed AIN-76A. In vitro reductive metabolism of [14C]nitrobenzene was significantly greater in the cecal contents of rats fed NIH-07 than that in the cecal contents of either of the groups fed the AIN-76A-based diets. These studies indicate that intestinal microfloral metabolism and red blood cell toxicity of nitrobenzene is markedly different in animals fed cereal-based versus purified diets. Furthermore, since inclusion of pectin into the purified diet diminishes the magnitude of these effects, differences in dietary composition of fermentable carbohydrates in cereal-based and purified diets may mediate differences in metabolism and toxicity of nitrobenzene.

Effects of ruminant digestion and metabolism on phenolic monomers of forages.

Immature and mature lucerne (Medicago sativa) and tall fescue (Festuca arundinacea) hays were analysed for their lignin and phenolic monomer (hydroxybenzoic and hydroxycinnamic acid derivatives) contents. These hays were given to four sheep with rumen, duodenal and ileal cannulas, and to a steer with rumen and abomasal cannulas, to investigate the extent and sites of digestion of lignin and phenolic monomers. The hays and digesta samples were analysed for alkali and nitrobenzene-extractable phenolic monomers. Lanthanum oxide was used as an external marker in the digestion studies. Urine was also collected for estimates of total phenolic balance. Grass hays contained greater concentrations of alkali-labile phenolic monomers than did legume hays, whereas legume hays had higher levels of nitrobenzene oxidation products. Mature tall fescue hay had greater concentrations of phenolic monomers than did immature tall fescue hay, but corresponding differences with maturity were not observed with lucerne hays. There were quantitative differences between diets in the digestibilities and in the sites of digestion of phenolic monomers. Digestibilities ranged from 0.191 to 1.004 for alkali-labile compounds and from -1.137 to 0.868 for nitrobenzene oxidation products. Composition of lignin was altered during its passage through the digestive tract. The proportion of phenolic monomers not recovered in urine and faeces varied from -1.014 to 0.871 for individual compounds and differed between diets. Soluble phenolic monomers decreased in concentration as digesta passed from the rumen to the abomasum and duodenum, but increased again in the ileum; levels varied with diet. Phenolics were rapidly solubilized in the rumen after feeding. Results indicate that forages differ in their phenolic monomer content and in the digestibility of these compounds. Therefore, lignin is not an inert compound in the digestive tract of ruminants as has been previously reported.

Effects of pectin-containing diets on the hepatic macromolecular covalent binding of 2,6-dinitro-[3H]toluene in Fischer-344 rats.

The influence of diets varying in pectin content on intestinal microfloral metabolic capacity of rats has been investigated as a possible mechanism for the alteration of toxicity of 2,6-dinitrotoluene (2,6-DNT) produced by these diets. Male F-344 rats were fed a purified diet (AIN-76A), AIN-76A plus 5% or 10% citrus pectin, or either of two cereal-based diets that vary in pectin content, NIH-07 or Purina Chow 5002. After 28 days, rats were given tritium-labeled 2,6-DNT (10 or 75 mg/kg po) and killed 12 hr later. Total hepatic macromolecular covalent binding (CVB) was determined by exhaustive extraction. The CVB of 2,6-DNT was found to be independent of diet at 10 mg/kg. However, at 75 mg/kg CVB was increased 40% by feeding 5% pectin in the purified diet and 90% by feeding 10% pectin in the purified diet. Animals fed Purina 5002 and NIH-07 had 135 and 150% higher CVB, respectively, than animals fed the purified diet alone and significantly greater CVB than animals fed the pectin supplemented diets. Elevated (two- to threefold) beta-glucuronidase and nitroreductase activities, microfloral enzymes proposed to be involved in the activation of 2,6-DNT to a toxicant, were found in the cecal contents of animals fed the pectin-containing diets which correlated with a two- to threefold increase in total number of cecal anaerobes. These results suggest that pectin-induced changes in microflora may enhance hepatoxicity after high doses of 2,6-DNT.

Mutagenicity of anti-cancer nitrobenzofuroxans.

Anti-leukaemically active benzofuroxans were tested for mutagenicity in Salmonella typhimurium. Mutagenicity was not found to be correlated to the previously established anti-leukaemic activity. One anti-leukaemically inactive compound after exposure to liver microsomal enzymes proved the most mutagenic of the derivatives for TA100, whereas after similar treatment, the mutagenicity of the most potent anti-leukaemic compound was reduced. All twelve derivatives tested were mutagenic in a base-substitution strain which was defective in excision-repair and also carried a plasmid-linked repair deficiency. Mutagenicity of five dervatives was undetectable in strains proficient for one or the other of the above repair pathways. Nine of the benzofuroxans could also be detected as mutagens in the frameshift tester strain TA98.