Simple and inexpensive Flow Injection Analysis for determination of sucrose using invertase and glucose oxidase immobilised on glass beads

A Flow Injection Analysis (FIA) for sucrose using invertase (E.C. 3.2.1.26), mutarotase (E.C.5.1.3.3) and glucose oxidase (E.C.1.1.3.4) was developed. The enzymes were immobilised on glass beads using glutaraldehyde. The sucrose concentration was related to oxygen saturation. Fall in O₂ concentration, as a result of sucrose oxidation, was detected by a low cost, home-made O₂ electrode. The system was able to measure sucrose from 0.025 to 100mM with a response time of 6min using 200 l of sample, with an apparent Km of 42mM of sucrose. The system has been operated satisfactorily for 50 days without loss any initial activity.     

Production of lipase free of citrinin by Penicillium citrinum

Lipase (Glycerol ester hydrolase E.G. 3.1.1.3) from a Brazilian strain of Penicillium citrinum free of the mycotoxin citrinin has been investigated. Citrinin production was inhibited by using culture medium containing olive oil, soybean oil and corn oil as carbon sources. Potassium concentration and pH play an important role in citrinin production. Potassium concentration lower than 30 mM and pH below 4.5 inhibited the mycotoxin production. P. citrinum produced lipase free of extraneous proteins and citrinin when cultured using, as nitrogen source, ammonium sulphate (lipase activity of 7.88 U/mg) and yeast extract (lipase activity of 4.95 U/mg) with olive oil as carbon source. This data is relevant to the larger scale production of lipases for food technology applications, from Penicillium citrinum.     

PCR amplification of the rDNA internal transcribed spacer region for differentiation of Saccharomyces cultures

Abstract The size of the internal transcribed spacer (ITS) region as measured by gel electrophoresis of PCR products, amplified by primers ITS1 and ITS4, was over 800 bp for all Saccharomyces sensu stricto species, but yeasts belonging to other Saccharomyces species had a shorter ITS region, making this characteristic potentially useful in the identification of Saccharomyces isolates. The ITS product length was homogeneous within the species Saccharomyces cerevisiae .     

Decrease of adenosine deaminase activity and increase of the lipid peroxidation after acute methotrexate treatment in young rats: protective effects of grape seed extract

The methotrexate (MTX) is an anti‐folate used to treat cancer and some inflammatory diseases. The efficacy of MTX is often limited by its severe toxicity. The present study was undertaken to determine whether Grape seed (Cabernet Sauvignon) extract (GSE) could ameliorate the MTX‐induced oxidative injury and the effect on adenosine deaminase activity (ADA) in rats. The rats were pretreated with 50 mg/kg of GSE, i.p., prior to MTX administration (10 mg/kg, i.p.) with a second dose given 4 h and a third dose 16 h after MTX administration. Biochemical parameters were investigated 48 h after the last MTX administration. The administration of MTX increased thiobarbituric acid reactive species (TBARS) levels in hippocampus, kidney and liver, whereas induced a significant decreased in the ADA activity in the cerebral cortex, kidney and liver tissues. MTX administration significantly increased the activity of ALT(alanine aminotransferase) and urea levels and decreased uric acid levels in the serum. Urinary uric acid levels decreased in the MTX group when compared to those of the control group. The GSE along with MTX‐administration significantly reversed these parameters toward to near normal. These results indicated that GSE could reduce hepatic and nephritic damage induced by MTX‐treatment in young rats therefore having free radical scavenging. Copyright © 2009 John Wiley & Sons, Ltd.     

The importance of leaf- and litter-feeding invertebrates as sources of animal protein for the Amazonian Amerindians.

At least 32 Amerindian groups in the Amazon basin use terrestrial invertebrates as food. Leaf- and litter-consuming invertebrates provide the more important, underestimated food sources for many Amerindian groups. Further, litter-consuming earthworms are also an important food resource for the Ye'Kuana (also known as Makiritare) in the Alto Orinoco (Amazonas, Venezuela). By selecting these small invertebrates the Amerindians are choosing their animal food from those food webs in the rainforest which have the highest energy flow and which constitute the greatest renewable stock of readily available nutrients. Here we show that the consumption of leaf- and litter-feeding invertebrates as a means of recovering protein, fat and vitamins by the forest-living peoples offers a new perspective for the development of sustainable animal food production within the paradigm of biodiversity maintenance.     

Development and Validation of a Biomarker for Diarrhea-Predominant Irritable Bowel Syndrome in Human Subjects

Diarrhea-predominant irritable bowel syndrome (IBS) is diagnosed through clinical criteria after excluding “organic” conditions, and can be precipitated by acute gastroenteritis. Cytolethal distending toxin B (CdtB) is produced by bacteria that cause acute gastroenteritis, and a post-infectious animal model demonstrates that host antibodies to CdtB cross-react with vinculin in the host gut, producing an IBS-like phenotype. Therefore, we assessed circulating anti-CdtB and anti-vinculin antibodies as biomarkers for D-IBS in human subjects. Subjects with D-IBS based on Rome criteria (n=2375) were recruited from a large-scale multicenter clinical trial for D-IBS (TARGET 3). Subjects with inflammatory bowel disease (IBD) (n=142), subjects with celiac disease (n=121), and healthy controls (n=43) were obtained for comparison. Subjects with IBD and celiac disease were recruited based on the presence of intestinal complaints and histologic confirmation of chronic inflammatory changes in the colon or small intestine. Subjects with celiac disease were also required to have an elevated tTG and biopsy. All subjects were aged between 18 and 65 years. Plasma levels of anti-CdtB and anti-vinculin antibodies were determined by ELISA, and compared between groups. Anti-CdtB titers were significantly higher in D-IBS subjects compared to IBD, healthy controls and celiac disease (P<0.001). Anti-vinculin titers were also significantly higher in IBS (P<0.001) compared to the other groups. The area-under-the-receiver operating curves (AUCs) were 0.81 and 0.62 for diagnosis of D-IBS against IBD for anti-CdtB and anti-vinculin, respectively. Both tests were less specific in differentiating IBS from celiac disease. Optimization demonstrated that for anti-CdtB (optical density≥2.80) the specificity, sensitivity and likelihood ratio were 91.6%, 43.7 and 5.2, respectively, and for anti-vinculin (OD≥1.68) were 83.8%, 32.6 and 2.0, respectively. These results confirm that anti-CdtB and anti-vinculin antibodies are elevated in D-IBS compared to non-IBS subjects. These biomarkers may be especially helpful in distinguishing D-IBS from IBD in the workup of chronic diarrhea.     

Gene and cell survival: lessons from prokaryotic plasmid R1

Plasmids are units of extrachromosomal genetic inheritance found in all kingdoms of life. They replicate autonomously and undergo stable propagation in their hosts. Despite their small size, plasmid replication and gene expression constitute a metabolic burden that compromises their stable maintenance in host cells. This pressure has driven the evolution of strategies to increase plasmid stability--a process accelerated by the ability of plasmids to transfer horizontally between cells and to exchange genetic material with their host and other resident episomal DNAs. These abilities drive the adaptability and diversity of plasmids and their host cells. Indeed, survival functions found in plasmids have chromosomal homologues that have an essential role in cellular responses to stress. An analysis of these functions in the prokaryotic plasmid R1, and of their intricate interrelationships, reveals remarkable overall similarities with other gene- and cell-survival strategies found within and beyond the prokaryotic world.     

3-Methylcrotonylglycine Disrupts Mitochondrial Energy Homeostasis and Inhibits Synaptic Na+,K+-ATPase Activity in Brain of Young Rats

Deficiency of 3-methylcrotonyl-CoA carboxylase activity is an inherited metabolic disease biochemically characterized by accumulation and high urinary excretion of 3-methylcrotonylglycine (3MCG), and also of 3-hydroisovalerate in lesser amounts. Affected patients usually have neurologic dysfunction, brain abnormalities and cardiomyopathy, whose pathogenesis is still unknown. The present study investigated the in vitro effects of 3MCG on important parameters of energy metabolism, including CO₂ production from labeled acetate, enzyme activities of the citric acid cycle, as well as of the respiratory chain complexes I–IV (oxidative phosphorylation), creatine kinase (intracellular ATP transfer), and synaptic Na⁺,K⁺-ATPase (neurotransmission) in brain cortex of young rats. 3MCG significantly reduced CO₂ production, implying that this compound compromises citric acid cycle activity. Furthermore, 3MCG diminished the activities of complex II-III of the respiratory chain, mitochondrial creatine kinase and synaptic membrane Na⁺,K⁺-ATPase. Furthermore, antioxidants were able to attenuate or fully prevent the inhibitory effect of 3MCG on creatine kinase and synaptic membrane Na⁺,K⁺-ATPase activities. We also observed that lipid peroxidation was elicited by 3MCG, suggesting the involvement of free radicals on 3MCG-induced effects. Considering the importance of the citric acid cycle and the electron flow through the respiratory chain for brain energy production, creatine kinase for intracellular energy transfer, and Na⁺,K⁺-ATPase for the maintenance of the cell membrane potential, the present data indicate that 3MCG potentially impairs mitochondrial brain energy homeostasis and neurotransmission. It is presumed that these pathomechanisms may be involved in the neurological damage found in patients affected by 3-methylcrotonyl-CoA carboxylase deficiency.