MODEL DEVELOPMENT AND PROCESS OPTIMIZATION FOR SOLVENT EXTRACTION OF POLYPHENOLS FROM RED GRAPES USING BOX–BEHNKEN DESIGN

The objective of the present study is to find out the optimum extraction conditions for extraction of polyphenols from red grapes using Box–Behnken design. Red grapes polyphenols were extracted using acid–ethanol solvent at various extraction temperature (40–60°C), extraction time (20–100 min) and different solid–liquid ratio (1:5–1:15 g:ml). The effect (main and interactive) of extraction conditions on total anthocyanin, phenolic and flavonoid content were studied using Box–Behnken design (three factors at three levels). The results showed that the contribution of the quadratic model was significant for all the responses. Second-order mathematical regression models were developed and were found to fit well with observed data. Derringer's desirability function methodology was performed to find out the optimal conditions based on both individual and combinations of all responses (extraction temperature: 57°C, time: 61 min, and solid–liquid ratio: 1:8.7 g:ml) were established. At this optimal condition, the anthocyanin yield, total phenolic and flavonoid content were 73.92 mg/100 g, 221.4 mg GAE/100 g, and 79.08 mg CE/100 g, respectively. A desirability value of 0.902 was achieved at this point.     

Partial Purification and Characterization of Chromate Reductase of a Novel Ochrobactrum sp. Strain Cr-B4

Hexavalent chromium contamination is a serious problem due to its high toxicity and carcinogenic effects on the biological systems. The enzymatic reduction of toxic Cr(VI) to the less toxic Cr(III) is an efficient technology for detoxification of Cr(VI)-contaminated industrial effluents. In this regard, a chromate reductase enzyme from a novel Ochrobactrum sp. strain Cr-B4, having the ability to detoxify Cr(VI) contaminated sites, has been partially purified and characterized. The molecular mass of this chromate reductase was found to be 31.53 kD, with a specific activity 14.26 U/mg without any addition of electron donors. The temperature and pH optima for chromate reductase activity were 40°C and 8.0, respectively. The activation energy (E_a) for the chromate reductase was found to be 34.7 kJ/mol up to 40°C and the activation energy for its deactivation (E_d) was found to be 79.6 kJ/mol over a temperature range of 50–80°C. The frequency factor for activation of chromate reductase was found to be 566.79 s^?1, and for deactivation of chromate reductase it was found to be 265.66 × 10³ s^?1. The reductase activity of this enzyme was affected by the presence of various heavy metals and complexing agents, some of which (ethylenediamine tetraacetic acid [EDTA], mercaptoethanol, NaN₃, Pb²⁺, Ni²⁺, Zn²⁺, and Cd²⁺) inhibited the enzyme activity, while metals like Cu²⁺ and Fe³⁺ significantly enhanced the reductase activity. The enzyme followed Michaelis–Menten kinetics with K_m of 104.29 µM and a V_max of 4.64 µM/min/mg.     

Phylogenetic relationships of Italian Bellevalia species (Asparagaceae), inferred from morphology,karyology and molecular systematics

The seven Bellevalia species and subspecies known from Italy, representing about 10% of the genus and three out of six sections, were studied. An integrated morphological, karyological and molecular approach was used to infer phylogenetic and systematic relationships among them. B. romana (the generitype) is the most distinctive species on karyotype asymmetry grounds. B. boissieri and B.dubia, usually considered as subspecies of one species (the latter endemic to Sicily), deserve specific status based on biparental nrDNA markers (internal transcribed spacer, ITS), since they do not form a single clade. The allotetraploid endemic B. pelagica, morphologically similar to B. romana, is sister to the latter under parsimony, both in morphological and ITS trees; it is also related with B. dubia, based on karyotype asymmetry and a uniparental cpDNA marker (trnL^(UAA)–trnF^(GAA) IGS (intergenic spacer)). A second allotetraploid endemic, B. webbiana, is closely related, on morphological, karyological and molecular grounds, with B. boissieri and B. ciliata, and also with B. trifoliata, three species that might all involved in its origin. B. sect. Conicae Feinbr. and sect. Nutantes Feinbr. are here typified, the former (type: B. ciliata) is most likely a synonym of the latter (type: B. trifoliata).     

Synthesis and characterization of 5,7-dihydroxyflavanone derivatives as novel protein tyrosine phosphatase 1B inhibitors

A series of 5,7-dihydroxyflavanone derivatives were synthesized and identified as reversible and competitive protein tyrosine phosphatase (PTP) 1B inhibitors with IC₅₀ values in the micromolar range. Compound 4k had the most potent in vitro inhibition activity against PTP1B (IC₅₀ = 2.37?±?0.37 μM) and the greatest selectivity (3.7-fold) for PTP1B relative to T-cell protein tyrosine phosphatase. Cell-based studies revealed that 4k was membrane-permeable and enhanced insulin receptor tyrosine phosphorylation in CHO/hIR cells.     

His22 of TLXI plays a critical role in the inhibition of glycoside hydrolase family 11 xylanases

Recently, a novel wheat thaumatin-like protein, TLXI, which inhibits microbial glycoside hydrolase family (GH) 11 xylanases has been identified. It is the first xylanase inhibitor that exerts its inhibition in a non-competitive way. In the present study we gained insight into the interaction between TLXI and xylanases via combined molecular modeling and mutagenic approaches. More specifically, site-specific mutation of His22, situated on a loop which distinguishes TLXI from other, non-inhibiting, thaumatin-like proteins, and subsequent expression of the mutant in Pichia pastoris resulted in a protein lacking inhibition capacity. The mutant protein was unable to form a complex with GH11 xylanases. Based on these findings, the interaction of TLXI with GH11 xylanases is discussed.     

Role for protein–protein interaction databases in human genetics

Proteomics and the study of protein–protein interactions are becoming increasingly important in our effort to understand human diseases on a system-wide level. Thanks to the development and curation of protein-interaction databases, up-to-date information on these interaction networks is accessible and publicly available to the scientific community. As our knowledge of protein–protein interactions increases, it is important to give thought to the different ways that these resources can impact biomedical research. In this article, we highlight the importance of protein–protein interactions in human genetics and genetic epidemiology. Since protein–protein interactions demonstrate one of the strongest functional relationships between genes, combining genomic data with available proteomic data may provide us with a more in-depth understanding of common human diseases. In this review, we will discuss some of the fundamentals of protein interactions, the databases that are publicly available and how information from these databases can be used to facilitate genome-wide genetic studies.     

Systemizing the structures and structuring the system

This Keystone symposium, entitled ‘Biomolecular Interactions and Networks: function and disease’, was held in Quebec City, Canada, 7–12 March 2010. The conference was distinctive in that it bridged two fields that may be perceived as having little in common: structural and systems biology. However, the growth in structural and omics data brings these two fields closer and closer. Indeed, in two sections of this article we cover talks on systematic analyses of protein structures, as well as systems level approaches that incorporate structural information. In two other sections, we report studies that aim at charting and analyzing cellular systems, and finally we discuss talks that pointed to the issue of promiscuity in biological networks.     

Tsetse flies,trypanosomes, humans and animals: what is proteomics revealing about their crosstalks?

Human and animal African trypanosomoses, or sleeping sickness and Nagana, are neglected vector-borne parasitic diseases caused by protozoa belonging to the Trypanosoma genus. Advances in proteomics offer new tools to better understand host–vector–parasite crosstalks occurring during the complex parasitic developmental cycle, and to determine the outcome of both transmission and infection. In this review, we summarize proteomics studies performed on African trypanosomes and on the interactions with their vector and mammalian hosts. We discuss the contributions and pitfalls of using diverse proteomics tools, and argue about the interest of pathogenoproteomics, both to generate advances in basic research on the best knowledge and understanding of host–vector–pathogen interactions, and to lead to the concrete development of new tools to improve diagnosis and treatment management of trypanosomoses in the near future.     

Emerging liquid chromatography–mass spectrometry technologies improving dried blood spot analysis

Dried blood spots (DBS), a micro blood sampling technique, has recently gained interest in drug discovery and development due to its inherent advantages over the conventional whole blood, plasma or serum sample collection. Since the regulatory authorities have agreed to the use of blood as an acceptable biological matrix for drug exposure measurements, its applications have been extended not only to therapeutic drug monitoring but also to toxicokinetic and pharmacokinetic studies. The pharmaceutical industry is keen to promote DBS as a prominent tool in bioanalytical applications due to the financial, ethical and organizational issues involved in clinical trials. This could be accomplished due to the latest advances in modern analytical technology, particularly liquid chromatography–mass spectrometry. The present review discusses some of the emerging liquid chromatography–mass spectrometry technologies in improving DBS analysis for its innovative applications in the development of new drugs.     

Proteomes of pathogenic Escherichia coli/Shigella group surveyed in their host environments

Proteomic studies on Shigella dysenteriae, Shigella flexneri, enterohemorrhagic Escherichia coli and uropathogenic E. coli (UPEC) are reviewed. UPEC causes infections in the urogenital tract, whereas the other species colonize and, to varying degrees, invade the intestinal tract. Type III secretion systems used to breach the mucosal barrier by the intestinal pathogens revealed distinct expression patterns in different host environments. Dynamic adaptations to changes in nutrient availability and oxygen were observed, including increased reliance on anaerobic respiration and mixed acid fermentation in vivo. Utilization of carbon and nitrogen resources by the bacteria varied considerably depending on the host model investigated. Shigellae and UPEC adapted to metal ion sequestration in the mammalian host by enhancing expression of various receptors and transporters for iron and zinc. This appears to reflect the preferred intracellular life stage of Shigella spp. and responses of UPEC to high levels of lipocalin and lactotransferrin in the urinary tract.