Phospholipase-mediated preparation of 1-ricinoleoyl-2-acyl-sn- glycero-3-phosphocholine from soya and egg phosphatidylcholine

1-Ricinoleoyl-2-acyl-sn-glycero-3-phosphocholine was prepared by incorporating ricinoleic acid completely in the sn-1 position of egg and soya phosphatidylcholine (PC) using immobilized phospholipase A₁ as the catalyst. The optimum reaction conditions for maximum incorporation of ricinoleic acid into PC through transesterification were 10% (w/w) immobilized enzyme (116 mg), a 1:5 mol ratio of PC (soya, 387 mg; egg, 384 mg) to methyl ricinoleate (780 mg) at 50 °C for 24 h in hexane.     

Diurnal variation of milk somatic and differential leukocyte counts of Murrah buffaloes as influenced by different milk fractions,seasons and parities

Milk cell counts are good indicators of a mammary infection and milk quality. The present study was done to record diurnal rhythmicity in the milk somatic and differential cell counts during different seasons, milk strips and parity in Murrah buffaloes. Milk somatic cell counts (SCC) were measured by SCC counter and milk differential cell counts were measured microscopically after making a milk smear and staining it to identify neutrophils, lymphocytes and macrophages. Maximum milk SCC was observed in the summer season. Milk neutrophils were lowest during thermoneutral (TN), intermediate during the winter season and highest during the summer season. Milk lymphocytes were highest during the winter season, intermediate in the TN and lowest in the summer season. Diurnal rhythm in the milk SCC and neutrophil percentage is noticed in the summer season only. Maximum milk SCC values were observed in the late strip but neutrophils were highest in the early strip. Diurnal rhythm was observed in the late strip for neutrophils and in mid strip for the lymphocytes. Milk SCC and milk neutrophils were found to be the highest in the multiparous buffaloes and diurnal rhythm was observed only in the lymphocytes of primiparous buffaloes. Milk macrophages were higher in the morning samples of primiparous as compared to the multiparous buffaloes. In this pioneer study, diurnal rhythms in the milk cell counts of buffaloes have been studied extensively. This will help in maintaining low milk cell counts in buffalo and thus help in getting more milk per buffalo during stress periods.     

Genetic and Chemical Diversity of High Mucilaginous Plants of <Emphasis Type="Italic">Sida</Emphasis> Complex by ISSR Markers and Chemical Fingerprinting

A method was developed based on multiple approaches wherein DNA and chemical analysis was carried out toward differentiation of important species of Sida complex that is being used for commercial preparation. Isolated DNA samples were successfully performed through PCR amplification using ISSR markers and degree of genetic diversity among the different species of Sida is compared with that of chemical diversity. For genetic fingerprint investigation, selected 10 ISSR primers generating reproducible banding patterns were used. Among the total of 63 amplicons, 62 were recorded as polymorphic, genetic similarity index deduced from ISSR profiles ranged from 12 to 51%. Based on similarity index, S. acuta and S. rhombifolia found to be most similar (51%). High number of species-specific bands played pivotal role to delineate species at genetic level. Investigation based on HPTLC fingerprints analysis revealed 23 bands representing to characteristic chemicals and similarity index ranged from 73 to 91%. Prominent distinguishable bands were observed only in S. acuta, while S. cordifolia and S. rhombifolia shared most bands making them difficult to identify on chemical fingerprint basis. This report summarizes the genotypic and chemotypic diversity and the use of profiles for authentication of species of Sida complex.     

Systematic comparison of label-free, metabolic labeling, and isobaric chemical labeling for quantitative proteomics on LTQ Orbitrap Velos

A variety of quantitative proteomics methods have been developed, including label-free, metabolic labeling, and isobaric chemical labeling using iTRAQ or TMT. Here, these methods were compared in terms of the depth of proteome coverage, quantification accuracy, precision, and reproducibility using a high-performance hybrid mass spectrometer, LTQ Orbitrap Velos. Our results show that (1) the spectral counting method provides the deepest proteome coverage for identification, but its quantification performance is worse than labeling-based approaches, especially the quantification reproducibility; (2) metabolic labeling and isobaric chemical labeling are capable of accurate, precise, and reproducible quantification and provide deep proteome coverage for quantification; isobaric chemical labeling surpasses metabolic labeling in terms of quantification precision and reproducibility; and (3) iTRAQ and TMT perform similarly in all aspects compared in the current study using a CID-HCD dual scan configuration. On the basis of the unique advantages of each method, we provide guidance for selection of the appropriate method for a quantitative proteomics study.     

Enhanced vincamine production in selected tryptophan-overproducing shoots of Vinca minor

Vinca minor is the sole source of vincamine, an alkaloid known to be used in a variety of cerebral disorders. Three stable variant shoot lines (V10, V20 and V30) with tolerance thresholds of 10, 20 and 30?mg/l 5-methyltryptophan (5-MT; analogue of tryptophan), respectively, were selected. These lines showed twofold to threefold increase in tryptophan content and 1.5- to 2-fold increment in the total alkaloids in comparison to the wild line shoots. A maximum of 16-fold enhancement in vincamine production was recorded in V30 line followed by eightfold in V20 line. Inter simple sequence repeat (ISSR)-PCR amplification of all the three lines showed total of 65 bands; out of which 60 were monomorphic (92.3?%) and 5 were polymorphic (7.7?%). Tryptophan being a limiting factor in the indole alkaloid pathway plays a crucial role in modulating the flux towards vincamine production and its over-production positively resulted into enhanced vincamine production.     

Gallic acid based steroidal phenstatin analogues for selective targeting of breast cancer cells through inhibiting tubulin polymerization

Phenstatin analogues were synthesized on steroidal framework, for selective targeting of breast cancer cells. These analogues were evaluated for anticancer efficacy against breast cancer cell lines. Analogues 12 and 19 exhibited significant anticancer activity against MCF-7, hormone dependent breast cancer cell line. While analogues 10-14 exhibited significant anticancer activity against MDA-MB-231, hormone independent breast cancer cell line. Compound 10 showed significant oestrogen antagonistic activities with low agonistic activity in in vivo rat model. These analogues also retain tubulin polymerization inhibition activity. The most active analogue 10 was found to be non-toxic in Swiss albino mice up to 300 mg/kg dose. Gallic acid based phenstatin analogues may further be optimized as selective anti-breast cancer agents.     

A temperature-sensitive mutation in the nodal-related gene cyclops reveals that the floor plate is induced during gastrulation in zebrafish

The floor plate, a specialized group of cells in the ventral midline of the neural tube of vertebrates, plays crucial roles in patterning the central nervous system. Recent work from zebrafish, chick, chick-quail chimeras and mice to investigate the development of the floor plate have led to several models of floor-plate induction. One model suggests that the floor plate is formed by inductive signalling from the notochord to the overlying neural tube. The induction is thought to be mediated by notochord-derived Sonic hedgehog (Shh), a secreted protein, and requires direct cellular contact between the notochord and the neural tube. Another model proposes a role for the organizer in generating midline precursor cells that produce floor plate cells independent of notochord specification, and proposes that floor plate specification occurs early, during gastrulation. We describe a temperature-sensitive mutation that affects the zebrafish Nodal-related secreted signalling factor, Cyclops, and use it to address the issue of when the floor plate is induced in zebrafish. Zebrafish cyclops regulates the expression of shh in the ventral neural tube. Although null mutations in cyclops result in the lack of the medial floor plate, embryos homozygous for the temperature-sensitive mutation have floor plate cells at the permissive temperature and lack floor plate cells at the restrictive temperature. We use this mutant allele in temperature shift-up and shift-down experiments to answer a central question pertaining to the timing of vertebrate floor plate induction. Abrogation of Cyc/Nodal signalling in the temperature-sensitive mutant embryos at various stages indicates that the floor plate in zebrafish is induced early in development, during gastrulation. In addition, continuous Cyclops signalling is required through gastrulation for a complete ventral neural tube throughout the length of the neuraxis. Finally, by modulation of Nodal signalling levels in mutants and in ectopic overexpression experiments, we show that, similar to the requirements for prechordal plate mesendoderm fates, uninterrupted and high levels of Cyclops signalling are required for induction and specification of a complete ventral neural tube.