Analysis of betalains from fruits of Opuntia species

Betalains are of great taxonomic significance in higher plants and occur only in 10 families of the order Caryophyllales (Centrospermae). They are water-soluble nitrogenous pigments. They can be divided into two major structural groups, the red to red-violet betacyanins and the yellow betaxanthins. Betalains are widely used as natural red food colorant as well as antioxidant potentials. Several methods have been published for the determination of betalain in fruits of Opuntia species. The purpose of the current review is to provide a systematic survey of the analytical techniques for the determination of betalain from fruits of Opuntia species.     

Spatiotemporal variability in archaeal communities of tropical coastal waters

Spatiotemporal variations in the archaeal community structure from four locations along the central west coast of India were analyzed by denaturing gradient gel electrophoresis (DGGE) profiling and sequencing of prominent bands on the DGGE gels. A total of 36 water samples were collected during pre-monsoon, post-monsoon and monsoon seasons from three depths (surface, mid-depth , and close to bottom ～20 m). The community DNA extracts from these samples were subjected to DGGE analysis. The results of this study clearly indicate a significant difference in the community structure between sampling locations and seasons. Location-wise variation was more pronounced during pre-monsoon and post-monsoon seasons. During monsoon, however, the depth-wise variation was pronounced, suggesting monsoon influence on archaeal community structure. Sequencing of DGGE bands suggested the presence of marine group I and II archaea. Canonical correspondence analysis indicated that nitrate, nitrite and chlorophyll a were the significant explanatory variables responsible for > 60 % variation in archaeal community in these coastal waters.     

Microbial profiles, in vitro gas production and dry matter digestibility based on various ratios of roughage to concentrate

The study assessed the effects of different roughage to concentrate ratios on enteric methane production, rumen fermentation and microbial counts. These ratios were 80:20, 50:50, and 20:80 for diets 1, 2, and 3, respectively. No significant differences were observed in total gas production among diets; however, methane emissions increased (P?<?0.05) with increased roughage in diet. The pH was greater (P?<?0.05) in diet 1 compared to diets 2 and 3 (6.38 vs 6.17 and 6.07). In vitro dry matter digestibility increased with decreased roughage ratios (47.67, 61.67, 67.33 % for diets 1, 2 and 3, respectively). Similarly, total volatile fatty acids (mM/100 mL) also increased with decreased roughage ratios [diet 1 (5.38); diet 2 (6.30); diet 3 (7.37)]. Methanogen counts, total bacterial counts and protozoal counts were lower (P?<?0.05) in diet 3 compared to diet 1 and 2. However, total fungal counts were higher in diet 1 compared to diet 2 and 3. The results indicate that methane emission, enteric fermentation patterns, and change in methanogens population appear only with higher level of roughage. These findings are important for reducing methane without any impact on rumen performance.     

Process optimization of xylanase production using cheap solid substrate by Trichoderma reesei SAF3 and study on the alteration of behavioral properties of enzyme obtained from SSF and SmF

This study aimed to assess the variability in respect of titer and properties of xylanase from Trichoderma reesei SAF3 under both solid-state and submerged fermentation. SSF was initially optimized with different agro-residues and among them wheat bran was found to be the best substrate that favored maximum xylanase production of 219 U (gws)^?1 at 96 h of incubation. The mycelial stage of the fungi and intracellular accumulation of Ca⁺⁺ and Mg⁺⁺ induced maximum enzyme synthesis. Inoculum level of 10 × 10⁶ spores 5 g^?1 of dry solid substrate and water activity of 0.6 were found to be optimum for xylanase production under SSF. Further optimization was made using a Box-Behnken design under response surface methodology. The optimal cultivation conditions predicted from canonical analysis of this model were incubation time (A) = 96–99 h, inoculum concentration (B) = 10 × 10⁶ spores 5 g^?1 of dry substrate, solid substrate concentration (C) = 10–12 g flask^?1, initial moisture level (D) = 10 mL flask^?1 (equivalent to a _w  = 0.55) and the level of xylanase was 299.7 U (gws)^?1. Subsequent verification of these levels agreed (97 % similar) with model predictions. Maximum amount of xylanase was recovered with water (6:1, v/w) and under shaking condition (125 rpm). Purified xylanase from SSF showed better stability in salt and pH, was catalytically and thermodynamically more efficient over enzyme from SmF, though molecular weight of both enzymes was identical (53.8 kDa).     

Cigarette Experimentation and the Population Attributable Fraction for Associated Genetic and Non-Genetic Risk Factors

Background

We, and others, have shown that experimenting with cigarettes is a function of both non-genetic and genetic factors. In this analysis we ask: how much of the total risk of experimenting with cigarettes, among those who had not experimented with cigarettes when they enrolled in a prospective cohort, is attributable to genetic factors and to non-genetic factors?

Methods

Participants (N = 1,118 Mexican origin youth), recruited from a large population-based cohort study in Houston, Texas, provided prospective data on cigarette experimentation over three years. Non-genetic data were elicited twice – baseline and follow-up. Participants were genotyped for 672 functional and tagging variants in the dopamine, serotonin and opioid pathways.

Results

In the overall model, the adjusted combined non-genetic PAF was 71.2% and the adjusted combined genetic PAF was 58.5%. Among committed never smokers the adjusted combined non-genetic PAF was 67.0% and the adjusted combined genetic PAF was 53.5%. However, among cognitively susceptible youth, the adjusted combined non-genetic PAF was 52.0% and the adjusted combined genetic PAF was 68.4%.

Conclusions

Our results suggest there may be differences in genotypes between youth who think they will try cigarettes in the future compared to their peers who think they will not and underscore the possibility that the relative influence of genetic vs. non-genetic factors on the uptake of smoking may vary between these two groups of youth.

Impact

A clearer understanding of the relative role of genetic vs. non-genetic factors in the uptake of smoking may have implications for the design of prevention programs.     

Metabolomic Analysis Reveals Extended Metabolic Consequences of Marginal Vitamin B-6 Deficiency in Healthy Human Subjects

Marginal deficiency of vitamin B-6 is common among segments of the population worldwide. Because pyridoxal 5′-phosphate (PLP) serves as a coenzyme in the metabolism of amino acids, carbohydrates, organic acids, and neurotransmitters, as well as in aspects of one-carbon metabolism, vitamin B-6 deficiency could have many effects. Healthy men and women (age: 20-40 y; n = 23) were fed a 2-day controlled, nutritionally adequate diet followed by a 28-day low-vitamin B-6 diet (<0.5 mg/d) to induce marginal deficiency, as reflected by a decline of plasma PLP from 52.6±14.1 (mean ± SD) to 21.5±4.6 nmol/L (P<0.0001) and increased cystathionine from 131±65 to 199±56 nmol/L (P<0.001). Fasting plasma samples obtained before and after vitamin B6 restriction were analyzed by ¹H-NMR with and without filtration and by targeted quantitative analysis by mass spectrometry (MS). Multilevel partial least squares-discriminant analysis and S-plots of NMR spectra showed that NMR is effective in classifying samples according to vitamin B-6 status and identified discriminating features. NMR spectral features of selected metabolites indicated that vitamin B-6 restriction significantly increased the ratios of glutamine/glutamate and 2-oxoglutarate/glutamate (P<0.001) and tended to increase concentrations of acetate, pyruvate, and trimethylamine-N-oxide (adjusted P<0.05). Tandem MS showed significantly greater plasma proline after vitamin B-6 restriction (adjusted P<0.05), but there were no effects on the profile of 14 other amino acids and 45 acylcarnitines. These findings demonstrate that marginal vitamin B-6 deficiency has widespread metabolic perturbations and illustrate the utility of metabolomics in evaluating complex effects of altered vitamin B-6 intake.     

Targeted Exome Sequencing Integrated with Clinicopathological Information Reveals Novel and Rare Mutations in Atypical,Suspected and Unknown Cases of Alport Syndrome or Proteinuria

We applied customized targeted next-generation exome sequencing (NGS) to determine if mutations in genes associated with renal malformations, Alport syndrome (AS) or nephrotic syndrome are a potential cause of renal abnormalities in patients with equivocal or atypical presentation. We first sequenced 4,041 exons representing 292 kidney disease genes in a Caucasian woman with a history of congenital vesicoureteral reflux (VUR), recurrent urinary tract infections and hydronephrosis who presented with nephrotic range proteinuria at the age of 45. Her biopsy was remarkable for focal segmental glomerulosclerosis (FSGS), a potential complication of longstanding VUR. She had no family history of renal disease. Her proteinuria improved initially, however, several years later she presented with worsening proteinuria and microhematuria. NGS analysis revealed two deleterious COL4A3 mutations, one novel and the other previously reported in AS, and a novel deleterious SALL2 mutation, a gene linked to renal malformations. Pedigree analysis confirmed that COL4A3 mutations were nonallelic and compound heterozygous. The genomic results in conjunction with subsequent abnormal electron microscopy, Collagen IV minor chain immunohistochemistry and progressive sensorineural hearing loss confirmed AS. We then modified our NGS approach to enable more efficient discovery of variants associated with AS or a subset of FSGS by multiplexing targeted exome sequencing of 19 genes associated with AS or FSGS in 14 patients. Using this approach, we found novel or known COL4A3 or COL4A5 mutations in a subset of patients with clinically diagnosed or suspected AS, APOL1 variants associated with FSGS in African Americans and novel mutations in genes associated with nephrotic syndrome. These studies demonstrate the successful application of targeted capture-based exome sequencing to simultaneously evaluate genetic variations in many genes in patients with complex renal phenotypes and provide insights into etiology of conditions with equivocal clinical and pathologic presentations.