Postcolumn Fluorimetric HPLC Procedure for Determination of Niacin Content of Cereals

Analytical procedures for determining niacin or vitamin B3 content of foods are tedious, require large quantities of toxic chemicals, and are timeconsuming. In addition, food matrices are difficult as samples because of their complex nature. A selective, sensitive HPLC technique was developed with postcolumn derivatization as well as fluorescence and spectroscopic detection systems. Niacin was separated and retained for 6.5 min on a polymeric column with an aqueous mobile phase containing sodium acetate buffer. A postcolumn system consisting of a stainless-steel pump and reaction coil allowed detection and quantitation of niacin. An acid-enzyme sample-extraction method was most compatible with HPLC and postcolumn derivatization with 5% each of acidified p-aminophenol and cyanogen bromide. Lower detection limit and mean recovery were 3.6 ng and 99.43%, respectively. Fluorescence response for nicotinamide was half that of nicotinic acid. A lower response for nicotinamide was also noted with conventional spectroscopy. However, the new method yielded comparable values for six of eight ready-to-eat commercial cereal samples. No significant difference was observed between the AACC reference and HPLC fluorimetric methods. Chemical derivatization was done within a reaction coil with reagents at half strength, limiting exposure to hazards and minimizing waste-disposal problems.     

Molecular detection of WA‐CMS restorers from tropical japonica‐derived lines,their evaluation for fertility restoration and adaptation

Hybrid rice based on wild‐abortive cytoplasmic male sterility (WA‐CMS) is important in boosting rice production, which requires diverse parents to harness heterosis. For this, exploiting the diversity of japonica through tropical japonica (TRJ) lines is an excellent route. In this study, 310 TRJ‐based new plant type (NPT) lines were developed and evaluated for Rf3 and Rf4 genes. Gene‐based (DRRM‐Rf3‐5 and DRRM‐Rf3‐10) and functional marker (RMS‐SF21‐5) targeted Rf3 locus, while gene‐linked (RM6100) and functional marker (RMS‐PPR9‐1) targeted the Rf4 locus. The frequency of the restorer allele of Rf3 gene was lower when compared to that of Rf4. Combined phenotypic and molecular screening using gene‐based and functional markers identified 42 lines that carried Rf3 and/or Rf4 genes. All the selected lines produced fertile F₁s when crossed to a WA‐CMS line, “Pusa 6A”, but with varying levels of spikelet fertility. This is the first report of a marker‐cum‐phenotype‐based restorer selection using TRJ‐derived lines. Multilocation evaluation of these lines at three locations indicated better adaptation for grain yield in some of the lines.     

Compositional changes of selected amino acids,organic acids,and soluble sugars in the xylem sap of N,P, or K‐deficient tomato plants

Xylem sap plays a major role in long‐distance transport of water, nutrients, and metabolites. However, there is little information on the behavior of metabolites in mineral‐deficient xylem sap. For this reason, the time‐dependent changes in selected metabolites (amino acids, organic acids, and soluble sugars) from tomato xylem sap in response to nitrogen (N), phosphorus (P), or potassium (K)‐deficient condition were investigated. Tomato plants (Solanum lycopersicum L.) were grown hydroponically in liquid culture under three different mineral regimes: N‐deficient [0.5 mM Ca(NO₃)₂ and 0.5 mM KNO₃], P‐deficient (0.05 mM KH₂PO₄), and K‐deficient (0.5 mM KNO₃), respectively. Xylem sap was collected at 10:00 am after 1, 5, 15, and 30 d, and the selected metabolites were analyzed with liquid chromatography. All N, P, or K deficiencies led to a substantial increase in metabolites in the xylem sap. The predominant amino acid in the xylem sap was glutamine and, interestingly, all mineral deficiencies resulted in a substantial amount of γ‐aminobutyric acid (GABA). Additionally, organic acids (citrate and malate) and soluble sugars were strongly increased in all mineral deficiencies, and, in particular, the level of shikimate was greatly affected by N deficiency. Based on these data, it is necessary to clearly elucidate an unknown event taking place in xylem loading in a variety of environmental impacts, and we are now studying to expand our knowledge on metabolic and proteomic responses using GC‐MS and LC‐MS.     

Inhibitory effect(s) of polymeric black tea polyphenol fractions on the formation of [(3)H]-B(a)P-derived DNA adducts

Five polymeric black tea polyphenol fractions (PBP-1-5) were isolated from a popular brand of black tea. The effect of these PBPs and epigallocatechin gallate (EGCG), a major green tea polyphenol, was studied on the formation of [(3)H]-B(a)P-derived DNA adducts in vitro, employing rat liver microsomes. PBP-1-3 inhibited microsome-catalyzed [(3)H]-B(a)P-derived DNA adduct formation in vitro in a dose-dependent manner. This inhibition was further enhanced on preincubation of microsomes with each of the PBPs. PBP-4 was not effective per se and required preincubation with microsomes to exhibit its inhibitory effect, whereas PBP-5 remained ineffective with or without preincubation with microsomes. Further investigations revealed that the observed decrease in [(3)H]-B(a)P-DNA adduct formation was due to inhibition of isozymes of CYP450s by PBPs. Overall, results suggest that polymeric black tea polyphenol fractions retain one of the chemopreventive effects exhibited by the monomeric green tea polyphenol EGCG in vitro.     

Differential expression of ISG 15 mRNA in peripheral blood mononuclear cells of nulliparous and multiparous pregnant versus non‐pregnant Bos indicus cattle

Embryonic mortality is found to be the main source of reproductive wastage in domestic ruminants. Many genes are involved in the growth and development of the embryo, and the interferon‐stimulated gene 15 (ISG 15) is one of the major gene stimulated by interferon tau, the maternal recognition of pregnancy signal in ruminants. In this study, both genomic and cDNA sequences of ISG 15 from Bos indicus (Deoni breed) were amplified and characterized. The genomic sequence of Deoni ISG 15 exhibited 99% identity with Bos taurus and 97% identity with that of Bos mutus and Bubalus bubalis. Moreover qRT‐PCR analysis revealed constitutive expression of the ISG 15 mRNA in peripheral blood mononuclear cells of Deoni heifers and multiparous cows during early pregnancy. Fourteen Deoni heifers and fifteen multiparous Deoni cows were synchronized for timed AI by CIDR‐Ovsynch protocol, and six animals were kept as cyclic control in each group. Blood samples were collected on days 7, 14, 16, 18, 21, 30 and 45 from the day of AI. Pregnancy was confirmed by plasma progesterone level through ELISA. A significantly higher expression of ISG 15 mRNA was found on day 16 (p < .05) and day 18 (p < .05) of pregnancy in nulliparous heifers. Although in multiparous Deoni cows ISG 15 expression was greater in pregnant cows, difference was statistically non‐significant. The result of this study indicates that ISG 15 gene expression is upregulated during 16–18 days of pregnancy and could be used as an early pregnancy marker in dairy cows especially in heifers.     

Water productivity and nutrient status of rice soil in response to cultivation techniques and nitrogen fertilization

Three methods of rice cultivation were compared in a field experiment at New Delhi, India during 2012 for their water use and changes in nutrient availability of soil. The experiment was laid out in a split plot design with conventional transplanting (CT), system of rice intensification (SRI), and aerobic rice (AR) cultivation technologies. Five doses of nitrogen included 100 % (120 kg N ha^?1), 125, and 150 % recommended dose of N(RDN) through urea, 75 % of RDN through urea (90 kg N ha^?1) + 25 % of RDN (30 kg ha^?1) through farm yard manure (FYM), and 100 % of RDN through FYM. Results revealed that status of available N in soil under rice at 45 and 90 days after sowing (DAS) was significantly higher in CT and SRI compared to AR method. Application of the highest dose of nitrogen through urea resulted in the highest availability of N (188.9, 174.2, and 135.2 kg ha^?1 for 45 and 90 DAS and at harvest stage, respectively). The soil under AR recorded significantly low availability of phosphorus and iron. However, availability of K in soil was not affected significantly under adopted production techniques and nitrogen management. The recorded irrigation water productivity was maximum in AR cultivation (9.16 kg ha mm^?1) followed by SRI (7.02 kg ha mm^?1) with irrigation water saving of 54 and 36 %, respectively compared to CT.     

Identification of parameters in transient water quality models from stochastic data

An approach is presented for the identification of parameters in time-varying water quality models from stochastic data measured at two points along a stream (input-output type data). The models are plug flow models described by linear-first order partial differential equations. The method employed is to reduce the partial differential equations to a set of ordinary differential equations with the method of characteristics from which an analytical solution is obtained. The time-lag phenomenon presented in plug flow models is used to correlate the input-output data. A gradient method (Bard's method) is employed to identify the parameters in the correlation equation.     

Quantitative trait loci for stay-greenness and agronomic traits provide new insights into chlorophyll homeostasis and nitrogen use in rice

Functional stay-green (FSG) is characterized by delayed senescence during the reproductive stage in rice. A recombinant inbred line (RIL) population derived from ‘Pusa 677’/‘PSG16’ was used to study the genetics of FSG in a rice mutant, ‘PSG16’. The RILs exhibited significant variation under two N regimes for agro-morphological traits, leaf chlorophyll content, flowering time, yield components and nitrogen (N) use. Using a genome wide linkage map spanning 1910.8 cM with 104 polymorphic markers, we have mapped six major quantitative trait loci (QTLs). One robust QTL on chromosome 1 was associated consistently across different N regimes with chlorophyll content and flowering time. The QTL on chromosome 7 was associated with grain number, whereas the QTL on chromosome 6 was found related to N harvest index and spikelet fertility. Although ‘PSG16’ showed a clear advantage in grain yield as well as having better N assimilation, we could not establish a direct genetic relationship SG trait and N use. Based on the QTL information, FSG trait of ‘PSG16’ could be useful in deciphering multiple stress responses in rice.     

Enhanced grain yield in rice hybrids through complementation of fertility restoration by Rf3 and Rf4 genes as revealed by multilocation evaluation of tropical japonica derived rice (Oryza sativa) hybrids

Thirty-one tropical japonica derived Rf gene carrying rice hybrids were classified into three classes as Rf3, Rf4 and Rf3 + Rf4 hybrids. These hybrids were tested under three different mega-environments. Between Rf3 class and both the classes of hybrids possessing Rf4 genes, significant variation for spikelet and pollen fertility and grain yield was found. The pollen fertility was five times higher among Rf4 hybrids than that of hybrids carrying Rf3 alone. Likewise, spikelet fertility among Rf4 hybrids was two times higher than that of Rf3 hybrids. Parallel grain yield increase in Rf4 hybrids was 2.4 times than in Rf3 hybrids. However, Rf3 gene was found complementing Rf4 to truncate the range of pollen and spikelet fertility. Hybrids possessing Rf3 alone exhibited partial pollen and spikelet sterility, and significant negative standard heterosis for grain yield. The high yielding hybrid, “HYB36” carrying both the Rf genes was found to be widely adapted. The present study established that Rf4 gene is essential either alone or in combination with Rf3 for fertility restoration to achieve enhanced grain yield in WA-CMS based hybrids.     

The host background of rice influences the resistance expression of a three genes pyramid (xa5 + xa13 + Xa21) to bacterial blight (Xanthomonas oryzae pv. oryzae) pathotypes of Indian mainland and Bay islands

Bacterial blight (BB) is a major disease of rice for which host resistance is the only effective solution. The three genes pyramid xa5 + xa13 + Xa21 is recently the most utilized combination for developing resistant varieties through marker‐assisted breeding. Our study was carried out to elicit the detailed response of twenty lines possessing these three genes in five genetic backgrounds to twelve diverse BB pathotypes in India. The lines developed from ADT 47 variety showed incomplete resistance to most of the pathotypes, whereas susceptibility varied from 8.3% to 16.6% in ADT 43 and IR24, respectively. However, in IMP ASD16/60 and Improved Samba Mahsuri, complete resistance against all pathotypes was observed. The overall results confirmed that genetic background plays crucial role for the effective expression of xa5 + xa13 + Xa21 combination. Molecular studies did not reveal correlation between origin of pathotypes and their virulence potential. It is suggested to deploy Improved Samba Mahsuri, IMP ASD 16/60 and AD1306 varieties in the bacterial blight prone areas or use them as donors for realizing wider and durable resistance.