Synthesis and in vitro antioxidant functions of protein hydrolysate from backbones of Rastrelliger kanagurta by proteolytic enzymes

Every year, a huge quantity of fishery wastes and by-products are generated by fish processing industries. These wastes are either underutilized to produce low market value products or dumped leading to environmental issues. Complete utilization of fishery wastes for recovering value added products would be beneficial to the society and individual. The fish protein hydrolysates and derived peptides of fishery resources are widely used as nutritional supplements, functional ingredients, and flavor enhancers in food, beverage and pharmaceutical industries. Antioxidants from fishery resources have attracted the attention of researchers as they are cheaper in cost, easy to derive, and do not have side effects. Thus the present investigation was designed to produce protein hydrolysate by pepsin and papain digestion from the backbones of Rastrelliger kanagurta (Indian mackerel) and evaluate its antioxidant properties through various in vitro assays. The results reveal that both hydrolysates are potent antioxidants, capable of scavenging 46% and 36% of DPPH (1,1-diphenyl-2 picrylhydrazyl) and 58.5% and 37.54% of superoxide radicals respectively. The hydrolysates exhibit significant (p < 0.05) reducing power and lipid peroxidation inhibition. Among the two hydrolysates produced, pepsin derived fraction is superior than papain derived fraction in terms of yield, DH (Degree of hydrolysis), and antioxidant activity.     

Arachidonic acid is involved in the regulation of hCG induced steroidogenesis in rat Leydig cells

Phospholipase C (PLC), an enzyme involved in the hydrolysis of membrane phospholipid- phosphatidylinositol-bisphosphate to inositol triphosphate and diacylglycerol, and Phorbol 12, myristate 13, acetate (PMA), a tumor promoting agent, could significantly stimulate testosterone (T) secretion from Leydig cells. Arachidonic acid (AA) stimulated T secretion by about 2 fold. The steroidogenic effect of PLC and AA was biphasic. At low concentrations both PLC and AA (100 mU and 12.5 microM, respectively) augmented hCG induced T secretion, while at higher concentrations (PLC: 500 mU and AA: 200 microM) they inhibited steroid production. AA also had a biphasic effect on hCG induced cyclic AMP secretion. 5, 8, 11, 14 Eicosatetraynoic acid (ETYA), a general inhibitor of AA metabolism, and Nordihydroguaiaretic acid (NDGA), an inhibitor of the lipoxygenase pathway of AA metabolism, inhibited hCG induced T secretion while indomethacin, an inhibitor of cyclo-oxygenase pathway, had no effect on hCG induced T secretion. We conclude from these data that AA plays a role in the regulation of hCG induced steroidogenic responses in rat Leydig cells and that the metabolite(s) of AA that are involved are not cyclooxygenase products.     

Purification and activation of brain sulfotransferase.

Galactosylceramide sulfotransferase (EC 2.8.2.11) catalyzes the biosynthesis of sulfatide from galactocerebroside and adenosine 3'-phosphate 5'-phosphosulfate (PAPS). This enzyme is developmentally controlled, reaching a maximum activity in the brains of mice corresponding to that of maximum myelination. The product, sulfatide, is an important component of myelin. This transferase from mouse brain has been purified 2600-fold using a combination of pyridoxal 5'-phosphate- and ATP-ligated columns. The purified enzyme yielded a single band following SDS-polyacrylamide gel electrophoresis with an apparent M(r) of 31,000. The entire purification procedure can be completed in 1 day. The pH optimum for the enzyme is 7.0. The Km for PAPS is 1.2 x 10(-6) M, and the Km for cerebroside is 2.6 x 10(-5) M. Cerebroside concentrations > 80 pmol/ml are inhibitory. Enzyme preparations were associated with several lipids. Vitamin K+P(i) activated purified preparations of the sulfotransferase and maintained enzyme activity during storage at -80 degrees C.     

Unique regulation of Na‐glutamine cotransporter SN2/SNAT5 in rabbit intestinal crypt cells during chronic enteritis

The only Na‐nutrient cotransporter described in mammalian small intestinal crypt cells is SN2/SNAT5, which facilitates glutamine uptake. In a rabbit model of chronic intestinal inflammation, SN2 stimulation is secondary to an increase in affinity of the cotransporter for glutamine. However, the immune regulation of SN2 in the crypt cells during chronic intestinal inflammation is unknown. We sought to determine the mechanism of regulation of Na‐nutrient cotransporter SN2 by arachidonic acid metabolites in crypt cells. The small intestines of New Zealand white male rabbits were inflamed via inoculation with Eimeria magna oocytes. After 2‐week incubation, control and inflamed rabbits were subjected to intramuscular injections of arachidonyl trifluoromethyl ketone (ATK), piroxicam and MK886 for 48 hrs. After injections, the rabbits were euthanized and crypt cells from small intestines were harvested and used. Results: Treatment of rabbits with ATK prevented the release of AA and reversed stimulation of SN2. Inhibition of cyclooxygenase (COX) with piroxicam did not affect stimulation of SN2. However, inhibition of lipoxygenase (LOX) with MK886, thus reducing leukotriene formation during chronic enteritis, reversed the stimulation of SN2. Kinetic studies showed that the mechanism of restoration of SN2 by ATK or MK886 was secondary to the restoration of the affinity of the cotransporter for glutamine. For all treatment conditions, Western blot analysis revealed no change in SN2 protein levels. COX inhibition proved ineffective at reversing the stimulation of SN2. Thus, this study provides evidence that SN2 stimulation in crypt cells is mediated by the leukotriene pathway during chronic intestinal inflammation.     

Breeding lines of the Indian mega-rice variety,MTU 1010, possessing protein kinase <Emphasis Type="Italic">OsPSTOL</Emphasis> (<Emphasis Type="Italic">Pup1</Emphasis>), show better root system architecture and higher yield in soils with low phosphorus

MTU 1010 is a high-yielding mega-variety of rice grown extensively in India. However, it does not perform well in soils with low phosphorus (P) levels. With an objective to improve MTU 1010 for tolerance to low soil P, we have transferred Pup1, a major quantitative trait locus (QTL) associated with tolerance from another mega-variety, Swarna, through marker-assisted backcross breeding (MABB). Foreground selection of the F₁ and backcross plants was performed with the co-dominant, closely linked CAPS marker, K20-2, while two flanking markers RM28011 and RM28157 were utilized for recombinant selection. At each backcross generation, positive plants were also analyzed with a set of 85 parental polymorphic SSR markers to identify the QTL-positive plants possessing maximum introgression of MTU 1010 genome. At BC₂F₁, the best backcross plant was selfed to generate BC₂F₂s. Among them, the plants homozygous for Pup1 (n?=?22) were reconfirmed using the functional marker for Pup1, viz., K46-1, and they were advanced through pedigree method of selection until BC₂F₆ generation. A total of five elite BC₂F₆ lines, possessing Pup1 and phenotypically similar to MTU 1010, were screened in the low soil P plot and normal plot (with optimum soil P levels) during wet season, 2016. All the selected lines showed better performance under low P soil with more number of productive tillers, better root system architecture, and significantly higher yield (>?390%) as compared to MTU 1010. Further, under normal soil, the lines were observed to be similar to or better than MTU 1010 for most of the agro-morphological traits and yield. This study represents the successful application of marker-assisted selection for improvement of tolerance to low soil P in a high-yielding Indian rice variety.     

Composition and functional property of photosynthetic pigments under circadian rhythm in the cyanobacterium <Emphasis Type="Italic">Spirulina platensis</Emphasis>

Circadian rhythm is an important endogenous biological signal for sustainable growth and development of cyanobacteria in natural ecosystems. Circadian effects of photosynthetically active radiation (PAR), ultraviolet-A (UV-A) and ultraviolet-B (UV-B) radiations on pigment composition have been studied in the cyanobacterium Spirulina platensis under light (L)/dark (D) oscillation with a combination of 4/20, 8/16, 12/12, 16/8, 20/4 and 24/24 h time duration. Circadian exposure of PAR?+?UV-A (PA) and PAR?+?UV-A?+?UV-B (PAB) showed more than twofold decline in Chl a, total protein and phycocyanin (PC) in light phase and significant recovery was achieved in dark phase. The fluorescence emission wavelength of PC was shifted towards lower wavelengths in the light phase of PAB in comparison to P and PA whereas the same wavelength was retrieved in the dark phase. The production of free radicals was accelerated twofold in the light phase (24 h L) whereas the same was retrieved to the level of control during the dark phase. Oxidatively induced damage was alleviated by antioxidative enzymes such as catalase (CAT), peroxidase (POD), superoxide dismutase (SOD) and ascorbate peroxidase (APX) in the light phase (0–24-h L) whereas the dark phase showed significant inhibition of the same enzymes. Similar characteristic inhibition of free radicals and recovery of PC was observed inside cellular filament after circadian rhythm of 24/24 h (L/D). Circadian exposure of P, PA and PAB significantly altered the synthesis and recovery of pigments that could be crucial for optimization and sustainable production of photosynthetic products for human welfare.     

Improvement of blast resistance of the popular high-yielding,medium slender-grain type,bacterial blight resistant rice variety,Improved Samba Mahsuri by marker-assisted breeding

Improved Samba Mahsuri (ISM) is a popular, high-yielding, bacterial blight resistant rice variety possessing medium-slender grain type. As ISM is highly susceptible to blast disease of rice, through the present study we have transferred two major blast resistance genes, Pi2 and Pi54 into the elite variety by marker-assisted backcross breeding. The two blast resistance genes were transferred to ISM through sets of backcrosses. In every backcross generation, PCR-based markers, specific for the blast resistance genes (Pi2 and Pi54) and bacterial blight resistance genes (Xa21, xa13 and xa5) were utilized for foreground selection, while a set of 144 parental polymorphic SSR markers were used for background selection and backcrossing was carried out until BC₂ generation. A solitary BC₂F₁ plant possessing Pi2 or Pi54 along with Xa21, xa13 and xa5 and >?90% recovery of ISM genome was selected from the two sets of backcrosses were crossed and the intercross F₁s (ICF₁s) thus obtained were selfed to generate ICF₂s. Homozygous ICF₂ plants carrying all the five resistance genes were identified through markers and advanced through selfing till ICF₅ generation by adopting pedigree method of selection. Three best lines at ICF₅, possessing excellent resistance against bacterial blight and blast and closely resembling or superior to ISM in terms of grain quality: yield and agro-morphological traits have been identified and advanced for multi-location trials.