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ThesisItem Open Access Studies on micropropagation with metabolite profiling for pharmacogenosic efficacy of Adina cordifolia and biochemical investigations of flowering in Dendrocalamus giganteus(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2019-02) Raypa, Pratima; Dubey, AshutoshAdina cordifolia a Rubiaceae family member is deciduous threatened tree and for in vitro propagation, axillary buds were augmented on MS medium supplemented with naphthalene acetic acid and 6- benzyl amino purine. Shoot length of propagated explants were found maximum on medium supplemented with 2 mg/L BAP or 0.5 mg/L NAA while maximum callus volume was induced in MS medium supplemented with 1.0 mg/L NAA, 1.0 mg/ L 2, 4- D and 0.5 mg/L BAP. On the metabolite profiling of Adina cordifolia different extracts through GC-MS, dominating constituents in the leaf extracts were trans squalene, vitamin E, phytol, and neophytadiene, while in bark extracts 7-hydroxycoumarin, nhexadecanoic acid, methyl commate, ergost-5-en-3.beta.-ol, -sitosterol. In root extracts major compounds, acetophenone, benzoic acid, 7-hydroxycoumarin, n-hexadecanoic acid, ergost-5-en-3.beta.-ol, beta.-sitosterol and gamma.-sitosterol were found. Through HPLC 7-hydroxycoumarin (7-HC), pharmaceutically important compound was found in high quantity in methonolic extracts of root, bark and calluses. Reducing power activity was positively correlated with all phytochemicals but best correlation was with phenols and flavonoids. FRAP activity were found positively correlated with phenolic content and flavonoid. Adina cordifolia extracts showed good cytotoxicity against human pathogenic strains i.e., Salmonella typhaemurium 52, Salmonella typhaemurium 149, Salmonella typhaemurium ATCC 51812, Salmonella entiritidis ATCC 13076, E. coli ATCC-25922 and E coli strain-13. Allelopathic effect of leaf extract of Adina cordifolia was studied on wheat genotypes as H2O2 content increment in wheat genotypes due to salt stress. Aqueous leaf extract of Adina cordifolia treatments decreased H2O2 content with increase in proline content. Flowering gene expression analysis in bamboo genotype Dendrocalamus giganteus was carried out by RT-PCR analysis. The transcript accumulation of homolog of CRABS CLAW, SEPALLATA1, AGAMOUS and PISTILLATA in flowering bud of Dendrocalamus giganteus was 3.6, 1.41, 3.02 and 5.46 folds higher respectively as compared to non-flowering bamboo leaves of Dendrocalamus giganteus. Metabolite profiling of D. giganteus revealed the presence of dominating constituents such as -thujene, vitamin E, 6-methyl-2-(5-methyl-5-vinyltetrahydro-2-furanyl)-5-hepten-3-one, elemol, -eudesmol, 7-epi- -eudesmol, n-hexadecanoic acid, n-heneicosane, n-tetracontane, campesterol, -sitosterol. Thus these results on A. cordifolia would be used to set new standard of prescribing the dosage of herbal drugs to propose an alternative to modern system of medicines and bamboo flowering gene expression analysis provide good understanding of flowering gene expression pattern.ThesisItem Open Access Investigation on biochemical basis of resistant starch with respect to physiochemical properties in cereals-millets, rice and wheat(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2018-12) Verma, Vivek Chandra; Agrawal, SanjeevThe present investigation was carried out to assess the physio-chemical properties of starch attributing towards resistant starch in millets (Barnyard millet; VL Madira-207, Finger millet; VL Mandua-352 and Foxtail millet; DHFT-109-3), rice (PB-2 and PD-19) and wheat (PBW-343 and UP-262). The study emphasize on the factors which influences the resistant starch in cereals. In the present study starch isolated from different cereals were viewed under SEM to analyse the shape and size of starch granules. The size of starch granules were diverse range in cereals like millets (4.6-12.23 μm) rice (6.22-177.13 μm) and wheat (2.50-198.23 μm). Small and large spherical, irregular and polygonal shape of starch granules was observed. The FTIR bands at 1047 and 1022 cm-1 gives ordered and amorphous structure of starches and absorbance ratio at 1047/1022 gives indexes of the short-range order of double helices ranged from 0.94-1.24. Total starch (57-70%), amylase (31-45%) and amylose-amylopectin ratio (0.46-0.83) was determined in different cereals under observation. The amylose content was positively correlated with resistant starch at P < 0.01 while total starch had non-significant impact on resistant starch. The other nutrient of cereals like dietary fibre (2.66-6.98%), total soluble protein (9.33-16.29%) and soluble protein fraction like albumin (1.41-2.44%), globulins (1.25-2.23%) and prolamines (2.04-2.88%) was estimated to establish the correlation with resistant starch. These other nutrient of cereals showed positive correlation with resistant starch at P < .01. In vitro digestion of native starch, gelatinized starch and retrograded starch was done at time interval of 30, 60, 90, 120 min and it was found that the rate of enzymatic digestion follows the order; Gelatinized starch > Native starch > Retrograded starch. During endosperm developmental stages at S1, S2, S3, S4 (7, 14, 21, 28 DAA) the key enzymes for starch biosynthesis like AGPase, SSS and GBSS were estimated. The activity of starch biosynthetic enzymes follows the order AGPase > GBSS > SSS in all these varieties during endosperm development. The higher activity of AGPase, GBSS and SSS found in rice comprises higher accumulation of amylose and total starch during endosperm development. The accumulation of amylose and total starch in millets was found at par to the wheat varieties which is related to the activities pattern of enzymes. All three enzymes under study showed the positive correlation with each other during endosperm development. Accumulation of amylose showed positive correlation with AGPase and GBSS, while non-significant correlation with SSS. Total starch accumulation showed positive correlation with AGPase.ThesisItem Open Access Optimization of pretreatment parameters and nanoparticle based enzyme immobilization for lignocellulosic biomass saccharification(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2018-09) Singh, Archana; Verma, A.K.Pretreatment is a prerequisite technology to facilitate the recovery of fermentable sugars from lignocellulosic biomass. Various lignocellulosic biomass viz. sugarcane leaves, rice straw, wheat straw, orange peel and pomegranate peel were used for pretreatment with ionic liquid EmimAc (1-ethyl-3-methylimidazolium acetate), aqueous ammonia and a combination of both followed by saccharification with immobilized cellulase and β-glucosidase (BGL) on magnetic nanoparticles. Magnetic nanoparticles (Fe3O4) and cellulase/BGL immobilized nanoparticles were characterized by TEM, SEM, FTIR and Zeta potential. Immobilization efficiency of cellulase and BGL were found to be 86.36% and 87.73%, immobilization yield were 83.38 and 85.18% respectively. The immobilized cellulase had optimum pH 5.0 and optimum temperature 50°C. The immobilized cellulase retained 90% of initial activity after 60 min incubation at 50°C while free cellulase had only 49% of its initial activity. Km value for free and immobilized cellulase was found to be 0.625 mM and 0.714 mM and the Vmax was 35.71 and 38.16 μmole/min/mg respectively. Immobilized cellulase retained up to 97% of residual activity on 6th day when stored at 4°C and 45% at 25°C whereas free cellulase had 86% of residual activity at 4°C and 20% at 25°C. Immobilized cellulase retained about 87% of residual activity after 10 cycle of reuse. The immobilized BGL had optimum pH 5.0 and optimum temperature 50°C. The immobilized BGL retained 91% of initial activity after 60 min incubation at 50°C while free BGL had only 51% of its initial activity. Km value for free and immobilized BGL was found to be 0.555 mM and 0.779 mM and the Vmax was 38.46 and 43.47 μmole/min/mg. Immobilized BGL retained up to 95% of residual activity on 6th day when stored at 4°C and 42% at 25°C whereas free BGL had 87% of residual activity at 4°C and 18% at 25°C. Immobilized BGL retained about 85% of residual activity after 10 cycle of reuse. The combined use of EmimAc and aqueous ammonia in 1:1 for pretreatment at 6 hrs with <2 mm size of biomass followed by saccharification with 3.2 U of immobilized cellulase showed a synergistic effect on biomass with release of 93% glucose. Combined pretreated biomass were analysed for saccharification by addition of 3.2 U BGL with 3.2 U cellulase and 97% of glucose released. Furthermore, EmimAc and immobilized cellulase were recycled for 6 times with 89% of efficiency; EmimAc, immobilized cellulase and immobilized BGL were recycled for 6 times with 90% of efficiency. Thus, compared to the other pretreatment method, this combine method is more efficient and cost effective. To identify the cheapest source for enzymes biogas slurry was analysed for different enzymes. These were cellulase (0.41), BGL (0.30), Xylanase (0.31), α-amylase (0.06), β-amylase (0.20), Dehydrogenase (0.37), and Protease (0.29) U/mL. Saccharification was done with crude enzyme isolated from biogas slurry and found 69% of glucose release in combined pretreated biomass.ThesisItem Open Access Effect of micro/macro environment of rhizogenesis for optimization of withanolide contents in Withania somnifera(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2018-08) Singh, Manali; Agrawal, SanjeevIndia is a varietal emporium of medicinal plants and is one of the richest countries in the world with regard to genetic resources of medicinal plants. Withania somnifera is a revered herb of Indian medicinal system. Owing to the presence of pharmacologically bioactive compounds. The roots and leaves of Ashwagandha contain various alkaloids viz. Withanolides and withaferin A. Among withanolides, withaferin A and withanolide A has been reported to be dominant metabolite distributed among various tissues of this plant in varying concentrations. Since the location and environmental factors play a key role in the biosynthesis of the bioactive metabolites so the agro-climatic conditions are conductive for introducing and domesticating new exotic plant varieties with enhanced and consistent withanolide A/withaferin A contents. Thus keeping in mind the pharmacological significance of the bioactive ingredients, the present study focuses on the different strategies employed for their enhanced production. MS medium supplemented with Kinetin 1.5 mg/l was found to have highest withaferin A content i.e. 0.93 mg/g (DW) while withanolide A content was highest in MS medium supplemented with Kinetin 1.5 mg/l i.e 0.348 mg/l (DW). Agrobacterium rhizogenes transformed Poshita recorded highest total withaferin A content i.e 5.969 mg/g (DW) and total withanolide content i.e 2.017 mg/g(DW). In aquaponics the total withaferin A content of 6 month grown seedling, was higher in Jawahar-20 i.e 2.904 mg/g (DW) while withanolide content was higher in Poshita i.e 4.019 mg/g (DW). Moreover use of different elicitors at very low concentrations were found to enhance the withaferin A /withanolide A content. Thus both the varieties i.e. Jawahar-20 and Poshita are the promising varieties of Withania somnifera, with higher withaferin A /withanolide A production. Thus implementation of tissue culture techniques, Hairy root culture, usage of elicitors, bio augmented soil and aquaponic system can offer promising approach for enhanced growth and sustainable production of pharmacologically bioactive ingredients in perspective of improving cultivation of Ashwagandha.ThesisItem Open Access Characterization and comparison of bioactive constituents in the in vitro regenerated and wild roots of Himalayan medicinal herb Valeriana jatamansi Jones(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2018-08) Saini, Soniya; Kumar, Vandana A.The medicinal herb Valeriana jatamansi Jones is a native of Northeastern Himalayan region belonging to family Valerianaceae and growing at altitudes of 1500-3000 m asl. The in vitro direct-, indirect rhizogenesis and plantlet regeneration was achieved from leaf explant in MS medium supplemented with ppm quantities of NAA (2.0)+BAP (2.0); IBA (1.0)+NAA (1.0)+BAP (0.1) and BAP (2.0)+NAA (0.2), respectively. Multiple shoots and plantlet regeneration was achieved from nodal explant in MS medium supplemented with ppm quantities of NAA(0.1)+TDZ(1.0) and IBA(1.0)+AgNO3(1.0), respectively. Phytochemical screening in crude and methanolic root extract by FTIR confirmed the presence of alkaloids, saponins, tannins, flavonoids, steroids and terpenes, and polyphenols. Methanol was the best solvent followed by dichloromethane and ethylacetate for extraction of phytochemicals from root and leaf of wild plant. The highest phytochemicals like phenol (351.0mg GAE g-1 extract), flavonoids (84.0mg QE g-1 extract), tannin (814.0mg TAE g-1 extract) and orthodihydroxy phenol (47.0mg CE g-1 extract) content and antioxidant activities like total antioxidant (935.4 AAE g- 1), FRAP (810.1 AAE g-1 extract), DPPH radical scavenging (IC 50=11.8μg ml-1), Superoxide radical scavenging (IC50=18.4μg ml-1), Hydroxyl radical scavenging (IC50=18.1μg ml-1) and Metal ion chelating (IC50=11.5μg ml-1) were found to be highest in methanolic extract of 6 months grown wild roots at 35ºC. The methanolic extracts of 6 months grown roots at 35ºC showed lowest MIC (62.50 μg) and maximum efficacy at par to gentamycin standard against 6 human pathogenic strains i.e., S. typhaemurium-52, S. typhaemurium-149, S. entiritidis-ATCC 13076, S. typhaemurium-ATCC 51812, E. coli-ATCC-25922 and E coli strain-13. Effect of maturity of roots (in vitro- and wild grown) and thermal stress indicated that the phytochemical, antioxidant, antioxidant enzymes and antimicrobial activities in 6 months grown in vitro roots were comparable to 3 months grown wild roots indicating adaptation towards higher temperature tolerance, since V. jatamansi was a temperate herb. Micronutrient analysis showed that Fe (0.254 ppm) was found to be significantly highest than Zn (0.137 ppm) in wild leaf and root. The third largest element in root was Mn (0.052 ppm) and Ca (0.118 ppm) in wild leaf because Mn facilitated the uptake of other nutrients like Fe and inorganic phosphate in root and Ca acted as binding agent in cell wall fusion in leaf. GC-MS analyses identified 38 constituents comprising 78.33% of total compositions of which 28% were contributed by patchouli alcohol confirming V. jatamansi as a patchouli chemotype. Valerenic acid content quantified by HPLC in the in vitro and wild roots was 8.6 mg g-1 and 25.09 mg g-1 dry weight, respectively.ThesisItem Open Access Evaluation of physiological and biochemical parameters for drought tolerant in barnyard millet from Kumaun region of Himalaya(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2017-08) Singh, Dipti; Verma, A.K.ThesisItem Open Access Studies on diosgenin production in fenugreek (Trigonella foenum-graecum L.)(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2017-08) Dwivedi, Himanshu; Agrawal, SanjeevFenugreek genotypes were evaluated for nutritional, antinutritional , antioxidant activity and diosgenin contents. Pant Ragini and Pusa Early Bunching varieties showed balanced proportion of nutrient and antinutritional contents with high diosgenin contents. The effect of different processing methods viz. soaking, germination, boiling, autoclaving and roasting on nutritional, antinutritional, antioxidant activity and diosgenin contents were studied. Germination process was found to be better as it enhanced antioxidant activity and reduced anti-nutrients. To study the diosgenin content Pant Ragini, a variety developed by G.B. Pant University was selected to study the effect of different elicitor at seedling level as well as field level. The effect of in vitro germination of fenugreek seed for 5 day with different elicitors (copper, cobalt, chitosan and ethrel) on growth and production of diosgenin was studied. The maximum enhancement in diosgenin content i.e. 70% (relative %) was observed when seed was soaked with 11.77 ppm cobalt treatment followed by 51% with 4.77 ppm copper treatment. The foliar application of these elicitors were also studied at field level for seed yield and diosgenin production. The highest seed yield i.e. 3.06 T/ha was recorded when 100 ppm chitosan was sprayed twice. However, highest diosgenin content i.e. 14.77 mg/g seed dry wt. was recorded when 11.77 ppm cobalt was sprayed. The highest diosgenin production i.e. 38 kg/ha can be obtained when 100 ppm chitosan sprayed twice. The production of diosgenin through tissue culture was also initiated. The highest diosgenin content i.e. 2.95 mg/g dry wt. was recorded in presence of 1ppm BAP and 1ppm 2-4 D for 40 days.ThesisItem Open Access Elucidation of degradative biomechanisms of pendimethalin using indigenous soil bacterial isolates(G.B. Pant University of Agriculture and Technology, Pantnagar (Uttarakhand), 2017-01) Trivedi, Neha; Dubey, AshutoshThesisItem Open Access Phytochemical and antimicrobial studies in Gentiana kurroo Royle(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2016-08) Joshi, Karishma; Kumar, Vandana A.