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  • ThesisItemOpen Access
    Ultrasound assisted nanocellulose extraction from microwave pre-treated rice straw to develop bio-nanocomposite for active packaging of horticultural produce
    (G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2022-04) Bhat, Mohd Ishfaq; Shahi, Navin Chandra
    Lignocellulosic biomass (especially agricultural residue) is a sustainable feedstock for production of value-added materials through specific processing techniques and sequences as per the desired end-product. The present study aimed to utilize one such agro-residue viz. rice straw by converting it into high crystalline nanocellulose to develop a bifunctional bio-nanocomposite for active packaging of horticultural produce. The first objective involved the application of a modified microwave assisted sodium chlorite pretreatment for intensive and quick delignification of rice straw. The effect of four critical process parameters on the delignification percentage of the rice straw was studied. The delignification was manifested by characterization using Fourier transform infrared spectroscopy, x-ray diffraction and scanning electron microscopy. Using a Box-Behnken experimental design (BBD), the effect of four critical process parameters, viz. microwave power (480-800 W), microwave treatment time (4-12 min), bleaching solution concentration (0.4-3.0 %), and bleaching time (1-5 h) on the delignification (%) was investigated, and the process was optimised using response surface methodology (RSM). The experimental data best fitted a quadratic model with an R2 of 0.9964. The optimized value of process parameters in aforementioned sequence was found to be 671 W, 8.66 min, 2.67 %, and 1h respectively, for the best delignification of 93.51 percent. The absence of lignin peaks (1516 and 1739 cm-1) was corroborated by deconstructed morphological structure and higher crystallinity index in the optimised delignified sample (53.7 %). The optimized delignified rice straw powder was then subjected to ultrasound assisted acid hydrolysis for nanocellulose extraction, preceded by the removal of hemicellulose and silica. The effect of sulfuric acid concentration (40-70%), ultrasonic treatment time (30-120 min) and ultrasound power (150-250 W) on the crystallinity index (%) and the mean particle size (nm) of the nanocellulose crystals, was studied through BBD and optimized through RSM. The optimum process parameters were obtained as 51.62 % sulfuric acid concentration, 48.55 min ultrasonic treatment time and 238. 80 W ultrasound power. The mean particle size and the crystallinity index at these optimum parameters were 69.01nm and 89.72 %, respectively. The spectral characterization revealed the increased intensity of peaks corresponding to the cellulose. The transmission micrograph of the nanocellulose revealed a rod like structure with average length and width of 190 ± 6 nm and 35 ± 3 nm, respectively. The nanocellulose crystals at 1, 5 and 10% concentration levels were used for reinforcement of chitosan biopolymer along with the addition of the titanium dioxide (TiO2) nanoparticles at its 1, 3 and 5% concentration levels. The overall performance of the developed bio-nanocomposite was measured in terms of thermal, mechanical, water vapor permeability (WVP) and water sensitive properties. Thickness of the developed films varied from 0.12 to 0.17 mm. The WVP and the tensile strength were reduced and increased by 27 % and 78 %, respectively for bio-nanocomposite with 10 % cellulose nanocrystals and 3% TiO2 concentration. The thermal stability of the developed bio-nanocomposite was better as compared to neat chitosan film in terms of higher glass transition and melting temperature. The best bio-nanocomposite was used for storage study of fresh-cut apple slices. The developed bio-nanocomposite showed promising results in terms of preserving the firmness, color and the radical scavenging activity of fresh-cut apple slices for a storage study period of 11 days.
  • ThesisItemOpen Access
    Process standardization for microwave and ultrasound assisted extraction of black cumin (Nigella Sativa L.) oil
    (G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2021-07) Iftikhar Alam; Shahi, N.C.
    Black cumin (Nigella sativa L.) seed belong to Ranonculaceae family and has been cultivated for use in spices and traditional medicine system of Unani and Ayurveda and has a long history. The seed is known for its oil content but the best method of extraction is still the point of concern. Mechanical expression and solvent extraction are currently used commercially methods of oil extraction of black cumin oil but both of the methods have their shortcomings of less oil recovery, degradation of oil quality and more time consumption. The present research work was thus done to overcome these problems. In the present study, the standardization and optimization of microwave, ultrasound and their combined extraction methods were evaluated to enhance the oil recovery and the quality of extracted oil from black cumin. Experiment were planned using box Behnken design considering parameters of microwave power (320, 480 and 640 W), microwave time (3, 5 and 7 min) and solvent volume (100, 150 and 200 ml/sample) for MAE, ultrasound power (150, 200, 250 W), ultrasound time (15, 30 and 45 min) and solvent volume (100, 150 and 200 ml/sample) for UAE and for combination was done at the optimized value of MAE and UAE. Particle size of 1 mm and the solvent, hexane, were kept constant for all the experiments. As per the results obtained from MAE the oil recovery obtained ranged from 83.27 to 97.45 %, Specific gravity ranged from 0.9 to 0.81, Color ranged from 44.18 to 61.66, DPPH Antioxidant activity ranged from 32.21 to 70.38 %, Free fatty acid ranged from 6.12 to 7.22 % and Peroxide value ranged from 7.54 to 9.37 meq O2/kg. In UAE the oil recovery obtained ranged from 85.51 % to 94.80 % Specific gravity ranged from 0.73 to 0.89, Color ranged from 36.24 to 53.88, DPPH Antioxidant activity ranged from 25.50 to 78.18 %, Free fatty acid ranged from 5.83 to 6.92 % and Peroxide value ranged from 6.63 to 8.24 meq O2/kg. Optimization of these methods was done based on their respective responses by means of Response surface methodology using Design Expert 10.0.1.0 software. The optimized parameters of MAE for black cumin oil gave the maximum oil recovery of 92.26 %, specific gravity 0.81, color 43.11, antioxidant 67.29, FFA 6.46 % and PV of 8.15 at microwave power 320W, microwave time 5.5 min and solvent to solid ratio 10 ml/g. and the optimized parameters of UAE for black cumin oil gave the maximum oil recovery of 87.88 %, specific gravity 0.73, color 38.8, antioxidant 76.32, FFA 6.06 % and PV of 6.92 at ultrasound power 158.1 W, ultrasound time 25.2 min and solvent volume (100, 150 and 200 ml/sample). Combined study was conducted at the optimized condition of parameters from MAE and UAE methods. A single experiment was run at these parameters to find the results. The maximum oil recovery 91.12 %, specific gravity 0.84, color 44.28, antioxidant 58.46, FFA 6.51 % and PV of 8.37 were found at ultrasound power 158 W, ultrasound time 25.2 min and solvent volume 100ml/sample followed by microwave power 320 W, microwave time 5.5 min. Results showed that the microwave assisted extraction proved very effective approach for the extraction of black cumin oil for the better oil recovery and the best quality.
  • ThesisItemOpen Access
    An innovative approach for microbial production of pyruvate using agro-industrial waste
    (G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2021-02) Pant, Manish; Omre, P.K.
    Pyruvic acid (pyruvate) is a cellular metabolite found in the biochemical link between glycolysis and the tricarboxylic acid cycle. The microbial production of pyruvate from yeasts or bacteria is based on limiting the natural catabolism of pyruvate and limiting the accumulation of its many potential by-products. The industrial pyruvate production methodology involves the use of organic chemicals along with standardised protocol which increases the cost of the end product. In this process, pyruvic acid is distilled from a mixture of tartaric acid and potassium hydrogen sulphates at 220°C; the crude acid obtained is then distilled under vacuum. This process is simple to realize but not cost-effective. Hence, to realise a cheaper and efficient methodology alternative nutrient sources are explored. Development of media formulations is key in any bio-transformation involving micro-organisms. Establishment of optimal culture constraints and process development is considered crucial in this regard. Since, microbial enzyme activity is important to pyruvate accumulation in isolated strains, advances in pyruvate production can be achieved by media optimization. In this study, the media formulation involved various carbon sources viz. glycerol, rice straw and jackfruit rind with various proportions of nitrogen source, corn steep liquor (diluted with distilled water). The amount of nutrient sources is also very critical for formulation of standard media composition. Thus, the screening experiments were focused on standardizing the carbon and nitrogen levels for final experiments. In screening experiments, only incubation time and incubation temperature had significant effect on pyruvate production at a confidence level p<0.05. Finally, the factors compromised for main optimization experiments were screened as follows: carbon source 25%g/g, CSL 12%v/v, pH 5.0, agitation speed 220rpm, KH2PO4 1.1%g/g, thiamine 1.3%μg/g, biotin 1.4%μg/g, MgSO4.7H2O 0.3%g/g and CaCO3 43%g/g. The final experiments were based on full factorial design on different levels of independent variables. With glycerol as carbon source, the highest effect of CSL concentration on pyruvate concentration was observed and was highly significant (p<0.01) because it had high calculated F-value (519.60). The effect of temperature (340.32) followed by time (324.60) was also found significant (p<0.01). As per the results, the optimum solution was obtained when the CSL concentration was 0.992601, time was 0.509991, and temperature was 0.317417. Similarly with rice straw as carbon source, the highest effect of CSL concentration on pyruvate concentration was observed and was highly significant (p<0.01) because it had high calculated F-value (444.80). The effect of time (252.25) followed by temperature (143.40) was also found significant (p<0.01). As per the results, the optimum solution was obtained in terms of coded values when the CSL concentration was 0.999997, time was 0.882823, and temperature was 0.363031. Similarly with jackfruit rind as carbon source, the highest effect of CSL concentration on pyruvate concentration was observed and was highly significant (p<0.01) because it had high calculated F-value (1199.93). The effect of time (925.66) followed by temperature (255.70) was also found significant (p<0.01). As per the results, the optimum solution was obtained when the CSL concentration was 0.999986, time was 0.851276, and temperature was 0.438559. In supervised learning approach, the highest accuracy corresponds to 68.8889% for multilayer perceptron under 10X cross validation fold-maker, and the lowest is 0% for IBk. In fact, in this experimental comparison, we can say that multilayer perceptron was the best scheme in all applicable classifiers, with highest accuracy. Moreover, in unsupervised learning approach feature selection preprocessing was considered essential and principal component analysis was performed prior to each clustering algorithms application. The paired t-test analysis of the three carbon sources show a draw when compared with glycerol as a standard source. This sustains the fact that, organic replacements instead of conventional organic source i.e. glycerol can be utilized for pyruvate production with no random aberrations in pyruvate yield. The SEM results show that the diametric dimensions of the pyruvate produced from carbon sources were in the range of 30μm to 300 μm and is comparable with that of industrially produced sample.
  • ThesisItemOpen Access
    Process optimization, characterization and modelling of microbially synthesized Poly-3-Hydroxybutyrate using damaged wheat grain
    (G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2020-06) Sirohi, Ranjna; Pandey, J.P.
    Polyhydroxybutyrate (PHB) is biodegradable, microbiologically produced biopolymers that is compatible with nature as compared to petrochemically derived plastics. PHB has advanced applications in medical sector, packaging industries, nanotechnology, agriculture, etc. It is produced using various feedstocks such as glycerol, dairy wastes, agro-industrial wastes, food industry waste, sugars, etc. Current focus on PHB research has been primarily on reducing the cost of production and, on downstream processing to isolate PHB from the cells. Selection of feedstock, therefore, plays a critical role in determining the economic feasibility and sustainability of the process. However, with increased demand for biobased products, raw materials for their production are at times in conflict with the supply of food provisions. This necessitates the identification of other potential feedstocks for PHB production. One of such alternative feedstocks could be damaged grains (discolored, broken, cracked, insect infested, chalky, smelly, damped and partial softened), which are available in substantial quantities in various parts of the world due to inadequate management of pre- and post-harvesting operations. Damaged wheat grains (DWG) is among these, which does not find any potential application and is often disposed off in environment, where it is decomposed naturally and causes environmental pollution. Thus, overall aim of this study was to explore the production of PHB employing DWG. Locally collected DWG after washing, drying and milling were subjected to hydrolysis using acids and enzymes, followed by fermentation. Initially, hydrochloric, phosphoric, nitric and sulphuric acid were screened for their relative potential for hydrolysis. Maximum production of reducing sugars (RS) and minimum production of inhibitory compounds were taken as performance parameters. Of the tested acids, HCl showed lowest concentration of inhibitors (furfural, 5-hydroxymethyl furfural, acetic acid, formic acid) while producing substantial quantities of reducing sugars (RS). Subsequent experiments with different DWG flour (10, 15, 20%; w/v) and acid concentrations (1.0, 3.0, 5.0%; w/v) were carried out to identify suitable hydrolysis conditions. Results revealed that 3.0% HCl with 15% substrate concentration produced highest RS (116.29 mg/mL) after 45 min of hydrolysis. Artificial neural network (ANN) and second-order models were applied to the experimental data of which ANN performed well in predicting RS after hydrolysis with good accuracy (R2=0.939). For enzymatic hydrolysis, DWG flour (10, 15, 20%; w/v) was treated using α-amylase (1.0, 3.0, 5.0%; v/v, 12 U/mL). Treatment period of 60 min resulted in 85.2 mg/mL RS using 5.0% α-amylase and 19.4% substrate. Reaction kinetics confirmed that substrate concentration higher than 10% enhanced the production of RS. The liquified and partially hydrolysed solution (α-amylase treated) was further treated using glucoamylase (1.0, 2.0, 3.0%; v/v, 46 U/mL) for the production of RS. At 2.4% glucoamylase concentration, 147.5 mg/mL RS was obtained in 103 min. This hydrolysate was used to grow Bacillus sp. NII2 for the production of PHB and the effect of processing parameters, viz., inoculum concentration (5.0, 7.5, 10%; v/v), sugar concentrations (20, 30, 40 g/L) and incubation time (48, 72, 96, 120 h) were evaluated. The optimum values of processing parameters for maximum PHB production was at 8.84% (v/v) inoculum concentration, 32.07g/L sugar concentration and 72 h incubation time. Analysis of the resulting polymer through 1H-NMR, 13C-NMR, FTIR, XRD and SEM confirmed the presence of PHB.