Agricultural Waste Management by Production of Second-Generation Bioethanol from Sugarcane Bagasse Using Indigenous Yeast Strain.

In the wake of rapid industrialization and burgeoning transportation networks, the escalating demand for fossil fuels has accelerated the depletion of finite energy reservoirs, necessitating urgent exploration of sustainable alternatives. To address this, current research is focusing on renewable fuels like second-generation bioethanol from agricultural waste such as sugarcane bagasse. This approach not only circumvents the contentious issue of food-fuel conflicts associated with biofuels but also tackles agricultural waste management. In the present study indigenous yeast strain, Clavispora lusitaniae QG1 (MN592676), was isolated from rotten grapes to ferment xylose sugars present in the hemicellulose content of sugarcane bagasse. To liberate the xylose sugars, dilute acid pretreatment was performed. The highest reducing sugars yield was 1.2% obtained at a temperature of 121 °C for 15 min, a solid-to-liquid ratio of 1:25 (% w/v), and an acid concentration of 1% dilute acid H2SO4 that was significantly higher (P < 0.001) yield obtained under similar conditions at 100 °C for 1 h. The isolated strain was statistically optimized for fermentation process by Plackett-Burman design to achieve the highest ethanol yield. Liberated xylose sugars were completely utilized by Clavispora lusitaniae QG1 (MN592676) and gave 100% ethanol yield. This study optimizes both fermentation process and pretreatment of sugarcane bagasse to maximize bioethanol yield and demonstrates the ability of isolated strain to effectively utilize xylose as a carbon source. The desirable characteristics depicted by strain Clavispora lusitaniae shows its promising utilization in management of industrial waste like sugarcane bagasse by its conversion into renewable biofuels like bioethanol.

Utilization of microalgae for agricultural runoff remediation and sustainable biofuel production through an integrated biorefinery approach.

Generally wastewater such agricultural runoff is considered a nuisance; however, it could be harnessed as a potential source of nutrients like nitrates and phosphates in integrated biorefinery context. In the current study, microalgae Chlorella sp. S5 was used for bioremediation of agricultural runoff and the leftover algal biomass was used as a potential source for production of biofuels in an integrated biorefinery context. The microalgae Chlorella sp. S5 was cultivated on Blue Green (BG 11) medium and a comprehensive optimization of different parameters including phosphates, nitrates, and pH was carried out to acquire maximum algal biomass enriched with high lipids content. Dry biomass was quantified using the solvent extraction technique, while the identification of nitrates and phosphates in agricultural runoff was carried out using commercial kits. The algal extracted lipids (oils) were employed in enzymatic trans-esterification for biodiesel production using whole-cell biomass of Bacillus subtilis Q4 MZ841642. The resultant fatty acid methyl esters (FAMEs) were analyzed using Fourier transform infrared (FTIR) spectroscopy and gas chromatography coupled with mass spectrometry (GC-MS). Subsequently, both the intact algal biomass and its lipid-depleted algal biomass were used for biogas production within a batch anaerobic digestion setup. Interestingly, Chlorella sp. S5 demonstrated a substantial reduction of 95% in nitrate and 91% in phosphate from agricultural runoff. The biodiesel derived from algal biomass exhibited a noteworthy total FAME content of 98.2%, meeting the quality standards set by American Society for Testing and Materials (ASTM) and European union (EU) standards. Furthermore, the biomethane yields obtained from whole biomass and lipid-depleted biomass were 330.34 NmL/g VSadded and 364.34 NmL/g VSadded, respectively. In conclusion, the findings underscore the potent utility of Chlorella sp. S5 as a multi-faceted resource, proficiently employed in a sequential cascade for treating agricultural runoff, producing biodiesel, and generating biogas within the integrated biorefinery concept.

Unraveling the Radioprotective Mechanisms of UV-Resistant Bacillus subtilis ASM-1 Extracted Compounds through Molecular Docking.

Radioresistant microorganisms possess inimitable capabilities enabling them to thrive under extreme radiation. However, the existence of radiosensitive microorganisms inhabiting such an inhospitable environment is still a mystery. The current study examines the potential of radioresistant microorganisms to protect radiosensitive microorganisms in harsh environments. Bacillus subtilis strain ASM-1 was isolated from the Thal desert in Pakistan and evaluated for antioxidative and radioprotective potential after being exposed to UV radiation. The strain exhibited 54.91% survivability under UVB radiation (5.424 × 103 J/m2 for 8 min) and 50.94% to mitomycin-C (4 µg/mL). Extracellular fractions collected from ASM-1 extracts showed significant antioxidant potential, and chemical profiling revealed a pool of bioactive compounds, including pyrrolopyrazines, amides, alcoholics, and phenolics. The E-2 fraction showed the maximum antioxidant potential via DPPH assay (75%), and H2O2 scavenging assay (68%). A combination of ASM-1 supernatant with E-2 fraction (50 µL in a ratio of 2:1) provided substantial protection to radiosensitive cell types, Bacillus altitudinis ASM-9 (MT722073) and E. coli (ATCC 10536), under UVB radiation. Docking studies reveal that the compound supported by literature against the target proteins have strong binding affinities which further inferred its medical uses in health care treatment. This is followed by molecular dynamic simulations where it was observed among trajectories that there were no significant changes in major secondary structure elements, despite the presence of naturally flexible loops. This behavior can be interpreted as a strategy to enhance intermolecular conformational stability as the simulation progresses. Thus, our study concludes that Bacillus subtilis ASM-1 protects radiosensitive strains from radiation-induced injuries via biofilm formation and secretion of antioxidative and radioprotective compounds in the environment.

Immobilization of ß-1,4-xylanase isolated from Bacillus licheniformis S3.

Industrial applications require enzymes to be highly stable and economically viable in terms of reusability. Enzyme immobilization is an exciting alternative to improve the stability of enzymatic processes. Immobilization of ß-1,4-xylanase produced by Bacillus licheniformis S3 is performed by using two polymer supports (agar-agar and calcium alginate). The maximum enzyme immobilization yield was achieved at a concentration of 3% agar, whereas a combination of sodium alginate, 4%, and calcium chloride, 0.3 M, was used for the formation of immobilized beads. The immobilization process increased the optimum reaction time from 10 min to 35 and 40 min for agar and calcium alginate, respectively, and the incubation temperature increased from 55°C to 60°C for agar, but it remained unchanged for calcium alginate. The pH profile of free and immobilized xylanase was quite similar in both cases. Both the techniques altered the kinetic parameters of immobilized ß-1,4-xylanase as compared with the free enzyme. The diffusion limit of high molecular weight xylan caused a decline in Vmax of the immobilized enzyme, whereas there was an increase in the Km value. However, calcium alginate-immobilized enzyme displayed broad thermal stability as compared with agar-agar-immobilized enzyme and retained 57.1% of its initial activity at 80°C up to 150 min. Biotechnological characterization showed that the reusability of enzymes was the most striking finding, particularly of immobilized xylanase using agar-agar as immobilization carrier, which after six cycles retained 23% activity.

Characterization and synthetic biology of lipase from Bacillus amyloliquefaciens strain.

Lipases with high tolerance to temperature play a significant role in industry from food manufacturing to waste management systems. Thus, there is a need to investigate these enzymes from different geographical areas to look out for a more thermo-stable one. Characterization of lipases through experimental approaches is time consuming process and sometimes the results are ambiguous due to errors. However, integration of computational technologies is quite useful for prediction of optimized conditions. Such technologies can be applied as synthetic biology, which has many major applications in engineered biological approaches for accurate prediction of effects of different physical and chemical parameters on the system. In this study, cloning and expression of a lipase gene from Bacillus amyloliquefaciens, isolated from a novel geographical region of Pakistan, in Escherichia coli DH5α cells followed by sequencing was carried out. To isolate thermostable lipase producing strains, all the samples were kept at 50 °C. Genomic DNA was isolated and signal peptide (1-32 residues) sequence was chopped (ΔSPLipase). The ΔSPLipase was amplified and expressed in Linearized p15TV-L vector. The purified lipase appeared as single band of approximately 26 kDa. Suitable conditions of factors required for maximum lipase activity such as temperature, pH, substrate, organic solvent, detergents and metal ions were predicted through synthetic biology approach and further confirmed in wet lab. The predicted suitable factors for enzyme were almost similar to those determined experimentally. The optimum enzyme activity was recorded at pH 8 and 50 °C temperature. Interestingly, the activity of enzyme was found on a number of solvents, metal ions, detergents, and surfactants. The predicted optimum values and their experimental confirmations highlights the importance of integrated synthetic biology approaches in wet lab experiments. The characterized lipase of B. amyloliquefaciens at molecular level from Pakistani strains displayed good activity on a range of factors that implies this strain to be used for application in industrial level production.

Lipolytic bacterial strains mediated transesterification of non-edible plant oils for generation of high quality biodiesel.

Biodiesel is one of the best alternative to depleting fossil fuels for transport sector. However, biodiesel production via lipase mediated transesterification has limitation of high costing microbial enzymes. In order to overcome this limitation, a process of sequential treatment of oil industry wastewater using isolated lipolytic bacterial strains and biodiesel production from non-edible plant oils was studied. In this study, efficient lipase producing bacteria were isolated and evaluated for production of biodiesel from mustard, soybean, jatropha and taramira oils utilizing methanol for the transesterification of oils and bioremediation. Selected strains were then identified, using 16s rRNA sequencing. Further, Bacillus subtilis strain Q1 KX712301 was optimized for biodiesel production from non-edible taramira oil via Plackett-Burman and central composite design. Highest volumetric yield of biodiesel obtained was 102% at optimized parameters. Finally, a sequential bioremediation of vegetable oil contaminated wastewater and then microbial production of biodiesel from non-edible taramira oil was carried out using efficient lipase producer B. subtilis strain Q1 at optimized conditions. During sequential process, complete chemical oxigen demand reduction of oil containing wastewater and theoretical volumetric yield of biodiesel was achieved. Gas chromatography/mass spectrometry chromatogram revealed that the total fatty acid methyl ester content of the produced biodiesel was >98% which is in accordance with the biodiesel quality standards specified by both ASTM and EU-14103.

Morningness-eveningness preferences, learning approach and academic achievement of undergraduate medical students.

Several studies have focused on determining the effect of chronotype and learning approach on academic achievement separately indicating that morning types have an academic advantage over the evening types and so have the deep learners over the surface learners. But, surprisingly none have assessed the possible relationship between chronotype and learning approach. So, the current study aimed to evaluate this association and their individual influence on academic performance as indicated by the Cumulative Grade Point Average (CGPA) as well as the effect of their interaction on academic performance. The study included 345 undergraduate medical students who responded to reduced Morningness-Eveningness Questionnaire and Biggs Revised Two-Factor Study Process Questionnaire. Morning types indulged in deep learning while evening types in surface learning. Morning and evening types did not differ on academic performance but deep learners had better academic outcomes than their counterparts. The interaction between chronotype and learning approach was significant on determining academic achievement. Our findings gave the impression that chronotype could have an impact on academic performance not directly but indirectly through learning approaches.