Biosynthesis of Zinc Oxide Nanoparticles Using Bacillus paramycoides for In Vitro Biological Activities and In Vivo Assessment Against Hepatorenal Injury Induced by CCl4 in Rats.

Recently, impressive developments in the field of nanotechnology have been achieved. The study aimed to synthetize zinc oxide nanoparticles (ZnONPs) from locally isolated terrestrial Bacillus paramycoides (MCCC 1A04098) bacteria and assess its role as antioxidant, antimicrobial, and anticancer agent. The antioxidant activity was done using the percentage of DPPH scavenging method. The antibacterial activity was evaluated against Escherichia coli, Staphylococcus aureus, Bacillus cereus, and Candida albicans. The anti-proliferation assay against hepatocellular carcinoma (HepG2) and human breast cancer (MCF-7) cell lines was estimated by neutral red assay. The apoptotic effect of ZnONP was measured by flow cytometry. The in vivo evaluation was carried out against hepatorenal injuries induced by carbon tetrachloride (CCl4) in rats comparing with silymarin as a reference drug. The oxidative stress markers, liver and kidney function enzyme indices, lipid profile, and the histological features of the liver and kidney were also examined. ZnONPs revealed antioxidant and antibacterial effects. It also exerted cytotoxic and apoptotic effect in a dose dependent manner without any toxicity on normal cell line. ZnONPs improved all the biochemical parameters under investigation to varying degrees, and the histological pictures of the liver and kidney confirmed the results. In conclusion, ZnONPs were successfully synthesized from the terrestrial Bacillus paramycoides and recorded in vitro antioxidant, anticancer, and antibacterial effects as well as in vivo anti-hepatorenal toxicity effects.

Rhizopus stolonifer biomass catalytic transesterification capability: optimization of cultivation conditions.

BACKGROUND: Using fungal biomass for biocatalysis is a potential solution for the expensive cost of the use o enzymes. Production of fungal biomass with effective activity requires optimizing the cultivation conditions. RESULTS: Rhizopus stolonifer biomass was optimized for transesterification and hydrolysis of waste frying oil (WFO). Growth and biomass lipolytic activities of R. stolonifer improved under shaking conditions compared to static conditions, and 200 rpm was optimum. As biomass lipase and transesterification activities inducer, olive oil was superior to soybean, rapeseed, and waste frying oils. Biomass produced in culture media containing fishmeal as an N-source feedstock had higher lipolytic capabilities than corn-steep liquor and urea. Plackett Burman screening of 9 factors showed that pH (5-9), fishmeal (0.25-1.7%, w/v), and KH2PO4 (0.1-0.9%, w/v) were significant factors with the highest main effect estimates 11.46, 10.42, 14.90, respectively. These factors were selected for response surface methodology (RSM) optimization using central composite design (CCD). CCD models for growth, biomass lipase activity, and transesterification capability were significant. The optimum conditions for growth and lipid modification catalytic activities were pH 7.4, fishmeal (2.62%, w/v), and KH2PO4 (2.99%, w/v). CONCLUSION: Optimized culture conditions improved the whole cell transesterification capability of Rhizopus stolonifer biomass in terms of fatty acid methyl ester (FAME) concentration by 67.65% to a final FAME concentration of 85.5%, w/w.

Biocatalysis of triglycerides transesterification using fungal biomass: a biorefinery approach.

BACKGROUND: The use of microbial biomasses, such as fungal biomass, to catalyze the transesterification of triglycerides (TG) for biodiesel production provides a sustainable, economical alternative while still having the main advantages of expensive immobilized enzymes. RESULTS: Biomasses of Aspergillus flavus and Rhizopus stolonifera were used to catalyze the transesterification of TG in waste frying oil (WFO). Isopropanol as an acyl-acceptor reduced the catalytic capability of the biomasses, while methanol was the most potent acyl-acceptor with a final fatty acid methyl ester (FAME) concentration of 85.5 and 89.7%, w/w, for R. stolonifer and A. flavus, respectively. Different mixtures of the fungal biomasses were tested, and higher proportions of A. flavus biomass improved the mixture's catalytic capability. C. sorokiniana cultivated in synthetic wastewater was used as feedstock to cultivate A. flavus. The biomass produced had the same catalytic capability as the biomass produced in the control culture medium. Response surface methodology (RSM) was adopted using central composite design (CCD) to optimize the A. flavus biomass catalytic transesterification reaction, where temperature, methanol concentration, and biomass concentration were selected for optimization. The significance of the model was verified, and the suggested optimum reaction conditions were 25.5 °C, 250 RPM agitation with 14%, w/w, biomass, 3 mol/L methanol, and a reaction duration of 24 h. The suggested optimum conditions were tested to validate the model and a final FAME concentration of 95.53%. w/w was detected. CONCLUSION: Biomasses cocktails might be a legitimate possibility to provide a cheaper technical solution for industrial applications than immobilized enzymes. The use of fungal biomass cultivated on the microalgae recovered from wastewater treatment for the catalysis of transesterification reaction provides an additional piece of the puzzle of biorefinery. Optimizing the transesterification reaction led to a valid prediction model with a final FAME concentration of 95.53%, w/w.

Green method for improving performance attributes of wool fibres using immobilized proteolytic thermozyme.

Wool has the tendency to turn into felt during agitation in washing machines. Thus, a benign non-polluting method for the production of machine-washable wool was developed herein. Initially, a proteolytic bacteria was isolated from hot region soil. The bacterial isolate was identified as Bacillus safensis FO-36bMZ836779 according to the 16S rRNA gene sequencing. Afterwards, the extracellular protease produced by this isolate was covalently immobilized in order to enhance its stability under non-ambient conditions which are usually adopted in industrial sectors like textile industries. Sericin, which is usually discharged into degumming effluent of natural silk, was utilized to prepare the immobilization carrier. Box-Behnken design was adopted in order to hone the preparation of the sericin-polyethylene-imine-glutaraldehyde activated agar carrier. The pH and temperature profiles of the free and immobilized proteases were compared. Later, wool fibres were bio-treated with both the free and the immobilized enzymes. The effect of process conditions on the resistance of the bio-finished wool to felting was investigated. The alteration in the fibre morphology was monitored using SEM. Amino acid analysis and alkali solubility tests were adopted to assign any change in the chemical structure of the bio-treated wool. The influence of bio-treatment of wool on its inherent properties was assigned. Results revealed that bio-treatment of wool with the said enzyme led to production of machine-washable wool without severe deterioration in the fibres' properties. In an energy- and water-consuming process, the hot solution from bio-treatment bath was used successfully in dyeing of wool. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03323-y.

Bioscouring of wool fibres using immobilized thermophilic lipase.

The hydrophobic nature of wool induced by its surface lipid barrier hinders its wettability during processing. Scouring of wool is conducted to remove this lipid barrier and facilitate any wet processes. Scouring of wool is conducted using soda ash followed by rinsing with huge amount of water to ensure complete removal of alkali. This work aimed at utilization of thermophilic lipase enzyme for removal of wool surface lipid barrier without deterioration on the fibre interior. A thermally stable lipase enzyme was produced from thermophilic microorganism; namely Bacillus aryabhattai B8W22, and was utilized in bio-scouring of wool. The produced enzyme was immobilized on sericin-based discs to enhance its stability and to make it reusable. The activity of both free and immobilized lipase enzymes at different conditions was assessed. The effects of bio-scouring of wool on its dyeability with acid, basic, and reactive dyes, as well as on some of its inherent properties, were monitored. Results showed that the bio-scoured wool exhibits enhanced dyeability with the said classes of dyes more than that of conventionally scoured samples. One-bath scouring and dyeing of wool fibres in two successive steps was conducted to reduce consumption of water and energy during wet processing of wool.

Structural characterization, catalytic, kinetic and thermodynamic properties of Keratinase from Bacillus pumilus FH9.

Bacillus pumilus FH9 keratinase was purified to homogeneity with a 59.9% yield through a series of three steps. The purified enzyme was a monomeric protein with a molecular mass around 50kDa and containing 7.3% carbohydrates. The pure B. pumilus FH9 keratinase was optimally active at pH 9.0 and 60°C. The calculated activation energy for keratin hydrolysis was 24.52kJmol-1 and its temperature quotient (Q10) was 1.19. The calculated values of thermodynamic parameters for keratin hydrolysis were as follows: ΔH*=21.75kJmol-1, ΔG*=65.86kJmol-1 ΔS*=-132.46Jmol-1K-1, (ΔG*E-S)=4.74kJmol-1 and ΔG*E-T=-11.254kJmol-1. The pure keratinase exhibited Km, Vmax, kcat and kcat/Km of 5.55mg/ml keratin, 5882Umgprotein-1 323.54s-1 and 58.28 (s-1/mgml-1). The calculated half-life time at 50, 60, 70 and 80°C was 90.69, 59.1, 16.62 and 9.48min, respectively. Similarly, the thermodynamic parameters for irreversible thermal inactivation at temperature ranging from 50 to 80°C were determined. The pure enzyme was stimulated by Ca2+ and Mg2+. However, Zn2+, EDTA, Co2+ and Hg2+ significantly inhibited the enzyme activity. The purified enzyme was able to hydrolyze different substrates showing its higher proteolytic activity on casein, bovine serum albumin, and collagen, followed by feather, horn and wool.

Comparison of Free and Immobilized L-asparaginase Synthesized by Gamma-Irradiated Penicillium cyclopium.

Gamma irradiation is used on Penicillium cyclopium in order to obtain mutant cells of high L-asparaginase productivity. Using gamma irradiation dose of 4 KGy, P. cyclopium cells yielded L-asparaginase with extracellular enzyme activity of 210.8 ± 3 U/ml, and specific activity of 752.5 ± 1.5 U/mg protein, which are 1.75 and 1.53 times, respectively, the activity of the wild strain. The enzyme was partially purified by 40-60% acetone precipitation. L-asparaginase was immobilized onto Amberlite IR-120 by ionic binding. Both free and immobilized enzymes exhibited maximum activity at pH 8 and 40 degrees C. The immobilization process improved the enzyme thermal stability significantly. The immobilized enzyme remained 100% active at temperatures up to 60 degrees C, while the free asparaginase was less tolerant to high temperatures. The immobilized enzyme was more stable at pH 9.0 for 50 min, retaining 70% of its relative activity. The maximum reaction rate (V(max)) and Michaelis-Menten constant (K(m)) of the free form were significantly changed after immobilization. The K(m) value for immobilized L-asparaginase was about 1.3 times higher than that of free enzyme. The ions K+, Ba2+ and Na+ showed stimulatory effect on enzyme activity with percentages of 110%, 109% and 106% respectively.

Cyclodextrin glucosyltransferase production by Bacillus megaterium NCR: evaluation and optimization of culture conditions using factorial design.

Statistically-based experimental designs were used to optimize the production of cyclodextrin glucosyltransferase (CGTase) from a local isolate of Bacillus megaterium using shack culture fermentation. Seven cultural conditions were examined for enzyme production and specific activity using Plackett-Burman factorial design. Fermentation time and K(2)HPO(4) level were the crucial for factors improving enzyme production process. The steepest ascent design was adopted-based on the results recorded with Plackett-Burman design. Maximal enzyme estimates (activity 56.1 U/ml, and specific activity 62.7 U/mg protein) were achieved. A verification experiment was carried out to examine model validation of this optimization.