Enhance the bacterial nitrate reductase production using mathematical and statistical model to formulate the affordable Silver nanoparticle for the production of nanofinished fabric



Nitrate reductase (NR) is the critical enzyme to formulate the silver nanoparticles (AgNPs). The specific characteristics with defined high concentration are necessary to achieve the maximum yield of AgNPs to meet the industrial requirement. Bacteria are well reported to produce the NR in variable amounts depending on the types of the species and cultural conditions. Each bacterial cell has its unique circumstances to provide the higher NR enzyme. The conventional OVAT strategy used in media engineering since last few decades but due to its single dimensional task, it does not explain interaction effects among the variables on the enzyme production process. Moreover, it is a time consuming and laborious practice due to the need for a large number of experiments and often fails to identify the optimal conditions for each factor in the process. The present study was conducted to overcome these limitations. The response surface methodology (RSM) based primary screening was performed using a statistical model to screen the 4 media component out of 5 responsible for the enzymes production and growth of Ornithinibacillus californiensis using Plackett–Burman design (PBD). Total 13 experiments were conducted separately based on PBD. The positive values of the regression coefficient and Lower P values indicated the rejected null hypothesis, so linear model fit into the conducted study. The levels of the 4 significant variables, i.e., Glucose, Yeast extract, Casein hydrolysate, and KNO3 were further optimized using 30 experiments generated by Central Composite Design (CCD). The optimized media offered the 121 fold enhancement in NR production. Favorable analysis of variance, less P-values and model terms for both responses were found significant. Thus the experimental verification of the model was performed successfully and further analyzed by the 3D contour surface plots to test the interaction among the 4 variables.  The crude NR produced by optimized media was applied for the formulation of the AgNPs, and subsequently, AgNPs were characterized by the UV-Vis spectrophotometry, DLS with zeta potential, SEM-EDX, XRD, and TEM. Stable, confined-size, circular, crystalline and functionalized AgNPs were formulated using 4mM AgNO3 at the 60ºC temperature after 30 minutes. Antibacterial properties of AgNPs were also tested against the 10 human pathogens. So based on the AgNPs functionality; finally, the nanosilver coated cotton fabric was developed using the colloidal AgNPs and nanocoating was verify by SEM and FTIR. The study provides the prototype to develop the affordable biosynthesis of AgNPs and development of the surgical antimicrobial cotton.  

Keywords: Nitrate reductase, silver nanoparticles, Antibacterial, nonosilver








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