Preparation and Evaluation of the ZnO NP-Ampicillin/Sulbactam Nanoantibiotic: Optimization of Formulation Variables Using RSM Coupled GA Method and Antibacterial Activities.

Nanoparticles (NPs) possessing antibacterial activity represent an effective way of overcoming bacterial resistance. In the present work, we report a novel formulation of a nanoantibiotic formed using Ampicillin/sulbactam (Ams) and a zinc oxide nanoparticle (ZnO NP). 'ZnO NP-Ams' nanoantibiotic formulation is optimized using response surface methodology coupled genetic algorithm approach. The optimized formulation of nanoantibiotic (ZnO NP: 49.9 µg/mL; Ams: 33.6 µg/mL; incubation time: 27 h) demonstrated 15% enhanced activity compared to the unoptimized formulation against K. pneumoniae. The reactive oxygen species (ROS) generation was directly proportional to the interaction time of nanoantibiotic and K. pneumoniae after the initial lag phase of ~18 h as evident from 2'-7'-Dichlorodihydrofluorescein diacetate assay. A low minimum inhibitory concentration (6.25 µg/mL) of nanoantibiotic formulation reveals that even a low concentration of nanoantibiotic can prove to be effective against K. pneumoniae. The importance of nanoantibiotic formulation is also evident by the fact that the 100 µg/mL of Ams and 25 µg of ZnO NP was required individually to inhibit the growth of K. pneumonia, whereas only 6.25 µg/mL of optimized nanoantibiotic formulation (ZnO NP and Ams in the ratio of 49.9: 33.6 in µg/mL and conjugation time of 27 h) was needed for the same.

Purification and characterization of chitinase from Streptomyces violascens NRRL B2700.

Chitinase is one of the important enzymes as it is directly linked to Chitin that has wide applications in industrial, medical and commercial fields for its biocompatibility and biodegradability. Here, we report extracellular chitinase production by Streptomyces violascens NRRL B2700 under submerged fermentation condition. Chitinase production started after 10 h of incubation and reached to maximum level at 72 h of cultivation. Studies on the influence of additional carbon and nitrogen sources on chitinase production revealed that maltose, xylose, fructose, lactose, soybean meal and ammonium nitrate served as good carbon and nitrogen sources to enhance chitinase yield by 1.6 to 6 fold. Medium supplemented with 1% colloidal chitin produced high chitinase concentration (0.1714 U/mg). The enzyme chitinase was purified from the culture broth by 75% ammonium sulphate precipitation, DEAE-cellulose ion-exchange and sephadex G-100 gel filtration. The molecular mass of the purified chitinase was 65 kDa as estimated by SDS-PAGE. The apparent Michaelis constant (K(m)) and the maximum rate (V(max)) of the enzyme for colloidal chitin were 1.556 mg/mL and 2.680 µM/min/mg, respectively suggested high affinity towards-chitin. Possibly, it is the first report on production of chitinase from S. violascens NRRL B2700. The findings were encouraging, especially for cost effective production, and further warrants media and purification optimization studies for enhanced yield.