Skrotskyi S. Biotechnology of direct inoculation preparation for activation of acetone-butyl alcohol fermentation.

The dissertation is devoted to the development of technology to produce the direct inoculation biopreparation from concentrates of acetone-butyl bacteria (CABB) on the basis of newly isolated strains of bacteria of the genus Clostridium. Biopreparation is purposed to simplify and increase the efficiency of obtaining fermentation products from substrates. Bacteria-producers of solvents (acetone, butanol, ethanol) and organic acids were isolated from various environmental sources. Based on 16S rRNA gene analysis isolates were identified as belonging to species Clostridium beijerinckii and Clostridium acetobutylicum. Obtained information was provided to GenBank database. Passports of three Clostridia strains (C. beijerinckii - 2 strains, C. acetobutylicum – 1 strain) were composed and Certificates of deposit No. IMV B-7806, IMV B-7701, IMV B-7807 were granted by the Depository of D.K. Zabolotny Institute of Microbiology and Virology NAS of Ukraine. There was demonstrated a possibility of replacement of high cost grain substrates for the synthesis of butanol and organic acids by C. beijerinckii IMV B-7806, C. beijerinckii IMV B-7701 and C. acetobutylicum IMV B-7807 with common industrial wastes that are mass pollutants of the environment (chicken manure, whey). For the first time a bacterial concentrate (CABB) of direct inoculation was proposed for acetone-butyl fermentation of substrates. Proposed preparation does not require any specialized microbiological operations for use and can be directly inoculated into the substrate in liquid or dry granular forms. The CABB included strains (C. beijerinckii IMV B-7806, C. beijerinckii IMV B-7701 and C. acetobutylicum IMV B-7807). Reducing the number of stages with use of CABB fastens the fermentation process in 4 times compared to traditional technology. For the first time the interaction of nanoparticles of metal oxides and ABB was demonstrated. Changes in cell morphology were shown in bacteria-nanoparticles interaction. The specific effect of nanoparticles on the synthesis of fermentation products was revealed. There is a potential for further enhancement of CABB efficiency by the use of cerium, silver and iron oxide nanoparticles. Immobilization of CABB cells in sodium alginate increases the subsequent synthesis of butanol of 26%. Efficient and inexpensive CABB manufacturing technology has been developed, consisting of: 1) separate cultivation of C. beijerinckii IMV B-7806, C. beijerinckii IMV B-7701 and C. acetobutylicum IMV B-7807 on nutrient media of different composition; 2) immobilization of strains using sodium alginate at a concentration of 20 g/L; 3) formation of calcium-alginate granules at a concentration of calcium chloride not lower than 5 g / L; 4) drying (40-45 °C.). A temporary technological regulation for the production of liquid and dry form of CABB was developed to use it for fermentation of substrates by the method of direct inoculation.