Owuna, Gabriel2023-12-142023-12-142019-01-05A THESIS SUBMITTED TO THE SCHOOL OF POSTGRADUATE STUDIES, NASARAWA STATE UNIVERSITY KEFFI, IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE AWARD OF DEGREE OF DOCTOR OF PHILOSOPHY IN ENVIRONMENTAL MICROBIOLOGYhttps://keffi.nsuk.edu.ng/handle/20.500.14448/5992DissertationBiobutanol is an ecologically friendly alternative fuel that can be used in place of gasoline in car engines. The efficient utilization of waste papers into biobutanol by microbial processes is a worthwhile venture. This will eliminate the use of food crops such as maize for biofuel production. Biobutanol has similar physical and chemical properties with gasoline, and `can be used as fuel in car engines to serve the same purpose as gasoline. In addition to its use as a fuel, it has other uses such as the manufacture of printing inks, perfumes, fumigants, and moistening agents as well as plasticizers and pesticides. In this investigation, the soil environment of Keffi metropolis was screened for the presence of biobutanol-producing Clostridium species which were isolated using standard methods and identified using standard molecular techniques involving the use of 16S rRNA. The ability of the various isolates to produce biobutanol was determined using Lucas‟ Test while the quantitative determination of biobutanol produced by each isolate was carried out using Gas Chromatography and Mass Spectrometry (GC & MS). The isolates were later subjected to Hank‟s buffer and UV light treatment for the possibility of obtaining high yield biobutanol in biobutanol-producing Clostridium species. The results obtained showed that the different species of Clostridium isolated from the study area included Clostridium difficile, Clostridium botulinum, Clostridium cylindrosporum, Clostridium perfringens, Clostridium acetobutylicum and Clostridium celerecrescens. The investigation further revealed that Clostridium acetobutylicum (Ca2) produced the highest yields of 241,496.0±1.00μg/mL and 1,465,122.0±1.00μg/mL both before and after treatment of clostridial isolates with UV light/Hank‟s buffer followed by Clostridium perfringens (Cp1) with a yield of 326.0±1.41 μg/mL and 1,404,668.0±1.41 μg/mL both before and after treatment of clostridial isolates with UV light/Hank‟s buffer while Clostridium difficile (Cd2) had the lowest yield of 187.0±1.41 μg/mL and 2,127.0±1.41 μg/mL both before and after treatment of clostridial isolates with UV light/Hank‟s buffer. Furthermore, only Clostridium difficile (Cd2), Clostridium perfringens (Cp1) and Clostridium acetobutylicum (Ca2) produced biobutanol during GC & MS analysis of fermentation broth before treatment of clostridial isolates with UV light/Hank‟s buffer with the yields of 187.0±1.41 μg/mL, 326.0±1.41 μg/mL and 241,496.0±1.00 μg/mL respectively. Similarly, Clostridium difficile (Cd1), Clostridium botulinum (Cb1), Clostridium difficile (Cd2), Clostridium perfringens (Cp1), Clostridium difficile (Cd3), Clostridium acetobutylicum (Ca1), Clostridium celerecrescens (Cce1), Clostridium acetobutylicum (Ca2), and Clostridium cylindrosporum (Cc2) produced 241,496.0±1.41 μg/mL, 1,956.0±1.41 μg/mL, 2,127.0±1.41 μg/mL, 1,404,668.0±1.41 μg/mL, 191,130.0±1.00 μg/mL, 3,961.0±2.0 μg/mL, 535.0±1.24 μg/mL, 1,465,122.0±1.00 μg/mL and 152,685.0±1.00 μg/mL after treatment of clostridial isolates with UV light/Hank‟s buffer respectively. It was discovered that there was a statistically significant difference of the concentrations (μg/mL) between the Gas Chromatography & Mass Spectrometry (GC & MS) analysis before and after induced mutation as determined by one-way ANOVA (F = 3.0781; P-value = 0.3846). The yields of biobutanol produced were also found to be significant at a confidence level of 95%. This study therefore validates the alternate hypotheses that Clostridium species are present in the soil environment of Keffi metropolis. This study further demonstrated that paper wastes can be utilized as substrates for biobutanol production and the results of this study could be explored for the the management of wastes papers and other cellulose containing wastes for biobutanol production using species of ClostridiumenThesis