DEVELOPMENT OF A MODIFIED METHOD FOR DESULFURIZATION OF DBE` USING A RECOMBINANT BACTERIAL STRAIN

Abstract

Sulfur content in crude oil available from various sources ranges from 0.03 to values as high as 8.0 wt%. These significantly high quantities of sulfur must be removed before the crude oil processing otherwise the combustion of this oil results in severe environmental pollution like acid rain. Due to high utility and operating costs, the conventional Hydrodesulfurization process (HDS) is considered to be uneconomic. The biotechnological option, Biodesulfurization (BDS) seems to be the attractive low cost, environmentally benign technology. Here we reported the development of recombinant bacterial strain which was designed by introducing the plasmid pSAD 225-32 containing desulfurization dsz genes, which was isolated from Rhodococcus erythropolis IGTS8 into a gram negative solvent tolerant bacterium, Pseudomonas putida (MTCC 1194). The recalcitrant heterocyclic organic sulfur compound Dibenzothiophene (DBT) was used as model compound for the biodesulfurization studies. This recombinant bacterium can desulfurize DBT in the sulfur specific 4S-pathway with out breaking the carbon-carbon bond. It has been observed that for the same concentration of DBT, the recombinant strain's growth rate and the consumption of DBT are greater than that of parent strain. The growth parameters 1-tmax and Ks were determined to be 0.0466 111 and 8.6875 mg/1 respectively. This genetically modified bacteria desulfurized 73.1% of 1.2 mM DBT in 67 h during cultivation. The actual biodesulfurization reaction industrially occurs in two-phase mixture containing oil to be desulfurized and the bacterial cell suspension, in aqueous phase. The desulfurization studies were performed using model oil. DBT dissolved in n-hexadecane is known as model oil, hexadecane represents hydrocarbons present in the 40% of the diesel oils. The optimum oil to water (0/W) operating ratio was determined to be 1:2. At the optimum 0/W ratio the recombinant bacteria desulfurized 96.95% of 2 mM DBT with in iii 24 h. The effect of surfactant, Tween-80 in enhancing the end product, 2-Hydroxybiphenyl (2-HBP) formation was also studied. After 36 h of desulfurization reaction, the 2-HBP produced was 1.5 times higher with the reaction mixture containing 1 g/I than that formed without surfactant. At the optimum 0/W ratio containing optimum surfactant concentration of 1 g/l, the resting cells of recombinant bacterium desulfurized 97.88% of 25.2 mM of DBT dissolved in n-hexadecane (model oil) with in 56 h with the specific desulfurization rate of 0:4397 mg sulfur/g biomass/I and it also desulfurized the diesel oil containing 22.025 g/1 to 2.934 g/I in 90 h with desulfurization rate 6.67 mg sulfur/g biomass/I. We conclude that the developed recombinant bacteria can efficiently desulfurize DBT in both aqueous as well as two phase media and also the real fraction of petroleum, diesel. Further research on this area of biodesulfurization using genetically modified microorganism may remove the bottlenecks present in the way of commercialization of BDS process. iv