Optimization of uranium biosorption by pretreated citrobacter biomass

Toxic heavy metal contamination of industrial water is a significant universal problem. They accumulate in living tissues throughout the food chain which has humans at its top. These toxic metals can cause accumulative poisoning, cancer, and brain damage. Uranium is one of the most serious heavy metals because of its high toxicity and radioactivity. Excessive amounts of uranium have found their way into the environment through the activities associated with the nuclear industry (1). Conventional methods for removing uranium from wastewater include precipitation, evaporation, ion exchange, membrane processing, and adsorption. Nevertheless, these methods have several disadvantages, such as high installation and operating costs, the requirement of preliminary treatment steps, the difficulty of treating the subsequently generated solid waste, and low efficiency at low metal concentration (2,3). Owing to an increase in environmental awareness, there has been an emphasis on the development of new environmentally friendly ways to decontaminate waters using low-cost methods and materials. In this study, microbial biomass has emerged as a complementary, economic, and eco-friendly device for controlling the mobility and bioavailability of metal ions (2,4). The present work evaluates the performance of the Citrobacter freundii biomass to remove uranium ions from aqueous solutions. The effect of pH, temperature, initial concentration, and sorbent dose on biosorption capacity is also studied.

Materials: Citrobacter freundii bacteria used in this research with PTCC No. 1772 was purchased from the Scientific and Industrial Research Organization of Iran. Uranyl nitrate salt (UO2(NO3)2.6H2O) was obtained from the Research Institute of Nuclear Sciences and Technologies. Nutrient Broth culture medium, sulfuric acid, and sodium hydroxide and other materials used in this research were supplied from the Merck Company.Preparation of uranium solutions and biomass: A stock solution containing 1000 mg L-1 of U(VI) was prepared of UO2(NO3)4.6H2O. The working solutions were prepared daily from stock solutions. In this study, the biomass of Citrobacter freundii bacteria was heat treated in an autoclave at a temperature of 121°C for 15 minutes at a pressure of 1.5 atmospheres. Experimental design and batch biosorption studies: The design of the experiment was done using the response surface method by Design Expert software. Four variables, including initial uranium concentration (mg/l), temperature (°C), pH, and biosorbent dose (g/l), in five levels α-, -1, 0, +1, α+, 1 were used to design the experiment (Table 1). Therefore, 27 experiments were presented using a central composite design. The values of the variables and the obtained answers are shown in Table 2. Uranium biosorption experiments were performed by adding specified amounts of bacterial biomass in 20 ml Erlenmeyer flasks containing uranium solution with the concentration and pH corresponding to each experiment, with the specified temperature in the Shaker. After 90 minutes, each sample was centrifuged at 4500 rpm for 15 minutes at 4°C. Then, the remaining uranium in the solution was measured by ICP (Perkin Elmer/Optima 7300DV). The percentage of uranium removal (R) was calculated by equation 1:  Where C0 and Cf are the initial and the final concentrations of the metal ion solutions (mg/l), respectively.Table 1- Variables and Levels of the Central Composite Design Method

By using the RSM-CCD method, the optimization of the biosorption process was carried out. Table 2 shows the experimental results based on each point of the experimental design. Then, using analysis of variance (ANOVA), the obtained results were evaluated.The equation obtained for the biosorption efficiency of uranium by Citrobacter freundii is expressed as follows:Removal= +68.97045-1.43160 * C (ppm)+12.81296 * pH+1.08935 * T (0C)+2.89856 * M (g/l)+0.55737 * C (ppm) * pH+0.011459* C (ppm)* T (0C)+0.014961* C (ppm)* M (g/l)-0.32111 * pH * T (0C)-0.62783 * pH * M (g/l)-0.037633* T (0C) * M (g/l)-6.93488E-003* C (ppm)2-4.06361 * pH2Table 2- Values of Variables and Experimental Responses in the Response Surface Method.

The F-value and p-value of the proposed model are equal to 5.03 and 0.0027, respectively, reflecting the accuracy of the proposed model. This model with R2 equal to 0.81 shows that the proposed model can well predict the experimental values. The results showed that the factor of initial uranium concentration and pH statistically affect (p-value‹ 0.05) the uranium biosorption process. In contrast, temperature and sorbent dose factor (p-value› 0.05) have no statistically effect on uranium removal by Citrobacter freundii. With increasing uranium concentration from 10 mg/l to 77.5 mg/l, the removal increases from %66.5 to %99/92. Then, with increasing uranium concentration from 77.5 mg/l to 100 mg/l, the removal decreases to %97.34. On the other hand, one of the most important effective parameters in biosorption is the pH of the solution. With increasing the pH from 2 to 5, the removal decreased from %96.82 to %79.01 due to the formation of uranyl complexes (5). In this research, the results indicated that the pre-treated biomass under the conditions suggested by Design Expert software (19.84 g/l of biomass, temperature 28.92 OC, pH 2.89 and initial uranium concentration 53.71 mg/l) is able to remove approximately 99.99 percent of uranium from the contaminated area, which shows valuable potential Citrobacter freundii in bioremediation applications of uranium from contaminated wastewaters.