| Heavy metal contamination of soils is one of the major environmental effects of urbanization and industrialization. Excessive heavy metals in soil will reduce crop production, influence the quality of agricultural products and groundwater, and be accumulated in human bodies and organisms through food chain which endangered the health of human beings. In recent years, fungus were reported to have the capability of producing organic acids that can serve as leaching agents for removing heavy metals from environment.This paper attempted to isolate an acid-producing heterotrophic fungi Aspergillus niger which was used for leaching heavy metals from contaminated soil. Then batch experients were performed to compare the leaching effiency of different culture methods and pulp density. At last, kinds of agricultural wastes as substrates were investigated and evaluated. The main conclusions are as follows.(1) An acid-producing heterotrophic fungi named Y9 was isolated from soil and identified as Apergillus niger through morphological identification and analysis of ITS rDNA sequences. Results showed that the optimal pH for its growth was from 3.0 to 8.0. Also, A. niger Y9 had a strong tolerance to Zn2+, Cd2+ and Pb2+, and the 80% inhibitory concentration was 250 mg/L,130 mg/L,2700 mg/L, respectively. After incubating for 20 days, pH value of the pure culture system declined to 2.1 from initial 5.3. Meanwhile, this fungi excreted 54.8 mmol/L citric acid,20.1 mmol/L gluconic acid,15.9 mmol/L oxalic acid and 6.3 mmol/L malic acid.(2) A. niger Y9 exhibited a good potential for heavy metals removal from soil. Results showed that after one-step process (fungus was incubated together with soil), at 5%(w/v) of pulp density and 12 d of leaching time, maximum removals of zinc, cadmium and lead were 64%,86% and 19% achieved, respectively. After the two-step process (soil was added after 2 days of fungi incubation), at 5%(w/v) of pulp density and 15 d of leaching time, highest removal of 75%Zn,100% Cd,43% Pb were obtained. Besides, results of sequential extraction showed that A. niger Y9 was effective in removing the exchangeable, carbonate and Fe/Mn oxide fractions of zinc, cadmium and lead, and after both process the metals remaning in the soil were mainly found in the stable fraction which was unavailable to surrounding environment. Considering the leaching efficiency and operating costs, it is preferable to remediate the contaminated soil through one-step process.(3) In bioleaching, the pulp density of soil could highly influence the pH, organic acid production and heavy metals extraction. Results indicated that as the pulp density of soil was gradually increased, pH of the leaching system was increased, and a higher concentration of gluconic acid was achieved, but the concentration of citric acid was decreased and the production of oxalic acid tended to delay. Meanwhle, leaching efficiency of heavy metals decreased with increasing pulp density. When pulp density was 5%(w/v), the highest extraction yields of zinc, cadmium and lead were 84%,95% and 40%, respectively. While at 10%(w/v) pulp density of soil, the amount of leached metals were Zn 71%, Cd 94%, Pb 32%. Considering the capital costs,10%(w/v) pulp density was preferable. Furthermore, the effects of pulp density and biogenic orgainic acids on leaching effiency were analyzed through ridge regression analysis in SAS. It was found that pH, pulp density, as well as citric acid concentration were the dominate factors for extracting zinc from soil. Lower pH and higher concentration of oxalic acid and gluconic acid production could significantly imrpove cadium extraction yield. Meanwhile, the leaching rate of lead were greatly affected by pH and content of citric acid. In addition, Batch experiments of solid-liquid reflux and liquid reflux were performed at 10%(w/v) pulp density. Results showed that both methods could effectively remove zinc and cadmium from soil after twice reflux, but the extraction yield of lead was too low. Compared to the solid-liquid reflux, liquid reflux exhibitied a better leaching efficiency.(4) To lower the costs of bioleaching, some food byproducts and agricultural wastes such as sugarcane molasses, citrus peels, potato peels and alkali-hydrolyzed corn straw were used as alternative carbon souces instead of sucrose in A. niger cultivation. In pure culture system, pH decreased in the order of sucrose, potato peels, alkali-hydrolyzed corn straw, sugarcane molasses, citrus peels. Results of HPLC showed that A. niger Y9 mainly produced oxalic acid and small amount of citric acid, gluconic acid and malic acid. Fermentation of the alkali-hydrolyzed corn straw resulted in the highest concentration of oxalic acid (111 mmol/L). In the bioleaching system, the variation tendency of pH was similar to the pure culture, but the pH value was slightly higher. In addition, the higher concentration of oxalic acid corresponded to higher leaching yield of heavy metals, which indicated that the main lixiviant in bioleaching was shown to be oxalic acid. Fermentation of sucrose, potato peels and alkali-hydrolyzed corn straw achieved a good leaching efficiency of zinc 60-80%. The maximum removal of cadmium was 60% obtained at sucrose medium, followed by alkali-hydrolyzed corn straw leaching system which was about 43%. Fermentation of alkali-hydrolyzed corn straw could also resulted a highest extraction yield of lead (23%), while other carbon sources was inefficient for removal of lead. As for sugarcane molasses and citrus peels, the leaching yields of metals were almost minimum due to the higher pH and lower concentration of organic acid.The results of this research could provide useful imformation to researchers and practitioners working on the remediation of contaminated soil, and promote the application of bioleaching techonology by Aspergillus niger. |
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