Adsorption Of Cu²⁺ By Natural Biofilms Associated With Extracellular Polymeric Substances （EPS）

Copper（Cu） is the indispensable trace element which could maintain the normal life activities of organisms. But Cu will produce toxic to creature, to be a typical heavy metal pollutant in water when it reaches a certain concentration. The natural biofilom has been a hot area of research for several years as a kind of composite solid medium which could be effectively combined with the heavy metal. In this study, we cultivated biofilms in spring, summer and autumn（it was difficult to form biofilms in winter） in Lihu Lake of Jiangnan University. The effects of the various environmental factors, which could influence the adsorption of Cu²⁺ in biofilm and its extracellular polymers（EPS） were studied.The Scanning Electron Microscope（SEM）, Denaturing Gel Gradient Electrophoresis（PCR-DGGE）, Three-dimensional Fluorescence（EEM）,Fourier Infrared Spectrometer（FT-IR） and Energy Dispersive X-ray Spectroscopy（SEM-EDX） were used to further confirme the mechanism of the sorption of Cu²⁺ and the influence of Cu²⁺ on the structure of the biofilm and EPS. The results are as follows:（1） Determine the period of the mature biofilms cultivation in different seasons by measuring the contents of biofilm biomass, chlorophyll, TOC, Fe oxide, Mn oxide,dehydrogenase activity and EPS, The results show that the period of the mature biofilm was 21 d（spring）, 15 d（summer） and 35 d（fall） respcetively.（2） By comprison of the five different extraction methods of EPS from natural biofilms, which including the thermal treatment, EDTA treatment, NaOH+Na Cl treatment, ultrasonic+CER treatment and high speed centrifugation（the control）.NaOH+Na Cl treatment was the most effective extraction method.Under the optimized condition of NaOH concentration 2 mol·L-1, NaCl mass fraction 0.85%, extraction time 2.5 h and centrifugal force 7000 g, the extraction amount of EPS was 157.57mg·L-1. Analysis of FT-IR shows that the protein was the main components of EPS,closely followed by the polysaccharide, the extraction of protein increased with the addition of NaOH.（3） Through the single-factor experiments, the best absorbent effect of the mature biofilms in different seasons could be obtained in the initial Cu²⁺ concentration of 5 mg·L-1, pH of 6 and 20℃, associated with the adsorption capacitice were 29.12mg·g-1（Spring）, 44.68 mg·g-1（Summer） and 15.35 mg·g-1（Autumn）. The adsorption amount of Cu²⁺ was less when Pb²⁺, Cr2+and Cu²⁺ coexistent system than in the single Cu²⁺and the influence of Pb²⁺ was greater than Cr2+. The adsorption kinetics agreedwell with the pseudo-second-order model, and the rate of adsorption was Summer>Spring>Autumn.（4） The results indicate that Cu²⁺ stimulated the biofilm to secrete more extracellular polymers, which could be conducive to the adsorption of Cu²⁺. The microbial community similarity of the biofilm was 73%, impling that the Cu²⁺ stress had no significant effects on microbes, while the bacterial diversity index was lower.The contributions of major chemical components to Cu²⁺ enrichment: Mn oxide（46.03%）>Organic matter（32.67%）>Fe oxide（15.69%）>Others（5.61%）. Analysis of EEM demonstrates that Cu²⁺ could cause fluorescence quenching of protein and humus.（5） The best absorbent effect of S-EPS and B-EPS could be obtained in the initial concentration of 7 mg·L-1, pH of 6, m（S-EPS）:m(Cu²⁺)=1.2:1, m（B-EPS）:m(Cu²⁺) =2:1 and 30℃, the adsorption capacitice were 133.65 mg(Cu²⁺)·g-1（S-EPS）>115.34 mg(Cu²⁺)·g-1（B-EPS）. The adsorption isotherm and the adsorption kinetics agreed well with Langmuir model and pseudo-second-order model. Analysis of FT-IR demonstrates that hydroxyl and acylamino were the mian functional groups in the reaction between metal and EPS. SEM-EDX analysis reveales that the ion-exchange was an important mechanism in the Cu²⁺ sorption process. The best desorption ratios of Cu²⁺ could be obtained in the initial NaNO3 concentration of 0.05 mol·L-1, pH of 2together with the best recovery rate was 81.34%.