bacterial sorption of heavy metals

1、Vol. 55, No. 12 APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Dec. 1989, p. 3143-3149 0099-2240/89/123143-07$02.00/0 Copyright © 1989, American Society for MicrobiologyBacterial Sorption of Heavy MetalstM. D. MULLEN,lt* D

2、. C. WOLF,' F. G. FERRIS,2§ T. J. BEVERIDGE,2 C. A. FLEMMING,2 AND G. W. BAILEY3Department ofAgronomy, 115 Plant Science Building, University ofArkansas, Fayetteville Arkansas 727011; Department of Microbiology,

3、 College ofBiological Science, University of Guelph, Guelph, Ontario, Canada NIG 2 Wi2; and Environmental Research Laboratory, U.S. Environmental Protection Agency, Athens, Georgia 30613-77993Received 19 June 1989/Accept

4、ed 29 September 1989Four bacteria, Bacillus cereus, B. subtilis, Escherichia coli, and Pseudomonas aeruginosa, were examined for the ability to remove Ag+, Cd2+, Cu2+, and La3+ from solution by batch equilibration method

5、s. Cd and Cu sorption over the concentration range 0.001 to 1 mM was described by Freundlich isotherms. At 1 mM concentrations of both Cd2+ and Cu2+, P. aeruginosa and B. cereus were the most and least efficient at metal

6、 removal, respectively. Freundlich K constants indicated that E. coli was most efficient at Cd2' removal and B. subtilis removed the most Cu2+. Removal of Ag+ from solution by bacteria was very efficient; an average

7、of 89% of the total Ag+ was removed from the 1 mM solution, while only 12, 29, and 27% of the total Cd2+, Cu2+, and La3+, respectively, were sorbed from 1 mM solutions. Electron microscopy indicated that La3+ accumulated

8、 at the cell surface as needlelike, crystalline precipitates. Silver precipitated as discrete colloidal aggregates at the cell surface and occasionally in the cytoplasm. Neither Cd2' nor Cu2+ provided enough electron

9、 scattering to identify the location of sorption. The affinity series for bacterial removal of these metals decreased in the order Ag > La > Cu > Cd. The results indicate that bacterial cells are capable of bind

10、ing large quantities of different metals. Adsorption equations may be useful for describing bacterium-metal interactions with metals such as Cd and Cu; however, this approach may not be adequate when precipitation of met

11、alsoccurs.The fate of toxic metallic cations in the soil environment depends largely on the interactions of these metals with inorganic and organic surfaces. The extent to which a metallic cation interacts with these sur

12、faces determines the concentration of metal in solution and, consequently, the potential for movement into groundwater or uptake by plants. A considerable amount of work has been done to evaluate the adsorption or comple

13、xation of various heavy metals by soils (11) and soil constituents, such as clays (22) and organic matter fractions (28). One potentially important organic surface which has received little attention is that of the soil

14、microbial population. Soil microorganisms are typi- cally associated with the clay and organic fractions of the soil microenvironment (21) and would be expected to par- ticipate in the metal dynamics typically ascribed t

15、o these fractions. Bacteria have a high surface area-to-volume ratio (2) and, as a strictly physical cellular interface, should have a high capacity for sorbing metals from solution. There is evidence that bacterial cell

16、s are more efficient at metal removal than clay minerals on a dry-weight basis (31). Kurek and co-workers (17) observed that sorption of Cd2+ by dead cells of a Paracoccus sp. and Serratia marcescens was greater than tha

17、t of montmorillonite when the solid-to- solution ratio was the same for both bacteria and clay. Live cells accumulated about the same quantity of Cd2+ as did clay. Several investigations have shown that relatively large

18、quantities of metallic cations are complexed by algae (19),* Corresponding author. t Published with the approval of the Director of the Arkansas Agricultural Experiment Station. t Present address: School of Agriculture a

19、nd Home Economics, 132 Brehm Hall, University of Tennessee-Martin, Martin, TN 38238-5008. § Present address: Nova Husky Research Corp., Biosciences Group, Calgary, Alberta, Canada T2E 7K7.bacteria (29), and fungi (2

20、0). Metal binding by isolated gram-positive and gram-negative bacterial cell walls has also been evaluated (3, 5, 6, 10, 20). Cell walls of the gram- positive bacteria Bacillus subtilis and B. licheniformis were observed

21、 to bind larger quantities of several metals than cell envelopes of the gram-negative bacterium Escherichia coli(3). We are interested in the role of microorganisms in the behavior of various heavy metals in the soil env

22、ironment. The objectives of this work were to determine the metal- binding capacities of whole cells of two gram-positive and two gram-negative bacteria and to determine whether an equilibrium model, the Freundlich adsor

23、ption isotherm, would adequately describe bacterial metal sorption. B. cereus, B. subtilis, and Pseudomonas aeruginosa were ex- amined as representatives of common species frequently isolated from soils. E. coli was also

24、 used as a second gram-negative bacterium because it is a well-characterized microorganism and its cell envelope has been shown to bind less metal than do B. subtilis cell walls (3). The four metallic ions used in this i

25、nvestigation were Ag+, Cd2“, Cu2“, and La3“. Cadmium and copper are both toxic cations of envi- ronmental importance. Silver and lanthanum, representative of monovalent and trivalent heavy metals, respectively, are also

26、toxic but are less frequently found in the environment.MATERIALS AND METHODSBacteria and growth conditions. The bacteria used in these experiments were B. cereus ATCC 11778; P. aeruginosa ATCC 14886, both obtained from t

27、he American Type Cul- ture Collection; B. subtilis 168; and E. coli K-12 strain AB264, both from the University of Guelph. The bacteria were routinely cultured in 0.5x brain heart infusion broth (BBL Microbiology Systems

28、) amended with 2.4 g of HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid) li- ter-' and 2.0 g of MES (2-N-morpholinoethanesulfonic acid) liter-' to buffer the medium acidity. The medium acidity was3

29、143BACTERIAL SORPTION OF HEAVY METALS 3145100.010.0W 1.030%.% 0.1 0 E 3 100.0c)10 C (j at high equilibrium concentrations, P. aeruginosa and B. cereus were most and least efficient, respectively, at removing Cd2“ and Cu2

30、“ from solution. Freundlich isotherms were not useful for describing the removal of Ag+ and La3“ from solution because there were too few datum points over a wide range of concentrations.100001-1 E0 -oE 0Qi)+1001011Equil

31、ibrium concentrations of La3“ were below detection limits when the initial concentration was 10 ,uM. Several of the observations for Ag+ equilibrium concentrations were also below detection limits at the 10 ,uM concentra

32、tion. When the equilibrium concentration was below detection limits, the total metal bound on a dry-weight basis was calculated with the assumption that essentially all of the Ag+ or La3“ in solution was bound. A 10 mM t

33、reatment was included for these metals to extend the concentration range examined. A good relationship was found for removal of Ag+ from solution as a function of the initial Ag+ concen- tration from 10 to 1,000 p.M (Fig

34、. 2). There were no signifi- cant differences in Ag+ removal among bacteria. Saturation of the cells with Ag+ apparently occurred in the 10 mM Ag+ treatment, as the total Ag+ bound increased only about 2-fold over a 10-f

35、old increase in Ag+ concentration. Significant differences among the bacteria for La3“ re- moval were found in the 1 mM treatment when 33, 70, 114, and 144 p.mol g-1 were removed by B. cereus, E. coli, B.subtilis, P. aer

36、uginosa, respectively (Fig. 3). There were no significant differences among bacteria for La3+ removal from the 10 or 100 ,uM solutions. The bacterial cells were evi- dently saturated with La at the 1 mM concentration, as

37、 verylittle additional La3+ was bound by these cells from the 10 mM La3+ treatment (Fig. 3). Silver was removed from solution much more efficiently than were the other metals at the 1 and 0.1 mM concentra- tions (Table 2

38、). An average of 99% of the total Ag+ was removed from solution in the 0.1 mM treatment. Cadmium was bound by the cells to a much lesser extent, with only 12 and 23% of the total Cd2' removed from the 1 and 0.1 mM tr

39、eatments, respectively. Even in the 0.001 mM treatment, an average of 46% of the added Cd2' remained in solution (data not shown). Electron micrographs of metal-treated cells showed that Ag was associated with the ce

40、ll primarily as discrete parti- cles at or near the cell walls of the bacteria, whether they were gram positive or gram negative (Fig. 4). Energy-10 100 1000 10000Initial Ag+ Concentration (/zmol/L)FIG. 2. Removal of sil

41、ver from solution as a function of the initial silver concentration. The line derived from least-squares regression ofall datum points over the concentration range of 10 to 1,000 ,umol liter-' is log y = -0.446 + 0.9