Environ. Sci. Technol. 1992, 26, 1606-1610
pool of water around the shower drain. Also, the physical characteristics and hydrodynamics of the shower system, including water-flow rate, can affect the volatilization process through their impacts on dropsize distribution and drop-residence time. It can be concluded from this work that the mass transfer of VOCs from the shower spray should not be modeled simply as occurring from a monodispersed, spherical drop-size distribution.
Acknowledgments We thank Drs. John E. Borrazzo (Carnegie Mellon University) and Albert Post (University of Pittsburgh) for their helpful comments and suggestions. We also thank Dr. Borrazzo for providing still photographs of the shower droplets. Registry No. Trichloroethylene, 79-01-6.
Literature Cited (1) Andelman, J. B.; Wilder, L. C.; Meyers, S. M. Indoor Air Pollution From Volatile Chemicals in Water. In Proceedings, 4th International Conference on Indoor Air Quality and Climate, Berlin, Aug 1987; Vol. 1, pp 37-41. (2) Angelo, J. B.; Lightfoot, E. N.; Howard, D. W. AIChE J. 1966, 12 (4), 751-760. (3) Handlos, A. E.; Baron, T. AIChE J . 1967,3 (l),127-136. (4) Ruckenstein, E. Znt. J. Heat Mass Transfer 1967, 10, 1785-1792.
(5) Altwicker, E. R.; Lindhjem, C. E. AZChE J. 1988, 34, 329-332. (6) Haney, P. D. J. Am. Water Works Assoc. 1954,46,353-378. (7) Hinds, W. C. Aerosol Technology; John Wiley and Sons: New York, 1981; p 111. (8) Adams, S. J.; Bradley, S. G.; Stow, C. D.; de Mora, S. J. Nature 1986, 321 (26), 842-844. (9) Mieure, J. P. J. Am. Water Works Assoc. 1977,69,60-64. (10) Glaze, W. H.; Rawley, R.; Burleson, J. L.; Mapel, D.; Scott, D. R. Further Optimization of the Pentane Liquid-Liquid Extraction Method for the Analysis of Trace Organic Compounds in Water. In Advances in the Identification a n d Analysis of Organic Pollutants in Water; Keith, L. H., Ed.; Ann Arbor Science Publishers: Ann Arbor, MI, 1981; Chapter 7, p p 267-280. (11) Giardino, N. J. Assessment and Modeling of Shower Emissions of Volatile Organic Chemicals. Doctoral Dissertation, University of Pittsburgh, 1990. (12) McKone, T. E.; Knezevich, J. P. J. Air Waste Manage. ASSOC.1991, 41, 832-837.
Received for review January 3,1992. Revised munuscript received April 22,1992. Accepted April 23,1992. This research has been funded in p a r t under cooperative agreement CR 812761-01 between the U.S.Environmental Protection Agency (EPA) a n d the University of Pittsburgh. This manuscript has not been subjected to EPA peer and administrative review policy and does not necessarily reflect its views, a n d no official endorsement should be inferred.
Anaerobic Oxidation of Elemental Metals Coupled to Methanogenesis by Methanobacterium thermoautotrophicum William H. Lorowltz, David P. Nagle, Jr., and Ralph S. Tanner" Department of Botany and Microbiology, University of Oklahoma, Norman, Oklahoma 730 19
Methanobacterium thermoautotrophicum Marburg, a thermophilic, obligately hydrogenotrophic, methanogenic bacterium, was used as a model organism to investigate anaerobic metal oxidation coupled to H2 utilization. Methane was produced with elemental aluminum, cobalt, copper, indium, iron, magnesium, manganese, nickel, tin, titanium, or zinc as the sole electron source. The extent of corrosion, and concomitant methanogenesis, was greatest in medium reduced with sulfide and buffered with carbonate under 100% COzheadspace. Comparisons between the concentrations of reducing equivalents generated in inoculated and uninoculated media revealed that M . thermoautotrophicum increased corrosion of copper, nickel, and zinc. Iron and magnesium (0.1 g/culture) were completely oxidized abiotically under 100% C02; M . thermoautotrophicum converted all reducing equivalents to methane. Under 20% COz, H2 utilization by M . thermoautotrophicum doubled the rate and increased the extent of iron oxidation. Incomplete oxidation of the other metals with M. thermoautotrophicum implied inhibition of methanogenesis by metal ions. Ni2+ (2.5 mM) completely inhibited methanogenesis, but Mg2+(200 mM) only slowed the rate. Cell numbers increased in cultures of M. thermoautotrophicum with aluminum, cobalt, iron, magnesium, manganese, or tin as the sole energy source, demonstrating that oxidation of these metals can support autotrophic growth of a methanogen. Introduction Corrosion can be defined as the degradation or wearing 1606
Environ. Sci. Technol., Vol. 26,
No. 8 , 1992
away of materials, usually metals. Metal corrosion in the United States results in several billion dollars of damage each year (1-3). In general, metal degradation is due to reaction with specific chemical and/or physical agents in the environment. Biocorrosion is a consequence of the activity of organisms indirectly through hostile metabolites or directly through electrochemical reactions. The most widespread problem of biocorrosion is the deterioration of iron and steel structures buried in soils and sediments. Several mechanisms may contribute to metal corrosion by anaerobic bacteria in these environments. Upon submersion in an aqueous environment, iron spontaneously undergoes an electrochemical corrosion (eq 1);iron is anodically dissolved by water acting as a neutral electrolyte (eq 2), resulting in the formation of a ferrous species and molecular hydrogen (eq 3). The reaction is self-limiting, and a protective layer of hydrogen forms on the metal surface. Corrosion is inhibited as long as the hydrogen layer remains intact.
-
Fez+ 2e-
+
(1)
2H+ + 20H-
(2)
FeO + 2Hz0
Fe(OH),
FeO 2H20
-
+ H2
(3)
According to the theory of cathodic depolarization (4, 5), Hz-oxidizing bacteria play the role of the depolarizer in the electrochemical iron corrosion process, removing the protective hydrogen layer and allowing oxidation to proceed. Anaerobic respiration with hydrogen as the electron
0013-936X/92/0926-1606$03.0010
0 1992 American Chemical Society
H2 -Utilizing Merhanogens
Table I. Effect of Reducing Agent on Methanogenesis with Nickel or Iron by Methanobacterium tbermoautotrophicuma
Co CH4 + 2H20 2 T
reducing agent, mM 2.6 cysteine 3.3 cysteine 1.7 sulfide 3.3 sulfide 5.0 sulfide
Iron Flgure 1. Cathodic depolarization of iron by an H2-utiiizing methanogenic bacterium, drawn after Daniels et al. (8).
donor may reduce nitrate (eq 41, sulfate (eq 51, or COz (eq 6). HNO, + 4H2 NH, + 3H20 (44 2HN03 + 5H2 N2 + 6Hz0 (4b)
-s2-+ - ++
+ 4Hz 4Hz0 COZ + 4Hz CH4 2H20 2C02 + 4H2 CH3COOH 2Hz0
(5) (64 ---* (6b) By convention, sulfate-reducing bacteria have been considered the primary microorganisms involved in anaerobic metal corrosion. In addition to cathodic depolarization by H2 utilization, the sulfides and phosphides produced by these bacteria can stimulate the cathodic reaction through formation of ferrous salts (6). Sulfide production coupled with the oxidation of steel wool by hydrogenase-positive Desulfovibrio strains was demonstrated by Cord-Ruwisch and Widdel(7), but occurred only in the presence of lactate. Desulfovibrio desulfuricans grew poorly with mild steel as the sole energy source (8). Cathodic depolarization coupled to nitrate reduction has been investigated (B), but corrosion episodes are largely anecdotal. Reduction of COz to acetate by acetogenic bacteria using hydrogen derived by cathodic depolarization (eq 6b) is thermodynamically feasible but has not been implicated in a corrosion episode or well-documented in the laboratory. The use of cathodically derived hydrogen from iron as the sole electron source for growth was demonstrated with methanogens (9) and is consistent with the mechanism proposed by von Wolzogen Kuhr and van der Vlugt (4) (Figure 1). Methanogenesis was demonstrated with iron and other metals (9, 10). In elegant experiments using a two-bottle system, Daniels and co-workers demonstrated the production of molecular hydrogen from metals in one bottle, with concomitant methane production by methanogenic bacteria in another bottle connected through the headspace. We have further investigated the mechanism of cathodic depolarization by a methanogen to elucidate the parameters influencing the rate and extent of corrosion of elemental metals. Methanobacterium thermoautotrophicum Marburg, a thermophilic, obligately autotrophic, methanogenic bacterium was used as the model organism. M. thermoautotrophicum is well-characterized and, as a methanogen, allows investigation of cathodic depolarization without the complication of sulfide production. s04’-
-+
Experimental Section Cultivation. M. thermoautotrophicum Marburg was maintained on a mineral medium with Hz + CO, as the energy and carbon source, respectively (11).Maintenance medium was buffered at pH 7.4 with 60 mM NaHCO, and reduced with 2.6 mM cysteine plus 1.7 mM sulfide. Except where noted, media for metal corrosion studies contained
+ 1.7 sulfide
CHI, rmol nickel iron 29.4 24.3 50.0 55.5 62.5
159 101 227 225 215
aDuplicate cultures contained 0.1 g of metal in 20 mL of medium,under 80% N2-20% C02. Incubation was at 60 “C for 200 h. Concentrations of CHI are per vessel.
0.1 g of metal, were buffered at pH 7.2 with 79 mM Na2C03 under 100% C02,and were reduced with 3.3 mM sulfide. Media were prepared anaerobically under C02, using a modified Hungate technique (12,13), and dispensed in 20-mL aliquota per 160-mL serum bottle (Bellco, Vineland, NJ). Cultures were inoculated with 1 mL M. thermoautotrophicum and incubated upright at 75 rpm on an orbital shaking incubator set at 60 “C. Metals. Powders of elemental metals were obtained from Bsar (Ward Hill, MA), Aldrich Chemical (Milwaukee, WI), or Fisher Scientific (Pittsburg, PA). Purity (%) and size (mesh) of the individual metals were as follows: antimony, 99.5,100, arsenic 99.99,100; beryllium, 99+, 200; bismuth, 99.5,100; cadmium, 99.5,325; chromium, 99,200; cobalt 99.5, 300; copper 99, 200; gallium, 99.999, 100; germanium, 99.999, 100; indium 99,325; iron, 99.9+, 325; lead, 99.5,100, magnesium, 99+, 50; manganese, 99+, 325; molybdenum, 99+, 100; nickel, 99.99, 100; niobium, 99.8, 325; palladium, 99.95, 200; rhenium, 99.997, 325; ruthenium, 99.95,325; selenium, 99.5, 100; tellurium 99.8, 200; vanadium, 99.5, 325; zinc, 99+, 325. Aluminum was described as “finest powder” (Fisher Scientific); silver and tungsten powders were 99.9%, 0.7-1.3 pm, and 99.9%, 12 pm, in purity and size, respectively. Electrician’s grade mercury was used. Ion Measurements. EM Quant test strips (EM Science, Gibbstown, NJ) were used for semiquantitative measurements of aluminum, cobalt, copper, iron, nickel, manganese, tin, and zinc cations. More precise quantitation of ferrous iron was done using a Spectroquant test kit (EM Science). Gas Measurements. Methane in the headspace was identified and measured by using a gas chromatograph equipped with a l/a-in. X 6-ft Porapak Q column (Alltech Associates, Deerfield, IL) and flame ionization detector. A reduction gas analyzer equipped with a mercury vapor detector (Trace Analytical, Menlo Park, CA) was used to measure hydrogen. Gas pressures were measured by using a hand-held pressure transducer system designed and constructed by P. F. Concannon (14). Cell Enumeration. Direct cell counts were made by using an improved Neubauer chamber (C. A. Hausser & Son, Philadelphia); 64 squares were counted for each sample.
Results and Discussion Medium Optimization. Our initial studies demonstrated methanogenesis by M. thermoautotrophicum coupled to oxidation of elemental iron, manganese, or nickel in the maintenance medium (with Nz replacing H,) (15). The apparent corrosion of nickel was surprising, as it is known to inhibit methane production in sewage digestors (16).Reduction in the effective concentration of Ni2+due to reaction with sulfide in the medium was Envlron. Sci. Technol., Voi. 26, No. 8, 1992
1607
Table 11. Range of Elemental Metals Examined as Source of Reducing Equivalents for Methane Production by Methanobacterium thermoautotrophicunP
metal
CH,,pmol
metal
CH,, pmol
aluminum antimony arsenic beryllium bismuth cadmium chromium cobalt copper gallium germanium indium iron lead magnesium manganese
2.41 -0.746 0.115 0.092 -0.235 -0.230 -0.091 10.8 4.02 0.185 0.055 1.38 415 0.301 954 65.2
mercury molybdenum nickel niobium pal1adium rhenium ruthenium selenium silver tellurium tin titanium tungsten vanadium zinc
0.597 -0.001 16.4 -0.132 -0.091 -0.047 -0.035 -0.082 -0.105 -0.23 2.67 5.69 0.667 -0.113 85.2
Table 111. Effect of Dissolved Cations on Methanogenesis from H2 C02"
+
cation, mM
CHI, pmol
cation, mM
CH,, pmol
none 0.5 Ni2+ 2.5 Ni2+
769 845 7
20 Mg2+ 100 Mg2*
1068 529
nMethanobacteriumthermoautotrophicum was grown in 20 mL of medium under 140 mL of COP(101 kPa at 60 "C) pressurized to 202 kPa with Hz. Cations were added as the chloride salt. Duplicate cultures were incubated at 60 OC, 80 rpm, for 5 days. Concentrations of CHI are per vessel.
'Cultures comprised 20 mL of medium plus 0.1 g of elemental metal. Concentrations of CHI are per vessel and represent the mean of at least duplicate cultures, corrected for methane produced by controls unamended with elemental metals. Controls contained an average of 0.756 pmol of CH,. 400
,
I 1600
0
50
100 150 200 250 300 350 400 4 5 0 5 0 0 550
Time ( h r )
+
Flgure 3. Effect of Mg2+on methanogenesis from H, C02 by M . thermoautotrophicum Concentrations of CH4 are per vessel and represent the mean of triplicate 20-mL cultures, grown in 160-mL serum bottles under 80% H,-20% COP(101 kPa at 60 OC), with and without 200 mM Mg2+. Media were buffered at pH 7.4 with 60 mM NaHCO, and repressurized with H2-C02, as required.
.
Flgure 2. Time course of iron oxidatlon with M . thermoautotr~hicum . Each value represents the mean of triplicate cultures grown with 0.1 g of iron as sole electron source. Concentratlons of CH, and Fe2+are per vessel.
examined. Methane produced by cultures with 1.7, 3.3, or 5.0 mM sulfide as sole reducing agent was twice as great as with cultures reduced with 2.6 mM cysteine 1.7 mM sulfide or 3.3 mM cysteine alone (Table I). Methanogenesis with iron was similarly affected, although to a lesser extent. Reduction of the media with cysteine alone resulted in the lowest methane yields with both metals. Furthermore, 2.6 mM cysteine + 1.7 mM sulfide gave lower yields than 1.7 mM sulfide alone. Thus, cysteine appeared to inhibit methanogenesis coupled to oxidation of nickel or iron. The sulfhydryl group of cysteine may allow it to act as an adsorption-type inhibitor, attaching itself to the metal surface and effectively blocking dissolution of metal ions (2). On the basis of these results, 3.3 mM sulfide was used as reductant for the following experiments. A separate investigation revealed that methanogenesis from iron increased more than 3-fold after replacing the 80% Nz-20% C02 headspace with 100% COz and altering the buffer from 60mMNaHC03(pH 7.4) to79 mMNa2C0,(pH 7.2). Range of Metals. Methanogenesis by M.thernoautotrophicurn was tested with a range of metals (Table 11). Methane concentrations showed a concomitant increase with ferrous iron concentrations during a time course analysis (Figure 2). Typically, all of the elemental iron
+
1808 Envlron. Sa. Technol., Voi. 26, No. 8, 1992
(0.1 g) was transmuted to ferrous ions and methane within 9-10 days of incubation. Magnesium metal was also an excellent source of electrons; all reducing equivalents were recovered as methane after 10 days (Table 11). In addition, methanogenesis occurred with aluminum, cobalt, copper, indium, manganese, nickel, tin, titanium, and zinc. Methane production from copper, nickel, and zinc was especially noteworthy in that these metals are considered highly toxic to the methanogenesis (16). Final cation concentrations of these three metals, as well as those of aluminum, cobalt, and manganese, agreed with the methane yields that were predicted by the cathodic depolarization model (Figure 1). Methane production was not observed with antimony, arsenic, beryllium, bismuth, cadmium, chromium, gallium, germanium, lead, mercury, molybdenum, niobium, palladium, rhenium, ruthenium, selenium, silver, tellurium, tungsten, or vanadium. Inhibition of Methanogenesis by Nickel and Magnesium Cations. The effect of divalent cations on methanogenesis from the physiological substrates H2 + C02 was tested with nickel and magnesium (Table 111). Methanogenesis was entirely inhibited by 2.5 mM Ni2+, a concentration stoichiometric with the amount of methane produced in incubation with the elemental metal (Table 11). With magnesium, methanogenesis appeared to be stimulated with 20 mM Mg2+;the yield was about 37% greater than in the unamended controls (Table 111). However, with 100 mM Mg2+,cultures produced only 69% of the methane in the controls after 5 days of incubation. A time course analysis of methane production from Hz + COz revealed that 200 mM Mg2+ (the concentration equivalent to complete oxidation of 0.1 g MgO) slowed the rate almost 50-fold but did not prevent methanogenesis (Figure 3).
Table IV. Production of Reducing Equivalents from Elemental Metals and Use by Methanobacterium thermoautotrophicum"
metal none aluminum cobalt copper indium iron magnesium manganese nickel tin titanium zinc
reducing equivalents recovered, @mol inoculated uninoculated in Hz in Hz, in CHI, Oh 200 h 200h 200h 1.60 10.8 3.11 1.58 2.32 145 7410 450 14.4 6.82 2.21
775
1.63 11.5 85.7 5.16 20.6 2660 7450 484 73.1 6.24 2.20 1080
3.33 11.8 5.74 3.70 7.87 4.93 3.00 1.25 4.14 4.00 3.03 1050
cells/mL f SE (XlO4)
4.52 7.52 86.2 24.6 27.8 2980 7630 521 131 7.70 7.00 682
9.81 f 0.51 14.3 f 0.64 12.4 f 0.50 8.06 f 0.49 10.3 f 0.56 15.4 f 0.70 51.2 A 1.49 12.6 f 0.76 9.13 f 0.53 13.4 i 0.61 9.81 f 0.51 11.0 k 0.59
Hydrogen and methane were measured initially and after 200-h incubation. Uninoculated media produced hydrogen (2 reducing equiv/mol); M. therrnoautotrophicum cultures produced methane (8 reducing equiv/mol) and trace hydrogen. Cells were enumerated by direct count at 200 h. All treatments were performed in duplicate. Concentrations of reducing eauivalents are Der vessel.
Abiotic H2 Production. Copper is frequently included in alloys as a corrosion inhibitor and generally used in cathodic depolarization studies as a negative control because of its chemical properties (IO). The standard reduction potential for the two-electron oxidation of elemental copper is negative (-0.342 V), indicating that under standard conditions the reaction will not occur. After several observations of methanogenesis, abiotic hydrogen production from copper under cultivation conditions was investigated (Table IV). Hydrogen concentrations were also measured with the other metals supporting methanogenesis. Initially, there was no difference between the concentrations of hydrogen in the copper medium and the control (no added metal) medium. After 10 days of incubation at 60 "C, the control still contained about 800 nmol of hydrogen, but the concentration in the headspace of the copper medium had increased to 2.58 pmol of hydrogen. All the other metals tested had higher initial concentrations of hydrogen. After incubation, increases in hydrogen concentration were observed with cobalt, indium, iron, nickel, and zinc in uninoculated medium. No increases were observed with aluminum, magnesium, manganese, tin, or titanium. With magnesium, the initial hydrogen concentration was consistent with complete oxidation of all metal present. At the end of the incubation period, the hydrogen concentration with iron indicated complete dissolution of the metal in uninoculated medium. The abiotic production of hydrogen (Table IV)was likely driven by the production of sulfide and carbonate salts. Free energies of formation (AG,")(Table V) indicate that formation of the salts is more thermodynamicallyfavorable than simple dissolution of the metal to the divalent cations. In the case of iron, media conditions favored complete dissolution of metallic iron to hydrogen (Table IV) and a precipitate solubilized by heating and acidification, tentatively identified as ferrous sulfide and ferrous carbonate. The thermodynamically driven formation of ferrous carbonate would help explain the increase in methanogenesis observed after increasing C02 in the headspace from 20% to 100%. Final hydrogen concentrations in the inoculated medium tended to be low, with most less than 3 pmol (Table IV). A notable exception to this was zinc, which contained 525 pmol of hydrogen, virtually the same concentration as that in the uninoculated medium. However, 85.2 pmol of methane was produced, equivalent to an additional 340 pmol of hydrogen. These data implied cessation of
Table V. Free Energy of Formation for Metal Species ( 17, 18 )
AG?, kJ/reaction metal
cation
carbonate
sulfide
copper iron magnesium manganese nickel tin zinc
15.66 -18.85 -108.7 -54.5 -10.9 -6.5 -35.14
nfn -159.35 -525.7 -195.2 -146.4 nfn -232.0
-12.8 24 -81.7 -52.2 -19 -23.5 -48.11
" Indicates value was not found. methanogenesis after an accumulation of cations, perhaps in a manner similar to inhibition with nickel, while chemical constraints limited abiotic hydrogen production to approximately 500 pmol. The addition of M. thermoautotrophicum increased the extent of oxidation with copper, nickel, tin, and zinc. In each instance, the methane concentrations were greater than predicted from the concentration of abiotically produced hydrogen. Toxic cations were likely sequestered as insoluble salts, permitting limited cathodic depolarization to occur. These results suggest that biocorrosion could be increased by secondary activities, such as metal interactions with metabolic products or biofilms, which could remove toxic cations from cellular contact. Although only low levels of corrosion occurred in batch culture, natural situations more closely mimic continuous culture; continual washout of free cations would likely result in increased oxidation of the metals. Methane concentrations from the other metals were close to those predicted from abiotic hydrogen yields. Under cultivation conditions, iron and magnesium were entirely oxidized abiotically (Table IV). A time course experiment comparing abiotic and biotic oxidation was performed with 0.3 g of iron in 20 mL of medium (pH 7.4) under 80% N2-20% COS (Figure 4). After 200-h incubation at 60 "C, the M. thermoautotrophicum cultures had produced approximately twice the concentration of reducing equivalents from iron at about twice the rate as that which occurred in the uninoculated medium. Thus, M. thermoautotrophicum enhanced the extent, as well as the rate, of iron oxidation. Significant 0) >0.95) increases in cell numbers, compared to the control (Table IV), were observed with aluminum, cobalt, iron, magnesium, manganese, and tin. This Environ. Sci. Technol., Vol. 26, No. 8, 1992
l60Q
remaining in the headspace of zinc cultures (Table IV) suggested a mechanism similar to inhibition with nickel. The different modes of inhibition could be important for controlling bacterial corrosion. H2 utilization by M. thermoautotrophicum doubled the rate and extent of iron oxidation and increased corrosion of copper, nickel, and zinc. The increase in cell numbers observed with aluminum, cobalt, magnesium, manganese, and tin (Table IV) indicated novel sources of energy for autotrophic growth of bacteria.
OV' 0
'
"
100
"
"
"
200
"
300
"
'
I 400
Time ( h r ) Figure 4. Effect of M . thermoautotrophicum on anaerobic iron oxidation. Total reducing equivalents (2/H2 and 8/CH4) were compared between uninoculated medium and inoculated cultures incubatedat 60 "C. Medium containlng 0.3 g of iron was buffered at pH 7.4 with 60 mM NaHCO,, under 80% N,-20% COP (101 kPa at 60 "C). Concentratbns of CH, are per vessel and represent the mean of triplicate cultures.
is the first report of autotrophic growth linked to aluminum, cobalt, magnesium, manganese, or tin as the sole energy source. Although this suggests an initial benefit to H2-utilizingbacteria, it must be considered against the concomitant increase in toxic cations resulting from oxidation of these metals. Manipulation of in situ oxidation of metals may provide a mechanism to limit overall corrosion within selected systems. Conclusions It is evident from this and other work (9,lO) that elemental metals can provide reducing equivalents for reduction of C02to CH4 by methanogens. Less clear are the parameters that influence the extent and rate of cathodic depolarization. A range of metals was examined and methane was produced by M. thermoautotrophicum with aluminum, cobalt, copper, indium, iron, magnesium, manganese, nickel, tin, titanium, or zinc (Table 11). Reducing media with sulfide and wing a carbonate/C02 buffer increased methane production from nickel and iron, respectively. The explanation for these observations may lie in the formation of insoluble sulfide and carbonate salts. With copper, nickel, and zinc, the formation of insoluble salts probably lowered the toxicity of the metal cations. The inhibitory effects of Ni2+and Mg2+were different (Table 111); Ni2+entirely inhibited methanogenesis from hydrogen, while Mg2+only appeared to slow the reaction (Figure 4). The relatively high concentration of hydrogen
1610 Envlron. Scl. Technol., Vol. 26, No. 8, 1992
Literature Cited (1) Fontana, M. G. Corrosion Engineering, 2nd ed.; McGrawHill Book Co.: New York, 1986; pp 1-9. (2) Iverson, W. P. In Microbial Iron Metabolism; Neilands, J. B., Ed.; Academic Press: New York, 1974; pp 477-479. (3) Tiller, A. K. In Corrosion Process; Parkins, R. N., Ed.; Applied Science Publishers: New York, 1982; pp 115-117. (4) von Wolzogen Kuhr, C. A. H.; van der Vlugt, L. S. Water 1934, 18, 147. (5) von Wolzogen Kiihr, C. A. H. Corrosion 1961, 17, 119. (6) Iverson, W. P. Nature 1968,217, 1265. (7) Cord-Ruwisch, R.; Widdel, F. Appl. Microbiol. Biotechnol. 1986, 25, 169. (8) Rajagopal, B. S.; LeGall, J. Appl. Microbiol. Biotechnol. 1989, 31, 406. (9) Daniels, L.; Belay, N.; Rajagopal, B. S.; Weimer, P. J. Science 1987, 237, 509. (10) Belay, N.; Daniela, L. Antonie van Leeuwenhoek 1990,57, 1. (11) Tanner, R. S.; McInerney, M. J.; Nagle, D. P., Jr. J. Bacteriol. 1989, 171, 6534. (12) Bryant, M. P. Am. J. Clin. Nutr. 1972, 25, 1324. (13) Balch, W. E.; Wolfe, R. S. Appl. Enuiron. Microbiol. 1976, 32, 781. (14) DeWeerd, K. A.; Concannon, F.; Suflita, J. M. Appl. Enuiron. Microbiol. 1991, 57, 1929. (15) Lorowitz, W. H.; Tanner, R. S. Abstracts of the General Meeting, 91st Annual Meeting, American Society for Microbiology, Dallas, TX, May 1991; American Society for Microbiology: Washington, DC, 1-121, p 210. (16) McCarty, P. L. Public Works 1964, 75, 91. (17) Handbook of Chemistry and Physics, 66th ed.; W e a t , R. C., Astle, M. J., Beyer, W. H., Eds.; CRC Press: Boca Raton, FL, 1985; p p D50-D93. (18) Lunge's Handbook of Chemistry, 13th ed.; Dean, J. A., Ed.; McGraw-Hill Book Co. New York, 1985; Section 9, pp 4-69.
Received for review February 24,1992. Accepted April 21,1992. This work was supported by grants from the Oklahoma Mining and Minerals Resource Research Institute and the Officeof Naval Research.
