Materials Transactions, Vol. 45, No. 7 (2004) pp. 2422 to 2428
#2004 The Mining and Materials Processing Institute of Japan
Fundamental Study on the Removal of Mn2þ in Acid Mine Drainage
using Sulfate Reducing Bacteria*
Kyoungkeun Yoo1 , Keiko Sasaki2 , Naoki Hiroyoshi1 and Masami Tsunekawa1
1
2
Graduate School of Engineering, Hokkaido University, Sapporo 060-8628, Japan
Faculty of Engineering, Kyushu University, Fukuoka 812–8581, Japan
The optimum conditions for Mn2þ removal from acid mine drainage was studied by a SRB (sulfate reducing bacteria) bioreactor. Chemical
experiments with Na2 S as a S2 source were conducted to investigate the effects of pH, coexisting metal ions, and the components in a growth
medium for SRB on MnS formation from Mn2þ solutions. The amount of Mn removed from the Mn2þ solutions decreased with decreasing pH.
The Zn2þ or Fe2þ coexisting in the solutions consumed S2 by forming ZnS or FeS, and this inhibited Mn removal. Sodium citrate, a component
of the growth medium for SRB, formed a complex with Mn2þ and suppressed MnS formation. Biological experiments using the SRB reactor
were carried out at 37 C and it was confirmed that the Mn2þ concentration decreased to less than 10 gm3 from 100 gm3 at neutral pHs (pH 5–
7) after 100 hours when other metal ions and sodium citrate were absent. The formed precipitate was identified to be metastable -MnS with a
band gap of about 3.8 eV by XRD, XRF, and UV-VIS.
(Received July 28, 2003; Accepted May 7, 2004)
Keywords: Mn2þ removal, sulfate reducing bacteria, coexisting metal ions, citrate complex
1.
Introduction
Abandoned Mn mines in Hokkaido, Japan, leak acid mine
drainage (AMD) containing 100 to 200 gm3 of Mn2þ .1) The
removal of Mn2þ has been achieved by a conventional
method precipitating Mn2þ at around pH 10 with alkaline
reagents. This method requires subsequent neutralization of
the supernatant, which consumes a substantial amount of
reagents and so is costly. Therefore, much effort has been
made to develop alternative methods.
The removal and recovery of heavy metal ions using a
sulfate reducing bacteria (SRB) reactor have attracted
interest in the treatment of AMD.2–5) The SRB are obligate
anaerobes, and the bacteria oxidize organic matters using
sulfate instead of oxygen. With the SRB, hydrogen sulfide is
generated and reacts with heavy metal ions in AMD to
precipitate metal sulfides.
Hammack and Dijkman5) investigated the SRB treatment
of AMD mainly containing soluble Cu, Zn, and Fe, and
reported that Mn, a minor component in the AMD, was
removed together with Fe during the treatment. This result
suggests that Mn2þ in AMD can be removed by SRB,
although the detailed mechanism of the Mn removal was not
discussed in the paper. However, other researches have
reported that Mn2þ in AMD was not removed in SRB
reactors.2,6,7)
The apparent inconsistency may be due to differences in
experimental conditions such as pH and coexisting compounds, which would affect the efficiency of metal removal
by SRB.8) To establish the conditions for Mn2þ removal
using an SRB bioreactor, the present study investigated the
effects of pH, coexisting metal ions, and components of the
medium for SRB on Mn2þ removal.
*This Paper was Presented at the Annual Meeting of MMIJ, held in Tokyo,
on March 28–30, 2002 and on March 27–29, 2003.
2.
Experimental
2.1 Chemical experiments for Mn removal using Na2 S
Metal ion solutions containing set concentrations of Mn2þ ,
Zn2þ , and Fe2þ were prepared with reagent grade
MnSO4 5H2 O, ZnSO4 7H2 O, FeSO4 7H2 O, and deionizeddistilled water, and Na2 S solutions were prepared with
Na2 S9H2 O in deionized-distilled water. The initial pH of the
solutions was adjusted to 4–5 for the metal ion solutions and
10 for the Na2 S solutions by adding 1 moldm3 NaOH or
H2 SO4 .
The 100 cm3 of Na2 S solution and 100 cm3 of the metal
solution were mixed in 500 cm3 -Erlenmayer flasks, which
were immediately sealed with a silicon rubber cap. In the
following, the concentration of the metal ions (or Na2 S)
added to the mixture is termed as the ‘‘initial concentration of
metal ions (or Na2 S)’’. The solution in the flasks was stirred
with a magnetic bar at room temperature for 1 hour, and the
supernatant was collected by filtration using a membrane
filter (pore size, 0.2 mm). Metal concentrations in the supernatant were analyzed with a SEIKO SPS 7800 inductively
coupled plasma atomic emission spectroscope (ICP-AES).
Heavy metal solutions containing 17 molm3 of sodium
citrate dihydrate (Na3 C6 H5 O7 2H2 O), a component of the
medium for SRB as described in detail below, were
investigated to determine the effects of complex formation
with sodium citrate and metal ions on the removal of metal
ions by Na2 S.
2.2
Biological experiments for Mn removal using sulfate
reducing bacteria
2.2.1 Bacterial strains and culture
The obligate anaerobe, Desulfovibrio desulfuricans
(ATCC 7757) was used, and grown in the modified ATCC
1249 medium (2 g of MgSO4 7H2 O, 5 g of Na3 C6 H5 O7 2H2 O (sodium citrate dihydrate), 1 g of CaSO4 2H2 O, 1 g of
NH4 Cl, 1 g of yeast extract, 0.01 g of K2 HPO4 , and 3.5 g of
NaC3 H5 O3 (sodium lactate) in 1 dm3 of distilled water).
Fundamental Study on the Removal of Mn2þ in Acid Mine Drainage using Sulfate Reducing Bacteria
2500PC UV-VIS recording spectrophotometer (Japan) at
room temperature.
Gas
Outlet
Sampling
port
Filter
3.
pH
Electrode
ORP
Electrode
Results and Discussion
The effects of Na2 S, pH and coexisting metal ions on
MnS formation.
The AMD contains heavy metal ions and sulfate in high
concentrations, and using organic matter such as lactate, SRB
reduce sulfate to sulfide, which reacts with heavy metal ions
to form metal sulfides. The process can be expressed as
follows;9)
3.1
Nitrogen
(Oxygen free)
pH and ORP
Recorder
Impeller
Temperature and
Agitation controller
2CH3 CHOHCOOH þ SO2
4
! 2CH3 COOH þ 2HCO
3 þ H2 S
2þ
Me þ H2 S ! MeS # þ2Hþ
Configuration of the SRB bioreactor used in this study.
2.2.2 Apparatus and procedure
Biological experiments for the Mn removal were carried
out using a cylindrical glass bioreactor (height, 160 mm;
internal diameter, 100 mm) as shown in Fig. 1. The ORP
(redox potential) and pH electrodes, N2 sparger and a gas
outlet tube to maintain anaerobic condition, a solution
sampling tube, thermo-controller, and magnetic stirring
system were installed in the bioreactor. All instruments were
autoclaved at 121 C for 20 min before inoculation of SRB.
Two mediums were used in the experiments: one was the
modified ATCC 1249 medium, and the other was the
modified ATCC 1249 medium without sodium citrate. In
both mediums, 0.44 gdm3 of MnSO4 5H2 O was added as a
Mn2þ source (Mn2þ concentration, 100 gm3 ), and 2 105
cellsdm3 of SRB was inoculated. The initial pH of the
medium was adjusted to 6.7 with 1 moldm3 NaOH and
H2 SO4 .
The Mn removal experiments were carried out with the
bioreactor containing 1 dm3 of the modified ATCC 1249
medium with or without sodium citrate under the following
operating conditions: N2 gas flow rate, 0.2 dm3 min1 :
stirring, 200 rpm; and temperature, 37 C. The N2 gas was
introduced only during sampling. The solution was sampled
through the sampling tube and was filtered using a membrane
filter (pore size, 0.2 mm), then the Mn and sulfate concentration were analyzed by ICP-AES and a SHIMADZU ion
chromatograph (IC, Japan) using IC shim-pack A3, respectively.
After the operation of the bioreactor was terminated, the
precipitate in the reactor was collected by filtration using a
membrane filter (pore size, 0.2 mm), and dried overnight at
55 C. The dried precipitate was ground in a mortar, and then
characterized using a JEOL JSX-3210A energy diffusive type
X-ray fluorescence analyzer (Japan) and a JEOL JDX-3500
X-ray diffractometer (Japan), respectively. The X-ray diffraction (XRD) data were collected with a monochromator
under the following conditions: radiation, Cu K, 30 kV,
200 mA; step scanning method; time constant, 0.5 seconds;
angle range, 2:565 degree/2, and The X-ray fluorescence
(XRF) was measured under the following conditions; Rh,
30 kV; current, auto; time, 300 seconds. To examine the
energy band gap of the ground precipitate, the optical
absorption spectra were measured with a SHIMADZU UV-
ð1Þ
ð2Þ
where Me2þ and MeS represent the heavy metal ions and
metal sulfides, respectively.
The effects of H2 S concentration, pH, and coexisting
compounds on the reaction in eq. (2) were investigated by
chemical experiments using Na2 S as the H2 S source. Figure 2
shows the effect of the concentration of the Na2 S added on
the Mn removal at the final pH 6.3–6.4. The initial
concentration of Mn2þ was 4.4 molm3 . With increasing
initial Na2 S concentrations, the amount of Mn removed from
the solution increased.
The effect of pH on MnS formation was investigated in the
mixture containing 2.2 molm3 Mn2þ (120 gm3 ) and 7.8
molm3 Na2 S. As can be seen in Fig. 3, 37.5% (80 gm3 ) of
Mn remained at the final pH 5.2, whereas nearly all of the
Mn2þ was removed at the final pH 6.2–6.4.
Figure 4 shows the effects of coexisting metal ions on the
Mn2þ removal. The Mn removal ratio decreases with
increasing concentrations of added Fe2þ or Zn2þ . After the
experiments, no soluble Zn and Fe was detected in the
solutions, indicating that the added Zn2þ and Fe2þ were
precipitated during the experiments.
100
Removal ratio of metal ions / %
Fig. 1
2423
80
60
40
20
0
0
10
20
30
Initial H 2S concentration, [I.S.] / mol . m-3
Fig. 2 Mn2þ removal ratio (open symbol) as a function of Na2 S
concentration at final pH 6.3–6.4 at 20 C with an initial 4.4 molm3
Mn2þ concentration. The volumes of gas and liquid phase are 0.3 dm3 and
0.2 dm3 . Solid and dotted lines indicate the calculated removal ratios of
Mn2þ and Zn2 .
2424
K. Yoo, K. Sasaki, N. Hiroyoshi and M. Tsunekawa
amount of metal sulfide as MeS, MH2 SðgÞ the mole amount of
H2 S in gas phase, Vaq the volume of liquid phase,
respectively.
The mole amount of H2 S in gas phase is expressed by
100
Mn removal ratio / %
80
MH2 SðgÞ ¼
60
20
0
5.6
6
6.4
½H2 SðaqÞ Vaq þ MMeS
Final pH
Fig. 3 The effect of pH on MnS formation in chemical experiments using
Na2 S as the S2 source. Initial conditions: Mn2þ 2.2 molm3 , Na2 S
7.8 molm3 .
ð6Þ
where Ka1 ð¼ 107:0 Þ and Ka2 ð¼ 1013:9 Þ are equilibrium
constants for the reactions in eqs. (7) and (8).
H2 S ¼ HS þ Hþ ;
Ka1 ¼
100
HS ¼ S2 þ Hþ ;
Mn removal ratio / %
ð5Þ
where PH2 S represents a partial pressure of H2 S in gas phase,
Vg volume of gas phase, R gas constant (8.314 JK1 mol1 ),
T temperature (K), HH2 S Henry constant of H2 S
(8:67 102 Pam3 mol1 ).10) Using eq. (5), Eq. (4) is
rearranged as follows.
Ka1
Ka1 Ka2 HH2 S Vg
MT.S. ¼ 1 þ þ þ
þ
½H [Hþ ]2
RT Vaq
40
5.2
PH2 S Vg HH2 S Vg
¼
½H2 SðaqÞ RT
RT
Ka2 ¼
½HS ½Hþ ½H2 SðaqÞ ½S2 ½Hþ ½HS ð7Þ
ð8Þ
80
The total mole amount of metal ions added (MT.Me. ) to
solution is given by
60
MT.Me. ¼ MMe2þ þ MMeS
40
20
With Zn ions
With Fe ions
0
0
2
4
6
8
10
Co-existing ion concentration / mol . m -3
Fig. 4 The effect of Fe and Zn ion concentration on Mn removal. Initial
conditions: Mn 5.4 molm3 , Na2 S 15.6 molm3 , and final pH 6.2–6.3.
The effects of the Na2 S concentration, pH, and coexisting
metal ions on the Mn removal can be explained thermodynamically as follows. The equilibrium constant of eq. (2) can
be expressed as eq. (3).
[Hþ ]2
K¼
[Me2þ ][H2 S]
ð3Þ
where [Hþ ], [Me2þ ], and [H2 S] represent the concentrations
of hydrogen ion, metal ion, and hydrogen sulfide, respectively.
The material balance for H2 S species is given by
MT.S. ¼ MH2 SðaqÞ þ MHS þ MS2 þ MMeS þ MH2 SðgÞ
¼ ð[H2 S(aq) ] þ [HS ] þ ½S2 ÞVaq þ MMeS þ MH2 SðgÞ
ð9Þ
where MMe2þ represents the mole amount of Me2þ in a
system.
The combination of eqs. (3), (6) and (9) gives
HH2 S Vg
1þ
[Hþ ]2 þ Ka1 ½Hþ þ Ka1 Ka2
RT Vaq
K¼
MT.S. MMeS
½Me2þ Vaq
HH2 S Vg
2
1þ
[Hþ ]2 þ Ka1 ½Hþ þ Ka1 Ka2 Vaq
RT Vaq
¼
MMe2þ ðMT.S. MT.Me þ MMe2þ Þ
HH2 S Vg
þ 2
þ
2
1þ
[H ] þ Ka1 ½H þ Ka1 Ka2 Vaq
RT Vaq
¼
MMe2þ ðVaq ½I.S. Vaq ½T.Me. þ MMe2þ Þ
ð10Þ
The equilibrium constants (K) for Cu, Zn, Fe, and Mn
are calculated on the basis of the thermodynamic data for
the related species shown in Table 1, and the values are
106:96 , 102:01 , 100:48 and 106:58 for Cu, Zn, Fe and Mn,
respectively.
Using eq. (10), the ratio of removed metal ions
(ðMT.Me MMe2þ Þ=MT.Me ) was calculated as a function of
the initial H2 S concentrations ([I.S.] (¼ MT.S. =Vaq )). In the
ð4Þ
where MT.S. represents the total mole amount of H2 S in a
system, MH2 SðaqÞ and [H2 SðaqÞ ] the mole amount and the
concentration of H2 S in liquid phase, MHS and [HS ] the
mole amount and the concentration of HS , MS2 and [S2 ]
the mole amount and the concentration of S2 , MMeS the mole
Table 1 Standard Gibbs free energy changes (G0 ) of formation for the
species involved in eq. (2) at 298 K at 1 atm (unit: kJ/mol).11Þ
H2 S
Mn
2þ
MnS
27:83
Fe2þ
78:90
ZnS
228:10
FeS
100:40
Cu2þ
147:06
CuS
218:40
2þ
Zn
201:29
65.49
53:60
Fundamental Study on the Removal of Mn2þ in Acid Mine Drainage using Sulfate Reducing Bacteria
100
Mn removal ratio / %
0
Mn
-4
Fe
-8
Zn
2+
Log(M Me / mmol)
2425
80
60
40
20
0
SC
-12
SL
MS
CS
AC
YE
Components of culture for SRB
Cu
Fig. 6 Mn2þ removal as MnS by adding components of the modified
ATCC 1249 medium in solutions including 2.2 molm3 Mn2þ and
22.4 molm3 Na2 S at pH 6.8 and room temperature. SC, Sodium Citrate;
SL Sodium Lactate; MS, Magnesium Sulfate; CS, Calcium Sulfate; AC,
Ammonium Chloride; YE, Yeast extracts.
-16
-20
0
2
4
6
8
pH
100
calculation, [T.Me.] (¼ MT.Me. =Vaq ), pH, temperature, and
volumes of gas and liquid (Vg and Vaq ) were assumed to be
4.4 molm3 , 6.3, 293 K, 0.3 dm3 and 0.2 dm3 , respectively.
The calculated results are shown in Fig. 2 as a solid line for
Mn2þ and as a dotted line for Zn2þ . The calculated result for
Mn shows a good agreement with the experimental data. The
experimental and calculated results indicate that more than
12 molm3 of H2 S is required to remove 95% of
4.4 molm3 Mn2þ due to a higher solubility of MnS, i.e.
low value of K, although the equilibrium amount of H2 S
(4.4 molm3 ) can remove 4.4 molm3 Zn2þ .
Figure 5 shows the equilibrium amount of various metal
ions in the presence of metal sulfide as a function of pH. In
the calculation, MT.S. , MT.Me. , temprature, Vg and Vaq are
assumed to be 250 mg (7.8 mmol), 2.2 mmol, 293 K, 1 dm3
and 1 dm3 . The calculated results indicate that a higher pH is
required for the Mn removal than for the removal of the other
metal ions, and that Cu2þ , Zn2þ , or Fe2þ may be removed in
preference to Mn2þ at any given pH. Therefore, coexisting
metal ions such as Zn2þ an Fe2þ consume the H2 S in the
solution, which would result in a suppression of Mn removal
by the coexisting metal ions.
3.2
The effects of components in the medium for SRB on
MnS formation
In the SRB treatment of AMD, nutrients such as the
components in the medium for SRB are required to maintain
high SRB activities in the reactor. If a nutrient suppresses the
MnS formation in eq. (2), the efficiency of Mn removal
would be decreased even when SRB were thriving. To
determine such nutrient element suppression of Mn removal,
the effects of the nutrients on MnS formation were
investigated by chemical experiments using Na2 S as the
S2 source.
Figure 6 shows the effects of various components in the
modified ATCC 1249 medium on Mn removal. Each
component of the same concentration as designated in the
modified ATCC 1249 was added to the mixture of Mn2þ and
80
Mn removal ratio / %
Fig. 5 Equilibrium concentrations of metal ions at an initial 2.2 mmol as a
function of pH in the presence of 7.8 mmol H2 S at 20 C. The volumes of
gas and liquid phase are 1 dm3 and 1 dm3 . MMe2þ : the mole amount of
metal ion.
60
40
Without citrate
With citrate
20
0
0
4
8
12
16
20
Na2S concentration / mol . m-3
Fig. 7 The effect of citrate addition on MnS formation using Na2 S as the
S2 sources in 2.2 molm3 Mn2þ solution at pH 7. Citrate concentration
added is 17 molm3 .
Na2 S, and, as shown in Fig. 6, sodium citrate suppressed Mn
removal, whereas the other components did not affect Mn
removal.
Figure 7 shows the Mn removal ratio in the modified
ATCC 1249 medium with or without citrate (17 molm3 ) as
a function of Na2 S concentration. Without citrate, Mn was
completely removed with more than 9 molm3 of Na2 S,
whereas, with citrate, Mn was not removed even with
17 molm3 of Na2 S.
To compare Mn2þ removal with that of other metal ions,
the effect of sodium citrate on Zn removal was investigated,
and the results are shown in Fig. 8. Citrate (17 molm3 ) also
suppresses Zn removal, but its suppressive effect was much
weaker than that in the Mn removal shown in Fig. 7, and Zn
was removed in both case independent of citrate addition.
Citrate acts as a ligand to various metal ions. The complex
formation with divalent metal ions (Me2þ ) and citrate (L) is
represented in eq. (11) and the equilibrium constant (KMeðm;nÞ )
for the reaction is given in eq. (12).
mMe2þ þ nL ¼ (Mem Ln )2mþ
ð11Þ
2mþ
KMeðm;nÞ ¼
½(Mem Ln ) [Me2þ ]m [L]n
ð12Þ
2426
K. Yoo, K. Sasaki, N. Hiroyoshi and M. Tsunekawa
0.017 mol. dm-3 Citrate
100
0
Log([S.Me.] / mol . dm-3)
Zn removal ratio / %
80
60
40
Without citrate
With citrate
20
-4
Mn
-8
Fe
-12
Zn
-16
Cu
0
0
1
2
3
-20
4
Na2S concentration / mol. m-3
-8
-6
Fig. 8 The effect of citrate addition on ZnS formation using Na2 S as the
S2 sources in 2.2 molm3 Zn2þ solution at pH 3.3. Citrate concentration
added is 17 molm3 .
Mn
Fe
m
1
1
1
1
2
n
1
1
1
2
2
3.7
4.4
4.9
5.9
log K Cu
14.8
The equilibrium constant between heavy metal ion and citrate.
MT.Me. ¼ MMe2þ þ Vaq
X
mKMeðm;nÞ [Me2þ ]m [L]n
þ MMeS
ð14Þ
MS.Me. ¼ MMe2þ þ Vaq
X
mKMeðm;nÞ [Me2þ ]m [L]n
or
m;n
where m and n are the numbers of metal ions and citrate that
form complexes. The numbers and the equilibrium constants
KMeðm;nÞ for Zn2þ , Fe2þ , Cu2þ , and Mn2þ are listed in Table 2.
The material balance for H2 S species is given by eq. (4),
and material balances of metal and citrate are given by
X
½T.L. ¼ ½L þ
n½(Mem Ln )2mþ ¼ ½L
þ
0
m;n
½S.Me. ¼ ½Me2þ þ
X
-2
Fig. 9 Equilibrium concentrations of heavy metal ions with different
citrate concentrations. Conditions: [I.S.], 15 molm3 ; [T.Me], 5 molm3 ;
pH, 7; T, 20 C; Vg , 1 dm3 ; Vaq , 1 dm3 . [S.Me], Soluble Me2þ concentration; [T.L.], Total citrate concentration.
Table 2 The constants in eqs. (11) and (12) and the equilibrium constants
between citrate and each heavy metal ion.12Þ
Zn
-4
Log([T.L.] / mol . dm-3)
X
mKMeðm;nÞ [Me2þ ]m [L]n
ð15Þ
m;n
where [T.L.] indicates the total concentrations of citrate,
MS.Me. the mole amount of soluble metal ions, and [S.Me.] the
concentration of soluble meal ions. Equations (3), (6), (10),
(14) and (15) were combined and arranged as the following
eq. (16):
m;n
nKMeðm;nÞ [Me2þ ]m [L]n
ð13Þ
m;n
HH2 S Vg
[Hþ ]2 þ Ka1 ½Hþ þ Ka1 Ka2
RT Vaq
K¼
P
½Me2þ ½I.S. ½T.Me. þ ½Me2þ þ mKMeðm;nÞ [Me2þ ]m [L]n
1þ
ð16Þ
m;n
Assuming a [I.S.] of 15 molm3 , [T.Me.] 5 molm3 , pH
7, temperature 293 K, Vg 1 dm3 , and Vaq 1 dm3 , [Me2þ ] at
various values of [L] was calculated by eq. (16). Using the
calculated value of [Me2þ ], [T.L.] and [S.Me.] were
calculated by eqs. (13) and (15), respectively. Figure 9
shows the total soluble metal concentration ([S.Me.]) as a
function of total citrate concentration ([T.L.]), where the
citrate concentration (17 molm3 ) in ATCC 1249 medium is
indicated with a dotted line. The effect of citrate on the total
Cu concentration was small and may be ignored; for Zn and
Fe, the total metal concentrations increase with citrate
concentration, however, with 17 molm3 citrate, the concentrations of Zn and Fe are less than 108 moldm3 . These
results indicate that Fe and Zn can be removed even in the
presence of 17 molm3 citrate. On the other hand, the Mn
concentration in the presence of 17 molm3 citrate is higher
than 1:8 104 moldm3 (10 gm3 ), the maximum contaminant level for wastewater including AMD, in Japan,
suggesting that Mn2þ removal is impossible in the presence
of 17 molm3 citrate.
The effect of components of medium has been scarcely
investigated for the formation of metal sulfide, which would
Fundamental Study on the Removal of Mn2þ in Acid Mine Drainage using Sulfate Reducing Bacteria
2427
Mn2+ concentration / g . m-3
(a)
120
80
40
Sulfate concentration / g . m-3
160
(b)
2000
1500
1000
500
0
0
0
40
80
0
120
40
80
120
Time / hours
Time / hours
200
8
(d)
0
ORP / mV
pH
6
4
2
-200
-400
(c)
0
-600
0
40
80
120
Time / hours
0
40
80
120
Time / hours
Fig. 10 (a) Mn2þ concentration, (b) sulfate concentration, (c) pH, and (d) ORP with time in the bioreactor. Open symbols indicate
experiments with citrate in the culture and solid symbols are without citrate.
result from the fact that the formation of FeS, ZnS, and CuS is
independent of complex formation related with components
of medium. In the case of Mn removal, however, the
solubility of MnS is relatively higher, and the effect of
complex formation should be considered: Citrate, investigated as a complexing agent here, is not always used in the
mediums for SRB.8,9) The ions of Mn can form complex with
not only citrate but with acetylacetone, 50 -adenosine triphosphate, catechol, glycylglycine, glycine, oxalic acid, and
picolinic acid. The use of waste materials as nutrients for
SRB has been proposed to decrease the cost of treatment.2,3,13,14) If the waste materials contain the ligands to
form Mn complex, Mn2þ could not be removed from
solutions.
3.3 Mn removal in the SRB bioreactor
As discussed above, coexisting metal ions such as Zn2þ
and Fe2þ and low pH suppresses the MnS formation in
eq. (2). Citrate, one of the nutrients for SRB growth, also
suppresses this reaction. As it can be expected that Mn is
removed by SRB in the absence of coexisting metal ions and
citrate at higher pH, Mn removal was attempted with the SRB
bioreactor. In this experiments, two mediums were used; the
modified ATCC 1249 medium and the modified ATCC 1249
medium without citrate.
Figure 10 shows the results of the experiments with the two
kinds of mediums. There were no notable effects of citrate on
the sulfate concentration, pH and ORP as shown in Figs.
10(b), (c), and (d). During the experiments, pH was kept in
the range between pH 5 to 7. As can be seen in Fig. 3, the
decrease of pH causes low removal ratios of Mn2þ . In Fig.
10(c), pH decreased to 5.3 around 20 hours, and this may
cause low removal ratio of Mn2þ . However, after 70 hours,
pH increased to about 7 which is closed to initial pH, and the
effect of the variation in pH can be negligible in considering
final removal ratio of Mn2þ . In Figs. 10(b) and (d), sulfate
concentration and ORP decreased with time, indicating that
sulfate was reduced by SRB, and that H2 S was generated.
As shown in Fig. 10(a), citrate affects Mn removal
considerably. In the presence of citrate, the Mn concentration
remained constant, and no Mn was removed from the
solution. Without citrate, the Mn concentration decreases
sharply after 48 hours, and most of the Mn was removed in
100 hours.
When the concentration of Mn and sulfate decreased in the
medium without citrate, a pink precipitate was observed in
the SRB bioreactor. The XRF analysis showed that this
precipitate consists of 47.6% Mn and 52.4% S and that the
molar ratio of Mn to S is close to 1:1. Figures 11 and 12 show
the XRD pattern and the UV-VIS absorption spectrum of the
K. Yoo, K. Sasaki, N. Hiroyoshi and M. Tsunekawa
4.
0
112
110
20
103
101
100
002
Intensity (a.u.)
2428
40
60
2θ / deg.
Fig. 11 XRD pattern of the pink precipitate in the bioreactor. Miller
indices are assigned according to JCPDS:40-1289 (Metastable -MnS).
Conclusions
The optimum condition for Mn removal using a SRB
bioreactor was investigated. Chemical experiments using
Na2 S as the S2 source shows that excess H2 S is required to
remove Mn2þ , and that low pH, coexisting Zn2þ and Fe2þ ,
and citrate (a component of medium for SRB) suppress the
MnS formation. Biological experiments using a SRB
bioreactor confirmed that Mn2þ can be removed as MnS at
pH 5–7 in the absence of the coexisting metal ions and citrate.
Acknowledgements
The authors express appreciation for support of this
research by a grant from Japan Society for the Promotion
of Science and the Arai Science and Technology Foundation.
Absorbance (a.u.)
REFERENCES
320
360
400
440
480
Wavelength / nm
Fig. 12 UV-VIS absorption spectrum of biogenic -MnS
precipitate. The XRD pattern corresponds to metastable MnS. The absorption of the precipitate edge in the UV-VIS
absorption spectrum was at about 326 nm, indicating that the
band gap of the precipitate was 3.8 eV.
There are three forms of MnS:15) -MnS (rock salt
structure) is a green stable form, and -MnS (sphalerite
structure) and -MnS (wurtzite structure) are pink metastable
forms. Metastable -MnS occupies a potential position in
short wavelength optoelectronic applications because of its
large band gap.16)
For the preparation of -MnS, Lu et al.17) and Zhang et
al.18) have reported that high temperatures are required to
generate H2 S from reagents, e.g. thiourea and thioacetamide,
but the present study indicates that the SRB bioreactor can be
used both in AMD treatment and in preparation of metastable
-MnS at ambient temperatures.
1) K. Sasaki, T. Haga, T. Hirajima, K. Kurosawa and M. Tsunekawa:
Mater. Trans. 43 (2002) 2778–2783.
2) W. J. Drury: Water Environ. Res. 71 (1999) 1244–1250.
3) D. H. Dvorak, R. S. Hedin, H. M. Edenborn and P. E. Mclntire:
Biotechnol. Bioeng. 40 (1992) 609–616.
4) P. Elliott, S. Ragusa and D. Catcheside: Wat. Res. 32 (1998) 3724–
3730.
5) R. W. Hammack and H. Dijkman: Proc. Copper 99-Cobre 99
International Conference (1999) Vol. 4, pp. 97–111.
6) T. Nihei, H. Hayashi, S. Tsuneda, A. Hirata and H. Sasaki: Proc. MMIJ
Annual Meeting (2002) Vol. 2, pp. 3–4.
7) J. S. Webb, S. McGinness and H. M. Lappin-Scott: J. Appl. Microbiol.
84 (1998) 240–248.
8) L. L. Barton and F. A. Tomei: Sulfate-Reducing Bacteria, (L. L.
Barton, eds.) (Plenum Press, New York, 1995) pp. 3–6.
9) J. R. Postgate: The sulphate-reducing bacteria (2nd ed.), (Cambridge
University Press, Cambridge, 1984) pp. 30–34, 57.
10) P. G. T. Fogg and C. L. Young: Hydrogen Sulfide, Deuterium Sulfide
and Hydrogen Selenide, Solubility Data Series (A. S. Kertes, eds.)
(Pergamon Press, Oxford and Tokyo, 1988).
11) D. D. Wagman, W. H. Evans, V. B. Parker, R. H. Schumm, I. Halow,
S. M. Bailey, K. L. Churney and R. L. Nuttall: The NBS Tables of
Chemical Thermodynamic Properties, J. Phys. Chem.. ref. Data 11,
Suppl. 2, (American Chemical Society, Washington. D. C. 1982).
12) A. E. Martell and R. M. Smith: Critical Stability Constants, Vol. 3,
(Plenum Press, New York, 1977).
13) M. A. Harris and S. Ragusa: Environ. Geol. 40 (2000) 195–215.
14) I. S. Chang, P. K. Shin and B. H. Kim: Wat. Res. 34 (2000) 1269–1277.
15) S. W. Kennedy, K. Harris and E. Summerville: J. Solid State Chem. 31
(1980) 355–359.
16) L. Wang, S. Sivananthan, R. Sporken and R. Caudano: Phys. Rev. B 54
(1996) 2718–2722.
17) J. Lu, P. Qi, Y. Peng, Z. Meng, Z. Yang, W. Yu and Y. Qian: Chem,
Mater. 13 (2001) 2169–2172.
18) Y. Zhang, H. Wang, B. Wang, H. Yan and M. Yoshimura: J. Cryst.
Growth 243 (2002) 214–217.