Minerals Engineering 39 (2012) 29–35

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Minerals Engineering
journal homepage: www.elsevier.com/locate/mineng

Performance of single substrate in sulphate reducing bioreactor for the treatment

of acid mine drainage
R.P. Choudhary, A.S. Sheoran ⇑
Department of Mining Engineering, Faculty of Engineering, Jai Narain Vyas University, Jodhpur 342 011, India

a r t i c l e i n f o a b s t r a c t

Article history: The focus of this study is to investigate efficiency of the sulphate reducing passive bioreactors for the
Received 29 December 2009 treatment of acid mine drainage. The performance of locally available substrates (manures, woodchips,
Accepted 7 July 2012 millet fodders and sugarcane waste) were examined for the treatment of acid mine drainage. The man-
Available online 4 October 2012
ures showed efficient removal of metals and other pollutants. With cow manure, buffalo manure and goat
manure pH increased from initial 2.70 to 6.25, 7.10 and 7.50. All cellulosic wastes also raised pH in a
Keywords: range of 4.83–5.32. Significant reduction was observed in Eh, acidity and sulphate concentration also.
Mining
It was observed that: 51.49–99.32% of Fe, 84.95–99.97% of Cu, 35.11–99.78% of Zn, 17.87–99.14% of Ni,
Acid rock drainage
Pollution
63.55–99.02% of Co and 12.68–73.86% of Mn were removed in maximum retention period of 10 days.
Bacteria Thus manures can be cost effective as a single substrate in a passive treatment method for removal of
Biotechnology metals from AMD in bioreactors. Also chemical makeup of substrate provided an insight on the ability
of sulphate reduction and metal removal.
Ó 2012 Elsevier Ltd. All rights reserved.

1. Introduction and sulphates. This anaerobic process is carried out by the sulphate
reducing bacteria (SRB) that uses sulphate as terminal electron
Acid Mine Drainage (also referred as AMD) is metal-rich water acceptor for the oxidation of organic waste and hydrogen (electron
formed when minerals exposed by mining activities undergo reac- donor), resulting in the production of hydrogen sulphide (Garcia et
tions with oxygen and water. AMD can result in severe environ- al., 2001). These bacteria are mainly controlled by the composition
mental stress, such as acidification and metal pollution of of the organic substrates which in turn should be easily biodegrad-
adjacent aquatic ecosystems (Hesketh et al., 2010). Abandoned able and available in long term in the bioreactors.
metal and coal mines around the world discharge acidic drainage Previous studies have evaluated the potential of individual and
with high concentration of iron, sulphate, and elevated levels of mixtures of organic materials (substrates) to promote and sustain
soluble toxic metals, which may persist for a very long time largely sulphate reduction in anaerobic biozones (Amos and Younger,
because of financial constraints (Feng et al., 2000; Mohan and 2003). It is now known that the organic substrates serve as an elec-
Chander, 2001; Bartzas et al., 2006). The mitigation of AMD is a tron donor to the sulphate reducing bacteria, like desulfovibrio and
serious environmental issue for the mining industry. If AMD is desulfotomaculum (Cohen and Staub, 1992; Siefert and Mutz, 2001).
discharged untreated, can adversely impact the ecosystem by acid- The activities of sulphate reducing bacteria in these systems con-
ifying soil and contaminates the ground and surface aquifers trol the efficiency of metal decontamination (Walker-David and Si-
(Champagne et al., 2005). grid, 2001; Willow and Cohen, 2003). An advantage of such system
Passive treatment options for AMD based on biogeochemical is that neither electrical power nor the frequent attention of an
processes are gaining ground over traditional (active) techniques operator is required.
such as limestone addition, which are expensive and require exten- Taking advantage of the fact that these microorganisms easily
sive operation and maintenance to function properly (Johnson and grow in the mining environment, bioreactors using anaerobic SRB,
Hallberg, 2005). Passive treatment systems have been used to has been used in the research study for the treatment of AMD.
mediate the environmental impact of abandoned acid mine drain- The microbiological diversity associated with the locally available
age (Behum et al., 2011). Sulphate reduction has become a suitable mining environment is very well proven fact. The effectiveness of
alternative for the treatment of wastewater that contains metals the SRB in treatment process depends on the organic substrates se-
lected as a food to the bacteria. The reaction is generally expressed
as (Widdel, 1988).
⇑ Corresponding author. Tel.: +91 291 2611512.
2CH2 O þ SO2 
4 ! 2HCO3 þ H2 S ð1Þ
E-mail addresses: [email protected], [email protected] (A.S. Sheoran).

0892-6875/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.mineng.2012.07.005
30 R.P. Choudhary, A.S. Sheoran / Minerals Engineering 39 (2012) 29–35

where CH2O represents a simple source of organic carbon. In these for biological decomposition and were most efficient. Percent of or-
passive systems, SRB oxidize the organic carbon source (electron ganic carbon was maximum 57.58 in babool woodchips and lowest
donar) into bicarbonate and reduce the sulphate (electron acceptor) in the cow manure with 23.36. Gross energy calculated based on
to hydrogen sulphides. The bicarbonate neutralizes the acidity, and ether extractives, protein and total carbohydrate ranged between
favours the precipitation of metal carbonate minerals. The produced 184 kcal/100 gm in cow manure to 442 kcal/100 gm in babool
hydrogen sulphide gas combines with metals to form insoluble me- sawdust.
tal sulphides.
2.2. Composition of synthetic AMD
Me2þ þ HS ! MeSð#Þ þ Hþ ðMe2þ  metal ionsÞ ð2Þ
Various organic wastes have been reported as the electron do- Simulated AMD was used to ensure a constant quality of feed
nors for the sulphate-reducers in the treatment of AMD including throughout this study. It was prepared in the laboratory taking into
fresh alfalfa, straw and hay, sawdust, woodchips, paper sludge, account the usual composition of AMD which contains high metals
peat, spent mushroom compost, cow manure, oak leaf (Gibert et concentration such as copper, zinc, iron, cobalt, manganese, and
al., 2004), poultry manure, sheep manures and cattle manure nickel. Simulated AMD composition was prepared in 500 L storage
(Amos and Younger, 2003). plastic tank for experiments. All the chemicals were of analytical
Straw, woodchips and sawdust supported active sulphidogene- reagent grade. The composition is given in Table 1.
sis when they were supplemented by easily assimilable electron
donors such as sucrose, peptone, lactate, pyruvate, or formate, 2.3. Experimental setup and sampling
whey, methanol and ethanol (Zagury et al., 2006; Sheoran et al.,
2010). Ten bench scale bioreactors were constructed with 1 litre trans-
In the present study, the potential role of SRBs inoculated in lo- parent narrow mouth glass bottles provided with inlet at the top
cally available natural organic substrates in the bioreactors have and outlet at lower part. The base of the reactor was filled with
been investigated on laboratory scale for the treatment of acid pebbles of about 5 mm size up to 25 mm height (to increase the
mine drainage. These bioreactors were functional for a period of porosity and to provide the solid surface to SRB’s) and each one
9 months to establish the sustainability of the organic electron do- was filled with 500 cc of different single substrate. Then each bio-
nors to drive sulphate reduction. The chemical characterization of reactor was inoculated with 100 ml whey and 5 g of fresh cow
a carbon source was correlated with the capacity to drive sulphate manure. Whey contains fermentable water-soluble proteins, lac-
reduction. This could be useful in predicting the capacity of a car- tose, and lactic acid. The fermentation products and lactic acid sup-
bon source to derive sulphate reduction as a substitute for expen- ply SRB with a suitable carbon and energy source (Widdel, 1988).
sive, time consuming continuous bioreactor set-ups. Fresh cow manure has been reported by various scientists to con-
tain SRBs (Reynolds et al., 1991; Christensen et al., 1996). The inoc-
2. Materials and methods ulated reactors were allowed to stand for an incubation period of
30 days (for proper development and multiplication of SRBs). After
2.1. Substrate incubation period, leading to the development SRBs as visible in
the form of black film layer and also strong smell of H2S, the biore-
All the organic materials were available at a very low cost and in actors were supplied with simulated AMD and the flow rate was
abundant quantity. All the manures and cellulosic wastes after col- controlled by I–V set used at outlet (Fig. 1).
lection were dried. Cellulosic waste (babool woodchips, babool After incubation period of 30 days to acclimatize SRBs to the
sawdust, mango woodchips, mango sawdust, sugarcane waste, conditions prevailing in bioreactors, then the water present was
pearl millet fodder, proso millet fodder) and organic waste (cow just drained out continuously without any observation for initial
manure, buffalo manure, goat manure) substrates were used in 10 days to avoid any false results as it contained excess of easily
the present study. The manures were collected from local dairy soluble fraction of the substrate. After that all the reactors were
farms. The woodchips and sawdust were collected from carpentry ready in continuous mode for observations e.g. after 40 days, sam-
factory. Sugarcane waste and Fodder (pearl millet and proso millet) ples of inlet and outlets were taken and analyzed for various
were collected from agriculture farms. parameters i.e. pH, Eh, acidity, sulphate and metal ions, continu-
Chemical characterization of the organic substrates used in the ously for 9 months. In the first phase the samples of inlet and out-
experimental setup is given in Table 4. Carbon availability and let of each bioreactor were taken after a retention period of 1 day,
nitrogen and protein content are among the main factors affecting 3 days, 5 days, 7 days and 10 days. This cycle was repeated four
the efficiency of AMD treatment by SRB. The pathway of anaerobic times, thus completing one cycle in 26 days. Then the reactors flow
degradation of organic matter involves the hydrolysis of large was stopped for 15 days. After that, again sampling was carried out
molecular weight compounds such as proteins, nucleic acids, car- for second phase and samples were analyzed after 1 day, 3 days,
bohydrates and lipids to lower molecular weight products such 5 days, 7 days and 10 days of retention period (cycle was repeated
as organic acids and alcohols. Manures have higher percent of pro- five times in second phase).
tein and nitrogen. Cow manure has 1.02 and 6.35 percent; goat
manure has 2.20 and 13.73 percent; buffalo manure has 1.46 and Table 1
9.15 percent of nitrogen and protein (Table 4). It was found that Composition of synthetic AMD.
the manures are rich source of biodegradable carbon source as
Component Concentration (mg/L) Source
comparison to other cellulosic waste. Organic matter ranged from
86.08% to 99.28%, except cow manure 40.28% buffalo manure Fe 188.9 FeSO47H2O
Cu 22.21 CuSO45H2O
56.08%, mango sawdust 52.03% and proso millet fodder 61% due Zn 21.44 ZnSO47H2O
to mixing of silica before sample collection. Mn 31.87 MnSO4H2O
C/N ratio (w/w), based on total dry organic matter, ranged be- Ni 10.43 NiSO46H2O
tween 22.22 and 374.6. The method of C/N computation is the Co 1.24 CoSO47H2O
Sulphate 4132–4960 –
most commonly used. Three manures having C/N ratios of 22–23
pH 2.70–3.35 –
indicated the easily biodegradable and/or available organic carbon
 R.P. Choudhary, A.S. Sheoran / Minerals Engineering 39 (2012) 29–35 31

centrations of sulphate ion using turbidity meter (Make- RI, Model

-211R). The acidity of the water samples was measured using
method 2310 B titration method (APHA, 1992). AMD and treated
effluent samples for metal analysis were filtered twice through fil-
ter no. 42 and acidified with ultra pure concentrated HNO3 imme-
diately after collection, to a pH less than 2 before storage. Only
dissolved (filtered) metals were analyzed. Metal concentrations
determined using Atomic Absorption Spectrophotometer (Make-
Thermo electron corporation Ltd., Model S2).

3. Results and discussion

Results of pH, Eh, acidity (in Figs. 2–4) sulphate and metal re-
moval (in Tables 2 and 3) of acid mine drainage are given for man-
ures (cumulative average of all manures), sugarcane, millet fodders
(average of pearl and proso) and wood materials (average of all
wood chips and saw dust).

Fig. 1. Experimental setup for bench scale single substrates bioreactors.

3.1. pH

2.4. Chemical characterization The pH value of the influent ranged from 2.70 to 3.35. The pH
increased to 7.50, 7.10, 6.25 with goat manure, cow manure and
After collection 10 organic materials were investigated. Study buffalo manure respectively in maximum 10 days retention time
was carried out to assess and evaluate the potential of individual during second phase. Cellulosic waste showed maximum increase
organic material to promote and sustain sulphate reduction in in pH up to 5.32 with sugarcane waste after 7 days of retention
anaerobic bio-zones. For all the experiments air dried samples of period (Fig. 2). The reduced microenvironment created in the sub-
the organic substrate were further dried in a forced air oven at strate forms an insoluble precipitate of metal sulphides and simul-
75 °C, after cooling and grinding the dried samples were passed taneously generates bi-carbonate ions leading to the increase in pH
through 1 mm sieve. These prepared samples were used for organ- value of the water. Goat, cow and buffalo manures provide a source
ic matter and other chemical analysis.
Moisture, dry matter and organic matter were determined in Table 2
accordance with standard gravimetric methods and weight differ- Change in sulphate (mg/L) of AMD in bench scale bioreactors (first and second phase).
ences (APHA, 1992). Nitrogen, protein and carbohydrate contents
Substrate Retention period
were determined by method described in AOAC (1980). Further,
1 Day 3 Day 5 Day 7 Day 10 Day
gross energy is derived from the components of organic matter like
protein, ether extractives and total carbohydrates (AOAC, 1980). AMD FP 4276 4248 4112 4259 4103
SP 4178 4132 4199 4150 4178
Manure FP 3922 3828 3733 3511 2771
2.5. Analytical techniques SP 2394 2344 2494 2140 2018
Woodchips FP 4212 4082 4040 4176 3976
pH and Eh of the samples were measured immediately after col- SP 3347 3286 3845 3293 3243
Millet fodder FP 4284 4066 3977 3951 3911
lection. pH was measured with a pH meter and combination pH
SP 3595 3520 3642 3472 3291
electrode, (Make- RI, Model 151R) using method 4500 H+–B. Eh Sugarcane FP 4220 4131 4055 4217 4006
of the samples were measured with combined platinum redox plus SP 3844 3499 3826 3400 3217
Ag/AgCl reference electrodes and Digital pH/Eh meter (Make- RI,
FP – First phase, SP – second phase.
Model 151R). Turbidimetric method was used to measure the con-

Fig. 2. Change in pH of AMD in bench scale bioreactors (first and second phase).
32 R.P. Choudhary, A.S. Sheoran / Minerals Engineering 39 (2012) 29–35

of carbon for microenvironment culture to develop in the bioreac- ity to 183 mg/L; remaining reactors reduced acidity ranging from
tors which is capable of raising pH significantly. Cellulosic wastes 400 to 616 mg/L as CaCO3 during the first phase of observations
(sugarcane, millet fodders and wood materials) contained some in 10 days of retention period (Fig. 4). In the second phase, acidity
of the non acid soluble contents and thus required more time to dropped to 18 mg/L, 175 mg/L as CaCO3 with goat manure and buf-
degrade to provide carbon source for SRBs, thus were not so effec- falo manure respectively in maximum 10 days of retention time.
tive in raising pH. Other reactors also reducing acidity ranging from 464 to 531 mg/
L as CaCO3 in 10 days of retention period (Fig. 4). Acidity is a key
3.2. Eh (redox potential) contaminant in mine water because metal ion mobility is critically
dependent on pH. The increase in pH, a common occurrence for
SRB needs an anaerobic medium and an anoxic and reduced bioreactors that are adequately buffered with bio-carbonate alka-
microenvironment with negative redox potential. The influent Eh linity, is predominantly attributed to the degassing of dissolved
value was maintained between 366 and 375 mV during whole per- CO2. When mine water contains sufficient alkalinity, metal re-
iod of observation. In the first phase of the experiment only man- moval occurs without a decrease in pH. Some of the earlier com-
ures reduced the Eh in negative range within seven and 10 days of prehensive studies also exist on the persistent microbial sulphate
retention time. Other bioreactors containing cellulosic wastes reduction in substrates receiving low pH, high oxidized waters
(sugarcane, millet fodders and wood materials) had positive Eh va- (Tuttle et al., 1969; Hedin et al., 1988).
lue (Fig. 3). In the second phase, reactors containing manures (or-
ganic wastes) gave promising results; bioreactors containing 3.5. Metal removal
manures observed negative Eh. Other bioreactors containing cellu-
losic waste observed positive Eh values showing the non genera- Comparison of the results among bioreactors containing differ-
tion or non-survival of SRB in these wastes if used as single ent organic and cellulosic waste indicated that good removal of all
substrate. metals was observed in reactors containing manures (Table 3). Per-
formance of the metal removal improved with the increase in
3.3. Sulphate retention time. All the reactors showed better performance in the
second phase. Copper was always the first element to be removed,
The influent and effluent values of sulphate are shown in Table followed closely by Fe, Ni, Zn and Co present in AMD. Copper sul-
2. Values ranged from 4103 to 4276 mg/L during both the phases of phide is highly insoluble over a broad pH range and sulphide pre-
the experiment. In the second phase, the sulphate concentration cipitation of copper ions is therefore a rapid and efficient process in
decreased to 1922, 1951 and 2183 mg/L with goat, buffalo and AMD (Sheoran and Bhandari, 2005). Copper concentration was re-
cow manure in maximum retention period of 10 days respectively. duced to 0.08–2.42 mg/L with cow, goat and buffalo manures. Re-
Bioreactors with cellulosic wastes decreased sulphate values rang- moval efficiency of copper ranged from 98.5% to 99.7% during first
ing from 3073 to 3509 mg/L after 10 days of retention period. In phase. Cellulosic wastes removed 68.4–75.6% of copper in the first
the second phase of the bench scale treatment the percentage of phase. Second phase revealed that all the reactors (organic and cel-
sulphate removal increased in the bioreactors. In the reactors con- lulosic waste containing reactors) removed copper efficiently. Iron
taining goat, buffalo and cow manure with reduced redox potential concentration dropped to 54.16 mg/L with goat manure; remaining
showed good growth of SRBs, The sulphate removal percentage in- nine reactors including cow and buffalo manure dropped iron from
creased from 43% to 45%, 43% to 44% and 47% to 54% in 1, 3, 5, 7 101.55 to 107.92 mg/L during the first phase of the observation. In
and 10 days of retention period respectively. Other bioreactors second phase iron concentration dropped to 1.30 mg/L, 27.94 mg/L
with cellulosic compost removed maximum up to 25 percent sul- and 92.28 mg/L with goat, buffalo and cow manures. Goat, cow and
phate concentration in this phase of experiment. buffalo manure showed iron removal above 99.3, 87.7, and 85.5
percent respectively.
3.4. Acidity Zn removal ranged between 88.2% and 99.5% with goat manure
during first phase, other reactors under performed during first
Influent acidity values ranged from 1132 to 1213 mg/L as phase showing maximum removal up to 40 percent. In the second
CaCO3. All the reactors reduced acidity, goat manure reduced acid- phase, Zn concentration dropped to 0.82 mg/L, 0.05 mg/L and

Fig. 3. Change in Eh (mV) of AMD in bench scale bioreactors (first and second phase).
 R.P. Choudhary, A.S. Sheoran / Minerals Engineering 39 (2012) 29–35 33

Fig. 4. Change in acidity (mg/L as CaCO3) of AMD in bench scale bioreactors (first and second phase).

goat manures. Remaining cellulosic waste reactors reduced Ni

between 5.13 and 11.82 mg/L in various retention periods. Input
Table 3
Percentage metal removal of AMD in bench scale bioreactors (10 days retention Co concentration was very low compared to other metals. Goat
period). manure achieves Co removal ranging from 64.8% to 69.3% in the
first phase of observations with different retention periods; with
% Metal removal Substrates
increased retention period all the reactors showed better Co re-
Manures Woodchips Millet fodders Sugarcane
moval. Similar to other metals Co show good removal with all
Copper FP 98.53 69.75 74.75 69.40 manures. Other cellulosic wastes improved removals with in-
SP 96.60 92.37 88.05 85.70 creased retention period but always under perform than manures
Iron FP 54.00 43.87 44.45 45.10
SP 90.83 52.10 52.15 52.00
containing bioreactors (Table 3). Successful passive treatment of
Zinc FP 53.80 15.17 19.80 37.10 mine water requires attention to both metal removal and acid neu-
SP 98.36 64.72 86.20 35.10 tralizing reaction. The greater Fe removal in the reactor indicates
Nickel FP 53.26 19.15 18.00 10.80 that sulphide precipitation or hydroxide precipitation contributed
SP 98.46 35.90 40.35 17.90
to the enhanced treatment (Peppas et al., 2000; Komnitsas et al.,
Cobalt FP 58.93 44.32 60.80 69.20
SP 95.63 80.72 69.60 74.30 2007). Garcia et al. (2009) confirmed by XRD analysis that the me-
Manganese FP 07.16 02.35 03.25 00.20 tal crystals were predominantly pyrite (FeS2), sphalerite (ZnS), and
SP 39.70 24.22 30.20 18.20 minor quantity of greenockite (CdS). Iron was present in major pre-
cipitation followed by zinc. The metal sulphides precipitate was
recovered from the system during different periods. With divalent
heavy metals the important metals hydroxides formed in following
0.29 mg/L with cow, goat and buffalo manures. Rest reactors per- manner (Feng et al., 2000).
formed only after 10 days of retention time. Apart from manures
the maximum removal was observed 87.2 percent in pearl millet M2þ þ OH ! MðOHÞþ ð3Þ
fodder. Similar to Zn, good removal was observed in Ni with goat
manure during first phase of the experiments. Ni concentration ðOHÞþ þ OH ! MðOHÞ2 ð4Þ
dropped to 3.21 mg/L, 3.22 mg/L, 4.47 mg/L, 1.43 mg/L and
0.29 mg/L in 1,3,5,7 and 10 days of retention respectively. Other
MðOHÞ2 þ OH ! MðOHÞ3 ð5Þ
reactors in first phase dropped Ni value between 5.82 and
11.83 mg/L. In second phase all the manures showed good removal The removal of Mn and Fe concentrations in the reactor are
of Ni ranging between 97.4 and 99.1 percent with cow, buffalo and indicative of exchange adsorption. Yu et al. (2000) reported that

Table 4
Chemical composition of organic substrate.

Substrates Chemical analysis

Moisture (%) Dry matter (%) Organic matter (%) Nitrogen Protein (%) Percent C (%) C/N (%) Silica (%) Total Gross energy
(%) carbo-hydrate (kcal/
(%) 100 gm)
Cow manure 3.13 96.87 40.28 1.02 6.35 23.36 23.00 53.8 32.48 184
Sugarcane waste 5.89 94.11 93.93 0.63 3.96 54.48 85.94 3.2 87.87 407
Goat manure 5.39 94.61 86.08 2.20 13.73 49.93 22.74 3.2 67.96 401
Buffalo manure 3.32 96.68 56.08 1.46 9.15 32.52 22.22 32 42.63 269
Babool woodchips 4.09 95.91 99.28 0.18 1.12 57.58 321.1 Nil 95.16 429
Mango woodchips 5.09 94.91 89.28 0.15 0.94 51.78 345.4 7.6 85.09 389
Babool sawdust 4.33 95.67 98.83 0.15 0.96 57.32 374.6 Nil 92.12 442
Mango sawdust 2.59 97.41 52.03 0.14 0.84 30.18 223.5 39.2 48.04 234
Pearl millet fodder 4.88 95.12 89.52 0.774 4.84 51.92 67.09 04 79.43 406
Proso millet fodder 5.97 94.03 61 0.787 4.92 35.38 44.95 4.2 45.78 315
34 R.P. Choudhary, A.S. Sheoran / Minerals Engineering 39 (2012) 29–35

ion exchange is probably one of the major adsorption mechanisms biodegradable composition but they definitely work for longer per-
for binding divalent metal ions to the sawdust. Adsorption is as iod. Hence, further research is needed to study the depletion rate of
much a function of pH as it is of the species affinity to sorbs, for the organic matter in passive bioreactors to accelerate the treat-
which manganese has perhaps the weakest affinity of all of constit- ment of acidic mine waste in the abandoned mine which is yet
uent metals cations (Willow and Cohen, 2003). key problem for the mining industry.
Influent manganese concentration was between 33.07 and
34.97 mg/L. Manganese removal rate was maximum 8.9 percent
in the goat manure containing bioreactor, undulating in the whole Acknowledgments
first phase of the experiments. In the second phase of the experi-
ments goat manure removed maximum manganese ranging from This research work was supported by the Council of Scientific &
60 to 73.9 percent. Remaining all reactors also removed about 30 Industrial Research (CSIR), Delhi, Grant # 9/98(0081)2K7-EMR-1.
percent manganese in 10 days of retention period. The metal re- We would like to thank CSIR which supported in kind, in sprit
moval mechanism changes during the life of bioreactor. Upon and financially.
startup of a passive bioreactor, the absorption of the dissolved met-
als onto organic sites in the substrate material is an important pro-
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