Hydrogen sulphide conversion to elemental sulphur in a suspended-growth continuous stirred tank reactor using Chlorobium limicola

PII: S0043-1354(97)00393-X

Wat. Res. Vol. 32, No. 6, pp. 1769±1778, 1998 # 1998 Elsevier Science Ltd. All rights reserved Printed in Great Britain 0043-1354/98 $19.00 + 0.00

HYDROGEN SULPHIDE CONVERSION TO ELEMENTAL SULPHUR IN A SUSPENDED-GROWTH CONTINUOUS STIRRED TANK REACTOR USING CHLOROBIUM LIMICOLA M M PAUL F. HENSHAW*, J. K. BEWTRA* and N. BISWAS*

Civil and Environmental Engineering, University of Windsor, Windsor, Ont., Canada N9B 3P4 (First received November 1996; accepted in revised form September 1997) AbstractÐA biological process employing green sulphur bacteria to remove sulphide (S2ÿ) from industrial wastewaters and convert it to elemental sulphur was investigated. This research was unique in that dissolved sulphide was present in the liquid in¯uent fed into a continuous-¯ow photosynthetic bioreactor. A suspended-growth once-through continuous-¯ow stirred-tank bioreactor was successfully operated under ®ve di€erent experimental conditions. For the ®rst three experiments, concentrated nutrient solution and sulphide stock solution were pumped separately into a 13.7 litre reactor at a hydraulic retention time of 45 h and S2ÿ loading rates of 2.1, 4.4, and 5.6 mg/h
l. At the lowest loading rate, nearly all in¯uent S2ÿ was oxidized to sulphate. The middle loading rate resulted in complete conversion of S2ÿ to elemental sulphur. Steady state conditions were not achieved at the highest loading rate, and S2ÿ accumulated in the bioreactor. In two more experiments, nutrient medium and S2ÿ stock solution were separately fed into a 12.0 litre bioreactor at S2ÿ loading rates of 3.2 and 2.7 mg/h
l and hydraulic retention times of 173 and 99 h respectively. In these trials, the loading rates were adjusted to maintain a residual of 20 to 30 mg S2ÿ/h
l in the bioreactor, and consequently, there was nearly complete conversion of the consumed S2ÿ to elemental sulphur. A parameter was developed to relate the results from these experiments to those reported in the literature, where smaller reactors and higher bacterial concentrations were used in batch reactors fed with H2S(g). This parameter described the capacity of the bioreactor to consume S2ÿ, and was calculated as the product of the radiant ¯ux per unit reactor volume and the bacteriochlorophyll concentration. # 1998 Elsevier Science Ltd. All rights reserved Key wordsÐphotoautotrophic, green sulphur bacteria, Chlorobium thiosulphatophilum, Chlorobium limicola.

INTRODUCTION

Hydrogen sulphide (H2S) is a highly toxic and malodorous gas (Cadena and Peters, 1988). Hydrogen sulphide is liberated in the process of re®ning crude oil and thus petroleum re®neries produce ``sour'' water containing H2S(aq). Process water in petroleum re®neries may contain up to 5000 mg S2ÿ/l (Nemerov, 1978). Sulphide (S2ÿ) has a high oxygen demand of 2 mol O2/mol sulphide and thus may cause signi®cant depletion of oxygen in receiving waters (Kobayashi et al., 1983). In Canada, re®neries are allowed to discharge a maximum of 0.3 kg S2ÿ/1000 m3 of oil re®ned/day (Losier, 1990). The addition of chemical oxidizers (hypochlorites, chlorine, potassium permanganate, hydrogen peroxide or oxygen) to sour water results in the formation of sulphate (SO2ÿ 4 ). The partial oxidation of H2S to elemental sulphur (S0) instead of SO2ÿ 4 has several advantages. Elemental sulphur is a nontoxic non-corrosive solid containing more sulphur *Author to whom all correspondence should be addressed.

per unit mass than any other form. Moreover, elemental sulphur is 3 to 8 times more valuable by mass than commercial H2SO4 (Cork, 1978; Kim and Chang, 1991). The main use of elemental sulphur is as a feedstock for the chemical, fertilizer and materials manufacturing industries (West and Duecker, 1974). For these reasons, petroleum re®neries recover H2S liberated in their processes by converting it to elemental sulphur. The chemical processes for sulphur recovery are expensive because of the need to replace poisoned and expired catalysts and contaminated reactor liquids (Cork et al., 1986). Some of the disadvantages of physical/chemical sulphur recovery can be overcome through the use of a biological sulphide removal process which converts H2S(aq) to S0. This process can be adapted to remove the S2ÿ remaining in wastewater streams after sour water stripping, where the sulphide concentration is approximately 80 mg/l (Lawlor, 1990), or potentially replace the combination of sour water stripping and sulphur recovery processes.

1769

1770

P. F. Henshaw et al. Table 1. Research conducted on continuous sulphide removal using chemoautotrophs

Ref.

Con®guration and oxygen source

Vol. (l.)

In¯uent [H2S]

S2ÿ loading (mg/l
h)

E€. rem. (%)

E€. con. (%)

Ass et al. (1983) Sublette and Sylvester (1987a) Sublette (1987)

FF, U, air SG, CSTR, NOÿ 3 SG, CSTR, air

Sublette and Sylvester (1987b) Buisman et al. (1990a)

SG, CSTR, NOÿ 3 FF, CSTR, O2 SG, CSTR, O2 FF, CSTR, O2 FF, biorotor, O2 FF, U, O2 FF, CC, air

47 1.44 1.44 2.0 1.44 5.5

30±60 mg/l in liquid 0.20±0.32 mM in gas 0.27±0.32 mM in gas " ? in gas 100 mg/l in liquid 55 mg/l in liquid 35±174 mg/l in liquid 45±203 mg/l in liquid 45±225 mg/l in liquid 0.17 mM in gas

21±96 32±74 38±51 32±33 58 129 7.9 104±521 208±938 208±1040 19±38

2100 94 2100 2100 2100 97 2100 100±60 100±69 100±73 77±69

NQ NQ <0.2 <0.2 0 83 SNM 2100 SNM 50±58 SNM 50±69 SNM 60±73 SNM NQ

Buisman et al. (1990b) Lizama and Sankey (1993)

8.3 3 20 0.34

Vol. = wet volume of reactor, FF = ®xed-®lm, SG = suspended-growth, CSTR = continuous stirred-tank reactor, U = up¯ow, CC = countercurrent contractor, electron acceptor: air = O2 in air, O2=pure oxygen and NOÿ 3 =nitrate, Vol. = wet volume of reac2ÿ 2ÿ 0 2ÿ tor, E€.rem. = removal eciency ((S2ÿ in ÿSout)/Sin ), E€.con. = conversion eciency (Sout/Sin ), NQ = not quanti®ed, SNM = results by subtraction since S0 not measured.

Therefore, the function of this biological process is twofold: to remove sulphide from wastewater and to produce elemental sulphur. LITERATURE REVIEW

Conversions between di€erent species of sulphur are accomplished by several types of naturally occurring bacteria in stagnant, sulphur-rich ponds. At the water surface, chemoautotrophs use the energy released in the spontaneous reaction of H2S or S0 with dissolved oxygen to form SO2ÿ 4 . In the upper anaerobic zone, photoautotrophs use infrared light energy to oxidize H2S or S0. Tables 1 and 2 summarize the results reported using chemoautotrophic and photoautotrophic bacteria respectively for sulphide removal. The desirable bacterium for the bioprocess under investigation should readily convert H2S to S0, require a minimum of nutrient inputs and produce S0 that is easily separable from the biomass. Comparing Tables 1 and 2, photoautotrophic bacteria in suspended-growth reactors have demonstrated the removal of sulphide at sulphide loadings up to 100 mg/h
l whereas chemoautotrophs have successfully removed sulphide at higher loadings. In the chemoautotrophic experiments, complete removal of sulphide and 100% conversion to S0 was not achieved. Buisman et al. (1990a) believed that oxygen di€used into the pores of the ®xed-®lm support medium faster than H2S and

2ÿ therefore S0 was oxidized to SO2ÿ could 4 before S be oxidized to S0. Cork et al. (1985) introduced the concept of the ``van Niel curve'' wherein the light radiated to a photosynthetic reactor was coupled to the conversion of sulphide to elemental sulphur or sulphate. Within photosynthetic green sulphur bacteria (GSB), the reverse citric acid cycle requires NADPH and ATP to assimilate CO2. The conversion of light to chemical energy by photophosphorylation occurs by cyclic (ATP producing) and noncyclic (NADPH producing) means. The total rate of photophosphorylation is dependant on the amount of light energy absorbed by the photochemical reaction centers (PRCs) on the membranes of the bacterial cells. NADPH production requires electrons from the oxidation of electron donor molecules such as S2ÿ or S0. The light energy absorbed by the PRCs is split between the two types of photophosphorylation such that the proper ratio of ATP to NADPH production occurs for CO2 assimilation. Thus a constant fraction of the light energy absorbed is used for NADPH production and this determines the number of electrons required. If the electron donor (S2ÿ) feed rate matches the light input, all of the S2ÿ will be converted to S0, releasing 2 electrons per molecule of S2ÿ oxidized. If the S2ÿ feed rate is lower at the same light input, the requirement for electrons is satis®ed by oxidizing S2ÿ to SO2ÿ 4 , yielding 8 electrons per molecule of

Table 2. Research conducted on continuous sulphide removal using photoautotrophs Con®guration

Vol. (l)

In¯uent [H2S]

S2ÿ loading (mg/L
h)

E€. rem. (%)

E€. con. (%)

Irradiance (W/m2)

Rad. ¯ux (W)

Kobayashi et al. (1983)

FF, U FF, plug

8 0.1

0.59±1.27 102±125

92±81 100

20 8-12

NQ NQ

NQ NQ

Cork (1984) Cork et al. (1985) Maka and Cork (1990) Kim et al. (1991) Kim et al. (1992)

SG, CSTR SG, CSTR SG, CSTR

0.8 0.8 0.8

16 mg/l in liquid 24±19 mg/l in liquid 4.1 mM in gas ? in gas 1±2 mM in gas

62 109±74 32±64

100 100 100±90

? 95±93 97±90

ID 2000±150 139

2.8 29±2.1 2

SG, CSTR SG, CSTR

4 4

2.1 mM in gas 2.1 mM in gas

61 64

99.5 100

35 63

490 714

122 178

Ref.

FF = ®xed-®lm, SG = suspended-growth, plug = plug ¯ow reactor, CSTR = continuous stirred-tank reactor, Vol. = wet volume of 2ÿ 2ÿ 0 2ÿ reactor, E€.rem. = removal eciency ((S2ÿ in ÿSout)/Sin ), E€.con. = conversion eciency (Sout/Sin ), Rad. ¯ux = radiant ¯ux, NQ = not quanti®ed, ID = insucient data to calculate.

H2S conversion to elemental sulphur using C. limicola

S2ÿ oxidized. If there is an excess of electron donors (i.e., a high S2ÿ feed rate) then all of the S2ÿ will not be oxidized and sulphide will accumulate in the bioreactor. Photoautotrophic bacteria have been shown to produce a higher percentage conversion of S2ÿ to S0 while simultaneously removing 100% of the sulphide (Table 2). In addition, the ability to quickly and automatically control the oxidation rate of sulphide by varying the light intensity has advantages when processing a waste stream of variable S2ÿ concentration (Kim et al., 1992, 1993). For photoautotrophs, the reactor must be designed to allow the maximum penetration of light, an uncommon characteristic for bioreactor design. The use of a suspended-growth photoreactor, fed with dissolved sulphide as in this work, has not been reported in the literature.

MATERIALS AND METHODS

Methods of analysis The concentrations of all sulphur species are reported as sulphur equivalents, in units of (mg S as species)/l. The uncertainty in the concentration of any sulfur species was calculated from its calibration curve by taking the di€er-

1771

ence in concentrations corresponding to the two 95% con®dence limits at a response of 0 and dividing by two. Sulphide. The methylene blue colorimetric method of Truper and Schlegel (1964) was used. Elemental sulphur. Aqueous samples were extracted into chloroform (CHCl3) prior to quanti®cation by high-performance liquid chromatography (HPLC; Lauren and Watkinson, 1985; Henshaw et al., 1997). Sulphate. The APHA et al. (1992) turbidimetric method was used. The turbidity of the bu€ered solution without barium chloride (BaCl2) was subtracted from that with BaCl2 to compensate for the turbidity produced by the addition of Bu€er A to samples with S2ÿ and S0 present. Thiosulphate. Ion chromatography using a 4.1  150 mm Hamilton PRP-X100 column and conductivity detector was employed with a run time of 30 min (Hamilton, 1992). The eluent was 4 mM p-hydroxybenzoic acid adjusted to a pH of 8.5 and pumped at 2.0 ml/min. Bacteria. Bacteriochlorophyll (bchl) was extracted into methanol and its absorbance was measured at 670 nm against methanol (Maka, 1986). The concentration of bchl was assumed to be proportional to the biomass concentration. Materials and apparatus Bacteria. The green sulphur bacterium Chlorobium limicola (originally deposited as Chlorobium thiosulphatophilum) was purchased from the American Type Culture Collection (ATCC, Rockville, Maryland; catalogue no. 17092) and subcultured weekly into sterile growth medium.

Fig. 1. Schematic diagram of apparatus from runs 3, 4 and 5.

1772

P. F. Henshaw et al.

Growth medium. The growth medium recommended by Madigan (1988) was used. It consists of three parts: a mineral salts solution (supplying nitrogen, phosphate, calcium, magnesium and trace elements), a bicarbonate solution and a sulphide stock solution containing sodium sulphide dissolved in deaerated deionized water. In the continuous¯ow bioreactor experiments, the components of the growth medium were split amongst several in¯uent streams. Due to equipment failures, only 5 of the 15 experiments performed maintained growing bacteria until steady state measurements could be taken. In Runs 3, 4 and 5, the in¯uent was composed of three streams: deionized water, a concentrated nutrient solution and a sulphide stock solution (Fig. 1). In Trials 9 and 10, there were only two feeds: a nutrient medium and a sulphide stock solution (Fig. 2). Apart from the subculturing tubes, the reactors and equipment were not sterile. The recipe for growth medium required that it be kept in the dark for 24 h so that the sulphide would consume any oxygen in the vessel and to prevent algal growth. Reactor. A New Brunswick Scienti®c model F-14 fermenter was used as a continuous reactor. The reactor was wrapped in a metallized mylar ®lm and two 105 mm high by 110 mm wide windows were cut, one above the other, into the ®lm so that the top of the upper window was at the water level of the constant temperature (308C) water bath. A second reactor was used as a submerged light source by removing the stir paddles and clamping two Philips IR 175 Watt R-PAR bulbs inside the vessel directed into the windows of the ®rst reactor. Both reactors were mounted in a New Brunswick Scienti®c model FS-314 fermenter drive assembly. This resulted in an average irradi-

ance of 258 W/m2 over both windows. The bioreactor stirring speed was approximately 200 rpm. Master¯ex (Cole-Palmer, Chicago) variable speed pumps with solid state speed controls and standard pump heads were used for all in¯uent and e‚uent reactor streams. The liquid level in the reactor was kept constant by a level probe inside the reactor which actuated the e‚uent pump. The pH inside the reactor was kept between 6.8 and 7.2 by an Omega (Omega Engineering, Stamford, CT) PHP-166 Chemical Metering Pump controlled by an Omega PHM 55 pH Regulator connected to an Ingold (Ingold Electrodes, Wilmington, MA) 465-35-K9 combined glass electrode inserted into the reactor. Procedure Start-up. The contents of a 70 ml culture tube containing Chlorobium limicola were poured into a 1.1 litre ¯ask containing 1.0 litre of growth medium. This was stirred under infrared light in a 308C water bath for three to four days. The contents of the 1 litre reactor were then poured into the fermenter containing 1 to 12 litres of growth medium where it was stirred and illuminated for an additional three to four days, after which the continuous feed pumps were turned on and adjusted. Operation. Feed pumps were adjusted by timing the revolutions until the proper speed (typically 2 to 10 rpm) was achieved. Occasionally, the reactor windows had to be cleaned of what was presumed to be elemental sulphur by inserting a nylon brush while the reactor was pressurized with CO2 or N2. Table 3 summarizes the reactor operating parameters. For Runs 3, 4 and 5, the concentration of the sulphide

Fig. 2. Schematic diagram of apparatus for trials 9 and 10.

H2S conversion to elemental sulphur using C. limicola

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Table 3. Variables in continuous reactor operation Experiment Run 3 Run 4 Run 5 Trial 9 Trial 10

Con®g.

RSL

Vol. (l.)

1 1 1 2 2

bottom bottom bottom top top

13.7 13.7 13.7 12.0 12.0

Dur. (h) 316 383 283 1680 1220

[S2ÿ]i (mg/l) 90 190 260 550 260

Loading (mg/h
l) 2.1 4.4 5.6 3.2 2.7

HRT (h) 45 44 46 173 99

pH adj. b b a a a

[S2ÿ]sss (mg/l) 1340 2760 4040 2730 2440

NI 1.02 0.99 1.05 1.03 1.01

[SO2ÿ 4 ]i (mg S/l) 29 30 32 37 31

Con®g. = number indicates Figure number for reactor con®guration, RSL = reactor sampling location, Vol. = reactor liquid volume, Dur. = duration of experiment, [S2ÿ]i=e€ective inlet sulphide concentration, Loading = average reactor sulphide loading, HRT = hydraulic retention time, pH adj. = acid (a) or base (b) used to adjust pH in reactor, [S2ÿ]sss=sulphide stock solution concentration, NI = nutrient index = the concentration of the nutrient solution times the dilution of this solution as it is mixed with the other in¯uent streams (1.00 is equivalent to the medium prescribed by Madigan, 1988), [SO2ÿ 4 ]i=e€ective inlet sulphate concentration (another indicator of the in¯uent nutrient concentration).

stock solution was increased in successive Runs while the hydraulic retention time (HRT) was held constant for all Runs (Table 3). Throughout each Run, a constant sulphide loading rate was maintained regardless of the condition in the reactor. Start-up was performed for Run 2, which was aborted due to equipment failures. However, the bacteria remained viable in the reactor for Runs 3, 4 and 5. In Trials 9 and 10, HRTs higher than those in Runs 3, 4, and 5 were used. The sulphide loading was adjusted so as to have a low but constant sulphide concentration in the reactor. Both Trials were started from scratch by inoculation from subcultured ATCC bacteria. RESULTS

Concentrations Toward the end of each experiment, when a steady state had been reached, the concentrations of all sulphur species were measured in the bioreactor input and output streams at time intervals ranging from 24 to 150 h (Table 4). Each set of measurements provided a ``snapshot'' of the reactor condition at that moment. The reactor was well mixed, so that the concentrations in the reactor and those leaving the reactor were assumed to be equal. A steady state was assumed to exist in the reactor when successive daily sulphide and bacteria measurements were relatively constant. The elemental sulphur and thiosulphate concentrations were analyzed at the end of each experiment, so the values of these parameters were not available at the time when the sets of sulphur species measurements were taken. At some measurement times, the total amount of sulphur going into the reactor did not equal the total amount of sulphur leaving the reactor. For example, in the measurements at 52 h in Run 3, a steady state in terms of elemental sulphur and sulphate had not yet been reached. In another example, at 382 h in Run 4, there was a transient decrease in the sulphide concentration in the inlet, which was not known at the time of the sulphur species measurements. In Run 3, a steady state was achieved after 150 h (3.3 HRTs). At this steady state, the sulphide leaving the reactor was measured to be less than 1 mg/l (Table 4). The S0 concentration was less than 2 mg/l, and the outlet sulphate concentration was roughly equal to the inlet sulphide concentration. In this

state, the bacteria were completely oxidizing the sulphide in the reactor to produce sulphate. Since elemental sulphur is the desired end-product from this process, Run 3 represents an underloaded condition in terms of sulphide. According to the theory that supports the van Niel curve, this condition would arise when, at the given light level, there was not enough sulphide being provided to satisfy the needs of the bacteria for electrons simply through the two-electron yielding conversion of S2ÿ to S0. In Run 4, the reactor was operated at nearly an optimum sulphide loading. Here, a steady state was reached after about 200 h or 4.5 HRTs, as indicated by a plateau in the S0 concentration. The S0 value in this steady state condition was equal to the e€ective S2ÿ input concentration, taking into account the uncertainty in the assays. Thus, the requirement for electrons in the reactor was satis®ed by oxidizing S2ÿ to S0. As in Run 3, the reactor S2ÿ concentration was less than 1 mg/l. Run 5 was operated at a high e€ective inlet sulphide concentration (Table 3). As a result after 210 h, the reactor S0 concentration slowly decreased and S2ÿ began to accumulate. Therefore, at this light and sulphide input, the reactor was overloaded with sulphide since the in¯uent sulphide was not being fully oxidized. The experiment did not reach a steady state condition as indicated by the increasing concentration of sulphide in the reactor e‚uent in Table 4. The reactor was operated at a nearly optimal condition in Trial 9. The elemental sulphur concentration in the e‚uent equalled or exceeded the e€ective sulphide in¯uent concentration. The sulphate concentration in the e‚uent was steady and nearly equal to the in¯uent SO2ÿ 4 concentration. These two conditions indicate that the reactor was being operated at a point on the van Niel curve and complete conversion of S2ÿ to S0 was occurring. At times, sulphide was present in the reactor, which normally indicates overloading, but the sulphide did not accumulate throughout the experiment. A balance between the total sulphur into and out of the reactor was only achieved at 1629 h. Trial 10 was also operated at nearly complete S2ÿ to S0 conversion. However, the steady state reactor S0 concentration was greater than the e€ective inlet

=not measured, used the average of the three later measurements in this experiment, bchl = bacteriochlorophyll 5 day running average, 2# = the uncertainty in the measurement of the value. The uncertainty in the Total value is the sum of uncertainties of the individual measurements.

Trial 10

Trial 9

Run 5

Run 4

a

total

862 14 115 2 14 114 2 18 762 18 822 18 552 18 422 18 372 14 392 14 222 0 282 0 312 14 312 14 302 14 262 26 322 0 292 0 312 0 Ð Ð Ð Ð Ð Ð Ð Ð Ð Ð 76a213 85 213 78 213 66 213 54a213 62 213 58 213 41 213 02 4 02 4 02 4 02 4 02 4 02 4 32 4 312 4 742 4 112 2 7 42 4 232 3 262 3 242 3 202 3 302 3 302 3 02 3 52 148 316 191 286 382 214 239 259 283 1343 1438 1629 1677 693 861 929 1028 Run 3

127 214 93 210 89 210 220 215 220 220 139 215 305 224 299 223 307 224 254 216 488 225 529 227 627 226 499 224 304 212 239 214 245 212 297 215

102 3 32 2 82 2 262 3 272 4 132 2 02 12 62 11 112 12 172 10 462 5 312 5 02 11 52 11 442 3 722 4 112 5 382 7

Ð Ð Ð Ð Ð Ð Ð Ð Ð Ð 43a213 632 70 272 67 402 64 81a213 107 2 54 372 37 100 2 48

302 1 292 1 302 4 292 4 282 4 302 4 302 5 292 5 292 5 272 5 302 13 372 13 312 13 352 13 282 13 292 13 322 7 272 6

167 218 125 213 127 217 275 222 275 228 182 221 335 240 334 239 347 240 298 231 606 257 660 2115 685 2117 579 2112 456 242 446 284 326 261 462 276

32 21 2 21 0 21 236 21 221 21 251 21 258 22 279 22 199 22 169 21 1051 21 988 21 622 22 612 22 240 21 457 21 388 21 402 22

SO2ÿ 4 S2O2ÿ 3

E‚uent concentration (mg S/l)

S0 S2ÿ total SO2ÿ 4

In¯uent concentration (mg S/l)

S2O2ÿ 3 S0 S2ÿ Time (h) Experiment

Table 4. Concentrations measured during steady state operation

118 2 19 117 2 19 114 2 23 312 2 23 303 2 23 306 2 23 303 2 24 347 2 20 312 2 20 303 2 8 11592 18 11272 32 757 2 33 733 2 33 339 2 18 581 2 17 505 2 18 474 2 18

bchl

P. F. Henshaw et al.

10.3 13.5 12.5 11.5 11.5 10.7 7.2 6.2 6.2 5.0 5.1 4.4 3.1 2.8 3.5 5.3 5.4 5.4

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sulphide concentration (Table 4). Total sulphur balance was only achieved at 1028 h. As in Trial 9, the reactor sulphate concentration was essentially steady and S2ÿ concentration ranged up to 30 mg/l. These concentrations were used to quantify the conversion of S2ÿ to S0. Table 5 shows the percentage of conversion for only those times where a total sulphur balance was achieved. Realistically, Run 3 showed essentially zero conversion of S2ÿ to S0. The conversion in Runs 4 and 5 averaged about 91% and 94% respectively. Trial 9 achieved essentially 100% conversion within the cumulative error of the measurements. The high conversion value at 1028 h in Trial 10 (123%), was due to the oxidation of some thiosulphate in the in¯uent to elemental sulphur. Rates The rates of change of each of the sulphur species were calculated by performing a mass balance on each sulphur species in the reactor between sulphur species measurements in each experiment (Table 6). The growth rate of bacteria was also calculated by a mass balance, and the speci®c growth rate, m, was calculated by dividing the calculated growth rate by the average bacteria concentration in that time period. Comparing experiments, no correlation was found between the concentration of any reactant and the rate of consumption of that reactant. The rate of consumption of sulphide was found to be equal to the rate of sulphide loading except for Run 5 where the bioreactor was overloaded. van Niel curve The data from Runs 3, 4 and 5 and Trials 9 and 10 (this work) were plotted onto a modi®ed version of the van Niel curve of Maka and Cork (1990) in Fig. 3. Only the infrared light experiments from Maka and Cork (1990) where at least 87% of the S2ÿ was converted to S0 were considered. The abscissa parameter of the van Niel curve was changed from irradiance (W/m2; Maka and Cork, 1990) to radiant ¯ux per unit reactor volume times bacteriochlorophyll concentration (W
mg/l2) to account for the larger reactor volumes and lower bacteria concentrations in this work. The region of Fig. 3 near the origin has been expanded in Fig. 4 to show how the van Niel curve derived from the data of Maka and Cork (1990) neatly divides the overloaded experiments in this work from the underloaded experiments in this work. According to the van Niel curve the light input determined the number of electrons transferred. For a sulphide loading ``on the curve'', the electron requirement of the GSB was met by the removal of two electrons from S2ÿ to form S0. If the electron requirement increased, further oxidation from S0 to SO2ÿ 4 yielded six more electrons per molecule of sulphur. The rate of electron transfer in an experiment was calculated from the rates of formation of the

H2S conversion to elemental sulphur using C. limicola

1775

Table 5. Conversion of sulphide to elemental sulphur for those measurement times where inlet sulphur equaled outlet sulphur Experiment

Time (h)

Run 3

148 316 191 286 214 239 259 283 1629 1028

Run 4 Run 5

Trial 9 Trial 10

[S2ÿ]iÿ[S2ÿ] (mg/l)

[S0] ÿ [S0]i (mg/l)

D[S0]/D[S2ÿ] (%)

93 214 89 214 220 219 220 224 302 228 268 227 233 228 142 222 601 229 297 218

ÿ1 23 ÿ8 23 210 23 194 25 258 213 273 213 188 213 152 211 622 213 364 29

ÿ1 22 ÿ9 22 952 9 882 11 852 36 102 2 15 812 16 107 2 24 103 2 8 123 2 10

2# = shows the uncertainty of the measurement of the value.

oxidized compounds in that experiment. Only one of these values was chosen from each experiment as indicated in Table 6, typically that with the best mass balance or rate data. The rates of formation in mg S/h
l (as di€erent species) were divided by 32 to give mmol S/h
l. Then the molar rates of for2ÿ mation of S0, S2O2ÿ 3 and SO4 were multiplied by 2, 4 and 8 respectively to give the rates of electron release (mmol eÿ/h
l) for each mmol of S2ÿ oxidized to each of the compounds. For the data from Maka and Cork (1990), the total sulphide loading rates in mmol/h
l were multiplied by the percentage of each species remaining at the end of the experiment and 2ÿ by 2, 4 and 8 for S0, S2O2ÿ 3 and SO4 respectively (sulphide remaining at the end of the experiment contributed no electrons to the bacteria). In either case, the rates of electron transfer were summed and plotted against radiant ¯ux per volume (Fig. 5). This relation describes the PRC. The PRC receives an electron from the electron donor molecule, absorbs light, and increases the energy of the electron. This high energy electron is then passed to NADP+ for the formation of NADPH. For each unit of light input, there are a certain number of electrons transferred. The number of electrons per unit of light energy (slope of curve in Fig. 5) decreases as the light energy increases. This may be attributed to the fact that the PRC becomes light

saturated at high light levels. The data plotted in Fig. 5 also shows that the experiments of this work are at much lower values of radiant ¯ux/volume as compared to those of Maka and Cork (1990). Unfortunately, the use of a conventional fermentor with lights shining from only one direction limited the amount of light that could be made available to the bacteria. From this, it can be concluded that higher S2ÿ loading rates can be achieved by increasing the light to volume ratio, increasing the GSB concentration or both. The former improvement suggests the use of a small diameter plug¯ow reactor. Higher bacteria concentrations may be achieved by recirculating the biomass leaving the reactor back into the reactor, or using a ®xed-®lm bioreactor. The energy change per electron released from the oxidation of S2ÿ to S0 is not the same as the energy change per electron released from the oxidation of S0 to SO2ÿ 4 . Table 7 illustrates the thermodynamic advantage of using electrons from S2ÿ as opposed to those from S0 or S2O2ÿ 3 . The rate of energy release was calculated by multiplying the electron yield for each sulphur conversion by the appropriate energy release per electron. This value was correlated to the radiant ¯ux/volume using an exponential function as shown in Fig. 6. The norm is de®ned as the sum of the squares of the di€er-

Table 6. Parameters calculated between steady state measurements Independent variables

Experiment Run 3 Run 4 Run 5 Trial 9 Trial 10

a

average S2ÿ average HRT (h) loading (mg/h
l) 2.3a 2.1 4.9a 4.1 6.7 6.6a 6.1 2.9 3.4a 3.3 2.7 2.4 2.7a

Values used in Figs 5 and 6.

48 45 45 44 45 46 46 175 172 172 99 100 100

Average concentrations [S2ÿ] (mg/l) 0 0 0 0 17 53 93 14 24 25 25 30 15

[S0] (mg/l) 17 1 229 236 269 239 184 1020 805 617 348 422 395

Rates of formation

[bchl] (mg/l)

S2ÿ (mg/h
l)

S0 (mg/h
l)

11.9 13.0 11.5 11.1 6.7 6.2 5.6 4.8 3.8 3.0 4.4 5.4 5.4

ÿ2.3 ÿ2.0 ÿ4.9 ÿ4.0 ÿ5.2 ÿ3.3 ÿ2.5 ÿ2.6 ÿ3.2 ÿ3.2 ÿ2.4 ÿ2.1 ÿ2.9

ÿ0.10 ÿ0.11 4.3 5.2 6.7 1.0 2.5 4.9 2.7 3.4 4.2 2.8 3.8

SO2ÿ Speci®c 4 (mg S/h
l) growth rate m (hÿ1) 1.8 1.9 1.2 0.61 0.021 0.30 ÿ0.64 0.013 ÿ0.019 ÿ0.036 0.040 ÿ0.035 0.022

0.024 0.022 0.022 0.022 0.016 0.022 0.013 0.0042 0.0040 0.0037 0.013 0.010 0.010

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Fig. 3. Sulphide loading as a function of radiant ¯ux per unit volume times bacteriochlorophyll. The solid line was obtained by ®tting the data of Maka and Cork (1990) to an equation of the form y = c(1 ÿ ekx) where c = 69.3 and k = ÿ 0.0119.

ences between the predicted function values and the data points. A lower norm indicates a better correlation. The norm for the energy vs. radiant ¯ux/ volume plot (Fig. 6) is higher than that for the elec-

Fig. 5. The calculated rate of electron transfer as a function of radiant ¯ux per unit volume.

tron transfer vs. radiant ¯ux/volume plot (Fig. 5). This is probably due to the greater ordinate values in Fig. 6. If the norms for these two plots are divided by the average ordinate value from the Maka and Cork (1990) data, the value for Fig. 6 is lower (0.43) than that for Fig. 5 (0.62). Figure 5 con®rms a relationship between the rate of electron transfer and the rate of light energy input. The better ®t of the parameter plotted in Fig. 6 to the data suggests the kinetics of the transfer of electrons from the electron donor molecule to the PRC are in¯uenced by the thermodynamics of the electron donor oxidation. Figure 6 can be used to make a rough calculation of the cost e€ectiveness of phototrophic sul®de conversion. For example, a light input of 4 W/l relates to a chemical energy release of 104 J/h
l. If this energy release was produced by converting S2ÿ to S0 and the cost of electricity was $0.06/kW
h then the energy cost of the sulfur produced would be $3,750/(tonne S0) calculated as follows: Table 7. Energy released from oxidation reactions of sulphur species

Fig. 4. Sulphide loading as a function of radiant ¯ux per unit volume times bacteriochlorophyll (Fig. 3 enlarged).

Half reaction

Number of electrons transferred

S2ÿ4S0 S0 4SO2ÿ 4 S2ÿ412S2O2ÿ 3 2ÿ 2ÿ 1 S O 4SO 4 2 2 3 S2ÿ4SO2ÿ 4

2 6 4 4 8

a

Free energya Energy per mole change at pH 7, electrons (kJ) DG0' (kJ/mol) ÿ52 ÿ115 ÿ71 ÿ96 ÿ167

26 19 18 24 21

Calculated by subtracting energies of formation of products minus reactants as found in Brock and Madigan (1988).

H2S conversion to elemental sulphur using C. limicola

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…4 W=l †…$0:06=kW
h†…10ÿ3 kW=W † ˆ $3750=tonne …104 J=h
l †=…26 J=mmol eÿ †…1 mmol S 0 =2 mmol eÿ †…32 mg S 0 =mmol S 0 †…1 tonne=109 mg† Based on prices reported in the literature (Cork, 1978; Kim and Chang, 1991), this cost must be reduced by at least one order of magnitude to make the process economical. Improved reactor eciencies will help. However, the greatest ineciency in this system results from the use of a light source in which only a small percentage of the energy input is utilized by the GSB. The use of light emitting diodes which deliver light energy at the speci®c wavelength for use by the GSB has been suggested (Cork, 1985) and tested (Kim et al., 1991) and found to give positive results.

CONCLUSIONS

The experiments in this work have shown that complete removal of S2ÿ from the in¯uent to a continuous stirred-tank photosynthetic bioreactor can be achieved with over 90% conversion of the removed sulphide to elemental sulphur. As was the case with chemotropic reactors (Buisman et al., 1990a), sulphate was not produced when S2ÿ was present in the reactor e‚uent. Under this condition, 100% of the S2ÿ that was removed was converted to S0. The reactor performance in these experiments can be compared to those reported previously for photobioreactors through the use of a parameter which takes into account the radiant ¯ux entering

Fig. 6. Calculated energy released by sulphur oxidation as a function of the radiant ¯ux per volume.

the reactor, the concentration of bacteria in the reactor and the volume of the bioreactor. Through this comparison, the concept of the van Niel curve was veri®ed, wherein the conversion of sulphide to elemental sulphur or to sulphate in a photosynthetic bioreactor is a function of the light radiated to the reactor and the feed rate of sulphide. AcknowledgementsÐThis research was supported by operating grants from the Natural Science and Engineering Research Council (NSERC) of Canada as well as the University Research Incentive Fund (Ontario, Canada).

REFERENCES

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