Fate and effect of silver on the anaerobic digestion process

PII: S0043-1354(00)00173-1

Wat. Res. Vol. 34, No. 16, pp. 3957±3966, 2000 7 2000 Elsevier Science Ltd. All rights reserved Printed in Great Britain 0043-1354/00/$ - see front matter

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FATE AND EFFECT OF SILVER ON THE ANAEROBIC DIGESTION PROCESS SPYROS G. PAVLOSTATHIS*M and SUNG KYU MAENG School of Civil and Environmental Engineering, Georgia Institute of Technology, Atlanta, Georgia 30332-0512, USA (First received 1 May 1999; accepted in revised form 26 January 2000) AbstractÐLaboratory assays were conducted to assess the anaerobic biodegradability of a silverbearing, waste activated sludge as well as the e€ect of silver compounds on the anaerobic digestion process. All assays were performed at 358C in the dark. The ultimate biodegradability of a silverbearing waste activated sludge (5.0 g silver/kg sludge dry solids) was 61% as compared to 59% for the control (i.e., silver-free) sludge. The rate and extent of methane production was similar for both sludge samples. Addition of either silver nitrate or silver sul®de to methanogenic, mixed cultures up to an equivalent concentration of 100 mg Ag/l did not a€ect the rate and extent of methane production. Silver thiosulfate when tested at an equivalent concentration of 100 mg Ag/l (and 1000 mg S/l), resulted in accumulation of ca. 28 mM of fatty acids (mainly acetate), 90% inhibition of methanogenesis and 39% inhibition of acidogenesis. However, when using silver-free, thiosulfate-amended controls, it was concluded that the observed inhibition in the silver thiosulfate-amended cultures was not attributed to the silver but rather to the excess thiosulfate (used as an alternative electron acceptor resulting in the production of soluble sul®de at inhibitory levels). Computer simulations under typical anaerobic digestion conditions using the geochemical equilibrium speciation program MINTEQA2 resulted in extremely low concentrations (<10ÿ14 M) of free silver ions (Ag+). The two predominant insoluble silver species were Ag2S and Ag8. Therefore, due to the high complexing capacity of the anaerobic digester mixed liquor as well as the reduction to elemental silver, relatively high levels of silver (at least up to 100 mg Ag/l) can be tolerated by anaerobic digestion systems. The results of this study have important implications on the biological treatment and management of photoprocessing wastewaters. 7 2000 Elsevier Science Ltd. All rights reserved Key wordsÐanaerobic digestion, inhibition, photoprocessing, silver, thiosulfate

INTRODUCTION

Photographic manufacturing accounts for more than 50% of the silver demand in the US. Silver is also used in electronics, jewelry, silverware, solder, bearings, and for medical and dental applications (Purcell and Peters, 1998). It has been estimated that approximately 2500 tonnes of silver are annually released into the environment, out of which 150 tonnes are associated with sludge produced in wastewater treatment plants (Ratte, 1999). Silver toxicity is associated with the free Ag+ ion. As a result, silver threshold concentrations range several orders of magnitude and depend on the silver compound, age and species of organism exposed, as well as the physical and chemical characteristics of the environment (Purcell and Peters, 1998; Ratte, 1999). For example, there was no toxicity to unacclimated activated sludge microorganisms by silver thiosulfate at silver levels of 100 mg/l, whereas 6.4 mg Ag/l added in the form of *Author to whom all correspondence should be addressed. Tel.: +1-404-894-9367; fax: +1-404-894-8266; e-mail: [email protected]

silver nitrate resulted in about 84% inhibition (Bard et al., 1976). During photoprocessing, the generated wastewater contains silver, typically in the form of soluble silver thiosulfate complexes such as [Ag(S2O3)n](2n ÿ 1)ÿ. The most common silver-thiosulfate species are those with n = 1, 2 or 3 with relatively low dissociation constants, K: 1  10ÿ9, 5  10ÿ14, and 1.26  10ÿ14, respectively (Pouradier et al., 1977). As a result, the concentration of free silver ions in photoprocessing wastewaters is extremely low and silver exists either as soluble, undissociated silver-thiosulfate complexes, relatively insoluble silver-thiocyanate complexes or as insoluble species (e.g., AgBr, Ag2S). Furthermore, silver recovery commonly used in photo®nishing facilities leads to as high as 98% removal of silver. As a result, very low silver concentrations (less than 5 mg/l) remain in the photoprocessing wastewaters (Dagon, 1973; Bober et al., 1992). In a survey of photoprocessing facilities, it was found that more than 99% of such facilities in the US discharge their e‚uents into municipal sewers leading to publicly owned treatment works (POTWs) (Versar Inc., 1981). A survey of photo-

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Spyros G. Pavlostathis and Sung Kyu Maeng

processing facilities found a 30-day average maximum total silver concentration equal to 1.1 mg/l for facilities which use conventional silver recovery and 0.4 mg/l for facilities using conventional silver recovery followed by ion exchange (Versar Inc., 1981). The silver concentration in the e‚uent from a POTW receiving photoprocessing wastewaters was usually less than 0.01 mg/l and silver removal eciencies of 95% were observed (JBF Scienti®c Inc., 1977). In a recent study on the fate of silver in ®ve POTWs, the in¯uent total silver concentration in these plants ranged from 0.004 to 0.10 mg/l and silver removal eciencies in excess of 94% were observed (Shafer et al., 1998). Previous work on the biological treatment of photoprocessing wastewaters has demonstrated that these wastewaters are amenable to both aerobic and anaerobic treatment and do not adversely a€ect the treatment processes (Dagon, 1973; Lytle, 1984; Bober et al., 1992; Pavlostathis and Sridhar, 1992, 1994; Pavlostathis and Morrison, 1994a,b; Pavlostathis and Jungee, 1994; Pavlostathis and Maeng, 1998). An experimental activated sludge system was able to treat silver-bearing photoprocessing wastewaters with a total silver concentration in the mixed liquor over 150 mg/l with no adverse e€ects (Dagon, 1973). The silver was present as silver sul®de (Ag2S) along with some elemental, metallic silver (Ag8). Both silver species were removed by sludge settling leading to a very high silver removal eciency (r90%). Silver-bearing waste activated sludge (1% Ag on dry weight basis; resulting from the treatment of a simulated photoprocessing wastewater with an in¯uent total silver concentration of 10 mg Ag/l) was anaerobically digested without any adverse e€ects on the anaerobic digestion process (Leonhard and Pfei€er, 1985). In contrast to the aerobic biological treatment of photoprocessing wastewaters, far less attention has been given to the e€ect of silver on anaerobic biological processes in spite the fact that anaerobic digestion of waste activated sludge is the most common method of sludge treatment and stabilization. To address this need, the present study was undertaken. The objective of the work presented here was to assess the anaerobic biodegradability of a silverbearing, waste activated sludge generated from the aerobic treatment of a photoprocessing wastewater as well as to assess the e€ect of three silver compounds on the anaerobic digestion process. MATERIALS AND METHODS

Activated sludge Activated sludge was generated by three laboratoryscale reactors fed with an organic mixture, organic mixture plus a simulated, ®xer-derived photoprocessing wastewater (a thiosulfate based ®xer), and organic mixture plus ®xer wastewater plus silver as previously described (Pavlostathis and Maeng, 1998). For a total silver feed concentration of 1:8520:05 mg/l, the reactor ®ltered (0.2 mm) e‚uent silver

concentration was below the method detection limit (0.01 mg Ag/l) and the steady-state mixed liquor total silver concentration was 1:8420:16 mg Ag/g mixed liquor suspended solids (dry weight basis)(values in mean 2 standard deviation; n r 5). The generated waste activated sludges were further digested aerobically for 60 days as reported elsewhere (Pavlostathis and Maeng, 1998). As a result of solids destruction, the ®nal total silver content of the aerobically digested sludge was 4.99 mg Ag/g total suspended solids (dry weight basis). When the silver-bearing, aerobically digested sludge was subjected to the toxicity characteristic leaching procedure (TCLP), the silver concentration in the sludge extract was equal to 0:1120:02 mg Ag/l (mean2standard deviation; n = 3) which is much lower than the 5 mg/l regulatory limit for silver (Code of Federal Regulations, 1996). Only 0.6% of the total, sludge-bound silver was leached out by the TCLP extraction. Therefore, silver in waste activated sludge, presumably present in the form of silver sul®de, is extremely stable and practically unextractable by the TCLP test (Pavlostathis and Maeng, 1998). At the end of the aerobic digestion, the sludge solids from each of the three reactors were concentrated by centrifugation (9000 rpm, 15 min), and then air-dried to a ®nal water content of 4:820:4, 3:320:1, and 3:920:3% (mean 2 standard deviation; n = 3), for the control, ®xeramended and ®xer plus silver-amended reactor, respectively. The sludge solids were then ground using a mortar and pestle until passed through a standard US sieve No. 60 (250 mm). The chemical oxygen demand (COD) content of the aerobically digested, dry activated sludge samples from the control, ®xer and ®xer-plus-silver reactors was 1.31, 1.36, and 1.45 mg COD/mg total solids (dry weight basis), respectively. Sludge ultimate digestibility assay The ultimate digestibility of the above described sludge samples was determined by using a serum bottle assay as previously described (Pavlostathis and Sridhar, 1994). Eight series were prepared in triplicate, 120-ml serum bottles (105 ml liquid volume) as follows. Dry, aerobically digested waste activated sludge samples were added to six serum bottle series (two series per sludge type) to arrive at an approximate initial sludge COD value of 4000 mg/l. The bottles were ¯ushed with helium gas for 5 min, then sealed with thick butyl rubber stoppers and aluminum crimps. Then, using a syringe, 100 ml of mixed liquor were anaerobically transferred from a laboratory-scale, anaerobic digester into each serum bottle. Details on the development and operation of the digester can be found elsewhere (Maeng, 1998). Two bottle series were prepared and used as controls: one contained only the anaerobic inoculum (referred to as seed blank) and the other (referred to as reference), in addition to the inoculum, contained an organic mixture composed of dextrin and peptone (initial concentration of 230 and 115 mg/l, respectively). In addition to the sludge, three bottle series (one per sludge type) contained the organic mixture at the same level as the reference. Incubation was carried out in the dark at 358C, and the bottles were agitated manually once a day. Anaerobic toxicity assay The objective of this test was to assess the e€ect of silver compounds commonly present in photoprocessing wastewaters on the anaerobic digestion process. The potential inhibitory/toxic e€ect of silver compoundsÐsuch as silver nitrate, silver thiosulfate, and silver sul®deÐon both acidogenesis and methanogenesis was assessed using a serum bottle assay similar to the above described ultimate digestibility assay. Silver thiosulfate complexes were prepared according to a procedure that simulates silver halide

Silver and the anaerobic digestion process photographic emulsions and avoids the formation of a certain amount of silver sul®de if silver nitrate is directly mixed with thiosulfate (Pavlostathis and Maeng, 1998). This procedure was carried out in two steps, as follows: Step IÐformation of silver halide: AgNO3 …aq† ‡ NaBr…aq† 4 AgBr…s† ‡ NaNO3 …aq† Step IIÐformation of silver-thiosulfate complexes: AgBr…s† ‡ nNa2 S2 O3 …aq† 4 Ag…S2 O3 †n…2nÿ1†ÿ …aq† ‡ Br ÿ …aq† ‡ 2nNa‡ …aq† The most common silver thiosulfate species are those with n = 1, 2 or 3 (Pouradier et al., 1977). To be conservative, n was taken as equal to three which results in the following silver thiosulfate complex: Ag(S2O3)5ÿ 3 (MW=444.2; S/Ag=1.78). For this experiment, in order to make sure that all silver was thiosulfate complexed, a ratio of S/Ag equal to 10 was used (i.e., about ®ve times more sulfur than needed based on the stoichiometric ratio). Concentrations of 1, 2, 10, and 100 g as S/l of sodium thiosulfate and 0.1, 0.2, 1 and 10 g as Ag/l of stock solutions were used to prepare the di€erent silver thiosulfate serum bottle series. Because of the high concentration of thiosulfate remaining after complexation with silver nitrate to prepare the silver thiosulfate complexes, another series of bottles with sodium thiosulfate was prepared. It was anticipated that the high concentration of thiosulfate and/or the resulting sul®de would have a negative impact on the anaerobic digestion process (e.g., depression of methane production due to COD consumption as a result of thiosulfate reduction and/or inhibition due to the production of relatively high sul®de concentrations). By using sodium thiosulfate alone in a separate series, the role of thiosulfate and sul®de in the silver thiosulfate series could be determined independently from silver. Six series were prepared in triplicate, 120-ml serum bottles (105 ml liquid volume) following the above described procedure for the ultimate digestibility assay, with the following amendments: none (seed blank), dextrin/peptone mixture (reference), silver nitrate, silver thiosulfate, silver sul®de, and sodium thiosulfate. The latter four series were also amended with the dextrin/peptone mixture at the same level as the reference series. Silver nitrate and silver thiosulfate were tested at four concentrations: 2, 10, 50, and 100 mg Ag/l (using concentrated stock solutions). Silver sul®de was tested at three concentrations: 10, 50, and 100 mg Ag/l (added as dry solid). Sodium thiosulfate was tested at four concentrations: 20, 100, 500, and 1000 mg S/l (using a concentrated stock solution). The silver thiosulfate and sodium thiosulfate series contained equal amounts of thiosulfate. Incubation was carried out in the dark at 358C, and the bottles were agitated manually once a day.

3959

The volume of the gas produced was measured by the displacement of an acid brine solution (10% NaCl w/v, 2% H2SO4 v/v) in a graduated burette connected to the serum bottles with a rubber tubing and a needle each time gas measurement was performed. The temperature and barometric pressure were recorded and the measured gas volumes were then converted to standard temperature and pressure (STP) conditions (08C, 1 atm) using the ideal gas equation. The gas composition (CO2, CH4 and H2S) was determined by gas chromatography (thermal conductivity detector) as previously reported (Prytula and Pavlostathis, 1996). Volatile fatty acids (VFAs) were analyzed using an HP Series II 5890 gas chromatography unit (Hewlett Packard, Palo Alto, CA, USA) equipped with a ¯ame ionization detector and a 30-m Stabilwax 0.53 mm i.d. column (Restek, Bellefonte, PA, USA). Helium was used as the carrier gas at a ¯ow rate of 10 ml/min. The injection port, oven and detector temperature were 210, 110 and 2208C, respectively. Calibration curves for acetic, propionic, isobutyric, butyric, isovaleric, and valeric acids were developed using standard stock solutions. The VFA method detection limit was 0.05 mM. Mixed liquor total sul®de was measured by converting the soluble and precipitated sul®des to H2S and determining the resulting gaseous H2S. The following procedure was used. Aliquots of 5 ml of 6N H2SO4 were transferred into triplicate 25-ml serum bottles. The bottles were sealed with thick rubber stoppers and aluminum crimps and ¯ushed with helium gas for 5 min. An appropriate volume of the mixed liquor or sodium sul®de (usually between 1 and 5 ml) was transferred by syringe into the serum bottle and the bottles were incubated at 358C for 1 h. Headspace H2S was determined by gas chromatography as described above. Sodium sul®de was used to prepare a calibration curve and was standardized using the iodometric method (American Public Health Association, 1995). RESULTS AND DISCUSSION

Sludge ultimate digestibility

Analyses

The sludge ultimate anaerobic digestibility assay of the aerobically digested sludges lasted for 63 days. The duration of this assay was more than three hydraulic retention times (HRT) of conventional municipal digesters which are usually operated at an HRT range of 10±20 days (Metcalf and Eddy Inc., 1991). Therefore, for all practical purposes, the duration of this assay represents the ultimate digestibility of the sludge samples. All samples were analyzed for pH, alkalinity and total COD at the beginning and the end of the incubation, and the results appear in Table 1. The cumulative volumes of methane and carbon dioxide produced during the incubation period are also

The following parameters were measured according to procedures outlined in Standard Methods (American Public Health Association, 1995): pH, alkalinity …pH ˆ 4:0), COD (dichromate re¯ux method), and oxidation-reduction potential (ORP). An Orion Research digital pH/millivolt meter model 611 was used in conjunction with a platinum electrode with an Ag/AgCl reference in 3.5 M KCl gel (Sensorex, Stanton, CA, USA) for ORP measurements. Thiosulfate was determined by ion chromatography as previously described (Pavlostathis and Maeng, 1998). Total silver was determined by acid digestion/atomic absorption according to standard procedures (US Environmental Protection Agency, 1986). The silver detection limit was 0.01 mg/l.

Fig. 1. Cumulative methane produced during the sludge ultimate digestibility assay.

3960

Spyros G. Pavlostathis and Sung Kyu Maeng Table 1. COD, pH and alkalinity change as well as cumulative gas production during the sludge ultimate digestibility assay

Parameter

Seed blank

pH Initial Finala Alkalinity, mg/l as CaCO3 Initial Finala COD, mg/l Initial Finala Cumulative gas, ml at STP CH4 CO2

Reference

Sludge without organic feed

Sludge with organic feed

Control

Fixer

Fixer+Silver

Control

Fixer

Fixer+Silver

7.6 7.8

7.4 7.2

7.5 7.3

7.7 7.6

7.7 7.6

7.2 7.2

7.4 7.4

7.4 7.4

4000 4350

4670 4850

4700 4945

4450 4680

4650 4750

5140 5320

5170 5400

5090 5145

1130 613

4790 1200

4240 2400

4260 2420

4285 2430

9030 3050

9050 3015

9075 3000

2.8 1.5

96.7 43.5

23.4 6.9

25.6 6.8

23.9 5.1

133.1 73.0

132.2 59.0

128.9 58.0

a

After 63 days of incubation at 358C.

shown in Table 1. The cumulative methane production pro®les during the incubation period are shown in Fig. 1. All three sludge samples produced the same amount of total gas and methane at 63 days of incubation (no statistical di€erence at a ˆ 0:05). Also, the volume of gas produced in the three sludge samples amended with the organic mixture was the same. It is noteworthy that when the volume of gas produced by the reference series (which initially contained only seed and the dextrin/ peptone mixture) was subtracted from that produced by the organic mixture-amended sludge series, on the average, resulted in 45% higher gas production in the latter samples as compared to those sludge series not amended with the organic mixture. Therefore, the addition of the dextrin/peptone mixture to the sludge samples resulted in an enhanced sludge biodegradability. Table 2 shows the digestibility, degradability ratio, speci®c methane production (SMP) as well as the COD balance data for the sludge digestibility assay. The degradability ratiosÐcalculated by dividing the amount of COD destroyed in each sample amended with the dextrin/peptone mixture by that of the reference sampleÐfor all three sludge samples were greater than unity (Table 2). Degrad-

ability ratios less than unity indicate inhibition. Therefore, all three sludge samples did not inhibit COD destruction and methane production resulting from the degradation of the added organic mixture. All three sludge samples had a very similar digestibility, ranging from 42.3 to 42.5%. When the digestibility data for samples containing the dextrin/ peptone mixture were corrected for the organic mixture contribution in terms of COD destruction, the sludge digestibility values were higher than for the same sludge samples without organic mixture amendment (ranging from 56.4 to 58.0%; see Table 2). In e€ect, the addition of the organic mixture resulted in a higher degradability of the three sludge samples. Again, the fact that the degradability ratios and the sludge digestibility values were very similar for all three sludge samples, indicates that neither the ®xer nor the ®xer plus silver had any adverse e€ect on the anaerobic degradation of activated sludge resulting from the aerobic treatment of laboratory-simulated photoprocessing wastewaters. Anaerobic digestion of a silver-bearing waste activated sludge (1% Ag sludge content on a dry weight basis) did not result in any adverse e€ect on the anaerobic digestion process (Leonhard and Pfei€er, 1985).

Table 2. COD balance, speci®c methane production and biodegradability data for the ultimate digestibility assaya Sample Reference Sludge without organic feed Control Fixer Fixer+Ag Sludge with organic feed Control Fixer Fixer+Ag a

COD balanceb (%)

Degradability ratioc

14.2

1.00

84.0

291

24.5 22.4 24.2

NAf NA NA

42.5 42.3 42.4

148 164 150

24.3 25.2 26.8

1.67 1.68 1.69

69.2 (56.4)g 69.7 (57.4) 70.0 (58.0)

227 223 216

Values corrected for seed blank contribution. (CODinÿCOD®nÿCODCH4)  100/CODin. Ratio of COD destroyed in the organic feed amended samples to that in the reference sample. d (CODinÿCOD®n)  100/CODin. e Speci®c methane production (ml CH4 produced at STP/g COD destroyed). f NA, not applicable. g Values in parenthesis corrected for the organic feed contribution in terms of COD destruction. b c

Digestibilityd (%)

SMPe

Silver and the anaerobic digestion process

The speci®c methane production (SMP; ml methane produced at STP per g COD destroyed) values for all samples varied from 148 to 291 ml CH4/g COD. These values are below the theoretical value of 350. The SMP and the COD balance values for the sludge-amended samples are signi®cantly lower than that for the reference sample (Table 2). Complete oxidation of thiosulfate to sulfate in a laboratory-simulated photoprocessing wastewater was achieved by the activated sludge process (Pavlostathis and Maeng, 1998). Therefore, reduction of any sulfate associated with the dry sludge during the anaerobic sludge digestibility assay could potentially lead to electron channeling away from methanogenesis. However, although neither the sulfate content of the dry activated sludge nor the sul®de possibly produced during the anaerobic sludge digestibility assay were measured for the ®xer and ®xer plus silver sludge series, it is noteworthy that the sludge control series had COD balance and SMP values comparable to those of the other two sludge series. Therefore, the reported higher COD balance and lower SMP values for all sludge-amended series as compared to those of the reference series may be attributed to either reductive processes not accounted for and/or experimental errors (e.g., sampling and COD quanti®cation). An ultimate digestibility of approximately 50% was reported for a biological sludge generated by a 10-day solids retention time (SRT) laboratory-scale activated sludge reactor (Gossett and Belser, 1982). The same researchers also demonstrated that the ultimate digestibility of sludge decreases with increasing SRT values of the activated sludge process. Pavlostathis and Gossett (1988) reported an ultimate digestibility of 61.5% for a biological sludge generated by a laboratory-scale activated sludge reactor operated at a 10-day SRT. Pavlostathis and Sridhar (1994) reported an ultimate digestibility of 83% for a biological sludge generated by a laboratory-scale activated sludge reactor fed with simulated photoprocessing wastewaters and operated at a 4-day SRT. The ultimate digestibility values of the biological sludges reported here are signi®cantly lower than previously reported values. However, it should be emphasized that, prior to the anaerobic digestibility assay, the three sludge samples were aerobically digested for 60 days where the sludge solids destruction reached values between 60 and 65% (Pavlostathis and Maeng, 1998).

3961

thiosulfate at the end of the incubation period are shown in Table 3. The cumulative methane production pro®les of the methanogenic cultures amended with di€erent levels of silver sul®de, silver nitrate, silver thiosulfate, and sodium thiosulfate, as well as that of the control series (seed blank and reference) are shown in Fig. 2. The degradability ratio values based on total COD destruction and methane production in each culture with the highest amendment level as compared to the reference culture (i.e., the culture without any silver amendment) are shown in Table 4. Note that for an accurate estimation of the organic mixture COD destruction (see Table 4), the measured COD depletion in the silver-thiosulfate and sodium-thiosulfate culturesÐ calculated as the di€erence between the total initial and ®nal COD valuesÐwas increased by the equivalent COD used for the reduction of thiosulfate to sul®de based on the observed extent of thiosulfate depletion. Nitrate reduction was not monitored in the silver nitrate-amended cultures. However, based on the excellent COD balanceÐwhich is based on the total COD destroyed and methane produced (see Table 4)Ðit is clear that methanogenesis was the main metabolic process accounting for the observed COD destruction. All levels of silver sul®de had no adverse e€ect on the gas production of the test cultures as compared to that of the reference culture (Fig. 2 and Table 3). Only acetate at 0.1 mM was detected at 72 days of incubation in the 100 mg Ag/l silver sul®de amended culture and was similar to the VFA content of the reference culture. The degradability ratios for the culture amended with the highest silver sul®de level (i.e., 100 mg Ag/l) were slightly above unity which further demonstrates that when silver was added to the methanogenic cultures in the form of the highly insoluble silver sul®de, it did

E€ect of silver compounds on the anaerobic digestion process The anaerobic toxicity assay lasted for 72 days The initial and ®nal pH values ranged from 7.2 to 7.4 and from 6.9 to 7.4, respectively. The initial and ®nal alkalinity values ranged from 4850 to 5100 and from 5030 to 6100 mg/l as CaCO3, respectively. The volume of methane and carbon dioxide produced, as well as the remaining total sul®de and

Fig. 2. Cumulative methane produced by the methanogenic cultures amended with di€erent levels of silver nitrate, silver sul®de, silver thiosulfate, and sodium thiosulfate (The cumulative methane production of the seed blank and reference cultures is repeated for the purpose of direct comparison of the data).

3962

Spyros G. Pavlostathis and Sung Kyu Maeng

Table 3. Methane and carbon dioxide produced as well as remaining total sul®de and thiosulfate after 72 days of incubation of methanogenic cultures amended with silver sul®de, silver nitrate, silver thiosulfate, and sodium thiosulfate Culture Seed blank Reference Ag±Sul®de (mg Ag/l): 10 50 100 Ag±Nitrate (mg Ag/l): 2 10 50 100 Ag±Thiosulfate (mg Ag/l): 2 10 50 100 Na±Thiosulfate (mg S/l): 20 100 500 1000

CH4 (ml)

CO2 (ml)

Total sul®dea (mg S/l)

Thiosulfateb (mg S/l)

23 135

7 75

12 13

NAc NA

138 145 148

72 77 80

25 27 28

NA NA NA

143 139 145 126

78 72 79 74

39 27 23 11

NA NA NA NA

145 137 86 13

83 84 67 46

38 84 445 553

5 12 20 195

143 143 96 21

77 75 78 54

39 87 423 638

5 20 27 214

a

Mixed liquor sul®de measured at 72 days. Remaining thiosulfate at 72 days. NA, not applicable.

b c

not result in any inhibitory e€ect. Silver nitrate levels as high as 100 mg Ag/l had no adverse e€ect on the methanogenic cultures as shown by the volume of gas produced (Fig. 2 and Table 3), degradability ratios very close to unity, and VFA content resembling that of the reference culture. Silver nitrate is known to be toxic to aerobic systems because of the toxicity of the free silver ion (Ag+) resulting from the dissolution of silver nitrate. For instance, compared to the free silver ion, silver sul®de was about 15,000 times less acutely toxic in 96h, ¯ow-through, acute toxicity tests using fathead minnows (Cooley et al., 1988). However, under the conditions of the anaerobic toxicity assay (measured Eh ˆ ÿ80210 mV), silver nitrate up to 100 mg Ag/ l did not adversely a€ect the methanogenic cultures as a result of silver complexation with various ligands in the culture media, precipitation as silver sul®de or reduction as metallic silver (i.e., elemental silver, Ag8), silver forms known to be non toxic. To further explain the lack of any inhibitory e€ect of

silver nitrate on the activity of the mixed methanogenic cultures which simulated the anaerobic digestion process, a close look at the silver speciation under conditions representing the anaerobic digestion process was undertaken using a geochemical equilibrium speciation computer program as discussed in a subsequent section. The methane production by the silver thiosulfateamended cultures over the incubation period is shown in Fig. 2. Silver thiosulfate at 2 and 10 Ag/l did not a€ect methanogenesis. An initial lag period of about 12 days with low methane production was observed for the 50 mg Ag/l silver thiosulfate amended culture, but methane production increased in the later part of the incubation period. However, although the 100 mg Ag/l silver thiosulfate amended culture produced methane up until day 12, methanogenesis was completely inhibited for the remainder of the incubation period. It is noteworthy that, due to the excess thiosulfate used to produce the silver-thiosulfate complexes, a large

Table 4. COD balance and degradability data for the reference as well as the methanogenic cultures with the highest amendment level (anaerobic toxicity assay) Culture

Reference Ag±Sul®de (100 mg Ag/l) Ag±Nitrate (100 mg Ag/l) Ag±Thiosulfate (100 mg Ag/l) Na±Thiosulfate (1000 mg S/l) a

COD balancea (%)

2.7 ÿ0.2 ÿ0.3 ÿ4.1 ÿ4.5

Degradability ratio based on: COD destructionb

Methane productionc

1.00 1.04 0.90 0.30d 0.36d

1.00 1.10 0.93 0.10 0.16

(CODinÿCOD®nÿCODCH4)  100/CODin. Ratio of COD destroyed in each test culture to that in the reference culture. Ratio of methane produced in each test culture to that in the reference culture. d COD destruction based on measured COD depletion plus the equivalent COD used for the observed extent of thiosulfate reduction (see text). b c

Silver and the anaerobic digestion process

portion of the thiosulfate was not complexed with silver. Based on stoichiometry (see Materials and Methods section), the uncomplexed thiosulfate levels were 16, 82, 411, and 822 mg S/l for the silver thiosulfate series of 2, 10, 50, and 100 mg Ag/l, respectively. However, a signi®cant portion of the initially added thiosulfate was not reduced in the methanogenic cultures by the end of the incubation period (Table 3). Signi®cantly low degradability ratios were observed for the silver thiosulfateamended culture at 100 mg Ag/l (Table 4). Table 5 shows the degree of inhibition of gas production in the 50 and 100 mg Ag/l silver thiosulfate amended cultures. Acetic acid was the predominant VFA in the silver thiosulfate-amended cultures at 50 and 100 mg Ag/l (Table 5). In contrast, only acetic acid at 0.10 mM was detected in the reference culture at the end of the 72-day incubation period. The concentrations of VFAs in the silver thiosulfate culture series at 2 and 10 mg Ag/l were very close to those of the reference culture (0.15 and 0.10 mM, respectively). Therefore, the addition of silver thiosulfate to the methanogenic cultures up to 10 mg Ag/l a€ected neither the production (i.e., acidogenesis) nor the utilization of VFAs (i.e., methanogenesis). Inhibition in terms of COD destruction was calculated by taking into account the methane produced and the amount of COD destroyed in the reference culture corrected by subtracting the equivalent amount of COD destroyed due to thiosulfate reduction in the thiosulfate-amended cultures (Table 5). The equivalent COD destruction due to thiosulfate reduction (i.e., 1 mg COD/mg thiosulfate-S reduced to sul®de) was calculated based on the following half-reaction 1=8S2 O32ÿ ‡ 6=8 H ‡ ‡ e ÿ 4 1=4S 2ÿ ‡ 3=8H2 O and the factor of 8 mg COD/milli-electron equival-

3963

ent. The calculated relatively lower degradability ratios based on methane production (Table 4) as compared to that based on the amount of COD destroyed in the thiosulfate-amended cultures is the result of thiosulfate reduction with the associated organic substrate COD destruction which in turn leads to electron channeling away from methanogenesis. The measured VFA concentrations (in equivalent COD units) in the 100 mg Ag/l culture accounted for 58% of the remaining degradable COD. By considering the sum of the VFAs±COD and methane± COD, and comparing it to the total degradable, initial COD, a 39% inhibition of acidogenesis was estimated (Table 5). The observed, but variable, inhibition of both acidogenesis and methanogenesis are related to the inhibitory e€ect of thiosulfate and more likely to that of sul®de as is discussed in the following section. All culture series amended with sodium thiosulfate had a performance similar to that of the respective, silver thiosulfate-amended cultures. Sodium thiosulfate at 20 and 100 mg S/l did not a€ect the methane production. However, an initial lag period with low methane production and signi®cant methane production inhibition and a severe methane production inhibition was observed with sodium thiosulfate at 500 and 1000 mg S/l, respectively, similar to that observed with silver thiosulfate at 50 and 100 mg Ag/l, respectively (Fig. 2). The degradability ratios, degree of inhibition based on gas production, COD destruction, and acidogenesis as well as the accumulation of VFAs were similar to those observed for the respective silver thiosulfate-amended cultures (see Tables 3, 4, and 5). Therefore, based on the silver speciation under anaerobic conditions (to be discussed in a subsequent section) and combined with the observed inhibition pattern of the sodium thiosulfate-amended cultures,

Table 5. Volatile fatty acids remaining at 72 days of incubation as well as the degree of inhibition in the silver and sodium thiosulfateamended cultures (two highest amendment levels) Parameter

Volatile fatty acids, mM Acetic Propionic Isobutyric n-Butyric Isovaleric n-Valeric Total Inhibition (%) based on: Total gas productionb Methanogenesisb COD destructionc Acidogenesis a

Ag±Thiosulfate culture at:

Na±Thiosulfate culture at:

50 mg Ag/l

100 mg Ag/l

500 mg S/l

1000 mg S/l

0.20 0.90 NDa ND 0.80 ND 1.90

22.60 2.30 0.31 0.26 0.90 0.09 26.46

0.08 0.09 ND ND 0.08 ND 0.25

25.00 2.60 0.46 0.21 1.07 ND 29.34

25 36 NAd NA

70 90 88 39

19 29 NA NA

ND, not detected. Based on the total gas or methane production of the reference culture. (CODReference, destroyedÿCODThiosulfate, reducedÿCODCH4)  100/(CODReference, destroyedÿCODThiosulfate, reduced). d NA, not available (COD was not measured for these culture series). b c

61 84 81 27

3964

Spyros G. Pavlostathis and Sung Kyu Maeng

we conclude that the observed inhibition in the silver thiosulfate-amended cultures is not related to the silver but to the sulfur (i.e., thiosulfate reduction channeling electrons away from methanogenesis and production of sul®de at toxic levels). Inhibitory e€ect of thiosulfate and sul®de In the case of the cultures amended with silverand sodium-thiosulfate at 50 mg Ag/l and 500 mg S/l, respectively, most of the methane production took place after an initial lag period of about 12 days during which time depletion of most of the initially added thiosulfate occurred. For cultures amended with 1000 mg S/l in both the silver thiosulfate and sodium thiosulfate series, low levels of methane were produced only initially (in less than 10 days; see Fig. 3), and the degradability ratios were very low (see Table 4), indicating a very high degree of inhibition. The e€ect of thiosulfate on mixed methanogenic cultures was studied by Zhang and Maekawa (1996). These researchers showed that the optimal thiosulfate concentration was 0.2±1 mM (13±64 mg S/l) and inhibition of gas production was signi®cant only at high thiosulfate levels. A thiosulfate concentration of 6 mM (384 mg S/l) was greatly inhibitory to the growth of the methanogens. Similarly, 5 mM (320 mg S/l) of thiosulfate were inhibitory to methanogenesis and the growth of two methanogens, Methanobacterium thermoautotrophicum and

Fig. 3. Silver speciation at varying sul®de (A) and silver nitrate (B) concentrations, as well as at varying redox potentials (C) based on the MINTEQA2 geochemical equilibrium speciation program under conditions representative of the methanogenic cultures used in the present study (see text).

Methanobacterium ivanov (Bhatanagar et al., 1984). Optimal growth of Methanococcus thermolithotrophicus and Methanobacterium thermoautotrophicum occurred at thiosulfate concentrations of 1±2 mM (64±128 mg S/) and 3±4 mM (192±256 mg S/l), respectively, and higher thiosulfate concentrations had no e€ect (Daniels et al., 1986). Inhibition of growth of Methanosarcina barkeri growing on methanol was not observed at a thiosulfate concentration of 10 mM (640 mg S/l) (Mazumder et al., 1986). The concentrations of thiosulfate in the silver and sodium thiosulfate series at 100 mg Ag/l and 1000 mg S/l, respectively, used in the present study were much higher than the thiosulfate concentrations found to be inhibitory to methanogens by the aforementioned researchers. Therefore, it is likely that the observed inhibition of methanogenesis in the present study was initially caused by the high thiosulfate levels. However, sul®de resulting from the reduction of thiosulfate may have further contributed to the observed inhibition as discussed below. Thiosulfate reduction resulted in total sul®de levels by the end of the incubation period (72 days) equal to 553 and 625 mg S/l in the silver and sodium thiosulfate cultures with the highest amendment level, respectively. By considering the thiosulfate levels remaining at the end of the incubation period, the total measured sul®de in the culture mixed liquor and that released as H2S with the produced gas, the fraction of the initially added sulfur which was not accounted for by these measurements was 0.12 and 0.13 for the silver- and sodiumthiosulfate amended cultures, respectively. Therefore, signi®cant levels of sul®de were produced in both the silver and sodium thiosulfate amended cultures at the two highest amendment levels (Table 3). In the case of the silver thiosulfate series, based on the concentration of silver added, only a very small fraction (<3%) of the sul®de produced was expected to be precipitated in the form of Ag2S. By considering the highest silver addition (100 mg Ag/ l) and the ferrous iron concentration of the culture media (0.5 mM), then the maximum amount of sul®de precipitated as both Ag2S and FeS is 30 mg S/l, or less than 6% of the total sul®de measured in the silver thiosulfate amended culture at 100 mg Ag/l. However, it is the free and un-ionized sul®de (H2S) which is considered to be toxic in anaerobic systems (Speece, 1996). For a culture pH of 7.2, the unionized sul®de concentration in the silver and sodium thiosulfate amended cultures with the highest amendment level was calculated as equal to 175 and 200 mg H2S(aq)/l, respectively. Inhibition of acetate consumption by methanogens was controlled by hydrogen sul®de (McCartney and Oleszkiewicz, 1991). A hydrogen sul®de concentration of 547 mg/l completely inhibited the growth of sulfate reducing bacteria (Reis et al., 1992). A wide range of sul®de concentrations lead-

Silver and the anaerobic digestion process

ing to 50% inhibition of methanogenesis has been reported (60±1200 mg H2S/l) (Speece, 1996). This variability may be explained by the di€erence in the response of the di€erent microorganisms predominant in di€erent systems, the type of anaerobic system (e.g., suspended vs attached growth), pH, metal concentrations and other conditions. Therefore, the above mentioned free, un-ionized sul®de concentrations attained in the silver and sodium thiosulfate amended cultures were in the inhibitory range for anaerobic processes which explains the observed high degree of inhibition for methanogenesis in the present study. Simulation of silver speciation In order to further explain the observed e€ect of silver compounds on the mixed, methanogenic cultures, computer simulations were carried out using the geochemical equilibrium speciation program MINTEQA2/PRODEFA2 (Allison et al., 1991) under the conditions of the methanogenic cultures used in the present study: pH ˆ 7:2, Eh ˆ ÿ80 mV and the following media composition (in mM): K+, 2+ ÿ + 14.0; HPO2ÿ , 4 , 5.2; H2PO4 , 3.7; NH4 , 9.4; Mg 2+ + ÿ 1.0; Fe , 0.5; Na , 101.0; Cl , 13.7; HCOÿ 3, 101.0; and Ca2+, 0.7 (ionic strength 1.245  10ÿ1 M). The e€ect of varying silver nitrate and sul®de concentrations as well as the culture redox potential on the silver speciation was assessed and results are shown in Fig. 3. In all cases, the free silver ion concentration (Ag+) is less than 7:976  10 ÿ15 M and the total soluble silver species concentration is less than 2:581  10 ÿ13 M. The relative distribution of silver in the predominant silver species, Ag2S and Ag8, is controlled by the available sul®de in the system. As seen in Fig. 3A, the concentration of Ag8 decreases linearly with increased sul®de concentration as a result of the formation of the highly insoluble Ag2S up until all added free silver (100 mg Ag/l) is removed (which stoichiometrically requires Table 6. MINTEQA2-calculated equilibrium silver speciationa Species Solids Ag2S Ag0 Total solids Ag Soluble Ag+ AgCl(aq) AgClÿ 2 AgCl2ÿ 3 AgCl3ÿ 4 AgHS(aq) Ag(HS)ÿ 2 AgOH(aq) Ag(OH)ÿ 2 AgNO3(aq) Total soluble Ag

Concentration (M) 3.119  10ÿ4 3.033  10ÿ4 9.271  10ÿ4 7.975  10ÿ15 9.808  10ÿ14 1.289  10ÿ13 3.981  10ÿ15 2.730  10ÿ16 1.878  10ÿ14 1.822  10ÿ23 9.277  10ÿ20 1.986  10ÿ24 2.120  10ÿ18 2.580  10ÿ13

Conditions: AgNO3=100 mg Ag/l (9.271  10ÿ4 M), Na2S=10 mg S/l, pH=7.2, Eh=ÿ80 mV, I=1.245  10ÿ1 M, and 358C.

a

3965

15 mg S/l). Similarly, for a constant sul®de concentration of 10 mg S/l, the predominant silver species is Ag2S until all sul®de is removed (which stoichiometrically requires 67.3 mg Ag/l) (see Fig. 3B), and then further addition of silver nitrate results in the formation of Ag8. For a constant silver and sul®de concentration (100 mg Ag/l and 10 mg S/l, respectively), almost equal molar concentrations of Ag2S and Ag8 are formed up until an Eh value of ÿ198 mV, beyond which almost all silver exists as Ag8. The silver speciation for the case of AgNO3=100 mg Ag/l (=0.9271 mM), Na2S=10 mg S/l, pH=7.2, Eh ˆ ÿ80 mV, I=1.245  10ÿ1 M and 358C is shown in Table 6. Based on these simulations, we conclude that under the experimental conditions of the anaerobic toxicity assay, the free silver ion concentrations were extremely low (less than 10ÿ14 M). Therefore, the observed inhibition is attributed to the thiosulfate/sul®de as discussed above and not to silver. Similar to our results, addition of silver compounds to sediments in previous studies resulted in either very low or no inhibition to freshwater organisms (Rodgers et al., 1997; Hirsch, 1998a,b; Call et al., 1999; Berry et al., 1999). These results are largely explained by the fact that ionic silver is rapidly complexed, precipitated, and sorbed to sediment components. In particular, sul®des largely control silver availability in sulfur-containing sediments as is the case with other metals (Di Toro et al., 1996).

CONCLUSIONS

In summary, neither the silver-bearing waste activated sludge generated by the aerobic biological treatment of a photoprocessing wastewater nor the silver compounds added to mixed, methanogenic cultures simulating the anaerobic digestion process resulted in any inhibitory e€ect. Due to the high complexing capacity of the anaerobic digester mixed liquor, free silver ion concentrations are extremely low. Therefore, relatively high concentrations of total silverÐat least 100 mg Ag/l, which far exceeds typical silver concentrations in sludgeÐ will not inhibit the anaerobic digestion process. Based on the results of our studies, it can be concluded that biological treatment of silver-bearing photoprocessing wastewaters is feasible by the activated sludge process followed by the anaerobic digestion of the resulting waste activated sludge without any operational problems.

AcknowledgementsÐWe gratefully acknowledge the assistance of the following Eastman Kodak personnel: T. Bober, J. Gorsuch, D. Vacco and D. Yeaw. This work was supported in part by the Eastman Kodak Company, Rochester, N.Y., USA.

3966

Spyros G. Pavlostathis and Sung Kyu Maeng REFERENCES

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