This article was downloaded by: [Canakkale Onsekiz Mart Universitesi]On: 29 October 2014, At: 14:32Publisher: Taylor & FrancisInforma Ltd Registered in England and Wales Registered Number: 1072954 Registered office: Mortimer House,37-41 Mortimer Street, London W1T 3JH, UK

Journal of Environmental Science and Health, PartA: Toxic/Hazardous Substances and EnvironmentalEngineeringPublication details, including instructions for authors and subscription information:http://www.tandfonline.com/loi/lesa20

Inhibition of Volatile Fatty Acid Production in GranularSludge from a UASB ReactorTurhan Dogan a , Orhan Ince b , Nilgun Ayman Oz a & Bahar Kasapgil Ince aa Institute of Environmental Sciences, Bogazici University , Istanbul, Turkeyb Department of Environmental Engineering , Istanbul Technical University , Istanbul, TurkeyPublished online: 06 Feb 2007.

To cite this article: Turhan Dogan , Orhan Ince , Nilgun Ayman Oz & Bahar Kasapgil Ince (2005) Inhibition of Volatile FattyAcid Production in Granular Sludge from a UASB Reactor, Journal of Environmental Science and Health, Part A: Toxic/Hazardous Substances and Environmental Engineering, 40:3, 633-644, DOI: 10.1081/ESE-200046616

To link to this article: http://dx.doi.org/10.1081/ESE-200046616

PLEASE SCROLL DOWN FOR ARTICLE

Taylor & Francis makes every effort to ensure the accuracy of all the information (the “Content”) containedin the publications on our platform. However, Taylor & Francis, our agents, and our licensors make norepresentations or warranties whatsoever as to the accuracy, completeness, or suitability for any purpose of theContent. Any opinions and views expressed in this publication are the opinions and views of the authors, andare not the views of or endorsed by Taylor & Francis. The accuracy of the Content should not be relied upon andshould be independently verified with primary sources of information. Taylor and Francis shall not be liable forany losses, actions, claims, proceedings, demands, costs, expenses, damages, and other liabilities whatsoeveror howsoever caused arising directly or indirectly in connection with, in relation to or arising out of the use ofthe Content.

This article may be used for research, teaching, and private study purposes. Any substantial or systematicreproduction, redistribution, reselling, loan, sub-licensing, systematic supply, or distribution in anyform to anyone is expressly forbidden. Terms & Conditions of access and use can be found at http://www.tandfonline.com/page/terms-and-conditions

Journal of Environmental Science and Health, 40:633–644, 2005Copyright C© Taylor & Francis Inc.ISSN: 1093-4529 (Print); 1532-4117 (Online)DOI: 10.1081/ESE-200046616

Inhibition of Volatile Fatty AcidProduction in Granular Sludgefrom a UASB Reactor

Turhan Dogan,1,† Orhan Ince,2 Nilgun Ayman Oz,1

and Bahar Kasapgil Ince1

1Bogazici University, Institute of Environmental Sciences, Istanbul, Turkey2Istanbul Technical University, Department of Environmental Engineering,Istanbul, Turkey

Inhibition of volatile fatty acids (VFA), namely acetate, butyrate, and propionate, on theactivity of acetoclastic methanogens within a full-scale upflow anaerobic sludge blan-ket (UASB) reactor was investigated using specific methanogenic activity (SMA) test.SMA tests were carried out at acetate concentrations in a range of 1000–25,000 mg l−1,butyrate concentrations in a range of 3000–25,000 mg l−1 and propionate concentra-tions between 500–10,000 mg l−1. Maximum potential methane production (PMP) rateswere obtained as 389 ml CH4 gTVS−1.d−1 at 3000 mg l−1 acetate concentration, 432 mlCH4gTVS−1·d−1 at butyrate concentration of 5000 mg l−1, and 162 mlCH4 gTVS−1.d−1

at 1000 mg l−1 propionate concentration. App. 50% and 100% inhibition occurred atacetate concentrations of 13,000 mg l−1 and 25,000 mg l−1, butyrate concentrations of15,000 mg l−1 and 25,000 mg l−1, and propionate concentrations of 3500 mg l−1 and5000 mg l−1, respectively.

Key Words: Specific methanogenic activity test; Acetate; Butyrate; Propionate; Anaero-bic sludge; VFA inhibition.

INTRODUCTION

Anaerobic waste treatment involves a complex interaction of diverse microbialpopulations. Stability of anaerobic digestion requires a balanced activity of themixed microbial population. This can be easily disturbed by different factorscausing a rapid increase in the concentration of volatile fatty acids (VFA) witha concurrent decrease in methane production.[1] Methanogens are the terminalmicrobial group and the most sensitive members of the microbial consortium

Address correspondence to Bahar Kasapgil Ince, Bogazici University, Institute of Envi-ronmental Sciences, Istanbul, Turkey; E-mail: bahar.ince@boun.edu.tr†Present address: Graduate School of Science, University of Tokyo, Japan.

Dow

nloa

ded

by [

Can

akka

le O

nsek

iz M

art U

nive

rsite

si]

at 1

4:32

29

Oct

ober

201

4

634 Dogan et al.

converting acetate and carbon dioxide into methane. Methanogens play a keyrole in the system by keeping the hydrogen partial pressure low—a conditionnecessary for the growth of many acetogenic bacteria.[2] Methane productionfrom anaerobic degradation of organic matter involves various biochemical andmicrobiological mechanisms in which VFAs are in principle intermediates.[3]

It is frequently reported that those VFAs tend to accumulate in digestersstressed, for example, by a substrate overload. Such an accumulation, resultingfrom a VFA production higher than the VFA consumption, can cause a severeinhibition of methane production. Except for acetic acid, which is directly de-graded to methane by acetoclastic methanogens, the VFA (propionic and bu-tyric) must first be degraded by obligate hydrogen-producing acetogenic bacte-ria to acetate, hydrogen, and carbon dioxide, which are utilized by methanogenicspecies.

The stability of the microbial ecosystem is very dependent on acetogenic ac-tivity since approximately two-thirds or more of methane formed during anaer-obic degradation of complex substrate results from acetic acid.[4–6] This activityis characterized by slow growth rates of acetoclastic methanogen and great sen-sitivity to inhibition processes.[7] For design and operation of anaerobic treat-ment processes, acetoclastic methanogenic “activity” is of great importance.Therefore, a significant point when starting up and operating anaerobic reac-tors is to maintain a sufficient quality of active acetoclastic methanogenic pop-ulation. Activity measurements of anaerobic sludge can be considered in twodifferent ways: an overall measurement, which gives information about thewhole degradative activity, and an activity measurement of each basic stageof the process.[8] In order to determine methanogenic activity, different tech-niques have been developed by a number of researchers.[9–11] In this study, theSMA technique developed by Monteggia[9] was used to determine inhibitioneffect that a specific substance or wastewater exerts on an anaerobic sludge.The specific methanogenic activity (SMA) test gives direct measurement of therate of methane production per unit of microbial biomass per unit of time.[12]

Therefore, this study attempts to evaluate inhibition of VFAs, namely acetate,butyrate, and propionate, on the activity of acetoclastic methanogens usingthe SMA test.

MATERIALS AND METHODS

Description of SMA Test EquipmentThe SMA test unit was initially developed by Monteggia.[9] This current

procedure used in this study was modified by Ince, Anderson, and Kasapgil.[12]

A schematic diagram of the SMA test is shown in Fig. 1. The SMA test unitconsisted of eight 1 L digestion flasks, which are placed into a water bath to con-trol the temperature stability. Mixing is provided by magnetic stirrers, which

Dow

nloa

ded

by [

Can

akka

le O

nsek

iz M

art U

nive

rsite

si]

at 1

4:32

29

Oct

ober

201

4

Inhibition of Volatile Fatty Acid Production 635

Figure 1: Experimental setup for SMA test unit.

run at a speed of 60 rpm. Gas measurement system contains a manometerand tubing for interconnection between the anaerobic reactor and the otherunits. This system has a solenoid valve, which has three ports. The valve iscontrolled with a pressure measurement device. There is a gas bulb for tem-porary storage of the gases and a line for interconnection anaerobic reactorand the units of the system. Pressure measurement device makes these portsopen or close to control the biogas pressure in the reactor. A PC connectedto the gas metering system has an eight channel analog input board modelDAS 800 (supplied by Metrabyte Corporation) which was used to simultane-ously monitor the gas production of the eight independent digesters. Detailsand the experimental procedure of the SMA test were given in a subsequentpaper.12

Seed and Feed for SMA TestsSMA test reactors were seeded with anaerobic sludge taken from a UASB

reactor treating alcohol distillery effluents. Due to the granular characteristicsof the sludge, instead of suspended solids (SS) and volatile suspended solids(VSS), total solids (TS) and total volatile solids (TVS) parameters were used.TS concentrations were 145,000 mg l−1 with a TVS/TS ratio of 0.91. Acetate,butyrate, and propionate were used as feed during the SMA tests.

Analytical MethodsGas composition was determined using a HP 6850 gas chromatograph (GC)

with a thermal conductivity detector (HP Plot Q column 30 m × 530 µm). VFAwere determined using a HP 5890 Model II gas liquid chromatograph (FFAPcolumn, 10 m × 530 µm × 1 µm). During the SMA tests pH, temperature, and

Dow

nloa

ded

by [

Can

akka

le O

nsek

iz M

art U

nive

rsite

si]

at 1

4:32

29

Oct

ober

201

4

636 Dogan et al.

Table 1: Characterization of the wastewaters containing anise seeds.

Parameter Range (mg l−1)

COD 25,000–33,000BOD5 12,000–16,000Total-N 350–450SO4− 50–100Total-P 150–250Ca++ 170–240pH∗ 5.5–6.0∗Unitless.

gas production were monitored. TS and TVS concentrations were measured.All analyses were carried out according to Standard Methods.[13]

Description of Treatment Plant at a Local Alcohol DistilleryThe wastewaters originating from the alcohol distillery have been treated

in a three-stage treatment plant. The wastewater characteristics are given inTable 1. In the first stage, wastewaters containing anise seeds have been treatedin physical and chemical treatment units. The raw wastewater containing ahigh amount of anise seeds is pumped through a screen having a pore size1 mm2 to remove the seeds. The wastewater is then collected into a tank andchemical treatment is carried out. A UASB reactor with a volume of 146 m3

has been used as second stage of treatment. Effluent from the UASB reactorcombined with domestic wastewaters is finally fed into a two-stage activatedsludge unit in order to comply with discharge standards.

Operation of the UASB ReactorThe long-term performance of this UASB reactor is shown in Fig. 2. The

UASB reactor performed well achieving COD removal efficiencies no lower

Figure 2: Changes in COD removal efficiency and OLR of UASB reactor in years 1996–2001.

Dow

nloa

ded

by [

Can

akka

le O

nsek

iz M

art U

nive

rsite

si]

at 1

4:32

29

Oct

ober

201

4

Inhibition of Volatile Fatty Acid Production 637

than 85% at OLRs in a range of 6–11 kg COD m−3 d−1 in years between 1996and 2001. The corresponding average methane yield was in a range of 0.15–0.30 m3CH4 kgCOD−1

removed.

RESULTS AND DISCUSSION

High concentrations of organic acids can inhibit anaerobic digestion. Normallyacetate, butyrate, and propionate are the predominating VFAs found in themethane reactors.[14,15] The accumulation of these intermediate acid productsleads to the inhibition of the growth of several microbial species with subse-quent decay in methane production. Although there had been several studiesabout the VFA on anaerobic reactors such as kinetics of VFAs, quantity predic-tions, etc., there is a lack of studies about effects of accumulations of VFAs onactivity of acetoclastic methanogenic archaea.[16,17]

SMA Test Results with AcetateIn first run, acetate concentrations of 1000, 2000, 3000, and 4000 mg l−1

were used. The SMA test results showed that maximum acetoclasticmethanogenic activity was found to be 214 ml CH4 gTVS−1 d−1, 311 CH4 gTVS−1

d−1, 389 CH4 gTVS−1 d−1, and 309 CH4 gTVS−1 d−1 at 1000 mg l−1, 2000 mgl−1, 3000 mg l−1, and 4000 mg l−1 acetate concentration, respectively. The max-imum potential methane production rate of enriched cultures cultivated onacetate has been reported to be approximately 1000 m3 CH4 gVSS−1 if all thebiomass (as VSS concentration) consists of acetoclastic methanogens.[18] There-fore, the PMP rate of 389 ml CH4gTVS−1 d−1 shows that nearly 39% of theUASB reactor sludge was composed of acetoclastic methanogens. When takinginto consideration the proportion of the reactor sludge such as TVS/TS ratioof 0.91, sludge volume index 15 ml g−1 TVS and comparing PMP rates of theanaerobic sludges (>200 ml CH4 gVSS−1 d−1) treating similar wastewaters inliterature, it can be said that the sludge has a good quality.[19] All SMA test re-sults of acetate concentrations between 1000–25,000 mg l−1 are plotted in Fig. 3.Decreases in PMP rates were considered as substrate inhibition on acetoclasticmethanogens. 4000 mg l−1 acetate concentration was considered as the begin-ning of inhibition. Further increases in acetate concentration from 4000 mg l−1

to 25,000 mg l−1 resulted in decreases in the acetoclastic methanogenic activ-ity. Increases in acetate concentrations resulted in PMPs rate to be reached atlonger test periods (Fig. 3). Figure 3 also shows that 50% inhibition occurredat acetate concentration of 13,000 mg l−1. Finally, acetoclastic methanogenicactivity of the anaerobic sludge ceased at acetate concentration of 25,000 mgl−1. According to the results of similar inhibition studies in literature, acetatehad been observed to be inhibitory at a concentration of 35,000 mg l−1.[20] Theanaerobic sludge used in our study was fully inhibited with acetate in relativelylower concentration than reported.

Dow

nloa

ded

by [

Can

akka

le O

nsek

iz M

art U

nive

rsite

si]

at 1

4:32

29

Oct

ober

201

4

638 Dogan et al.

Figure 3: Changes in methanogenic activity with different acetate concentrations usingSMA test.

Dow

nloa

ded

by [

Can

akka

le O

nsek

iz M

art U

nive

rsite

si]

at 1

4:32

29

Oct

ober

201

4

Inhibition of Volatile Fatty Acid Production 639

SMA Test Results with ButyrateAs a result of SMA tests that were conducted with butyrate concentration in

the range of 3000 mg l−1–25,000 mg l−1, it can be said that 5000 mg l−1 concen-tration of butyrate resulted in a maximum acetoclastic methanogenic activityof 432 ml CH4 gTVS−1 d−1. SMA test results of all butyrate concentrations,3000–25,000 mg l−1 are shown in Fig. 4. 3000 mg l−1 and 4000 mg l−1 butyrateconcentrations were found to be substrate limited. As shown in Fig. 4, 6000 mgl−1 butyrate concentration was the beginning of butyrate inhibition, thus higherconcentrations were tested for further investigation. Substrate amount was notrestricted with these concentrations of butyrate; any fall on the rate of PMP wasnow considered as inhibition due to excess butyrate concentration. Increasesin butyrate concentration above 5000 mg l−1 resulted in increases in degreeof inhibition and acetoclastic methanogenic activity was totally inhibited at25,000 mg l−1 butyrate concentration. At each butyrate concentration tested,different lengths of time to reach potential methanogenic activity and comple-tion of the tests were observed. It has been reported that significant inhibitoryconcentration of butyrate was found to be 15,000 mg l−1, which corresspondsto 50% inhibiton of the acetoclastic methanogenic population in the anaerobicsludge used in this study.[20] SMA test results of butyrate were in agreementwith results reported in the literature.

SMA Test Results with PropionateAs a result of SMA tests with propionate concentrations between 500–

10,000 mg l−1, 1000 mg l−1 concentration of propionate was found to be op-timum to reach maximum acetoclastic methanogenic activity, i.e., 162 ml CH4

gTVS−1 d−1. Decreases of PMP rate due to increase in propionate concentra-tions over 1000 mg l−1 were considered as beginning of inhibition. SMA testvalues with propionate concentrations between 500–6,000 mg l−1 are shownin Fig. 5. As seen, 2000 mg l−1 propionate concentration was the beginningof inhibition, thus for further investigation, higher concentrations were alsoapplied. 500 mg l−1 propionate concentration was considered to be insufficientsubstrate. Although it was reported that propionate concentration over 3000 mgl−1was critical,[20] the SMA test results with over 3000 mg l−1 propionate cor-responded to vital inhibition on potential acetoclastic methanogenic activity.Thus, it can be said that the anaerobic sludge used in this study was inhibitedwith propionate concentration close to the one stated in literature.

SMA test results shown in Fig. 6 indicated that propionate inhibition oc-curred at relatively lower concentrations than butyrate and acetate. Even-numbered carbon fatty acids were degraded more easily than odd-numberedones. This is related with the fermentation step where all VFAs are degradedto acetate. Even-numbered VFAs can be degraded more easily to acetate thanodd-numbered ones.[21] Results found for acetate and propionate in a study that

Dow

nloa

ded

by [

Can

akka

le O

nsek

iz M

art U

nive

rsite

si]

at 1

4:32

29

Oct

ober

201

4

640 Dogan et al.

Figure 4: Changes in methanogenic activity with different butyrate concentrations usingSMA test.

used aerobic waste-activated sludge for start-up seed of mesophilic and ther-mophilic anaerobic digestion correlate results of this study.[22] Results of thisstudy also match with the ones observed in a study that investigated perfor-mance of UASB reactor treating leachate from acidogenic fermenter in two-phase anaerobic digestion of food waste. The propionate concentration in the

Dow

nloa

ded

by [

Can

akka

le O

nsek

iz M

art U

nive

rsite

si]

at 1

4:32

29

Oct

ober

201

4

Inhibition of Volatile Fatty Acid Production 641

Figure 5: Changes in methanogenic activity with different propionate concentrations usingSMA test.

effluent was higher than any other acid. Among VFAs, propionate is known tohave the lowest tolerance level for anaerobic populations. When an anaerobictreatment system is overloaded, propionate tends to accumulate in the reac-tor and its removal is difficult during recovery. The degradation of propionateto acetate is thermodynamically infeasible unless the by-product hydrogen isremoved by the hydrogen-consuming bacteria.[21]

It has been reported that substrate inhibition of acetate degradation in apilot study was weak.[14] This is in accordance with other studies.[15,23] Pro-pionate, on the contrary, is frequently regarded as a stronger inhibitor.[24] Inpropionate inhibition tests, it was reported that the degradation of propionateis gradually inhibited concentrations over 800 mg l−1 with pH 7 and acetatecould be degraded with concentration below 2500 mg l−1.[14] In another study onVFA inhibition in anaerobic reactors, it is reported that propionate degradationtook a longer time compared to acetate and butyrate at same concentrations. It

Dow

nloa

ded

by [

Can

akka

le O

nsek

iz M

art U

nive

rsite

si]

at 1

4:32

29

Oct

ober

201

4

642 Dogan et al.

Figure 6: Overall specific methanogenic activity test results with different acetate,butyrate, and propionate concentrations.

was further reported that degree of inhibition of propionate was greater thanthat of other VFAs.[15] These results are also compatible with the results of thisstudy. It has been reported that the best conditions to produce methane wereobtained at the optimum acetate concentrations of 4500–6000 mg l−1, which isalmost parallel to results of this study.[25]

CONCLUSIONS

The SMA test results drawn from this study are comparable with the valuesreported in literature. However, variations in degree of inhibitions of acetate,

Dow

nloa

ded

by [

Can

akka

le O

nsek

iz M

art U

nive

rsite

si]

at 1

4:32

29

Oct

ober

201

4

Inhibition of Volatile Fatty Acid Production 643

butyrate, and propionate on methanogenic archaea could be due to the evalua-tion method adopted in the study. VFA inhibitions on methanogens in anaero-bic reactors are mainly evaluated based on actual methane production (AMP)rates. Considering only AMP rates cannot show potential inhibition effects ofVFAs on acetoclastic methanogens. However, SMA tests carried out at optimumconditions for acetoclastic methanogens determine potential methane produc-tion rates of these species. The SMA test can, therefore, be effectively used forevaluation of inhibition studies.

ACKNOWLEDGMENTS

Author Bahar Kasapgil Ince would like to acknowledge the project coded 01S101of the Research Fund of Bogazici University. Author Orhan Ince would also liketo acknowledge the project coded 844 of the Research Fund of Istanbul TechnicalUniversity.

REFERENCES

1. Ahring B.K.; Westermann P. Toxicity of heavy metals to thermophilic anaerobicdigestion. Eur. J. Appl. Microbiol. Biotechnology 1983, 17, 365–370.

2. Harper, S.R.; Pohland, F.G. Recent developments in hydrogen management duringanaerobic wastewater treatment. Biotechnol. Bioeng. 1986, 27, 585–602.

3. McCarty, P.L.; Mosey, F.E. Modeling of anaerobic processes. Water Sci. Technol. 1991,24 (8), 17–33.

4. Gujer, W.; Zehnder, A.J.B. Conversion processes in anaerobic digestion. Water Sci.Technol. 1983, 15, 49–77.

5. Fukuzaki, S.; Nishio, N.; Nagai, S. Kinetics of the methanogenic fermentation ofacetate. Appl. Environ. Microbiol. 1990, 56, 3158–3163.

6. Zinder, S.H. Physiological ecology of methanogens. In Methanogenesis: Ecology,Physiology, Biochemistry and Genetics; Ferry, G., Ed., Chapman & Hall: New York, 1993;128–206.

7. Boone, D.R. Terminal reactions in the anaerobic digestion of animal waste. Appl.Environ. Microbiol. 1982, 43, 57–64.

8. Soto, M.; Mendez, R.; Lema, J.M. Methanogenic and non-methanogenic activ-ity tests theoretical basis and experimental set-up. Water Res. 1993, 27 (8), 1361–1376.

9. Monteggia, L. The use of a specific methanogenic activity test controlling anaerobicreactors. Ph.D. diss. University of Newcastle upon Tyne, 1991.

10. James, A.; Chernicharo, C.A.L.; Campos, C.M.M. The development of a new method-ology for the assessment of specific methanogenic activity. Water Res. 1990 24 (7), 813–825.

11. Chiang, C.F.; Dague, R.R. Determination of acetoclastic methanogenic activity inanaerobic systems. In Proceedings of the 43rd Purdue Industrial Waste Conference, Pur-due University, 1989; 353–361.

Dow

nloa

ded

by [

Can

akka

le O

nsek

iz M

art U

nive

rsite

si]

at 1

4:32

29

Oct

ober

201

4

644 Dogan et al.

12. Ince, O.; Anderson G.K.; Kasapgil, B. Control of olr using the specific methanogenicactivity test during start-up of an anaerobic digestion system. Water Res. 1995, 29 (1),349–355.

13. Standard Methods for the Examination of Water and Wastewater. American PublicHealth Association/American Water Works Association/Water Environment Federation:Washington, DC, USA, 1995.

14. Mosche, M.; Jordening, H.J. Detection of very low saturation constants in anaer-obic digestion: Influences of calcium carbonate precipitation and pH. Appl. Microbiol.Biotechnol. 1998, 49, 793–799.

15. Aguilar, A.; Casas, C.; Lema, L.M. Degradation of volatile fatty acids by differentlyenriched methanogenic cultures: Kinetics and Inhibition. Water Res. 1995, 29, 505–509.

16. Kaspar, H.F.; Wuhrmann, K. Product inhibition in sludge digestion. Microbiol. Ecol.1978 4, 241–248.

17. Alonso, M.S. Influence of the acetate concentration on the recovery time of a per-turbed anaerobic digester. Biotechnol. Lett. 1992, 14, 535–538.

18. Valcke, D.; Verstraete, W. A practical method to estimate the acetolasticmethanogenic biomass in anaerobic reactors. J. Water Pollut. Con. F. 1983, 55, 1191–1195.

19. Ince, O.; Anderson, G.K.; Kasapgil, B. Use of the specific methanogenic activity testfor controlling the stability and performance in anaerobic digestion of brewery wastew-ater. In Proceedings of the 49th Purdue Industrial Waste Conference, Purdue University,West Lafayette, IN, USA 1994; 189–204.

20. Ianotti, E.L.; Fischer. J.R. Effects of ammonia, volatile acids, ph and sodium ongrowth of bacteria isolated from a swine manure digester. Proceedings of the 40th GeneralMeeting Society of Industrial Microbiology, Sarasota, Florida, USA, 1983, Vol. 1, pp. 14–19.

21. Boone D.R.; Xun L. Effects of pH, temperature, and nutrients on propionate degra-dation by a methanogenic enrichment culture. Appl. Environ. Microbiol. 1987, 53 (7),1589–1592.

22. Kim, M.; Speece, R.E. Aerobic waste activated sludge (was) for start-up seed ofmesophilic and thermophilic anaerobic digestion. Water Res. 2002, 36, 3860–3866.

23. Witty, W.; Markl, H. Process engineering aspects of methanogenic fermentationon the examile of fermentation of pencillium mycelium. German Chem. Eng. 1986, 9,238–245.

24. Barredo, M.S.; Evison, L.M. Effect of propionate toxicity on methanogen-enrichedsludge, Methanobrevibacter smithii, and Methanospirillum hungatii at different pH val-ues. Appl. Environ. Microbiol. 1991, 57, 1764–1769.

25. Munoz, M.A.; Sanchez, J.M.; Rodrıguez-Maroto, J.M.; Borrego, J.J.; Morinigo, M.A.Methane production in anaerobic sludges supplemented with two support materials anddifferent levels of acetate and sulphate. Water Res. 1997, 31 (5), 1236–1242.

Dow

nloa

ded

by [

Can

akka

le O

nsek

iz M

art U

nive

rsite

si]

at 1

4:32

29

Oct

ober

201

4