ART. CELLS, BLOOD SUBS., AND IMMOB. BIOTECH.

, 27(5&6), 4 6 1 4 7 (1999)
Artif Cells Blood Substit Immobil Biotechnol Downloaded from informahealthcare.com by National Environmental Engineering Research Institute on 07/03/12 For personal use only.

Biphasic Biomethanation of Wood-Hydrolysate Effluent

S.K. Chakrobarii, P.K. Roychoudhury * and P.K. Bojpai Thapar Cenlrefor bduslrlal Research & Deveiopmenl Bhadson Rood, Paliala - 147 001, Indio *Departmen/ of Biochemical Engineering dr Blolechnology Indian Inslllule of Technology New Delhl- 110 016. India
ABSTRACT

The dissolving pulp industry, spread throughout the world, is the principal source of wood-hydrolysate effluent rich in hemicelluloses. This effluent is the major source of pollution in the industry. COD and BODI values of the effluent range from 60,000 to 103,000 and 42,000 to 78,000 mg/l respectively. Biomethanation of this effluent is the best possible treatment option for reducing the COD load and recovering the bioenergy embedded in the effluent. This paper deals with the study on the biphasic biomethanation of the wood-hydrolysate in upflow acidogenic reactor coupled with anaerobic tiller methanogenic reactor. The two reactors were operated at organic loading rates of 69.6 and 30.1 g CODlVd respectively. The overall COD, hemicelluloses and lignin reductions, and methane generation were observed to be 88%, 92%, 82% and 6.5 VI reactor volume/d respectively. The relative size of the biphasic, anaerobic respectively. filter (mono-phasic) and upflow anaerobic sludge blanket (mono-phasic) reactors is found to be I :1.6:2.03
Key wordr

Wood-hydrolysate emuent, Dissolving pulp industry. Upflow acid reactor. Anaerobic filter methanogenic reactor, Methane yield, COD reduction, Hemicelluloses reduction, Lignin reduction.
INTRODUCTION

Since the tint commercial scale production of dissolving grade pulp in 1903 in Norway, the industry has grown throughout the world. The present global production of dissolving grade pulp is about 3 million tonnes per annum and half of it is produced by the haft process (International Pulp & Paper Directory, 1994). The wood-hydrolysate, which is rich in hemicelluloses and is generated at the rate of 5-6 m' per ton of pulp, is the major source of water pollution in the dissolving pulp industry. About 90% of the COD load in the pulp mill comes from the wood-hydrolysate itself. The major constituents of the wood-hydrolysate are hemicelluloses, acid soluble low molecular weight lignins and volatile fatty acids formed from the hemicelluloses moiety by autohydrolysis (Chakrabarti et al., 1995). These three components together mg/l) and amibute very high COD (60,000-103,000 BOD, (42,000-78,000 mg/l) to the effluent. Among the various treatment options that might be considered for treatment of wood-hydrolysate, biomethanation is the best one in respect of pollution abatement and energy recovery (Chakrabarti, 1998).
A few studies have been carried out on the anaerobic treatment of wood-hydrolysate effluent in mono-phasic anaerobic reactors (Good et al., 1982;McCarty et al., 1983;Chakrabarti et al., 1991 and 1995). Chakrabarti et al. (1995)reported maximum organic loading rates (OLR) of 11.6 and 14.7g COD/Vd that were attained during the prolonged study in UASB and anaerobic filter (AF) reactors respectively.

Separation of acidogenic and methanogenic reactions for biomethanation of different substrates is being carried out in different laboratories and in commercial installations for getting stability in the anaerobic process, and product advantages (Ghosh and Pohland, 1974;Cohen et al., 1979;Cohen et al.. 1984;Weiland and Thomsen. 1990). The objective of the present study is, therefore, set to reveal the biochemical complexity of phase separation of a complex substrate like wood-hydrolysate in order to develop an advanced technology for its treatment.
MATERIALS AND METHODS

Reactors: The upflow acidogenic reactor (UFR) is made of Plexiglas had an effective volume of 7.0 litre. It had the circular cross section and side feed entry arrangement. The anaerobic filter methanogenic reactor (AFM) is made of mild steel with exterior surface epoxy coated had an effective volume of 7.0 litre. Feed was distributed in the bottom of the reactor in the same way as in the case of the acidogenic reactor. Cross flow filter media (FB 10 27) was used in the AFM reactor. The reactors were placed in a thermostatically controlled room at 37f3"C. Inoculum: Anaerobically digested municipal sewage sludge (MSW) rich in acidogenic and methanogenic bacteria (Weiland, 1987) was used in both the reactors as the source of microbial biomass. Substrate: Wood-hydrolysate was treated with quick lime (available CaO : 65-70%) and supplemented with

46 1
Copyright Q 1999 by Marcel Dekker. Inc www.dekker.com

462
Artif Cells Blood Substit Immobil Biotechnol Downloaded from informahealthcare.com by National Environmental Engineering Research Institute on 07/03/12 For personal use only.
nutrients and micronutrients: nitrogen and phosphorus in the proportion o f I.O g : 0.2 g per 100 g of COD; Na, Mg, Fe concentrations o f 100, 5 and 2.0mg/l respectively, and Co, Ni, Mn, Zn, B,Cu concentrations of 1 mg/l each. Analytical methods Hemicelluloses were estimated by the method suggested by Deschatelets and Yu (1986). Lignin was estimated by the method o f Johnson et al., 1961. Volarile /a//y acids (VFA) were determined by CC technique using chromosorb 101 glass column with flame ionization detector; temperature of oven, injector and detector were maintained at 210, 230 and 2 5 0 T respectively; carrier gas (N2) flow was 30 mllmin. Chemical oxygen demand (COD) of centrifuged samples (at 4000 rpm) was determined by the colourimetric method as per HACH method no. 435. Biogus volirme was determined by downward displacement of acidified water ( 5 % wiv H2S04)with the help of a specially designed apparatus made of Plexiglas. Composirion o biogas was determined by f , GC technique and orsat apparatus; H,, N O2 and CHI were estimated by the former and C 0 2 by the latter. In GC, molecular sieve column (13X) and thermal conductivity detector was used, and temperature of oven, injector and detector was maintained at 90. 100 and I00"C respectively. Argon (30 ml/min) was used as carrier gas. RESULTS AND DISCUSSION Optimuni acidification in the first stage (or first phase) is the essential step for smooth operation of biphasic anaerobic treatment of soluble substrates. Acidification of wood-hydrolysatc Acidification of any substrate i s dependent upon environmental and operating conditions such as pH. temperature, hydraulic residence time, feed concentration and organic loading rate. The present study was conducted under mesophillic condition (37+30C). E/fect ofpH Acidification of wood-hydrolysate, which was carried out in batch mode with acidogenic bacterial biomass. grown by suppressing the growth o f methanogenes, The was found to depend on pH (Table I ) . acidification peaked in the pH range of 4.5-5.5. This has some similarity with that o f starch (pH 5.0) (Mudrack and Kunst, 1982) whereas, acidificalion of glucose was observed to be optimum at pH 6.0 (Cohen et al., 1979).

CHAKRABARTI, ROYCHOUDHURY, AND BAJPAI

rable I
Sr. Nu

Effect o f pH on acidification o f woodTold

Initial Acidificalion
( "10 1

hydrulysale pH AA BA range (rngil) (mgll)

COD
from acids

COD
(mgil)

(nigll)
I 2 3 4 5 6

4.0-4 4 5-5

s
o

5 0-5 5 5 5-60 6.0-6.5 6 5-70

1350 1621 1673 1196 YE

381 441
400 372

304
212

841

2150 2~50 2525 1964 1550 1290

sono
5000 5000
5000 5000 5000

41

51 n 50 5 393

31

25 8

Effect ofoperating condirions on acidi$cu/ton

Acidification of wood-hydrolysate was studied using simple upflow reactor to minimize the cost o f acidification reactor and to avoid operational hassles (Alexiou et al.. 1994). The continuous acidification reaction was studied for 331 days. Preacclimatized MSW sludge having 184.4 g total suspended solids and 77.4 g volatile suspended solids were taken in the reactor. Initial COD concentration was 10 pi1 when the pi1 was adjusted to 5.0 and duration o f batch mode o f operation was 7 days. During the 33 I days run, HRT and COD were changed from 1.16 to 0.31 days and 10.3 to 40.5 g/l respectively (Fig. I). Accordingly. organic loading rate varied from 9 2 to 92.6 g COD/l/d.

111

110 lW

110 (10 S YD N

Il*ytO

Fi*I:V~io&d

'

b i r u - d m e K

Even though the acidification reaction was started with a higher COD concentration and OLR, a higher methane content in the biogas was observed (Table 2). Similar observation has also been reported elsewhere (Alan et al., 1985; Dinopoulou and Lester, 1989). This of phenomenon continued upto an OLR 35. I g CODIVd. Even under extremely unfavourable conditions like low HRT, low pH and high feed concentration, growth of methanogens could not be suppressed because of strong symbiotic relationship between methanogens and acidogens. COD reduction was found to have an inverse relationship with the organic loading as indicated by Fig. 2.

BlPHASlC BIOMETHANATION
Artif Cells Blood Substit Immobil Biotechnol Downloaded from informahealthcare.com by National Environmental Engineering Research Institute on 07/03/12 For personal use only.

463
attached to lignin (at 1712 cm-l) and aromatic C-H bond (at I 5 10 cm-I) disappeared.

_ L

.
-.
I

No statistical correlation has been observed between the degree o f acidification and OLR. The value o f r'
was obtained to be 0.0215. This is in contrast with the observations o f other researchers who have reported inverse relationship o f acidification with OLR (Dinopoulou et al.. 1988). The striking feature o f the acidogenesis is that in spite o f frequent changes in OLR during the operation. it maintained a constant degree of acidification, which i s desirable for long term operation o f the reactor under actual conditions. Substrate removal rate constant k, which is a measure o f the removal of COD from the reactor during acidogenic reaction, should be independent of' OLR in an ideal case. But it has been observed that k has a negative relationship with OLR (Fig. 3). A close look at the value o f k indicates that i t is high as long as methanogenesis is significant. As the acidogenesis progresses, the value of k gradually decreased. I t has also been noticed that whenever the organic load in the reactor was reduced, the value of k instantaneously decreased. With respect to COD balance during the acidification o f wood-hydrolysate. i t has been found that COD loss through the generation of microbial biomass lies between 9.6-15.2% o f the feed COD Though statistical significance was not observed between the COD loss through sludge generation and organic loading rate, an inverse relationship exists between the two (Fig. 4).

0 0

.
25

.. .

50 75 OLR (g COD/lld) Fig. 2: Effect of OLR on performance of

100

acidogenic reactor 82.6-98.5% reduction of hemicelluloses took place during the first leg of increase in organic load. An inverse relationship was observed between hemicelluloses reduction and OLR (P = 0,001). I t is apparent that for a system where hydrolysis is not a rate limiting step, the break-down o f hemicelluloses to low molecular fractions can be attained by acidogenic culture. Low molecular weight soluble lignin, which is an important constituent o f the wood-hydrolysate, was reduced by 3567.4%. An inverse relationship was observed between lignin degradation and organic loading in the acidogenic reactor. Regression analysis o f lignin reduction against OLR points out that the latter is statistically significant (P = 0.01).
Tahlc 2 Hioga generalion and 11) cornpositloll

Sr

01.11

Ihogic\ (I/d)

Biueas consiiiueiil

L%]
SD
05 34 S4 29 47 31 83 61 37

No

1eCOI) /lid)
Valuc

SD

Hi Value
42 04 5 1 0 0

SD
03 03 2a 0 0 10 (4 40 24

CHI Value
371

07

.
40

.
07 25 73

3
4
5

6 7

a v
10 II 12 I3 14 IT 16

157 224 351 I(, I 324 50s 721 967 710 541 607 509 625 51 5 730

123
249 in0

23 01

an
70

157 1x7

.
27 33

IIO 98
88 74 73

172 152
168

14

1s

160 I64

31 I I
30

94
I05

13 16

362 390 464 667 114 16s 57 93 45 65 4 6 33 26

06

I.o
h I

09

OR

07 05

O . 7

;*:
25

(.:,
50
75
01.R (g CODllld)

*,
( ? n.os9)

IhY

.
I 5

103

1 1

oa

0.0

ioo

125

Failure o f mono-phasic reactors at a moderately high OLR, as reported by Chakrabarti et al. (1995) might be due to the toxic effect o f lignin or their derivatives as suggested by Sierra-Alvarez and Lettinga (1991). but these compounds do not inhibit acidogenesis upto the level o f organic loading used. The present study indicates that acidogenesis reduces molecular weights o f both hemicelluloses and lignins. Structural modification has also been observed in the lignin molecules during acidogenesis, e.g., carboxyl group

Fig. 3: Ef'feci oforganic loadingon firs1 order reaction rate constanf. k It happened due to the fact that both acidogenic and methanogenic bacteria are very strong in the initial stage o f the acidification, giving an upward push in the generation o f microbial sludge. A strong inverse correlationship has been observed between the COD loss through gas generation and OLR (Fig. 4). This establishes the fact that acidogenesis results in very low loss o f COD through gas generation; in the

464

CHAKRABARTI, ROYCHOUDHURY, AND BAJPAI

Artif Cells Blood Substit Immobil Biotechnol Downloaded from informahealthcare.com by National Environmental Engineering Research Institute on 07/03/12 For personal use only.

advanced state of acidogenesis. the loss is only 0.I0.2%. 30 ' COD loss (%)-pas (r2: 0.584) ' COD loss(%)-S!udge (r2:0.023)
20
10

g
z i;

0
-10

25

50

75
hhip((di

OLR (g COD/l/d) Fig. 4 : Effect of OLR on COD loss during acidogenic reaction Methanogenesis of emuent acidified wood-hydrolysate

Fig. 5: Variation in operating and envirOnmentalp m e t e n during methanogenesis (a) during start-up with synthetic acid m x u e itr (b) during operation w t acidified wood-hydrolysate ih reduction against the change of OLR (upto 30. I g COD/l/d) shows a direct relationship. Reduction (94.6-98.4%) of volatile fatty acids by methanogenic consortia conclusively indicates the growth of both acetogenic as well as methanogenic bacteria. Methanogenesis of acidic effluent having low to moderately high hemicelluloses content was studied

For faster start-up and growth of methanogenic consortia, synthetic acid mixture comprising of acetic, propionic and butyric acids in the COD proportion of 50:25:25 was used. The three acids were chosen as they are the major acid products in the acidogenic effluent of wood-hydrolysate. Acetic acid proportion was kept higher to generate Merhonosarcina and Methanofhrir in larger proportion; propionic and butyric acids help in the growth of acetogenic bacteria. Initially, after pouring the MSW sludge, the reactor was fed with volatile acid mixture. The resultant COD was kept at 2 gA and pH at 7.5. The reactor was kept in the batch mode for 10 days. This resulted in partial acclimatization of the methanogenic and acetogenic bacteria. Thus, when the reactor was started in the continuous mode at OLR of I .7 g COD/l/d (Fig. 5), it attained about 40% COD reduction within 10 days. Within next 110 days, the reactor biomass attained a capacity of 80% COD reduction when it was under operation at 7.62 g COD/l/d OLR. From 121 day of continuous run, substrate was changed to the acidified effluent of the first phase reactor while reducing the OLR from 7.62 to 4.57 g COD/l/d. A marginal fall in COD reduction (from 80 to 71%) was noticed in this condition. But the adaptation of microorganism did not take a long time. The faster start-up and adaptation helped the reactor to attain an OLR of 30.I g COD/l/d within 200 days of the continuous run. The reactor was kept under dormant condition for 24 days to observe the biomass behaviour. The recovery was very fast; at relatively lower OLR of 8.3g COD/l/d the reactor could attain 75% COD reduction within 5 days of operation. On 240th day, the organic loading rate was 18.8g COD/l/d. During the entire period of the study, the COD reduction ranged from 75 to 86.3% (Fig. 6). Most of the time the COD reduction was more than 80%. This shows the high stability of the biphasic methanogenic reactor with anaerobic filter configuration. The regression analysis of the COD

E 100-

COD redr. (r? 0.198) ' Acid redn. (r2: 0.079) ' Ligninredn. (r2: 0.01 I )

G a
50" 8

,
*

. .. . .
I

.-

20 30 40 OLR (g COD/l/d) Fig. 6: Effect of OLR on performance AFM reactor during the entire run. Reduction of hemicelluloses varied from 42.3 to 56.4%. This establishes that once the major amount of hemicelluloses is converted to volatile fatty acids in the acidogenic reactor, residual hemicelluloses can Further be reduced by the methanogenic-acidogenic consortia in the methane reactor. It is also noteworthy that minor amount of acidogenic bacteria alongwith methanogenic bacteria in the second phase (methanogenic) reactor are eventually beneficial for the overall treatment. Methanogenic consortia have also been found to degrade a substantial amount of lignin molecules. The lignin degrading ability of the methanogens is quite uniform at all levels of organic loading. A significant structural change of the lignin molecules has been observed during the methanogenesis. This is one of the most significant
0

10

BIPHASIC BIOMETHANATION
Artif Cells Blood Substit Immobil Biotechnol Downloaded from informahealthcare.com by National Environmental Engineering Research Institute on 07/03/12 For personal use only.
features of the phase separated anaerobic treatment of wood-hydrolysate wherein large amount of lignin molecules get degraded. It has also been established that low molecular weight lignin molecules (below 1000 Daltons) of forest based sources are not inhibitory to methanogenic bacteria once these are modified by acidogenic bacteria in the first phase of the biphasic reactor system. Biogas generated out of the methanogenesis is highly rich in methane, the concentration of which varied from 75.7 to 89.5% (Table 3 ). Methane content has an inverse relationship with the organic loading rate (Fig. 7). Specific methane yield (I CH4/I reactor volume/d), which is dependent on the generation of biogas as well as methane content in it, increases sharply with the increase of OLR. At the highest OLR of 30.1 g CODIVd, the specific methane yield was observed to be 9.77 VI reactor volume/d.
Table 3

465
the faster growth of methanogenic bacteria in the lower substrate concentration and the extremely low value of Ks for aceticlastic and carbon dioxide reducing methanogens (Pohland, 1992). At the peak loading rate of 30.1 g COD/l/d, the COD loss by sludge generation is only 6.7% based on input and 8.0% based on COD removed. Optimization of biphasic biomethanation of woodhydrolysste The volume of the acidogenic and methanogenic reactors has been selected based on the optimum hydraulic retention times and organic loading rates in the acidogenic and methanogenic reactors which are 0.42 and 0.82 d, and 69.6 and 30.1 gCOD/l/d respectively. The optimum organic loading rate in the acidogenic ractor has been selected based on the degree of acidification, reduction of hemicelluloses and lignin of wood-hydrolysate, and minimum COD loss through biogas. The overall performance of the biphasic reactor system has been computed to arrive at an optimum design ofthe reactor system (Table 4).

Biogas generation in AFM composition

reactor and its

Sr No
1

OLR
(gCOD/l/d)

Biogas (I/d) Value SD

2 3 4 5 6
7

8 9
10

457 680 950 I I 60 1340 1910 2360 2070 3010 I

104 167 280 31 6 388 521 709 592 894

D

077
135

CHI in biogas (%) Value SD 895 18

48 06 08 21 35 23

822 790 81 I

31
17 II lo 34 41 45 I5

100

901

. .
'

*. .

812
825 757 776 765
E

32
L

II 12

83
188

193 55 I

31

I7

806 81 3

08 09
-0

10

a
h

10090-

OLR (g CODIIW)

io

3'0

40

Methanein biogas

Methane yield
(r': 0.995

p; '

0.506)

9 0 I

70.50-01

y.

Fig 8 Etlect o f organic loading on COD released(blow) and COD removcd (sludge) during methanogenesin CODreleased * CODremo~da BS biogas (%) an sludge (%)on mpui inpul C O D h i s ( ' ) COD basis(') CODrcleased CODremowdas as b l o w (%) on sludge (%) on reduced reduced COD baris Ibi COD basis Id)

CONCLUSION
The biphasic reactor system comprising of upflow reactor and upflow filter reactor as acidogenic and methanogenic reactors respectively offers a better reactor configuration compared to mono-phasic UASB and AF reactors for the anaerobic treatment of woodhydrolysate emuent. Optimum hydraulic retention times and organic loading rates in the acidogenic and methanogenic reactors were 0.42 and 0.82 d. and 69.6 and 30. I g COD/l/d respectively. Acidogenic reactor 13.8% COD and converts 82.5% reduced hernicelluloses and 48.8% lignin to low molecular weight compounds. Methanogenic reactor reduces 86% COD resulting in 9.8 I methanell reactor volume/d with 75.8% methane in the biogas. The overall reductions of COD, hemicelluloses and lignin were 88%. 92% and 82% respectively and specific methane

Fig. 8 gives an account of COD balance in the methanogenic reactor. Larger amount of COD is released from the reactor as CHI and only a part of COD is utilized for cell syntheses. 70-77% of input COD or 86-93.5% of COD removed is released as biogas whereas, 6.5-10.8% of input COD or 7.3-12.7% of COD removed is converted to biosludge during the methanogenesis of acidified effluent of woodhydrolysate. The inverse relationship between the rate of release of COD as biogas and that of removal of COD as hiosludge with the increase of OLR is due to

466
Artif Cells Blood Substit Immobil Biotechnol Downloaded from informahealthcare.com by National Environmental Engineering Research Institute on 07/03/12 For personal use only.
generation was 6.5 I reactor volumeld. l h e relative size lof the biphasic, AF (mono-phasic) and UASB (mono-phasic) reactors was found to be I : 162.03 respectively.
lahle 4 Optimum conditions in the biphasic reacloi system Parameter Valuc Reactor vulun~c. I 20 7 Overall HRT. d I24 OLR. g COD/l/d 23 5 Inlet COD concentration. %/I 29 2 Effluent COD concentration, g/l 3 45 COD reduction. % 2 llcmicelluloses reduction. % 92 5 Lignin rcductioii. % 82 3 Specific methane yield. 1/1 reactur volumcid 6 5 Meihanc conteni i n tlie biugar. % 75 8 Overal hiogas un COD reinoval basis, RI 4 17 n COI) removal as sludge. % Volume ratio of mcthanogenic reactor to I Y5 I acidogenic reactor

CHAKRABARTI, ROYCHOUDHURY, A N D BAJPAI

Cohen, A,, Van Gemen, J.M., Zoetemeyer, R.J. and Main characteristics and Breure, A.M. (1984). strichiometric aspects o f acidogenesis o f soluble Prucrss carbohydrates containing wastewaters. Biocheni.. 19:228-232. Deschatelets, L. and Yu, E.K.C. (1986). A simple pentose assay for biomass conversion studies. Appl. Mcrobiol. Biotechnul, 24:379-385. Dinopoulou. G., Rudd, T. and Lester, J.N. (1988). Anaerobic acidogenesis o f a complex wastewater: I The influence o f operational parameters on reactor performance. Biotechnd. Bioengn. 3 1958-968. Dinopoulou, G. and Lester, J.N. (1989). Optimization o f a hvo phase anaerobic digestion system treating a complex wastewater. Environni. Technol Leu. I0:799-8 14. Chosh, S. and Pohland, F.G. (1974). Kinetics o f substrate assimilation and product formation in anaerobic digestion. J, Wafer Pollvt. Control. Fed 41748-756. Good, P., Moudry, R. and Fluri, P. (1982). Use of fixed film and CSTR reactor for anaerobic treatment of stillage o f wood-hydrolysate. Biotechnol. Leu., 4(9):595-600.

xn

ACKNOWLEDGEMENTS
The authors are thankful to the Director, Thapar Centre for Industrial Research and Development, Patiala for allowing the publication o f the research findings. Thanks are given to Dr. N.Tejo Prakash, Dr. G.S.Grewal, Dr. M.Roy, Mr. Sarabjeet Singh, Mr. Rajesh Kapoor, Mr. Sandip Chandra, Mr. Vishal fa the various technical assistence provided during the study.

REFERENCES
Alan, L., Sutlon, P.M., Corrado, J.J. and Kothari, D. (1985). Optimization of two-phase anaerobic fluidized bed reactors. lnt Con/. Fixed. Film Biol Processes. 2nd, 1984(Pub. 1985), 2:1741-1759. Alexiou. I.E.. Anderson, G.K., Evison, L.M. (1994). Design of preacidilication reactors for the anaerobic treatment of industrial wastewaters. Wat. Sci Technol. 29(9):199-204. Chakrabarti. S.K., Kothari, R.M. and Garg, R.P. (1991). Anaerobic treatment of wood-hydrolysate effluent of a rayon pulp industry. Proc. lnt. Workshop Efllu. Treat., New Delhi, 16-20 Sept., 1991, pp..S-l4. Chakrabarti, S.K., Bajpai, P.K. and Roychoudhury, P.K. ( I 995). Biomethanation of wood-hydrolysate in high-rate bioreactors. Proc. Eleventh National Convent Chem Engineers, Roorkee, 28-29 Sept., 1995, pp.lll 10-15. Chakrabarti, S.K. (1998). Ph.D. Thesis - Biphasic biomethanation of hemicellulose rich eifluent. Thapar Institute o f Engineering and Technology, Patiala, India. Cohen. A,, Zoetemeyer, R.J., Van Deursen, A. and Van Andel, J.G. (1979). Anaerobic digestion o f glucose with separated acid productioii and methane production. War Res 13:571-580.

lnrernarional P i i l p & Paper Direcrory (1994). Pulp & Paper International, New York, USA.
Johtisoii, D.B., Moore. W.E. and Zank, L.C (1961). The spectrophotometric determination o f ligniti in small wood samples. Tuppi. 44(11):793-798. McCarty, P.I.., Bough, K., Bachmann. A,. Owen, W . and Everhart, T. (1983). Autohydrolysis for increasing methane yields from lignocellulosic materials, in Fuel Gas Drvelopnient, Wise. D.L. (Ed.), CRC Press, Florida, USA, pp.49-72. Mudrack, K. and Kunst, S. (1982). Studies o f the twostage operation o f reactors for anaerobic treatment of carbohydrates. 2. Wassrr Abwasser. Forsch. 15(6):277-280 and 285-287. Pohland, F.G. (1992). Anaerobic treatment: fundamental concepts, applications and new horizons f f in Design o Anaerobic Processes for the Treatment o lndustriol find Municipal Wastes. Melina Jr.. J.F. and Pohland, F.C. (Editors). Technomic Publishing Camp. Inc., Lancaster, USA, 1-34. Sierra-Alvaraz, R. and Leninga. G. (1991). The methanogenic toxicity o f wastewater lignins and lignin related compounds. J Chem Technol. Biotechnol 50(4):443-455.

BIPHASIC BIOMETHANATION
Artif Cells Blood Substit Immobil Biotechnol Downloaded from informahealthcare.com by National Environmental Engineering Research Institute on 07/03/12 For personal use only.
Weiland, P. (1987). Development of anaerobic filters for treatment of high strength agro-industrial wastewaters. Bioprocess Engn.. 2:39-47.

461
Weiland. P. and Thomsen. H. (1990). Operational hehaviour of an industrial fixed bed reactor for hiornethanation of alcohol slops from different crops WaferSci Techno/. 22 (1-2):38S-394.