Process Biochemislry 28 (1993) 83-90

Kinetic Study of Anaerobic Digestion of Wine Distillery Wastewater

R. Borja,a A. Martin, b M. Luquea & M.M. Duriina n Instituto de la Grasa y sus Derivados (CSIC), Avda. Padre Garcia Tejero 4, E-41012 Sevilla, Spain b Departamento de Ingenieria Quimica, Facultad de Ciencias, Avda. San Albert0 Magno s/n, E- 14004 Cbrdoba, Spain

(Received 27 February 1992; accepted 21 March 1992)

A kinetic study was made of the anaerobic purification or biomethanation of wine distillery wastewaters ( ‘vinasses ‘), using bioreactors containing various suspended clayey supports (sepiolite, bentonite and saponite), on to which the microorganisms eflecting the puriJication were immobilised. Assuming that the overall anaerobic fermentation process conforms to.first-order kinetics, experimental data pairs, namely the methane volume yielded (G) and the time (t), jitted the fo/Iowing equation: G = G, x (I -exp (K, x t)). The specific rate constant, K,, was determined in each of the situations studied. The support used has a marked influence on the kinetic constant of the process,. the saponite support yielding sign$cantIy the highest values. On the other hand, the specific rate constant decreased over the chemical oxygen demand (COD) range studied (I-MI g/litre) when the volume of wastewater added or substrate concentration was increased; this showed an inhibition phenomenon. Also, the mean rate biogas production and the methanogenic activity decreased irrespective of the support used. The yield coeficient, Y,, was 0.32 litres CH., STP/g COD.

INTRODUCTION offer several advantages over aerobic treatment processesRm’3 for waste treatment. Because of the

Wine distilleries produce large volumes of wastes, low growth rates of anaerobic microorganisms, a know as ‘vinasses’, which have a low pH and a high variety of methods has been developed to retain organic content (chemical oxygen demand (COD) them within bioreactors and avoid their loss with values 10-60 g/litre) and cause considerable en- the effluent which would result in a slowing of the vironmental problems.’ 7 process. In the fluidised bed reactor, the bacteria

It is generally accepted that anaerobic processes colonise particles of support materials which in- creases the useable surface area for bacterial

Corresponding author: R. Borja growth. Due to a retained biomass these reactors

83 Process Biochemiswy 0032-9592/93/$MOO 0 1993 Elsevier Science Pubhshers Ltd. England.

84 R. Borja, A. Martin, M. Luque, M.M. Durrin

may be used with a higher volume load and are behaviour.lg Their chemical composition and suitable for treating wastewaters of a high organic features are summarised in Table 1. content.14

The results obtained from previous research on Inocullun

the microbiology and biochemistry of anaerobic The digesters were inoculated with biomass from an processes show the influences that different supports anaerobic reactor that processes piggery wastewater have on the immobilisation of the microorganisms and contained the methanogenic flora. The com- which carry out digestion.‘5m1” The aim of this work position of this biomass is summarised in Table 2. was to carry out a kinetic study of the anaerobic digestion of wine distillery wastewater in three bioreactors containing microorganisms im- mobilised on sepiolite, bentonite and saponite supports, in order to study the influence of these supports on the biokinetic parameters of digestion. Treatments involving fluidised beds require sturdy supports of low apparent density in order to reduce power consumption. The supports used in this study have these properties.

MATERIALS AND METHODS

Experimental design Anaerobic digestion units (ADUs) of 3-litre ca- pacity were magnetically stirred and immersed in a water bath at 37 “C. The biogas generated was passed through a solution of sodium hydroxide to retain carbon dioxide and the volume of methane was determined indirectly as the amount of water displaced by the gas.

Supports The materials used as supports for the anaerobic bacteria were commercially available micronised sepiolite, bentonite and saponite supplied by Tolsa, S.A. (Madrid, Spain). These clayey supports were selected on account of their favourable kinetic

Table 1. Composition and features of the bioreactor supports”

Saponitr Benronire Sepiolite

SiO Al,& Fe&), TiO Mgb CaO Na,O K-0

57.3 6D3 62.0 4.4 16.8 l-7 2.0 3-6 0.5 0.2 0.2 -

254 4.6 23-9 0.6 l-7 0.5 0.2 4.5 0.3 1.0 1.3 0.6

Glcination loss 8.3 6.7 10.5 Moisture content (%) 8.0 15.0 8.5 Bulk density (g/mi) 0.5 0.7 0.1

a Typical chemical analysis (% sample dried at 105 “C).

Wastewater Wine distillery wastewater was collected from the factory ’ Incamasa’ in Aguilar de la Frontera (Grdoba, Spain). The features of this wastewater are given in Table 3.

Chemical analyses The following parameters were analysed: COD, pH, total solids, volatile solids, mineral solids, total suspended solids, volatile suspended solids, mineral suspended solids, volatile acidity, ammonium ni- trogen and phosphorus. These analyses were carried out according to Standard Methods for the Exam- ination c$ Water and Wastewater.2o

The total phenol content was determined by the Folin-Ciocalteau method, while o-diphenols were assayed with sodium molybdate and sodium nitrite.‘l

Experimental procedure Experiments were conducted in three ADUs each of which contained a sepiolite, bentonite or saponite support. Each ADU was supplemented with 2500 ml of distilled water, 500 ml of the above- mentioned inoculum and 20 g/litre of the cor- responding support. While larger amounts of support allowed an increase of biomass, they could also increase the apparent viscosity of the medium and hence hinder mass transfer and decelerate the process of biodegradation.

Before the experiments were started, the biomass was conditioned by feeding it with gradually increasing volumes of the corresponding wastewater for 4 months. The added volume was modified every time methane production was completed, according to the rate of biogas production. The experiments were conducted batch-wise using 60, 120, 180, 240, 300, 360, 480 and 600ml of wastewater. In each, the methane volume produced per day, and the initial and final COD were determined. The wastewater volumes were added after separating the same volume of liquid from the bioreactor after settling (2 h) in order to avoid biomass losses. All experiments were conducted in duplicate.

Kinetic study of anaerobic digestion of wine distillery wastewafer 85

Table 2. Composition of the biomass used as inoculum

PH 7.5 Total solids (g/We) 65.9 Mineral solids (g/litre) 17.4 Volatile solids (g/litre) 485 Total suspended solids (g/litre) 61.3 Mineral suspended solids (g/like) 15.4 Volatile suspended solids (g/litre) 45-9

Table 3. Features of the distillery wine wastcwaters used

PH COD Total solids Mineral solids Volatile solids Total suspended solids Mineral suspended solids Volatile suspended solids Volatile acidity (acetic acid) 1 Nitrogen (NH,+) Phosphorus (POp3-) Total phenols (caffeic acid) O-Diphenols (caffeic acid)

3.8 40-O g/litre 32.0 g/litre

66 g/litre 254 g/litre

3,7 g/litre O-9 g/litre 2-8 g/litre

1600 mg/litre 140 mg/litre 130 mg/litre 290 mg/litre

45 mg/litre

RESULTS AND DISCUSSION a Support used-bentonite.

Tables 66 list the methane volumes accumulated at different times (days) for the different feed volumes used in the digesters containing sepiolite, bentonite

Table 4. Volume of methane accumulated (ml) as a function of time (days) for the different feed volumes used”

Time Feed volume (dvs) Cm0

1 2 3 4 5 6 7 8 9

10 11 12 13 14 15 16 17 18 19 20 21

60 120 180 240 300 360 480 600

750 1455 2010 1560 2330 2085 2510 1595 805 1875 2865 2660 3875 3510 3515 1950 840 2015 2870 3260 5195 5340 4865 2705 840 2160 2875 3420 5320 5605 6260 3815 - 2166 2880 3505 5345 5705 6475 4820

~~ 2880 3560 5425 5795 6505 5570 - 3567 5500 5910 6575 6530

- 5508 5970 6620 7190 - 6035 6685 8180 - 6036 6685 8180

- 6715 8793 - 6745 9200 - 6753 9370 - 6777 9485 ~ 6810 9535 - 6830 9540 - 6873 9573

~ 9600 -- 9696

- - 9702 - 9702

_ ” Support used-sepiolite.

Table 5. Volume of methane accumulated (ml) as a function of time (days) for the different feed volumes used”

Time Feed volume (days) (ml)

60 I20 180 240 300 360 480 600

1 840 1710 1950 2215 2125 1625 1970 1730 2 867 1870 2400 3348 2910 2210 3470 2980 3 867 1872 2409 3381 4200 2900 3830 4270 4 ~ 1875 2409 3426 4383 3965 4195 4865 5 _-- 3450 4395 4980 4910 5910 6--_ 3450 4410 5052 5100 7590 7 -~ ~ 4431 5097 5802 8265 8 - - - - 4431 5088 6665 8480 9__--_ 5088 7005 8780

10 _ _ _ ~ ~ ~~ 7035 9010 11 _ - _ _ - - 7070 9060 12 - 7100 9075 13 _ _ _ _ _ ~ 7105 9090 ,4 _ _ _ _ - - 7119 9100 15 ~ ~ ~ ~ _ 7140 9105 16 _ ~ _ - _ _ 7160 9120 17 ~ ~ _ _ _ - 7177 9126 18 _ _ _ _ ~ ~ 7191 9129 19 _ _ - _ - - - 9132 20 - 9132

Table 6. Volume .of methane accumulated (ml) as a function of time (days) for the different feed volumes used”

Time Feed volume (Ws) (ml)

60 120 180 240 300 360 480 600

1 795 1650 2040 2330 2850 2475 2370 1344 2 816 1800 2295 3200 3920 4662 3735 1555 3 816 1845 2298 3245 4070 4893 6030 1617 4 ~ 1845 2298 3290 4095 4965 6230 1735 5 __- 3303 4110 5010 6230 1885 6--- 3303 4119 5022 6230 2160 7 _ ~ ~ ~~ 4122 5022 6230 2755 8------ 6231 3790 9------ 6234 4375

10 _ - _ _ ~ ~ 6237 4840 11 _ _ _ _ _ - 6291 5590 12 ~ - _ _ _ - 6459 6025 13 _ _ _ _ _ _ 6501 6325 14 _ _ _ _ _ _ 6513 6705 15 ~ - 6516 7190 16 _ _ _ _ ~ ~ 6516 7680 ,7 _ _ - _ - - - 7825 18 ~ ~ ~ ~ ~ _ _ 7830 19 _ _ - _ _ _ ~ 7854 20 _ _ _ _ _ _ _ 7854

D Support used-saponite.

and saponite supports, respectively. The mean rates of gas production, r, referred to the digester volume, were calculated after a preset time of 4 days. In the

86 R. Borja, A. Martin, M. Luque, M.M. Durrin

rho 8

Fig. 1. Variation of the mean rate of gas production (r/r,,) as a function of the substrate concentration (S/S,,).

equations below, S, denotes the lowest substrate concentration used in each experiment and r,, the corresponding mean rate. The plot of T/T,, versus S/S, (Fig. 1) shows a maximum which is well- defined in most instances and after which the relative rates decrease gradually. This phenomenon is observed in all cases, and suggests the occurrence of an inhibition process at concentrations slightly higher than those investigated. The different co- ordinates of the maximum reveal the influence of the support on the kinetics of the process; saponite and sepiolite appear to enhance the reaction rate.

In order to quantitate the extent of inhibition an analytical relationship was obtained between the volume of methane (reaction product) formed and the fermentation time. Thus, the Monod equation

P = P,,, x (S/K, + s> (1) applied to dilute solutions reads

P = ol,lK,) x S (2) where p (day-‘) is the specific growth rate of the microorganisms, pm is the maximum value, S (g COD/litre) is the substrate concentration and KS (g COD/litre) is a constant.

The coefficient of yield of the microorganism is defined as

Y, = -dX/dS (3)

where X (g VSS/litre) is the microorganism con- centration. Hence

Since - Y, x (dS/dt) = dX/dt (4)

dX/dt=pxX (3

by substituting into eqn (2) and taking into account that X = X,, as Y, is very sma11,22*23 one has

with -dS/S = K, x dt (6)

K, = PU, x X,/K, x Y, (7) where X0 (g VSS/litre) is the initial microorganism concentration. This hypothesis is valid only math- ematically and was fulfilled by all the digesters and experiments since the values obtained for X were the same in all cases and ranged between 6.9 and 7.1 g VSS/litre (mean value = 7-O).

Integration of eqn (6) on this assumption when t = 0 for S = S,, yields

S= S,xexp(-&xt) (8) The yield coefficient of the product formed (meth- ane) is defined by

Y, = -dG/dS (9) integration of which when G = 0 for S = S, yields

G= Y,x(S,-S) (10) By solving eqn (10) for S and substituting into eqn (S), one has

G= S,x Y,x(l-exp(-K,,xt)) (11) and hence

G=G,x(l-exp(-K,,xt)) (12) where

G, = S,, x Y,

Equation (12) coincides with that proposed by Roediger in Edeline24 to relate the volume of gas and time in a batch anaerobic digestion process. In this equation, G (ml) is the volume of methane gas accumulated at a given time, G, (ml) is the maximum volume accumulated at an infinite di- gestion time, K,, (day-‘) is the observed specific rate constant and t (days) is the digestion time.

By plotting the experimental data listed in Tables 4-6, curves were obtained coinciding with the predictions of eqn (12). Thus, G was zero at t = 0, and the rate of gas production became zero at t = co. Also, the slopes of the curves decreased with

Kinetic study of anaerobic digestion of wine distillery wastewater 87

Sepidi te

Q Ctred 6

7

6

6

4

a

2

1

0 1 6 10 16 20

Time (days)

- 80 ml. + 180 ml. * MOmL + P40 mL

* LOOmL + WOmL + 4BOmL

Fig. 2. Variation of the volume of methane accumulated, G (litres), with time (days) for the sepiolite digester.

6

6

~Iltrer)

Saponite

1

6 20 26

Fig. 3. Variation of the volume of methane accumulated, G (litres), with time (days) for the volumetric load of 600 ml of wastewater in the saponite digester.

Saponi te

n lOm/(Qm-011

0 1 2 9 4 6 6 7 6

Time Maya)

++120 mL + 240 mL

Fii. 4. Representation of the In (G,/(G, - G)) values versus time (days) for the saponite dtgester.

increasing time. Figure 2 shows the variation of the accumulated methane volume as a function of time for different feed volumes (60-480 ml) in the digester with sepiolite as support.

For feed volumes of 600 ml or greater, the curves G = j(t) no longer fit an exponential model, as Roediger’s equation predicts. Instead, they show regions of scarce or no production of methane, determining the existence of points of inflexion in the curves. Figure 3 shows the kinetic curve for this volumetric load in the reactor with saponite support. Thus, it does not seem appropriate to apply the proposed kinetic model for wastewater loads equal to or greater than 600 ml.

On the other hand, taking Napierian logarithms in eqn (12) and ordering the terms, the following is obtained :

ln(G,/(G,-G)) = K, x t (13) indicating that In (G,/(G, -G)) versus t should give a straight line of slope equal to K, with ordinate zero. As an example, Fig. 4 shows part of the experimental data for the saponite reactor. The value of G, has been considered equal to the volume of methane accumulated at the end of each experiment. On representing the experimental data as indicated, eqn (13) gives straight lines with the

88 R. Borja, A. Martin,

Table 7. K, values (days-‘) with 95 % confidence limits for each digester and experiments

Feed volume

Cm0

60 2.27 f 0.09 120 1.07 f 0.07 180 1.28 + 0.05 240 @63 k 0.08 300 0.63 f 0.07 360 0.5 I + 0.08 480 0.45 kO.05

Sepiolite Bentonite

3-44+@08 2-44 If: 0.09 1.65 + 0.09 1-12 kO.08 O-62 + 0.07 0.28 + 0.09 0.23 kO.04

Saponite

3.36-hO.05 2.25 + 0.03 2.16f0.02 1.28 + O-09 1.23 + 0.08 0.72 10.09 0.56 f 0.08

Table 8. G, values (ml CH,) with 95% confidence limits for each digester and experiments

Feed Sepiolite Bentonite Saponite

Cm0

60 833.25 120 2158k51 180 2920+ 125 240 3651k126 300 5597k205 360 6141 k281 480 6848f145

867+_2 816+_2 1868k 19 1838f25 2437 k 79 2307 + 73 3499+ 177 3322 + 73 4567&388 414Of63 5836k790 5163+275 7388 +295 6463+216

0 2 0

S;o COD/L) 8 lo

- 80plolllo h I)onlonlW

Fig. 5. Variation of the specific rate constant, K0 (days-‘), with the substrate concentration (g COD/litre) for the three supports studied.

M. Luque, M.M. Durrin

ordinate practically at zero. The same behaviour was observed in the other digesters for feed volumes less than 480 ml. Once it had been checked qualitatively that the experimental data (feed volumes between 60 and 480 ml) conformed to the proposed model, the parameters G, and K, were calculated analytically by using a nonlinear re- gression program.25

Tables 7 and 8 list the K, and G, values, respectively, with 95 % confidence limits.

The variation of K,, with the initial COD for each system was plotted and as can be seen from Fig. 5 and Table 7: the specific rate constant, K,, decreased with the substrate concentration, suggesting the occurrence of an inhibition process. The observed decrease depended on the kind of support used, saponite and sepiolite giving the lowest decrease. The values of K,, for each support tested differed both at high and low substrate concentrations, which suggests that the influences of the supports are quite different, saponite featuring the highest K,, (day-‘), at both high and low COD values.

On the other hand, the anaerobic biodegrad- ability of these wastes is high, whatever the load added, since the final COD of the digester was virtually constant (0.2 g/litre). Thus, the average fraction of biodegraded substrate was found to be > 90 %, showing effectiveness of anaerobic di- gestion as a purifying procedure for this wastewater. The yield coefficient of methane, Y,, was determined from the methane volume produced (Tables 4-6) and the final and initial COD, which were known in each case (1.0, l&2.6,3.4,4*2, 5*0,66 and 8.1 g/litre for 60, 120, 180, 240, 300, 360, 480 and 600 ml of feed volume added, respectively). The value of this coefficient was found to be 327 ml CH, STP/g COD.

Finally, the observed inhibition process can also be demonstrated via the following route. The mean rate of methane production can be calculated from the overall accumulated volume and the corre- sponding time. Since the amount of biomass present in each reactor is known (7.0 g VSS/litre), the methanogenic activity (MA) may be calculated from the following equation:

MA = G/(tx Xx Vx Y,,)

Figure 6 shows the plot of MA for the three digesters used against the initial COD. There is a maximum in each case after which the MA decreased markedly. This indicates the occurrence of an inhibition phenomenon, independently of the support used.

Kinetic study of unaerobic digestion of wine distillery wastewater 89

0.08

0.06

0.04

0.02

0

Mathmcgenic Aetiulty (gCOD/gVSS day) 0.1

I I I I

0 2 6

S;p COD/L) 8 lo

Fig. 6. Variation of the methanogenic activity (g COD/g VSS day) with the substrate concentration (g COD/litre) in the three digesters used. (VSS-volatile suspended solids.)

CONCLUSIONS

The anaerobic digestion of wine distillery waste- water conforms to first-order kinetics at substrate concentration equal to or lower than 6.55 g COD/ litre.

The specific rate constant, K,, decreases with the substrate concentration over the COD range stud- ied, revealing the occurrence of an inhibition phenomenon. This decrease in the kinetics of the anaerobic digestion was found in all the supports tested. However, saponite features the highest K,, at both high and low COD values.

This waste can be readily biodegraded by an- aerobic digestion, since over 90% of the initial COD is removed. The yield coefficient, Y,, is 0.327 litres CH, STP/g COD and is independent of the type of support used.

ACKNOWLEDGEMENTS 17.

The authors wish to express their gratitude to the ‘Consejeria de Education y Ciencia’ of the ‘Junta de Andalucia’ and to the CSTC for their financial support of this work. Thanks also to Carmen Sanchez for her help in the analysis and to the

factory ‘Incamasa’ for kind provision of the wastewater used.

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