Biotechnology Letters Vol ii No 3 219-224 (1989)

Received February 21

THE EFFECT OF CALCIUM ON ANAEROBIC DIGESTION

C A Jackson-Moss , J R Duncan

Department of Chemistry and Biochemistry, Rhodes Universi ty,

Grahamstown, 6140, South Afr ica

and

D R Cooper

Leather Industries Research Ins t i tu te

P 0 Box 185, Grahamstown, 6140, South Afr ica

SUMMARY

The a b i l i t y of methanogenic bacteria to adapt to high concentrations of

calcium was investigated using a 9 ~ Upflow Anaerobic Sludge Blanket (UASB)

reactor fed semi-continuously with a synthetic waste. I t was found that a

calcium concentration of up to 7000 mg/~ had no inh ib i to ry effect on

anaerobic digestion.

INTRODUCTION

The anaerobic digestion process, which is widely used for the treatment of

municipal waste sludges, is now being applied in a var iety of industr ia l

ef f luent treatment plants throughout the world (Callander and Barford,

1983a). However, being a microbiological process, the successful treatment

of organic wastes depends on the maintenance of an environment that is

favourable to the organisms involved, and thus the el iminat ion or control

of toxic materials is of major importance in maintaining such a favourable

environment.

Tannery wastewaters contain high concentrations of calcium ions. With the

poss ib i l i t y of anaerobic digestion being used as an a l ternat ive treatment

method for these wastewaters, the ef fect of these ions on the process

requires pr ior invest igat ion.

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Calcium (Ca 2+) is known to be essential for the growth of some strains of

methanogenic bacteria (Murray and Zinder, 1985). I t is also known to play

an important role in the formation of microbial pel lets in UASB reactors

(Sam-Soon et al, 1986). However, very l i t t l e is known about the tox i c i t y

of Ca 2+ to the anaerobic digestion process. One of the few tox i c i t y

studies that have been carried out showed that Ca 2+ is moderately

inh ib i tory at a concentration of 2500-4000 mg/~, but is strongly inh ib i tory

at a concentration of 8000 mg/e (Kugelman and McCarty, 1965).

In view of the l imited information available regarding the toxic effects of

Ca 2+ on anaerobic digestion, this study was carried out in order to

investigate whether an anaerobic bacterial population could be acclimatized

to high concentrations of Ca 2+.

MATERIALS AND METHODS

Anaerobic Digesters

The experiments were performed in g ~ perspex Upflow Anaerobic Sludge

Blanket (UASB) reactors. These were maintained at 35~ by passing water

from a constant temperature water bath through internal heating coi ls. The

digesters were covered in t i n f o i l in order to help maintain mesophilic

temperatures, as well as to prevent growth of photosynthetic anaerobes.

Biogas was collected in sealed 5 ~ aspirator bott les containing a 20 % NaCI

- 5 % H2SO 4 solution to minimize CO 2 so lub i l i t y (Mackie and Bryant, 1981).

The volume of gas was measured by f lu id volume displacement, and was

corrected to 760 mm Hg pressure and O~ (STP).

Sludge Innocula

The anaerobic sludge innocula were obtained from a completely mixed

anaerobic digester at the Grahamstown Municipal Sewage Works, which treats

mainly domestic sewage.

Synthetic Waste

The synthetic waste which was used throughout th is investigation had the

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fol lowing composition (mg/~): Glucose (2500), Urea (500), Yeast extract

(500), K2HPO 4 (500), MgSO4.7H20 (500), CaCl 2 (I00), NaCl (I00), FeCI 3 ( I0) ,

MnSO4.4H20 ( I ) , COC12.6H20 (0,5), CuSO4.5H20 (0,5), Na2B407.1OH20 (0 , I ) ,

ZnSO4.7H20 (0,5), NiSO4.6H20 (0,5), Na2MoO4.2H20 (0,2). The pH was adjusted to 7,0 • 0,I using NaOH. The rate of feeding was 1 l i t r e per day

which results in a hydraulic retention time of 9 days and a loading rate of

330 mg/~.day. Digester startup took place over a period of 4 weeks pr ior

to the start of the experiment. The incremental addit ion of Ca 2+ to the

digester was carried out over a period of I00 days in order to allow the

methanogens to adapt to the increasing concentration.

Analyt ical Methods

Volat i le fa t t y acids and percentage composition of the biogas were analyzed

as described previously (Jackson-Moss et al, 1988).

Calcium was determined by digesting a sample in a 20 m~ solution of I : I

concentrated HCI concentrated n i t r i c acid. A Varian 1275 atomic

absorption spectrophotometer was used to determine the concentration of

calcium.

Chemical oxygen demand (COD) and a l ka l i n i t y were determined according to

Standard Methods (APHA, 1980).

RESULTS AND DISCUSSION

The addition of Ca 2+ ions to the digester up to a concentration of 7000

mg/e (Figure l) had no inhibitory effect on the methanogenic microbes. The

volume of biogas produced remained constant over the experimental period

(Figure 2), with the methane content varying between 75 % and 80 % (Figure

3). The pH, volati le fatty acids and percentage COD removed (Table l) were

also maintained at a stable level, indicating that methanogens have the

abi l i ty to adapt to h igh concentrations of Ca 2+ and that after

acclimatization, the metabolic act ivi ty of the microbes was the same as at

low concentrations of Ca 2+ ions.

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Ca 2+

Concentration (mgle)

7000

6000

5000

4000

3000

2OOO

1000

M 0

Influent/

Ca2+ /

//,/ �9 I";//E f~l.e.t

Ca2+

20 40 60 80 [00

lime [Days)

FIGURE I - InfIuent and effluent Ca 2+ (Note: The control digester had an influent and effluent Ca 2. concentration of 74 and 70 mg/(respectlve]y) '

,,oo( 1 I

Biogas BOO Volume (me) 600

6.00

200

I 1 l J _

0 20 40 60 ~0 ', @t)

FIGURE 2 - Biogas volume

( . . . . Ca 2. digester; . . . . . cc)ntrol digester

Time (Days)

100

80

60

40

2O

% C11

yc co 2

I ! I I

20 40 60 80 I00

FICURs 3 - Percentage biog~s composition

( ....... Ca 2+ digester; . . . . control digester)

Time (Days)

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Unlike heavy metals which tend to prec ip i ta te out and accumulate in

anaerobic digesters (Jackson-Moss et al, 1988), a large proportion of the

Ca 2+ ions passed through the digester and were present in the ef f luent .

There was, however, some accumulation of Ca 2+ ions at the base of the

digester (Table 2) as a resul t of prec ip i ta t ion of Ca 2+ ions as Ca(OH) 2.

The so lub i l i t y product of Ca(OH) 2 at pH 7,0 is 4 x 10 -6, indicat ing that a

proportion of the Ca 2+ ions w i l l prec ip i ta te as the hydroxide at neutral pH

(Callander and Barford, 1983b).

Table I: Characteristlcs of the Digester Effluents at Various Stages During the Experiment

Control Digester ,Ca 2+ Digester

Parameter Time (Days) Time (Days)

I 25 50 75 I00 1 25 50 75 100

pH 7,58 7,57

Alkal in i ty (mg/e CaC03) 1224 1479

COD Removed 84,2 85,3

Volatile Fatty Acids as Acetic (mg/~) 39,7 45,9

Ca 2+ (mg/e) 64 72

7,16 7 , 4 4 7,44 7,23 7 ,44 7 , 4 3 7 , 2 0 7,10

1428 1377 1224 1232 1219 1035 1127 920

86,4 8 9 , 7 85,3 84,8 88 ,2 9 0 , 4 8 9 , 5 87,7

30,8 3 9 , 5 56,5 77,7 87 ,0 4 9 , 2 7 0 , 8 61,4

68 70 74 44 154 934 1894 3596

Table 2: Stratigraphic Analysis of the Digester Contents on Completion of the Experiment

Height of Sampling Port Above Inlet

(cm)

Control Digester Ca 2+ Digester

pH Ca 2+ Total Volatile pH Ca 2+ (mg/e) Fatty Acids as (mg/e)

Acetic (mg/e)

Total Volatile Fatty Acids as Acetic (mg/e)

58 40 27 14

7,44 74 56,5 7,10 3594 7,32 76 30,3 8,54 3394 7,10 111 48,2 6,55 3894 7,05 104 58,8 6,57 8994

61,4 31,0 11,0 28,5

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CONCLUSION

From the results of this investigation, i t can be concluded that anaerobic

digestion can be used for the successful treatment of wastewaters

containing high concentrations of calcium ions, provided the microorganisms

have suff icient time to adapt to the presence of the Ca 2+ ions.

ACKNOWLEDGEMENTS

This work was financed by scholarships from the Council for Scienti f ic and

Industrial Research and the Leather Industries Research Inst i tute.

REFERENCES

APHA, AWWA, WPCF (1980). Standard Methods for the Examination of Water and Wastewater. 15th ed. Washington, D.C.

Callander, l .J . and Barford, J.P. (1983a). 24-30/37.

Process Biochem. 18 (4),

Callander, l .J. and Barford, J.P. (1983b). Biotechnol. Bioeng. 25, 1959-1972.

Jackson-Moss, C.A., Duncan, J.R. and Cooper, D.R. (1988). J. Am. Leather Chem. Ass. (in press).

Kugelman, l .J. and McCarty, P.L. (1965). J. Water Poll. Control Fed. 37, 97-I16.

Mackie, R.I. and B~yant, M.P. (1981). 1363-1373.

Appl. Environ. Microbiol. 41,

Murray, P.A. and Zinder, S.H. (1985). Appl. Environ. Microbiol. 50, 49-55.

Sam-Soon, P.A.L.N.S., Loewenthal, R.E., Dold, P.L. and Marais, G.V.R. (1986). Proc. Anaerobic Digestion Symposium 22-24 September, University of Orange Free State.

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