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Page 1: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

Biological Wastes 27 (1989) 1-14

Effects of Total Ammonia on Anaerobic Digestion and an Example of Digestor Performance from Cattle

Manure-Protein Mixtures*

J. E. Robbins, S. A. Gerhardt & T. J. Kappel

Chemistry Department, Montana State University, Bozeman, Montana 59717, USA

(Received 13 October 1987; revised version received 11 March 1988: accepted 15 March 1988)

A B S T R A C T

Anaerobic digestion of dairy cattle manure slurries was performed at different total ammonia concentrations, c. 50, 100, 150, 200 and 300mM. Digestor performances, i.e. daily methane yields, volatile acid concentrations, pH and relative rates of utilization of acetate, propionate and glucose were monitored. Rates of utilization were determined prior to, during initial time periods following a change in total ammonia concentration and after acclimation had occurred. Digestor performances indicated inhibition occurred, followed by return to steady-state performance. Relative rates (~[acetate utilization were most affected by changes in total ammonia concentrations. Thus, we concluded that methanogenic metabolism was primarily inhibited by a high concentration of total ammonia. Anaerobic digestion of manure-bovine serum albumin as an example of manure containing high concentrations of proteinaceous material was also performed at three diff'erent added protein concentrations. Digestor instability was proportional to protein concentration.

I N T R O D U C T I O N

A n a e r o b i c digest ion has been utilized as a waste t r ea tment and a process for p r o d u c t i o n o f me thane f rom a variety o f wastes. Wi th the inevitable

* Contribution from the Agricultural Experiment Station, Montana State University Published as Journal Series No. J1456.

1

Biological Wastes 0269-7483/89/$03.50 © 1989 Elsevier Science Publishers Ltd, England. Printed in Great Britain

Page 2: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

2 J.E. Robbins, S. A. Gerhardt, T. J. Kappel

shortage of energy, more emphasis should be placed on the production of methane. Recent studies of anaerobic digestion have been concerned with optimizing the methane yield by focusing on the efficiency of turnover rates of organic matter and stability of the process. Studies by Hills (1979), Hills & Roberts (1980), and Robbins et al. (1983a) indicated methane yield and conversion efficiency were improved by adding cellulosic material to manure. These mixtures produced optimum methane yields and conversion efficiency when the C:N ratios of the mixtures were in the range of 30-40:1. Hashimoto (1983) studied the effects of temperature, hydraulic retention time (HRT), and composition of straw-manure mixtures on methane yield. The data of Hashimoto indicated that the methane yield declined proportionately with straw content. Contrasting the results of Hills & Roberts (1980) and Robbins et al. (1983a) with those of Hashimoto (1983) indicates that the nature of the substrates used in anaerobic digestion may be more important than a simplistic C:N ratio.

Originally this study was intended to assess the efficacy of methane production and solids conversion of anaerobic digestion of wastes containing high nitrogen content, such as slaughterhouse wastes. The preliminary studies (unpublished data) involved anaerobic digestion of manure-protein mixtures where bovine serum albumin (BSA) was used as a model nitrogenous material. These studies experienced instability and reduced conversion efficiency. However, in trying to assess the cause of poor digestion performance, we were confronted with the possibilities of NH~ or NH 3 inhibition, volatile acid inhibition or that digestion of serum albumin led to metabolic problems. A clear cause-effect relation could not be indicated.

The effects of ammonium ion or ammonia on methane production from mixed population digestors have been addressed in some aspect in several previous studies (McCarty, 1964; Melbinger & Donnellon, 1971; Converse et al., 1977; Kroeker et al., 1979; van Velsen, 1977, 1979). The toxic level of NH,~ or NH 3 was a point of variance among the earlier studies (McCarty, 1964; Melbinger & Donnellon, 1971; Converse et al., 1977). Van Velsen (1977) and Kroeker et aL (1979) demonstrated methanogenesis was inhibited by high concentrations of NH~ but continued at NH~ concentrations much higher than the levels previously thought to be toxic. Van Velsen (1979) clearly showed that acclimation to high concentrations of NH~ was possible but could not indicate which digestion activity, fermentative, methanogenic or acetogenic was most inhibited. Hobson & Shaw (1976) showed that NH~ inhibited CH4 production in defined cultures of M e t h a n o b a c t e r i u m f o r m i c i c u m , an H2-utilizing methanogen. However, a mixed population digestor presents a more complex system where the populations are metabolically interdependent.

Page 3: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

Anaerobic digestion of cattle manure 3

We formulated this study to address the question of what effect high levels of total ammonia (NH2 + NH3) had on utilization of glucose, propionate and acetate. This study did not address which species, NH~ or NH 3, was the inhibitory substance. The processes of glucose fermentation, propionate oxidation and acetate splitting represent metabolic processes of fermenta- tive, acetogenic and methanogenic organisms, respectively. Thereby, measuring the relative rates of utilization of glucose, propionate and acetate in digestors of different total ammonia concentrations would indicate the effect on these metabolic processes. The results of these studies indicated that inhibition was more pronounced with methanogenic activity than with the fermentation of glucose or propionate oxidation. Data from digestion of manure-serum albumin mixtures are offered as a specific example of digestor performance from waste containing proteinaceous material. The results should not be generalized to indicate the performance to be expected from all manure-protein mixtures.

Experimental rationale and design

The basic rationale and experimental design was based on monitoring rates of substrate utilization and digestor performance prior to a change in total ammonia concentration, during a transition period and after acclimation to the change had occurred. The transition period and acclimation was judged by digestor performance indicators of pH, gas production and volatile acid concentrations of effluent. Two experimental trials were performed to assess the effect of changes in total ammonia concentration. In the first trial three digestors were used to assess the effects caused by changes from 50 to 100 mM, 50 to 200raM, 50 to 300mM and 200 to 300mM, where the digestor that was changed from 50 to 200mM was allowed to acclimate and then changed to 300 mM. This latter change was performed in order to compare the effects of a change from 50 to 300 mM with a change from 200 to 300 mM. The second trial was performed to provide replicate data from two digestors that were changed from c. 50 to 150 mM followed by a change from 150 to 300mM after acclimation to 150mM and two other digestors that were changed from 50 to 300 mM.

METHODS

The general procedures of manure collection and dilution of dairy cattle manure feedstock to appropriate solids content, homogenization by blending, packaging and storage were as previously described (Robbins et al., 1979, 1983a,b). The cattle were fed 40% grain mix and 60% hay. The

Page 4: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

4 J.E. Robbins, S..4. Gerhardt, T. J. Kappel

grain mix was 50% barley, 25% wheat, 10% soymeal, 10% sugar beet pulp and 5% mustard meal. The design of the 64 ml working-volume digestors was as previously described (Robbins et al.; 1979, 1983a,b).

The digestors were fed daily with 4 ml of feedstock following removal of 4 ml of digest. The hydraulic retention time or turnover volume was 16 days. The temperature was maintained at 37°C in a water bath. Inoculum for digestors was obtained from a larger (3.2-1itre) digestor operated under the same conditions.

Ammonium chloride was obtained from Baker (reagent grade). 13C-1- glucose, 13C_2_sodium acetate and 13C_3_sodium propionate were obtained from Stohler/Kor Stable Isotopes. These compounds were 99% enriched. The bovine serum albumin was obtained from Sigma Chemical Co. (St. Louis, Missouri, USA).

The total ammonia concentrations were adjusted to different levels by the addition of NH4C1 to the daily aliquot of feed and the level of serum albumin was adjusted by addition of the protein to the daily feed aliquot. In the experiments with serum albumin a single digestor was used and the level of protein was initially added as 1% additional volatile solids (VS) (40 mg) and after seven turnover volumes the added protein was increased to 2% (80mg) additional VS for 10 turnover volumes and this was followed by addition of 3% (120mg) protein. The latter addition was continued for approximately 24 turnover volumes. For the studies of the effects of total ammonia, three digestors were used in Trial 1. One digestor received manure feedstock only initially, the second digestor received 60 mg NH4C1 per day and the third digestor received 100 mg NH4C1 per day. When effluent from the second digestor reached 100mM total ammonia the digestor was maintained at that level by addition of 12 mg NH4C1 per day and when the third digestor effluent reached 200mM total ammonia that level was maintained by addition of 65 mg NH4C1 per day. The total ammonia levels were 50 _+ 10mM (digestor 1), 110 _+ 10mM (digestor 2) and 200 _+ 20mM (digestor 3). After 320 and 290 days the total ammonia concentrations in digestors 1 and 3 were raised to 300 _+ 20 mM, respectively.

In Trial 2 all four digestors were allowed a period of 70 days of operation on feedstock only. This period was used to obtain baseline data on utilization rates. The changes in total ammonia concentration were made by addition of NH4C1 to the daily feed in a manner similar to Trial 1. The rates of glucose, propionate and acetate utilization were determined within 48 h following the change in total ammonia for each digestor to assess the rates during transition. These determinations were repeated 80 days after the change in total ammonia to assess rates after acclimation. The data from this second trial were subjected to a statistical t-test. The changes in total ammonia concentration in this trial were: digestors A and B, 50 to 150mM

Page 5: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

Anaerobic digestion o f cattle manure 5

followed by a change from 150 to 300 mM after acclimation to the change to 150 mM and digestors C and D, 50 to 300 mM.

The manure feedstock was analyzed for total solids and volatile solids by standard methods (Standard Methods for the Examination of Water and Wastewater). The fiber content was determined as described by Goering & van Soest (1970) and the nitrogen content was determined by the Kjeldahl method. Volatile acids were determined (twice a week) by gas-liquid chromatography (GLC) using a Varian 1400 with a flame ionization detector and a glass column (1.8 m × 2.0 mmi.d.) packed with 10% SP1200 acidified with 1% phosphoric acid (Supelco). Helium was used as the carrier gas (26ml min-1). Retention times were determined by comparison with volatile acid standards (Supelco) and quantitative estimates were made with an integrating recorder (Hewlett-Packard, 3392A). Gas volumes were determined daily by the solution-displacement method (0-1 N HC1) and the CO2 and CH4 content was determined twice a week by gas chromatography (GC) using a Carle 8000 GC equipped with a thermal conductivity detector and a stainless steel column (1.8 m × 2-0 mm i.d.) packed with Porapak N (Waters). Helium was used as the carrier gas. A standard gas mixture of 50-10% CH4 and 49.90% CO/(Matheson) was used for calibration. Total ammonia concentrations were determined every 32 days and just prior to rate analyses by the enzymatic method of Robbins & Weber (1977) but higher dilutions were necessary (150 × ) for samples containing 100 mM or more. Concentrations of NH 3 were calculated as described by Hashimoto (1983). Samples of effluent from digestors were analyzed only for pH, VA and total ammonia.

The kinetics of the utilization of glucose, propionate and acetate were determined by 13C-nuclear magnetic resonance spectroscopy (13C-NMR). The ~3C-1H decoupled N M R spectra were obtained in Fourier transform mode at 62.8 MHz on a Bruker WM-250 spectrometer. The spectra were Fourier transforms of accumulated radio frequency pulses {45 ° pulse angle, 45s repetition time). Chemical shifts were assigned relative to tetramethyl- silane (0ppm) as determined from an external reference, dioxane. N M R measurements were performed on 2 ml samples removed from the digestors anoxically (10-12 h after feeding), placed into 10ram N M R coaxial tubes (Wilmad) containing 7 mg of 1 -x aC-glucose, or 4 mg 3 -13C-propionate and 3 mg 2 -13C-acetate. The digester sample was transferred to the N M R tube in a glove bag (Aldrich) filled with O2-free N 2. A 4 mm coaxial tube (Wilmad) containing 2H20 and dioxane was inserted into the 10 mm tube. The coaxial tube provided a seal to the external atmosphere and the 2H20 a lock signal. The tube was then lowered into the probe in the N M R magnet and data acquisition begun. Data were accumulated in 1 h intervals for acid utilization and 10rain intervals for glucose utilization, over 12 and 2h

Page 6: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

6 J .E. Robbins, S. A. Gerhardt, T. J. Kappel

respectively. The intensities or integrals of the resonance peaks were proportional to concentration and these values at time intervals were taken as relative concentrations of labeled substrates. The rate of utilization was then calculated based on the equation

dC/dt = k[C], (1)

where C is the total concentration ofsubstrate (labeled + unlabeled), t is time in h, and k the first-order rate constant. The pool of unlabeled acid was determined by GLC analysis on a separate sample taken at the same time as the sample used for kinetic analyses. The rate constant was estimated graphically by plotting In C (labeled) vs time and calculating the slope (k). The rates obtained were initial rates of utilization and were used for comparative purposes only. The kinetic data obtained from acetate and propionate utilization obeyed first-order kinetics. In experiments monitor- ing the utilization of glucose, the change in concentration versus time did not obey first-order kinetics but appeared to be mixed, zero and first-order. Thus, glucose utilization was estimated based on total change in glucose intensity over the duration of the experiments.

The per cent conversion of solids was calculated using the formula

12 (g/g atom carbon) x (moles gas/day) x 100 % conversion =

0.4 (fraction carbon in waste) x (g VS of waste)

This calculation provides an approximation since the fraction of carbon in the waste was assumed to be 40% and was used as a relative indicator of conversion of solids.

TABLE 1 Composition of Dry Matter of Manure and Volatile Acid

Concentration of Manure Feedstock

% Dr)' matter

Volatile solids 84 Ether extract 4 Cellulose 25 Hemicellulose 24 Lignin 9 Total nitrogen (crude protein) 2.8 (17.5)" Ammonia 0-4

Concentration (mM) Acetate 69 Propionate 10 Butyrate 5-5

Total nitrogen × 6.25 = crude protein.

Page 7: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

Anaerobic digestion o f cattle manure 7

RESULTS

The analysis of the manure feedstock is given in Table 1. The manure feedstock contained 5% TS of which 84% was volatile solids. The digestor performance profiles ofdigestors used in Trial 1 are shown in Figs 1, 2 and 3. The per cent conversion of volatile solids added and CH 4 yields for different periods for these digestors are given in Table 2. The utilization rates determined for acetate, propionate and glucose are given in Tables 3 and 4 for Trials 1 and 2 respectively. The digestor performance profiles for the digestion of manure-bovine serum albumin (BSA) mixtures are given in Figs 4, 5 and 6. The total ammonia concentrations in these digestors were in the range of 40-60mM for 1% BSA (bovine serum albumin), 130-150mM for 2% BSA and 240-340mM for 3% BSA.

DISCUSSION

The digestor performance profiles from Figs 1, 2 and 3 indicate the destabilizing effect of total ammonia where there were changes in pH, CH 4 production and volatile acid concentrations subsequent to an increase in total ammonia concentration. These performance data also show the acclimation to increased total ammonia concentration in that effluent volatile acids, CH 4 production and pH returned to a steady-state level in

%.-J

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A 1 _ o 6 0 - \ H 12o (D

2~; 40 O

4 P'~(5mM NH 3) ~'i (ImM NH 3)

20

O 0 k ~ J ~ x l _ ~ ~ .L.. I , _ . . a . _ L ~_ . .b ' " . . 100 200 3 0 0 400

TIME (DAY)

Fig. I. Digestor performance of a digestor that received no additional NH4CI until day 320. At day 320 the NH~- concentration was changed from 50 to 300mM by the addition of

NH4CI.

Page 8: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

8 J.E. Robbins, S. A. Gerhardt, T. J. Kappel

Fig. 2.

77 ~'Z6 [ - ~ p H ^ 7.4 ~/ ~ 7 . 3 7.2 -7d

100

CH 4 50 T 80 . ,-'. /:'f~-"'L.~ / A v - ~ 40

~ / ~ 20 __. 60

~: | - |Ace ~ 4 0 50 mM I | 0 _NH4 + ~ ~ I I lOOmM NH4+ ~.

20 (2 mM NH3) !../:':'":--'...'I'.. ~ (2 mM NH 3)

t i j . . . . P r 0 ] ~ . , . _ . . . . . I , , , _ , . . . _ t . . . . . . . . . . . . .

O, }00 200 300 400

TIME (DAY)

Digestor performance of a digestor that received addit ional N H 4 C I on day 60 to change the NH~ concentration from 50 to 100mM.

digestors 2 and 3 at 100 and 200 mM total ammonia. Acclimation was also indicated by the fact that when the total ammonia concentration was changed from 200 to 300 mM in digestor 3 the digestor performance was not as disturbed as in digestor 2 when the concentration of total ammonia was changed from 50 to 100 mM and in digestor 1 when the change from 50 to 300 mM occurred. These observations on dairy cattle manure digestors are similar to those o f van Velsen (1979) in studies of municipal sludge and piggery wastes. Also these observations are consistent with the studies of van

7.7

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7,2 3'. I

100 A ! Ace ,~ CH, ]5O -

80-- I I ! L /~ /V~ ... / \ . - '~ -t40

oo- ',I," A t " IO -J _5oo, f' 'l , , H . . . . . . . . ?o = 40 4-NH4+ ~ ~1 I 200m 4 ~.~--ouumMNH4 " (2raM NH3) "/ :.~ ::'-. I?. :-.. ..." ":'" (4mM NH 3) {hmM NH3] I

I00 200 300 400

TIME (DAY)

Fig. 3. Digestor performance of a digestor that received additional NH4C1 on day 60 to change the NH2 concentration from 50 to 200 mM and subsequently from 200 to 300 mM.

Page 9: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

Anaerobic digestion of cattle manure 9

Velsen (1979) in that the time required for complete acclimation (i.e. return to steady state as noted by effluent volatile acid concentrations) was longer for digestor 3 when total ammonia concentration was changed from 50 to 200mM compared to the 50 to 100ram for digestor 2. However, it is interesting to note that the major recovery from acetate accumulation occurred in approximately the same time period (c. 70 days) in these two

TABLE 2 Comparison of Per cent Conversion and Daily C H 4 Production

Digestor and ml CH45 ml CH4 per % CH 4 % time period" g VS added Conversion

Digestor 1 1-64 29_+3 168+ 17 62_+2 36_+4 65-128 35 _+ 3 203 _+ 20 65 + 3 40 _+ 4 129 192 38_+4 220-+22 70-+3 42-+4 193-256 37 -+ 4 215 _+ 21 70 __%_ 3 40 -+ 4 257-320 35 +_ 4 203 __%_ 20 66 -+ 6 40 _+ 4 321-384 a 20_+ 11 116+58 58-+8 26-+ 13 385-400 28 _+ 3 163 _+ 16 62 _+ 6 34 -+ 3

Digestor 2 1-64 32-+4 186_+23 61 _+3 40_+4 65 128" 20+ 14 116-+80 52_+8 29_+ 16 129-192 3 9 + 3 227-+ 17 65-+5 46-+5 193-256 38_+3 221 _+ 17 63_+4 46-+5 257-320 38-+3 221 _+ 17 61 _+4 47_+5 321 384 34 -+ 3 198 -+ 17 58 _+ 5 44 _+ 5 385-400 34 -+ 3 198 -+ t7 59 _+ 5 44 _+ 5

Digestor 3 1 64 28_+5 163_+29 61 _+3 35_+4 65-128" 18 -+ 10 105 -+ 58 51 _+ 7 27 -+ 10 129-192 38 _+ 8 221 _+ 46 65 _+ 6 44 _+ 8 193 256 36 _+ 6 209 _+ 35 62 _+ 6 44 + 4 257 320" 35 -+ 8 203 _+ 46 60 _+ 6 44 _+ 4 321-384 30_+3 174-+ 17 62-+6 37_+6 385-400 30 _+ 3 174 +_ 17 65 -+ 6 35 -+ 5

a For digestor 1 the time period of 321 384 days was a transition period when the digestor NH,~ concentration was changed from 50 to 300mM. For digestor 2 the transition period was 65-128 days, when the NH + concentration was changed from 50 to 100raM. For digestor 3 there were two changes in NH,~ concentration; viz. from 50 to 200mM after 4-HRT, transition: 65-128 days, and from 200 to 300mM after 18 HRT, transition: 290-360 days (HRT = hydraulic retention t ime= 16 days). b Data reported as mean_+ standard deviation, these were based on 18 determinations for each time period except the last period where four determinations were made. Gas volumes were corrected to STP.

Page 10: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

10 J . E . Robbins, S. A. Gerhardt, T. J. Kappel

digestors. There was one difference in results from these studies with those of van Velsen's (1979), in that in these studies the CH 4 yields following acclimation were the same regardless of the total ammonia concentration, whereas van Velsen's data indicated a decreased CH 4 yield with increased total ammonia concentration. The data for CH 4 yields from this trial are given in Table 2. It is clear that CH 4 yield and conversion efficiency decreased in the transition period (i.e. the time required for acclimation) but these indices returned to levels similar tO those prior to the change in total ammonia concentration.

Inspection of these data demonstrates the difficulty in attempts to indicate the specific microbial metabolic activities that were inhibited by the changes in total ammonia from typical digestor performance data. The utilization rates obtained in Trial 1 (Table 3) and Trial 2 (Table 4) indicate that the methanogenic activity was most affected, whereas the acetogenic and fermentative activities were much less affected. It is important to recognize the variance in rate values obtained for utilization of acetate, propionate and glucose in digestors in steady-state at the normal total ammonia

TABLE 3 Comparative Utilization Rates a

Digestor and Total ammonia Acetate Propionate Glucose time period (mM) (mM h- 1) (mM h- l) (ram h- 1)

Digestor 1 1-320 50 + 10 3.4 + 0"3 (6) 0.56 + 0"09 (6) 370* 300 + 40 0 (1) 0 (1)

Digestor 2 110" 100___20 1.5_+0.1 (1) 0.4_+0-1 (1) 260 100_+20 3"6_+0'3 (1) 0'4_+0"1 (1) 61" 100_+20 76* 100 ___ 20 108" 100 _+ 20

Digestor 3 90* 200 _+ 20 1.6 _+ 0.1 (1) 0-7 _+ 0.1 (1) 240 200+__30 3 .7+0.3 (1) 0 .4+0.1 (1) 291 * 300 _+ 40 320* 300 _+ 40 354 300 _+ 40

6.0 -+ 0.5 (3)

5'3 + 0"5 (1) 6"0 + 0"5 (1) 6"6 + 0-5 (1)

5.8 +0'5 (1) 5"5 + 0'5 (1) 6 -0+0 '5 (1)

a The time periods indicate the rate analyses were performed during that time period or on the given day following day zero. The numbers in parentheses designate the number of determinations. The standard deviations for rates based on a single determination were calculated from the variance of the rate constant within the time periods of the experiment. The asterisks denote determinations during transition.

Page 11: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

Anaerobic digestion o f cattle manure 11

concentration of c. 50mM. The ~3C-NMR analyses in Trial 1 showed that the ranges of kinetic values were 3"0-3-7mMh -1 for acetate, 0"47- 0-66 mM h- ~ for propionate and 5.5-6.5 mM h- 1 for glucose. In Trial 2 the ranges were 2.2-3.8, 0.47-1-8, and 5.2-6"1 mMh-~ for acetate, propionate and glucose, respectively. These latter kinetic values were obtained from all four digestors. The data in Table 3 indicate that acetate utilization was inhibited following each change in total ammonia concentration but returned to a normal value after acclimation. There is some question as to whether total ammonia inhibition of propionate utilization occurred in digestor 1 following the change from 50 to 300 mM or if some other factor may have caused loss ofpropionate utilization. No appreciable inhibition of propionate utilization occurred in the other digestors nor was there any appreciable changes in glucose utilization. The data in Table 4 show that

T A B L E 4 Comparative Utilization Rates of Replicates

Digestor Total ammonia Glucose Acetate Propionate (mM)

Baseline kinetics: rates mM h-1 A 49 6-1, 5'4 3" 1, 2-9 B 38 5"5 3"7, 3"6 C 46 52 3"2, 2-3 D 44 5'8 2"2, 3"8, 2"7

Mean values for all digestors 5.6 _+ 0.3 30 -+ 0"5

Transition kinetics (48 h after the change in total ammonia)"

A B 50-150mM 50-150mM 4"4 5"8} (0"19) 2'3 1"8} (0"02) 1"3 1"1} (0.34)

C 50-300mM 5-7 1"4 1"2 } (0"23) D 50-300rnM 35} (0"05) 0"12} (0"0) 0-43

A 150-300 mM 4'9 ~ (0"24) 1'8"~ (0-01) 0"70} (0"45) B 150-300 mM 5"5J 2"1J 1"5

0"83, 0"56, 1"0 0-47, 0"60 l'l, 1-7, 1-1 1"8, 1-5 1'1 +0"5

Kinetics after acclimation (80 days after the change in total ammonia)

AB 50-50-150 raM150 mM 5356)") (0'39) 392"9) (0"23) 1"12}1 (0"40)

C 50-300raM 5"9"~ (0"43) 2'8} (0-10) 09} (0"36) D 50--300 mM 5"5 ) 2' 1 1 "0 A 150 300mM 5"0} (0'19) 2-3} (0.16) 0.81; (0"23) B 150-300 mM 5'2 2"9 0.83)

" The P-values in parentheses are given for comparison of means between the baseline mean value and the means obtained from the different treatments. The P-values are for negative correlation since inhibition was expected, thus the values are half that of the standard P- value.

Page 12: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

12 J. E. Robbins, S. A. Gerhardt, T. J. Kappel

Fig. 4.

Oq "T'" I i "i..-"r = " l " l i i t 5o ioo

TIME (DAY)

50 7- 40 ,~

30

2o ~

o

I I I 150

Digestor performance of a digestor that received 1% additional VS as BSA. The N H + concentration range was 4 0 - 6 0 m M .

acetate utilization was significantly inhibited (i.e. P<0.10) , propionate utilization was not significantly inhibited (P>0-10), however, glucose utilization showed significant inhibition (P < 0-10) in digestors C and D following the change from 50 to 300 mM total ammonia. Individually, the rate in digestor C was not inhibited, whereas the rate in digestor D was significantly lower• These observations suggest that some inhibition may have occurred with utilization of glucose and propionate but clearly indicate that methanogenic activity of acetate utilization was most affected by changes in total ammonia. It may be that methanogenic activities in general

Fig. 5.

A

7o

u) 60

5O

4O

E 3O

20

i0 ..,,"

,.~,"

1 I 1 1 I I I I I I I I I I I I 5 0 I 0 0 1 5 0

TIME (DAY)

5O 7 -

40

30 e-~ ~ r

20 N Io -J

0

t I 2OO

Digestor performance of a digestor that received 2% additional VS as BSA. The N H ~ concentration range was 130 -150 riM.

Page 13: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

Anaerobic digestion o f cattle manure 13

Fig. 6.

140 ~. -e.O i" i:i i pH

r20 -

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-t t4o "~ 8 0 - t ~ ,,, i ~ . . - ! .~! "'t;'k ,,,\ 30 j

o ,,~ - 20

~ 60-- - i

40 ~."

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O " ~ J I ~ J ~ , I , , J I , , , ,

0 IOO 20O 300 4O0

TIME (DAY)

Digestor performance of a digestor that received 3% additional VS as BSA. The NH~ concentration range was 240-340mM.

are more subject to inhibition from total ammonia since Hobson & Shaw (1976) showed inhibition of H2 utilization in Methanobacterium formicium by high concentrations of total ammonia.

The digestor performance data from digestion of BSA-manure mixtures (Figs 4, 5, 6) indicate that there was metabolic stress caused by the protein, since the instability was greater with higher BSA content. The total ammonia concentrations in the digestors that received BSA-manure were similar to those used to study the effects of total ammonia concentrations. However, the fluctuations in volatile acid levels persisted throughout the experimental period in the BSA-manure digestions. The pH was higher in digestors that received protein. These observations indicate that the instability encountered was not due solely to inhibition by total ammonia concentration, and other factors must have caused the prolonged instability.

A C K N O W L E D G E M E N T

This work was supported by the Montana Department of Natural Resources and Conservation, Energy Bureau, Grant No. RAE 403-722 and the Montana Agricultural Experiment Station.

REFERENCES

American Public Health Association (1971). Standard Methodsjbr the Examination of Water and Wastewater, 13th edn, American Public Health Association, Washington, DC.

Page 14: Effects of total ammonia on anaerobic digestion and an example of digestor performance from cattle manure-protein mixtures

14 J. E. Robbins, S. A. Gerhardt, T. J. Kappel

Converse, J. D., Evans, G. W., Verhoeven, C. R., Gibbon, W. & Gibbon, M. (1977). Performance of a large size anaerobic digestor for poultry manure. ASAE paper no. 77-0451.

Goering, H. K. & van Soest, P. J. (1970). Forage Fiber Analysis. Agricultural Handbook No. 379, US Department of Agriculture, Washington, DC.

Hashimoto, A. G. (1983). Conversion of straw-manure mixtures to methane at mesophilic and thermophilic temperatures. Biotechnol. Bioeng., 25, 185-200.

Hills, D. J. (1979). Effects of carbon :nitrogen ratio on anerobic digestion of dairy manure. Agric. Wastes, 1,267-78.

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