World
Journal
of Microbiology
and Biotechnology,
9, 4!3-49
Thermophilic aerobic treatment potato-processing wastewater B. Malladi
of
and S.C. Ingham*
Four batches of potato-processing wastewater were collected every two weeks. After sedimentation for 1 h and subsequent decanting, the supernatant had a s-day biochemical oxygen demand (BOD) ranging from 620 to 1743 mg/l and total suspended solids ranging from 0.31 to 0.49 mg/ml. Thermophilic aerobic digestion (SST, 0.6 1 air/min) decreased the BOD and total suspended solids of the supematant by 98% and 75%, respectively, in 96 h. rCey words: Biochemical solids.
oxygen
demand,
potato-processing
In 1987, over 2.9 x lo9 kg of potatoes were produced in Canada, with a per capita consumption of 78 kg (Statistics Canada 1990). A major portion of the potatoes harvested are processed into such foods as french fries, potato chips, mashed potatoes, dehydrated potatoes, and perogies. The production of these foods entails the use of large volumes of water for unit operations such as washing and peeling. Waste-waters from potato-processing operations are characterized by high biochemical oxygen demand (BOD) and total suspended solids (TSS). A variety of biological processes have been studied to treat potato-processing wastewaters, many of which are anaerobic (Landine et al. 1983, 1986; Guo & Lin 1990; Wambeke et al. 1990). To our knowledge, thermophilic aerobic digestion of potato-processing wastewater has not been investigated. There are several advantages to thermophilic treatment of wastewater compared with treatment at lower temperatures. These advantages include a higher rate of microbial metabolic activity, leading to a shorter retention time, the ability of thermophilic microbes to metabolize a wide range of substrates, and the prevention of growth by pathogenic microorganisms (Slapack et al. 1987). The following report describes the effective decrease of BOD and TSS of potato-processing wastewater (PPW) by use of a thermophilic aerobic batch digestion system.
B Malladi and S.C. lngham are with the Department of Applied Microbiology and Food Science, University of Saskatchewan, Saskatoon, Saskatchewan. S7N OWO. Canada: FAX: 306-966-6896. *Corresponding author. @ 7993 Rapid
Communrcations
of Oxford
wastewater,
thermophilic
Materials
aerobic treatment,
total suspended
and Methods
Microorganisms
Microorganisms indigenous treatment process.
to the PPW were utilized in the
Digestion
Effluent (1.2 1) was collected in a pre-sterilized container from a potato-peeling machine at a local food plant. The container was sealed and transported to our laboratory. The PPW was allowed to settle for 60 min and the supematant was aseptically decanted to a sterile 2-1, jacketed Wheaton jar which was then sealed with a headplate modified to hold an autoclavable pH electrode, a stainless steel tube for aeration, and stainless steel wells for holding a temperature sensor and heating element. The fermenter was maintained at 55°C and aerated at 0.6 Urnin. The PPW was not mechanically stirred but some mixing resulted from the aeration. When the temperature of the PPW in the fermenter reached 55°C (in approximately 10 min), the treatment run began and the zero time sample was taken immediately. Thereafter, samples were taken at 24, 48, 72, and 96 h. Four runs, using PPW collected at two-week intervals, were done in this manner. Microbial
Enumwation
At each sampling time, 0.1 ml of PPW (diluted in sterile 0.1% (w/v) peptone water as necessary) was analysed for numbers of aerobic, amylolytic, and thennophilic spore-forming bacteria. All microbial enumerations were done in triplicate. The aerobic plate count was determined using the spread plate method on Plate Count Agar (Association of Official Analytical Chemists 1984). Bacteria with amylolytic activity were enumerated using a medium containing (w/v): 1.0% potato starch, 0.3% beef extract, and 2.0% agar. Spread plates were prepared and then incubated at 3i’OC for 2 days. Each
Ltd World ~oumal of Minobiology
and Biotechnology, Vof 9. 1993
45
B. Malladi
and S.C. lngham
plate was flooded with iodine and each colony with a surrounding clear (not blue) zone was counted as one ‘Zone Forming Unit’ (ZFU). Thermophilic, spore-forming bacteria were enumerated using tryptonelglucose/yeast extract/agar pour-plates. Plates were prepared after the sample was heat-shocked at 100°C for 20 min to induce activation of thermophilic spores. The prepared plates were incubated at 55°C for 48 h. For each sampling time, four colonies each were selected at random from the aerobic plate count, the amylolytic plate count, and the thermophilic spore-former count. A total of 60 bacterial isolates from the second run were identified by standard morphological and biochemical characteristics (Sneath 1989) with biochemical tests being performed using the API 20E System (API Analytab Products, New York). In the case of thermophilic isolates, the API test strips were incubated at 55°C. Effluent Charurferizufion Five-day BOD (BOD,) values for the effluent were determined on all samples using the azide modification (American Public Health Association 1985). Concentrations of total Kjeldahl N, total P, NO;, Cl-, SO:-, Ca’+, Mg2+ and Mr?+ were determined by the Saskatchewan Soil Testing Laboratory. Starch was measured enzymatically (Fleming & Reichert 1980) using a test kit (Sigma Chemical Company, St Louis, MO, USA) in which the sample was first extracted with ethanol to remove free sugars and then the starch was enzymatically digested by cc-amylase and amyloglucosidase. Glucose released was then analysed with a glucose electrode (Yellow Springs Instrument Co., Yellow Springs, OH, USA). TSS were measured by filtering 10.0 ml of the sample through a pre-weighed glass microfibre filter disc (Whatman 934AH; Whatman International Ltd, Maidstone, UK) followed by drying at 105°C to constant weight. TSS were calculated from the difference in weights.
Results BOD Figure
I shows
BOD
measurements
the
BOD profiles of runs 1 to 4, over 96 h. were made daily after the run had
A
0
I
20
40
80
Time
100
(hy”
Figure 1. Biochemical oxygen demand (BOD) reduction of potatoprocessing wastewater digested aerobically at 55°C for 96 h, during runs 1 (m), 2 (O), 3 (A) and 4 (A). Each point represents the mean of four replicates. Error bars representing SD are included in each point symbol; in most cases, the error bars are smaller than the point symbols and are not visible.
46
World Journal of Microbiology and
Biotechnology, Vol 9. 1993
0.5
0.4 = E
0.3
E 0.2 F 0.1
0.0 0
20
40 Time
60
60
1
(h)
Flgure 2. Reduction of total suspended solids (TSS) in potatoprocessing wastewater in 96 h, during runs 1 (0) 2 (O), 3 (m) and 4 (0). Each point represents the mean of duplicate values.
commenced. The major portion of the BOD decreases occurred in the first 48 h. The run was continued for another . 48 h whereupon further decrease of BOD was observed. During the first 48 h, an average BOD decrease of 82% was achieved, with a high of 95% and a low of 56%. Over 96 h, the fall in BOD averaged 98%. with a range of 96% to 99.8%. TSS Figure 2 shows that TSS gradually decreased over the first 48 h. An average TSS decrease of 75% occurred over 96 h, with a range of 69% to 80%. Starch Figure 3 shows the disappearance of starch during the four runs. On average, 89% of the starch disappeared during the first 48 h (range of 83% to 93%). By 96 h, an average of 96% of the starch in the PPW had disappeared (range of 95% to 98%). PH Figure 4A shows the pH profiles of the four runs. With one exception, pH of the PPW was close to neutrality at the start of the runs. pH profiles of the four runs were similar and showed a steep drop over the first 24 h, followed by an equally steep rise over the next 24 h. Thereafter, the pH plateaued between 8.5 and 9.0. Microbial Counts Figures 48 and 4C show the aerobic and amylolytic plate counts obtained during the four runs. These counts rose rapidly and peaked at 24 h. With the exception of run I, the counts did not appear to change between 72 and 96 h. Figure 4D shows the thermophilic spore-former counts obtained. An increase in counts was observed in all runs between 24 and 96 h, with the exception of run 4, which showed a decline after 72 h.
Thermophilic aerobic treatment of wastewafers
9 6 I
a 7
6 5 106 107 -i 2 5 0
20
40
60
Time Figure digestion 1 (A), values. symbol; symbol
60
106 105
100
(h)
3. Starch reduction profiles during thermophilic aerobic of potato-processing wastewater in 96 h, during runs 2 (n), 3 (0) and 4 (0). Each point represents triplicate Error bars representing SD are included in each point in most cases, the error bars are smaller than the point and are not visible.
104 103 10’ 107 E 3 K
106 105
Microbial Characfetizafion Characterization of 60 isolates from run 2 showed that 46 belonged to the genus Bacillus, seven to the genus Lacfobmillus, and the remaining seven were unidentifiable, Gram-positive rods (Table I). Effluent Characterization A summary of the nutrient and mineral analyses of the PPW is given in Table 2. The ratios of BOD: total Kjeldahl N: total P for the four runs are shown in Table 3.
104 103 109 108 107 < 106 3 2 106 104 103 102 10’
Discussion The results of this study indicate that thermophilic, aerobic digestion of PPW using indigenous microbes reduced BOD, TSS, and starch concentration by an average of 98%, 75% and 96%, respectively. It was observed that the decrease of BOD during the first 24 h of digestion was greatest in the PPW samples having the highest initial BOD. A similar trend was seen for the disappearance of starch. Samples with higher BOD and starch levels also had the greatest increases in aerobic and amylolytic plate counts over the first 24 h. No such relationship was observed between TSS reduction and microbial growth. Our findings suggest that this treatment system is more efficient at higher BOD and starch-loading rates.
0
20
40
60
Time
60
1 10
(h)
Figure 4. pH profiles (A), aerobic plate counts (B), in colony-forming units (c.f.u.), amylolytic plate counts (C), in zone-forming units (z.f.u.), and counts of thermophilic sporeformers (D), of potato-processing wastewater digested aerobically, during runs 1 (0) 2 (0) 3 (W) and 4 (0). Each point represents triplicate values. Error bars representing SD are included in each point symbol; in most cases, the error bars are smaller than the point symbol and are not visible.
It has been reported that efficient biological wastewater treatment requires a BOD : N : P ratio of 100 : 5 : 1 (Eckenfelder & O’Connor 1961). Although the BOD:N:P ratio in two batches of PPW was less than 1005: 1, because of low
World ]oumal
of
Microbiology and Bmtechnology, Vol 9, 1993
47
B. Malladi
and SC. lngham
Table 1. Microbial Isolates from potato-processing wastewater during thermophlllc aerobic dlgestlon at 55°C uslng standard test procedures [see Sneath (1989) and Methods and Materials]. Taxon
No. of Isolates
Bacillus stearothermophilus Bacillus brevis Bacillus licheniformis Bacillus coagulans Bacillus acidocaldarius Lactobacillus spp. Unidentifiable
17 14 6 5 4 7 7
Total
60
Table 2. Nutrient and mineral content (mgll) of potato-processing wastewater prior to thermophlllc aerobic dlgestlon at 55°C. Run no.
TKN
TP
NO;
Cl-
SO:-
Ca2+
Mg2+
Mn2+
1 2 3 4
100 200 100 100
6.1 16.2 3.1 13.3
3.2 9.2 17.7 6.0
61.3 98.1 71.0 77.0
62.4 86.5 75.1 79.7
19.0 17.9 15.5 19.3
14.5 21 .o 20.7 20.7
0.01 0.02 0.01 0.05
TKN-total
Table water
Kjeldahl
nitrogen;
3. Nutrlent balances prior to thermophlllc
TP-total
(mgll) aerobic
phosphorus.
In potato-processing digestion at 55°C.
counts were also obtained at 24 h (Figures 4B and 4C). The decrease in aerobic and amylolytic plate counts after 24 h coincided with the observed increase in pH (Figure 4A) and low levels of starch remaining in the PPW (Figure 2). The organisms predominating after 24 h may have been using carbon sources other than starch, with the resultant release of alkaline byproducts. Thermophilic spore-former counts peaked at either 72 or 96 h (Figure 4D). The predominant genus of bacteria throughout the digestion was Bacillus. Thus, the number of bacilli sporulating increased as the concentration of starch decreased. If a bacterial species is to maintain the same population in a given environment, it must be able to successfully compete with other species that may be present. Although aerobic plate, amylolytic plate and thermophilic sporeformer counts fluctuated during the digestion, several species of bacteria were isolated at each sampling time. It is possible to conclude that each of these species had effectively competed for a niche in the PPW environment. Further studies are needed to determine whether the relationship between simultaneously isolated species was mutualism or commensalism. The major drawback of an industrial-scale, thermophilic, aerobic digestion is the energy input necessary to maintain digester temperature. A crucial step in scaling up this thermophilic, aerobic digestion procedure would be to determine if the steam used for peeling potatoes could also be economically used to maintain the digester temperature.
waste-
Conclusions Run no.
BOD
TKN
TP
BOD:TKN:TP
1 2 3 4
1017 1356 1743 620
100 200 100 100
6.1 16.2 3.1 13.3
100: 9.8:0.6 100:14.7:1.2 100: 5.7:0.2 100: 16.1:2.1
BOD-Five-day dahl Nitrogen;
biochemical oxygen TP-total Phosphorus.
demand;
TKN-total
Kjel-
phosphorus concentration (Table 3), the decrease of BOD and TSS in these samples did not appear to be adversely affected. It is possible that the dominant Bacillus spp. isolated from the PPW during thermophilic aerobic digestion have an optimum BOD:N:P ratio that is not 100:5:1. Further studies are being conducted to determine if this is the case. The concentrations of several ions varied between batches of PPW (Table 2). The range of concentrations appeared not to be inhibitory to BOD, TSS or starch concentration reduction during the 96 h digestion. The pH of the PPW during digestion was most acidic (near 5.0) at 24 h and most alkaline (near 9.0) at 96 h (Figure 4). It appears that low pH was associated with microbial growth, because the highest aerobic and amylolytic plate
48
World Jouml
of
Mimobiology and Biotechnology, Vol 9, 1993
Thermophilic aerobic digestion, on a laboratory-scale, can effectively decrease the BOD, TSS and starch content of PPW. The indigenous microflora of the PPW during this digestion is predominantly Bacillus spp. Nutrient and mineral levels in PPW are sufficient for these indigenous organisms to reduce BOD, TSS, and starch concentration by 98%, 75% and 96%, respectively, during a 96 h digestion.
Acknowledgements This study was supported by The University of Saskatchewan, College of Agriculture Trust Fund. We thank Dr F.W. Sosulski of the University of Saskatchewan, Department of Crop Science and Plant Ecology, for the use of the Model 27 Glucose Analyzer.
References American Public Health Association 1985 Sfudurd Methods Emminafion of Wafer and Wastewafer, 16th edn. Washington APHA.
for the DC:
Themophilic Association of Official Analytical Chemists 1984 Bacteriological Analyfical Man&, Division of Microbiology, Center for Food Safety and Applied Nutrition, U.S. Food and Drug Administration, 6th edn. Arlington, VA: AOAC. Eckenfelder, W.W. Jr. & O’Connor, D.J. 1961 Biological Waste Treatment. Oxford: Pergamon Press. Fleming, S.E. & Reichert, R.D. 1980 Note on a modified method for the quantitative determination of starch. Cereal Chemistry, 57, 153-154. Guo, J. & Lin, K.C. 1990 Anaerobic treatment of potato-processing wastewater by a UASB system at low organic loadings. Water, Air and Soil Pollution 53, 367-377. Ito, K. & Thompson, P.J. 1984 Thermophilic flat sour sporeformers. In Compendium of Methods for the Microbiological Examination of Foods, ed Speck, M.L. pp. 242-250. Washington DC: American Public Health Association. Landine, R.C., Pyke, S.R., Brown, G.J. & Cocci, A.A. 1986 Low-rate anaerobic treatment of a potato processing plant effluent. In Proceedings of the 41st Industrial Waste Conference, pp. 511-519. Lafayette, IN: Purdue University.
aerobic treafmenf
of wastewafers
Landine, R.C., DeGairie, CJ., Cocci, A.A., Steeves, A.L., Brown, G.J. & Viraraghavan, T. 1983 Anaerobic pretreatment facility also provides sludge disposal capability and source of renewable energy for food processor. In Proceedings of the 38th Indwtria2 Waste Conference, pp. 805-815. Lafayette, IN: Purdue University. Slapack, G.E., Russel, I. & Stewart, G.G. I987 Themrophilic Microbes in Ethanol Production. Boca Raton, FL: CRC Press. Sneath, P.H.A. 1989 Bacillus. In Bergey’s Manual of Systemafic Bacferiology, Vol. 2, ed Holt, J.G., pp. 1104-1139. Baltimore, MD: Williams and Wilkins. Statistics Canada 1990 Canada Year Book 1990. Ottawa: Government of Canada. Wambeke, M.V., Grusenmeyer, S., Versrtraete, W. & Longry, R. 1990 Sludge bed growth in an UASB reactor treating potato processing wastewater. Process Biochemisfry International 25, 181-186.
(Received 22 April
1992; accepted 20 June 1992)
World Journal of Microbiology
and Biotechnology. Vol 9. 1993
49
Journal
of Microbiology
and Biotechnology,
9, 4!3-49
Thermophilic aerobic treatment potato-processing wastewater B. Malladi
of
and S.C. Ingham*
Four batches of potato-processing wastewater were collected every two weeks. After sedimentation for 1 h and subsequent decanting, the supernatant had a s-day biochemical oxygen demand (BOD) ranging from 620 to 1743 mg/l and total suspended solids ranging from 0.31 to 0.49 mg/ml. Thermophilic aerobic digestion (SST, 0.6 1 air/min) decreased the BOD and total suspended solids of the supematant by 98% and 75%, respectively, in 96 h. rCey words: Biochemical solids.
oxygen
demand,
potato-processing
In 1987, over 2.9 x lo9 kg of potatoes were produced in Canada, with a per capita consumption of 78 kg (Statistics Canada 1990). A major portion of the potatoes harvested are processed into such foods as french fries, potato chips, mashed potatoes, dehydrated potatoes, and perogies. The production of these foods entails the use of large volumes of water for unit operations such as washing and peeling. Waste-waters from potato-processing operations are characterized by high biochemical oxygen demand (BOD) and total suspended solids (TSS). A variety of biological processes have been studied to treat potato-processing wastewaters, many of which are anaerobic (Landine et al. 1983, 1986; Guo & Lin 1990; Wambeke et al. 1990). To our knowledge, thermophilic aerobic digestion of potato-processing wastewater has not been investigated. There are several advantages to thermophilic treatment of wastewater compared with treatment at lower temperatures. These advantages include a higher rate of microbial metabolic activity, leading to a shorter retention time, the ability of thermophilic microbes to metabolize a wide range of substrates, and the prevention of growth by pathogenic microorganisms (Slapack et al. 1987). The following report describes the effective decrease of BOD and TSS of potato-processing wastewater (PPW) by use of a thermophilic aerobic batch digestion system.
B Malladi and S.C. lngham are with the Department of Applied Microbiology and Food Science, University of Saskatchewan, Saskatoon, Saskatchewan. S7N OWO. Canada: FAX: 306-966-6896. *Corresponding author. @ 7993 Rapid
Communrcations
of Oxford
wastewater,
thermophilic
Materials
aerobic treatment,
total suspended
and Methods
Microorganisms
Microorganisms indigenous treatment process.
to the PPW were utilized in the
Digestion
Effluent (1.2 1) was collected in a pre-sterilized container from a potato-peeling machine at a local food plant. The container was sealed and transported to our laboratory. The PPW was allowed to settle for 60 min and the supematant was aseptically decanted to a sterile 2-1, jacketed Wheaton jar which was then sealed with a headplate modified to hold an autoclavable pH electrode, a stainless steel tube for aeration, and stainless steel wells for holding a temperature sensor and heating element. The fermenter was maintained at 55°C and aerated at 0.6 Urnin. The PPW was not mechanically stirred but some mixing resulted from the aeration. When the temperature of the PPW in the fermenter reached 55°C (in approximately 10 min), the treatment run began and the zero time sample was taken immediately. Thereafter, samples were taken at 24, 48, 72, and 96 h. Four runs, using PPW collected at two-week intervals, were done in this manner. Microbial
Enumwation
At each sampling time, 0.1 ml of PPW (diluted in sterile 0.1% (w/v) peptone water as necessary) was analysed for numbers of aerobic, amylolytic, and thennophilic spore-forming bacteria. All microbial enumerations were done in triplicate. The aerobic plate count was determined using the spread plate method on Plate Count Agar (Association of Official Analytical Chemists 1984). Bacteria with amylolytic activity were enumerated using a medium containing (w/v): 1.0% potato starch, 0.3% beef extract, and 2.0% agar. Spread plates were prepared and then incubated at 3i’OC for 2 days. Each
Ltd World ~oumal of Minobiology
and Biotechnology, Vof 9. 1993
45
B. Malladi
and S.C. lngham
plate was flooded with iodine and each colony with a surrounding clear (not blue) zone was counted as one ‘Zone Forming Unit’ (ZFU). Thermophilic, spore-forming bacteria were enumerated using tryptonelglucose/yeast extract/agar pour-plates. Plates were prepared after the sample was heat-shocked at 100°C for 20 min to induce activation of thermophilic spores. The prepared plates were incubated at 55°C for 48 h. For each sampling time, four colonies each were selected at random from the aerobic plate count, the amylolytic plate count, and the thermophilic spore-former count. A total of 60 bacterial isolates from the second run were identified by standard morphological and biochemical characteristics (Sneath 1989) with biochemical tests being performed using the API 20E System (API Analytab Products, New York). In the case of thermophilic isolates, the API test strips were incubated at 55°C. Effluent Charurferizufion Five-day BOD (BOD,) values for the effluent were determined on all samples using the azide modification (American Public Health Association 1985). Concentrations of total Kjeldahl N, total P, NO;, Cl-, SO:-, Ca’+, Mg2+ and Mr?+ were determined by the Saskatchewan Soil Testing Laboratory. Starch was measured enzymatically (Fleming & Reichert 1980) using a test kit (Sigma Chemical Company, St Louis, MO, USA) in which the sample was first extracted with ethanol to remove free sugars and then the starch was enzymatically digested by cc-amylase and amyloglucosidase. Glucose released was then analysed with a glucose electrode (Yellow Springs Instrument Co., Yellow Springs, OH, USA). TSS were measured by filtering 10.0 ml of the sample through a pre-weighed glass microfibre filter disc (Whatman 934AH; Whatman International Ltd, Maidstone, UK) followed by drying at 105°C to constant weight. TSS were calculated from the difference in weights.
Results BOD Figure
I shows
BOD
measurements
the
BOD profiles of runs 1 to 4, over 96 h. were made daily after the run had
A
0
I
20
40
80
Time
100
(hy”
Figure 1. Biochemical oxygen demand (BOD) reduction of potatoprocessing wastewater digested aerobically at 55°C for 96 h, during runs 1 (m), 2 (O), 3 (A) and 4 (A). Each point represents the mean of four replicates. Error bars representing SD are included in each point symbol; in most cases, the error bars are smaller than the point symbols and are not visible.
46
World Journal of Microbiology and
Biotechnology, Vol 9. 1993
0.5
0.4 = E
0.3
E 0.2 F 0.1
0.0 0
20
40 Time
60
60
1
(h)
Flgure 2. Reduction of total suspended solids (TSS) in potatoprocessing wastewater in 96 h, during runs 1 (0) 2 (O), 3 (m) and 4 (0). Each point represents the mean of duplicate values.
commenced. The major portion of the BOD decreases occurred in the first 48 h. The run was continued for another . 48 h whereupon further decrease of BOD was observed. During the first 48 h, an average BOD decrease of 82% was achieved, with a high of 95% and a low of 56%. Over 96 h, the fall in BOD averaged 98%. with a range of 96% to 99.8%. TSS Figure 2 shows that TSS gradually decreased over the first 48 h. An average TSS decrease of 75% occurred over 96 h, with a range of 69% to 80%. Starch Figure 3 shows the disappearance of starch during the four runs. On average, 89% of the starch disappeared during the first 48 h (range of 83% to 93%). By 96 h, an average of 96% of the starch in the PPW had disappeared (range of 95% to 98%). PH Figure 4A shows the pH profiles of the four runs. With one exception, pH of the PPW was close to neutrality at the start of the runs. pH profiles of the four runs were similar and showed a steep drop over the first 24 h, followed by an equally steep rise over the next 24 h. Thereafter, the pH plateaued between 8.5 and 9.0. Microbial Counts Figures 48 and 4C show the aerobic and amylolytic plate counts obtained during the four runs. These counts rose rapidly and peaked at 24 h. With the exception of run I, the counts did not appear to change between 72 and 96 h. Figure 4D shows the thermophilic spore-former counts obtained. An increase in counts was observed in all runs between 24 and 96 h, with the exception of run 4, which showed a decline after 72 h.
Thermophilic aerobic treatment of wastewafers
9 6 I
a 7
6 5 106 107 -i 2 5 0
20
40
60
Time Figure digestion 1 (A), values. symbol; symbol
60
106 105
100
(h)
3. Starch reduction profiles during thermophilic aerobic of potato-processing wastewater in 96 h, during runs 2 (n), 3 (0) and 4 (0). Each point represents triplicate Error bars representing SD are included in each point in most cases, the error bars are smaller than the point and are not visible.
104 103 10’ 107 E 3 K
106 105
Microbial Characfetizafion Characterization of 60 isolates from run 2 showed that 46 belonged to the genus Bacillus, seven to the genus Lacfobmillus, and the remaining seven were unidentifiable, Gram-positive rods (Table I). Effluent Characterization A summary of the nutrient and mineral analyses of the PPW is given in Table 2. The ratios of BOD: total Kjeldahl N: total P for the four runs are shown in Table 3.
104 103 109 108 107 < 106 3 2 106 104 103 102 10’
Discussion The results of this study indicate that thermophilic, aerobic digestion of PPW using indigenous microbes reduced BOD, TSS, and starch concentration by an average of 98%, 75% and 96%, respectively. It was observed that the decrease of BOD during the first 24 h of digestion was greatest in the PPW samples having the highest initial BOD. A similar trend was seen for the disappearance of starch. Samples with higher BOD and starch levels also had the greatest increases in aerobic and amylolytic plate counts over the first 24 h. No such relationship was observed between TSS reduction and microbial growth. Our findings suggest that this treatment system is more efficient at higher BOD and starch-loading rates.
0
20
40
60
Time
60
1 10
(h)
Figure 4. pH profiles (A), aerobic plate counts (B), in colony-forming units (c.f.u.), amylolytic plate counts (C), in zone-forming units (z.f.u.), and counts of thermophilic sporeformers (D), of potato-processing wastewater digested aerobically, during runs 1 (0) 2 (0) 3 (W) and 4 (0). Each point represents triplicate values. Error bars representing SD are included in each point symbol; in most cases, the error bars are smaller than the point symbol and are not visible.
It has been reported that efficient biological wastewater treatment requires a BOD : N : P ratio of 100 : 5 : 1 (Eckenfelder & O’Connor 1961). Although the BOD:N:P ratio in two batches of PPW was less than 1005: 1, because of low
World ]oumal
of
Microbiology and Bmtechnology, Vol 9, 1993
47
B. Malladi
and SC. lngham
Table 1. Microbial Isolates from potato-processing wastewater during thermophlllc aerobic dlgestlon at 55°C uslng standard test procedures [see Sneath (1989) and Methods and Materials]. Taxon
No. of Isolates
Bacillus stearothermophilus Bacillus brevis Bacillus licheniformis Bacillus coagulans Bacillus acidocaldarius Lactobacillus spp. Unidentifiable
17 14 6 5 4 7 7
Total
60
Table 2. Nutrient and mineral content (mgll) of potato-processing wastewater prior to thermophlllc aerobic dlgestlon at 55°C. Run no.
TKN
TP
NO;
Cl-
SO:-
Ca2+
Mg2+
Mn2+
1 2 3 4
100 200 100 100
6.1 16.2 3.1 13.3
3.2 9.2 17.7 6.0
61.3 98.1 71.0 77.0
62.4 86.5 75.1 79.7
19.0 17.9 15.5 19.3
14.5 21 .o 20.7 20.7
0.01 0.02 0.01 0.05
TKN-total
Table water
Kjeldahl
nitrogen;
3. Nutrlent balances prior to thermophlllc
TP-total
(mgll) aerobic
phosphorus.
In potato-processing digestion at 55°C.
counts were also obtained at 24 h (Figures 4B and 4C). The decrease in aerobic and amylolytic plate counts after 24 h coincided with the observed increase in pH (Figure 4A) and low levels of starch remaining in the PPW (Figure 2). The organisms predominating after 24 h may have been using carbon sources other than starch, with the resultant release of alkaline byproducts. Thermophilic spore-former counts peaked at either 72 or 96 h (Figure 4D). The predominant genus of bacteria throughout the digestion was Bacillus. Thus, the number of bacilli sporulating increased as the concentration of starch decreased. If a bacterial species is to maintain the same population in a given environment, it must be able to successfully compete with other species that may be present. Although aerobic plate, amylolytic plate and thermophilic sporeformer counts fluctuated during the digestion, several species of bacteria were isolated at each sampling time. It is possible to conclude that each of these species had effectively competed for a niche in the PPW environment. Further studies are needed to determine whether the relationship between simultaneously isolated species was mutualism or commensalism. The major drawback of an industrial-scale, thermophilic, aerobic digestion is the energy input necessary to maintain digester temperature. A crucial step in scaling up this thermophilic, aerobic digestion procedure would be to determine if the steam used for peeling potatoes could also be economically used to maintain the digester temperature.
waste-
Conclusions Run no.
BOD
TKN
TP
BOD:TKN:TP
1 2 3 4
1017 1356 1743 620
100 200 100 100
6.1 16.2 3.1 13.3
100: 9.8:0.6 100:14.7:1.2 100: 5.7:0.2 100: 16.1:2.1
BOD-Five-day dahl Nitrogen;
biochemical oxygen TP-total Phosphorus.
demand;
TKN-total
Kjel-
phosphorus concentration (Table 3), the decrease of BOD and TSS in these samples did not appear to be adversely affected. It is possible that the dominant Bacillus spp. isolated from the PPW during thermophilic aerobic digestion have an optimum BOD:N:P ratio that is not 100:5:1. Further studies are being conducted to determine if this is the case. The concentrations of several ions varied between batches of PPW (Table 2). The range of concentrations appeared not to be inhibitory to BOD, TSS or starch concentration reduction during the 96 h digestion. The pH of the PPW during digestion was most acidic (near 5.0) at 24 h and most alkaline (near 9.0) at 96 h (Figure 4). It appears that low pH was associated with microbial growth, because the highest aerobic and amylolytic plate
48
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Thermophilic aerobic digestion, on a laboratory-scale, can effectively decrease the BOD, TSS and starch content of PPW. The indigenous microflora of the PPW during this digestion is predominantly Bacillus spp. Nutrient and mineral levels in PPW are sufficient for these indigenous organisms to reduce BOD, TSS, and starch concentration by 98%, 75% and 96%, respectively, during a 96 h digestion.
Acknowledgements This study was supported by The University of Saskatchewan, College of Agriculture Trust Fund. We thank Dr F.W. Sosulski of the University of Saskatchewan, Department of Crop Science and Plant Ecology, for the use of the Model 27 Glucose Analyzer.
References American Public Health Association 1985 Sfudurd Methods Emminafion of Wafer and Wastewafer, 16th edn. Washington APHA.
for the DC:
Themophilic Association of Official Analytical Chemists 1984 Bacteriological Analyfical Man&, Division of Microbiology, Center for Food Safety and Applied Nutrition, U.S. Food and Drug Administration, 6th edn. Arlington, VA: AOAC. Eckenfelder, W.W. Jr. & O’Connor, D.J. 1961 Biological Waste Treatment. Oxford: Pergamon Press. Fleming, S.E. & Reichert, R.D. 1980 Note on a modified method for the quantitative determination of starch. Cereal Chemistry, 57, 153-154. Guo, J. & Lin, K.C. 1990 Anaerobic treatment of potato-processing wastewater by a UASB system at low organic loadings. Water, Air and Soil Pollution 53, 367-377. Ito, K. & Thompson, P.J. 1984 Thermophilic flat sour sporeformers. In Compendium of Methods for the Microbiological Examination of Foods, ed Speck, M.L. pp. 242-250. Washington DC: American Public Health Association. Landine, R.C., Pyke, S.R., Brown, G.J. & Cocci, A.A. 1986 Low-rate anaerobic treatment of a potato processing plant effluent. In Proceedings of the 41st Industrial Waste Conference, pp. 511-519. Lafayette, IN: Purdue University.
aerobic treafmenf
of wastewafers
Landine, R.C., DeGairie, CJ., Cocci, A.A., Steeves, A.L., Brown, G.J. & Viraraghavan, T. 1983 Anaerobic pretreatment facility also provides sludge disposal capability and source of renewable energy for food processor. In Proceedings of the 38th Indwtria2 Waste Conference, pp. 805-815. Lafayette, IN: Purdue University. Slapack, G.E., Russel, I. & Stewart, G.G. I987 Themrophilic Microbes in Ethanol Production. Boca Raton, FL: CRC Press. Sneath, P.H.A. 1989 Bacillus. In Bergey’s Manual of Systemafic Bacferiology, Vol. 2, ed Holt, J.G., pp. 1104-1139. Baltimore, MD: Williams and Wilkins. Statistics Canada 1990 Canada Year Book 1990. Ottawa: Government of Canada. Wambeke, M.V., Grusenmeyer, S., Versrtraete, W. & Longry, R. 1990 Sludge bed growth in an UASB reactor treating potato processing wastewater. Process Biochemisfry International 25, 181-186.
(Received 22 April
1992; accepted 20 June 1992)
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