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Water and Environment Journal. Print ISSN 1747-6585

Effects of carbon dioxide deficiency and metal toxicity on biological oxygen demand
Mehmet Berkun
Department of Civil Engineering, Karadeniz Technical University, Trabzon, Turkey

Keywords autotrophism; bacteria; BOD; carbon dioxide; metal toxicity; nitrication; oxygen uptake; respirometer. Correspondence Mehmet Berkun, Department of Civil Engineering, Karadeniz Technical University, 61080 Trabzon, Turkey. Email: berkun@ktu.edu.tr doi:10.1111/j.1747-6593.2009.00193.x

Abstract
In this study, the effects of carbon dioxide (CO2) deciency on the biological growth under respirometric conditions, rst- and second-order biological oxygen demand (BOD) progressions using two different BOD measurement techniques and metal toxicity effects on the respirometric BOD are investigated. The effects of CO2 deciency in the growth of bacteria and related effects on the rst- and second-stage BOD progressions are investigated using various media with respirometers in comparison with the BOD dilution method. CO2 deciency causes signicant retardations on the growth of bacteria and the second-stage respirometric BOD values are suppressed. CO2 seems to be an essential nutrient for the growth of microorganisms and for the oxygen uptake progressing rates. HgCl2, HgSO4, CuSO4, K2Cr2O7, ZnSO4 and Al2(SO4)3 inorganic metal compounds cause signicant retardations in the respirometric BOD values obtained from a synthetic autotrophic medium. Effects are found to be dependent on the applied concentrations of these chemicals in the medium.

Introduction
The biological oxygen demand (BOD) test has its widest application in measuring waste loading to treatment plants, in evaluating the BOD removal efciency of such treatment plants and in river water quality modelling studies (Adrian & Sanders 1998; Adrian et al. 1999, 2004; Roider & Adrian 2007). The environmental signicance of biodegradation studies (Orhon et al. 1995) as well as the complexity of phenomena that are involved in biodegradation reactions have initiated research since the late 1800s. Determination of the 5-day BOD as an important water quality indicator using the standard BOD method has been the most popular experimental procedure for a number of years. Because BOD tests are usually designed for a 5-day period, formation of the rst-stage BOD phase including the lag and plateau, and its kinetics has been extensively investigated (Cailas & Gehr 1989a, b). Attempts have been made to eliminate some of the technical disadvantages of the standard BOD method and to develop alternative techniques (respirometers) to provide a more realistic approach to the measurement of oxygen demand in treatment plants (Tebbutt & Berkun 1976; Arthur 1984). The experimental procedures that have gained popularity and that are intimately related to the

BOD principle are the manometric and electrolytic respirometers to monitor the progression of biodegradation reactions in terms of oxygen uptake (Young & Baumann 1976; Larson & Perry 1980; Suschka & Ferreira 1986; Chandran et al. 1996; Ellis et al. 1996; Li & Zhang 1996; Cecen & Yangin 2000, 2001). In these techniques, carbon dioxide (CO2) produced should be sufciently removed by an alkali solution so that the BOD determinations are not affected. However, the possibility of the resulting CO2 deciency in the reaction asks is also a matter of concern because autotrophic activity especially in the rst- and second-order BOD phases is closely related to the existence of sufcient CO2 in the medium. Investigations that have been carried out on the assimilation of CO2 by various microorganisms can be divided into two main groups such as mixed or pure culture studies (Harris 1954; Mattman 1954; Gaffney 1965) and radioactive carbon studies (Abelson et al. 1952). Although these studies showed the vital importance of the presence of CO2 in the media for the growth of heterotrophic and autotrophic bacteria, information is lacking on the formation of the rst-stage and especially the second-stage BOD as being mainly an autotrophic phase, related to different BOD measurement methods. In respirometers, CO2 produced during the BOD reaction is removed by an alkali
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solution, but in the standard dilution method it remains in the reaction asks. Investigations carried out on the effects of toxicity on the biodegradability of wastes using large volume respirometers, besides their experimental convenience, questioned the meaning and reliability of these efforts (King & Dutka 1986; De Bel et al. 1996). Although some researchers investigated the effects of some inorganic metal compounds on BOD (Berkun 1982, 2005; Yetis et al. 1992; Albek et al. 1997; Gernaey et al. 1997), knowledge is lacking on the effects exerted on the autotrophic BOD including the second-stage BOD phase. For example, nitrifying bacteria, being autotrophic and the causative agent of nitrication, must use CO2 for their growth. The aim of this research is to investigate the BOD progression and growth of bacteria in autotrophic and other media using different BOD measurement methods and an aeration system having respirometric conditions related to CO2 deciency. The effects of metal toxicity in respirometric BOD measurements using an autotrophic medium were also relatively investigated.

Materials and methods

To provide information about the effects of CO2 levels on bacteriological activity, an aeration train was set up as shown in Fig. 1 to provide normal air and CO2-scrubbed air under otherwise identical conditions. Experiments were carried out with 300 mL samples. The apparatus could be considered as a constant pressure respirometer because excess air pressure in the reaction vessel is discharged via the water trap in the second vessel. Air was scrubbed of CO2 by bubbling through two traps containing 15% sodium hydroxide and a water trap prevented transfer of alkaline spray into the reaction vessel. The reaction vessel itself contained a conventional absorber boat having a 20% KOH solution. Synthetic medium (Table 1) and crude sewage samples were buffered at pH =7 with a phosphate system and inoculated with 1% by volume crude sewage. In the respirometric experiments, two sample bottles were used with KOH and other two without KOH so that a comparison could be made. The growth of bacteria was investigated by turbidity and plate count determinations with standard nutrient agar medium. In the tables, plate counts and turbidity

data were given as the mean value of three experiments. BOD experiments were carried out with a large volume respirometer (Hach Model 2173 A, Hach Chemical, Loveland, CO, USA) and with the dilution method so that a comparison could be made. Dilution BOD determinations were carried out on the same samples as those used for the respirometer studies using standard procedures. Dissolved oxygen (DO) measurements were made with an oxygen electrode. BOD experiments for mixed cultures were carried out using crude sewage samples and bacteriological-grade dissolved (1.5 mg/L) meat extract medium added to the autotrophic medium (Table 2) excluding (NH4)2SO4 to prevent the excessive stimulation of the growth of nitrifying bacteria. The pH value of this autotrophic medium is 8.2 and is adjusted to 8 to make it more suitable for the growth of nitrifying bacteria (Hoffman & Lees 1953; Siddigi et al. 1967). Variations of NO2 and NO3 values in the samples were measured with a spectrophotometer (Hach Model DR-EL/2, Hach Chemical, Loveland, CO, USA). A culture of autotrophic bacteria was prepared by seeding the autotrophic medium (Table 2) with the soil samples collected near grass roots. Inoculated samples were continuously stirred and held under aerobic conditions for 15 days by renewing the autotrophic medium after 7 days. Different concentrations of HgCl2, HgSO4, CuSO4, K2Cr2O7, ZnSO4 and Al2(SO4)3 were placed in the autotrophic medium in the respirometer sample bottles and were inoculated 0.2% with an autotrophic bacteria culture. All experiments were carried out at 20 1C.

Results and discussion

Conventional BOD measuring tests are problematically slow, typically intended to last 5 or 7 days. Conventional tests are also affected by factors such as dilution and sample preparation. On the other hand, conventional BOD tests have some benets as compared with new, rapid, automatic and analytical techniques and they do not require expensive equipment. As a result of the increasing demand considering, for example, biodegradability, it is very important to develop automatic, faster and more precise ways for BOD determination. BOD results obtained from different methods must subsequently be compared with the conventional methods in order to detect the differences and to nd the reasons for

Fig. 1. Aeration system.

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them. The effect of CO2 content in the BOD reaction vessels on the biological growth and on the rate of oxygen uptake is of considerable interest.

Effect of CO2 on the growth of mixed microorganisms

The growth of microorganisms in synthetic medium in the large volume respirometer was determined by plate

Table 1 Synthetic medium (NH4)2SO4 (NH4)2H2PO4 KH2PO4 K2HPO4 NaCl MgSO4 FeSO4 CaCl2 C6H12O6 H2O 0.4 g 1.0 g 2.0 g 2.0 g 1.5 g 0.2 g 0.02 g 0.5 g 1.0 g 1L

counts. The plate counts showed that CO2 deciency exerted a considerable effect on the growth of bacteria. The results obtained after 24 h from the samples without KOH were, in most cases, greater than those obtained from the samples with KOH (Table 3). The growth of bacteria was investigated by turbidity and plate count determinations using the aeration system (Fig. 1). In all cases, the normal air produced turbidity levels and plate counts were greater than those obtained with the CO2scrubbed air (Table 4). Repeated experiments carried out with the respirometer using sterilized crude sewage samples (boiled for 25 min and inoculated 1% with crude sewage) showed that growth was suppressed under CO2-decient conditions (Table 5).

Effect of CO2 deciency on the growth of heterotrophic microorganisms

This investigation was carried out with a synthetic medium consisting of glucose (1 mg/L) and adenine (0.4 mg/L) in a large volume respirometer. Adenine was used to supply the organic nitrogen and hydrogen requirements of the microorganisms. Inorganic chemicals were not used in order to prevent the growth of chemoautotrophic microorganisms. Plate counts obtained from the samples without KOH under the respirometric conditions were, in most cases, higher than the results obtained from the samples with KOH (Table 6).

Table 2 Autotrophic medium (NH4)2SO4 K2HPO4 MgSO4 FeCl3 CaCO3 NaHCO3 H2O 56.0 mg/L 3.0 mg/L 25.0 mg/L 0.125 mg/L 50.0 mg/L 250.0 mg/L 1L

Table 3 Plate counts obtained from respirometer using synthetic medium Initial plate counts per mL With KOH Sample volume (mL) 200 Test 1 2 3 4 22 1C o30 o30 o30 o30 Without KOH 37 1C o30 o30 o30 o30 After 24 h plate counts per mL With KOH 22 1C o30 4 300 70 128 37 1C o30 125 80 78 Without KOH 22 1C 4 300 4 300 4 300 4 300 37 1C 182 4 300 4 300 4 300

Table 4 Turbidity and plate counts obtained from synthetic medium using the aeration system Initial After 24 h Plate counts per mL Sample volume (mL) 300 Test 1 2 3 Atmosphere Normal air CO2-scrubbed air Normal air CO2-scrubbed air Normal air CO2-scrubbed air pH 7.0 7.0 7.0 7.0 7.0 7.0 Turbidity FTU 10 10 80 80 15 15 pH 7.0 7.0 7.0 7.0 7.0 7.0 Turbidity FTU 30 10 150 80 40 15 22 1C 271 120 144 57 78 22 37 1C 200 83 104 31 4 300 70

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Effect of CO2 deciency on the growth of chemoautotrophic microorganisms

This investigation was carried out with the synthetic medium in which only the inorganic chemicals (Table 2) were placed in the large volume respirometer. Organic materials were not used in order to prevent the growth of heterotrophs. In contrast to the results obtained previously from mixed and heterotrophic microorganisms, no growth as determined from plate counts was obtained after 24 and 48 h. But with sterilized crude sewage samples, the effect of CO2 deciency on the growth remained for 48 h (Table 5). The experimental results given in this research showed that the biological activity was delayed by a CO2 deciency. Although this effect on the growth of bacteria can be measured, the biological pathways of CO2 assimilation by microorganisms are still not completely known. Because of the fast oxidation rate of glucose and rapid multiplication of bacteria, this effect could not be studied beyond 24 h. For respirometry, a continuous and efcient removal of CO2 should be maintained in the reaction asks so that CO2 does not affect the pressure balances in the system. But this can cause a CO2 deciency and as a result growth and oxygen uptake are affected. Some investigators like Gaffney (1965) attempted to control CO2 variations in the reaction asks of small volume respirometers, but the applicability of chemicals used in these studies is still questionable. An experimental study using the aeration unit also showed that CO2 deciency affected the growth. Early investigations indicated that, when 0.31.0% CO2
Table 5 Plate counts obtained from sterilized crude sewage samples using respirometer Sample volume (mL) Test 300 1 2 3 After 24 h Atmosphere Normal air CO2-scrubbed air Normal air CO2-scrubbed air Normal air CO2-scrubbed air 22 1C 4 300 o30 4 300 165 4 300 o30 37 1C 118 o30 183 78 4 300 o30 After 48 h 22 1C 4 300 4 300 4 300 4 300 4 300 4 300 37 1C 4 300 123 140 50 135 69

was present in the sample bottles, which is more than normally present in the atmosphere, growth was stimulated. The results obtained from the heterotrophic medium are important because they show that CO2 is an essential nutrient for the growth of heterotrophs as well as autotrophs.

First- and second-stage BOD curves

Although formation of the BOD plateau was investigated using the standard dilution method and Warburg apparatus (Bush 1958; Bhatla & Gaudy 1966), no effort has been made for a comparative study using different BOD measurement methods including the formation of the secondstage BOD and plateau phases. The second-stage BOD values obtained from the large volume respirometer using crude sewage (Fig. 2) and synthetic medium (Fig. 3) were depressed in comparison with the values obtained from the standard dilution method. Experiments carried out under respirometric conditions using the autotrophic medium seeded with autotrophic bacteria showed that the growth of the autotrophic bacteria was delayed more than 6 days under CO2-decient conditions of the respirometer in comparison with the normal atmospheric conditions (Fig. 4). Conrming time-dependent variations of related NO3 (Figs 5 and 6) and NO2 (Figs 7 and 8) values obtained from autotrophic and crude sewage media were given.

Effects of inorganic metal compounds

The effects of inorganic metal compounds on the BOD values obtained from the autotrophic bacteria-seeded autotrophic medium (Table 2) showed the following order in terms of toxicity effects: HgCl2 4HgSO4 4CuSO4 4K2 Cr2 O7 4ZnSO4 4Al2 SO4 3 BOD of this medium was completely inhibited for 5 days due to the presence of the following concentrations of metal compounds: 1.25 mg/L HgCl2, 1.75 mg/L HgSO4, 5 mg/L CuSO4 and 5 mg/L K2Cr2O7, but ZnSO4 did not completely suppress the BOD for 5 days when used up to

Table 6 Plate counts obtained from respirometer using synthetic medium (glucose+adenine) Initial plate counts per mL With KOH Sample volume (mL) 200 Test 1 2 3 4 22 1C o30 o30 o30 o30 Without KOH 37 1C o30 o30 o30 o30 After 24 h plate counts per mL With KOH 22 1C 216 81 117 120 37 1C 56 o30 o30 o30 Without KOH 22 1C 4 300 4 300 4 300 4 300 37 1C 4 300 109 223 175

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900 800 700 600 500 400 300 200 100 0 0

BOD (mg/L)

From respirometer From dilution tec.

Time (days) 1 2 3 4 5 6 7 8 9 10 11 12 13

Fig. 4. Biological oxygen demand (BOD) values obtained from crude sewage samples. Fig. 2. Colony counts obtained from autotrophic medium using respirometer.

700 600

3.5 NO (mg/L) 3 BOD (mg/L) 3

Atmospheric conditions

2.5 500 400 300 From respirometer 200 100 Time (days) 0 0 1 2 3 4 5 6 7 8 9 10 11 12 13
Fig. 3. Biological oxygen demand (BOD) values obtained from meat extract media.

Respirometric conditions

1.5

From dilution tec.

0.5 Time (days) 0 4 5 6 7 8 9 10 11 12 13

Fig. 5. NO3 values obtained from autotrophic medium under atmospheric and respirometric conditions.

80 mg/L. Information indicating the dependence of the effects on the metal concentrations is presented in Fig. 9. These concentrations are lower than the values given for the rst-order BOD values in the literature. More complex, nutrient-rich and active microbial structure of the rst-stage BOD phase could be more suitable for the acclimatization and growth of microorganisms.

Conclusions
(1) In the existence of a CO2 deciency under normal atmospheric and respirometric conditions, growth of

microorganisms in mixed, heterotrophic and autotrophic culture media was drastically suppressed. In contrast to the rst-stage respirometric BOD values, second-stage BOD values of synthetic and crude sewage were lower than the BOD values obtained from the BOD dilution method. This indicated that nitrication effects that have been a matter of concern in BOD measurements will be less in respirometric measurements than in the dilution method. Although the dilution method and a number of developed respirometers have been successfully used in biodegradability investigations, the importance of the existing CO2 concentrations in the closed sample bottles of the dilution method and respirometers still exists as a
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NO3 (mg/L)

240 220

BOD5 (mg/L) ZnSO4

2.5
Atmospheric conditions Respirometric conditions

200 180 160 K2Cr2O7 CuSO4

1.5

140 120

1 100 0.5 Time (days) 60 0 5 6 7 8 9 10 11 12 13

Fig. 6. NO3 values obtained from crude sewage medium under atmospheric and respirometric conditions.

80 HgSO4

40 20 0 0 1 2 3 Metal concentration (mg/L) 4 5 HgCl2

Fig. 9. Effects of inorganic compounds on respirometric biological oxygen demand (BOD) obtained from autotrophic medium.

complicating and effective factor on the reliability of BOD and oxygen uptake rates. (2) Respirometric BOD values obtained from autotrophic synthetic medium were affected by the presence of inorganic metal compounds. Toxicity effects were dependent on the type and concentrations of the used inorganic chemicals. The concentrations of the inorganic chemical applied that completely inhibited the BOD values for 5 days were 1.25 mg/L HgCl2, 1.75 mg/L HgSO4, 5 mg/L CuSO4 and 5 mg/L K2Cr2O7. But ZnSO4 did not completely suppress the BOD for 5 days when used up to 80 mg/L.
Fig. 7. NO2 values obtained from autotrophic medium under atmospheric and respirometric conditions.

Acknowledgement
This research was supported by the research fund of Karadeniz (Black Sea) Technical University.
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References
Abelson, P.H., Bolton, E.T. and Aldous, E. (1952) Utilization of Carbon Dioxide in the Synthesis of Proteins by Escherichia coli. J. Biol. Chem., 198 (1), 165172. Adrian, D.D. and Sanders, G.T. (1998) Oxygen Sag Equation for Second Order BOD Decay. Water Res., 32 (3), 840848. Adrian, D.D., Sanders, T.G. and Roider, E.M. (1999) Oxygen Sag Equation for Three-Halves Order BOD Reaction. J. Am. Water Resour. Assoc., 35 (5), 11911200.

Fig. 8. NO3 values obtained from crude sewage medium under atmospheric and respirometric conditions.

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Adrian, D.D., Roider, E.M. and Sanders, T.G. (2004) Oxygen Sag Models for Multiorder Biochemical Oxygen Demand Reactions. J. Environ. Eng. ASCE, 130 (7), 784791. Albek, M., Yetis, U. and Gokcay, C.F. (1997) Effects of Ni(II) on Respirometric Oxygen Uptake. Appl. Microbiol. Biotechnol., 48 (5), 636641. Arthur, R.M. (1984) Twenty Years of Respirometry. In Bell, J.M. (ed). Proceedings 39th Purdue Industrial Waste Conference, pp. 861874. Lewis, Chelsea, MI. Berkun, M. (1982) Effects of Inorganic Metal Toxicity on BODI, Methods for the Estimation of BOD Parameters-II. Water Res., 16, 559564. Berkun, M. (2005) Effects of Ni, Cr, Hg, Cu, Zn, Al on the Dissolved Oxygen Balance of Streams. Chemosphere, 59, 207215. Bhatla, M.N. and Gaudy, A.F. (1966) Studies on the Causation of Phasic Oxygen Uptake in High Energy Systems. J. Water Pollut. Control Fed., 38, 14411451. Bush, A.W. (1958) BOD Progression in Soluble Substrates. Sewage Ind. Wastes, 30, 13361349. Cailas, M.D. and Gehr, R. (1989a) The Oxygen Uptake Rate Approach for Analysing Respirometric Biochemical Oxygen Demand Data-I. Method Dev. Water Res., 23 (8), 985992. Cailas, M.D. and Gehr, R. (1989b) The Oxygen Uptake Rate Approach for Analysing Respirometric Biochemical Oxygen Demand Data-II. Application and Evaluation. Water Res., 23 (8), 9931001. Cecen, F. and Yangin, C. (2000) Comparison of Results Obtained by Dilution and Nanometric Methods in Sanitary Landlls. J. Environ. Monit., 2, 628633. Cecen, F. and Yangin, C. (2001) Application of Various BOD Test Methods in Landll Leachates. J. Environ. Sci. Health, A336 (4), 545564. Chandran, K., Accashian, J.V. and Smets, B.F. (1996) Discussion of Oxygen Transfer Limitations of a Respirometer. Water Environ. Res., 68, 10841086. De Bel, M., Stokes, L., Upton, J. and Watts, J. (1996) Application of Respirometry Based Toxicity Monitor. Water Sci. Technol., 33 (1), 289296. Ellis, T.G., Barbeau, D.S., Smets, B.F. and Grady Jr., C.P.L. (1996) Respirometric Technique for Determination of Extant Kinetic Parameters Describing Biodegradation. Water Environ. Res., 68, 917926.

Gaffney, P.E. (1965) Carbon Dioxide Effects on Glucose Catabolism by Mixed Microbial Cultures. Appl. Microbiol., 13, 507510. Gernaey, K., Verschuere, L.S., Luyten, L. and Verstraete, W. (1997) Fast and Sensitive Acute Toxicity Detection with an Enrichment Nitrifying Culture. Water Environ. Res., 69, 11631169. Harris, J.O. (1954) The Inuence of Carbon Dioxide on Oxygen Uptake by Resting Cells of Bacteria. J. Bacteriol., 67, 476479. Hoffman, T. and Lees, H. (1953) The Respiration and Intermediary Metabolism of Nitrosomonas. Biochem. J., 54 (4), 579583. King, E.F. and Dutka, B.J. (1986) Respirometric Techniques. In Dutka, B.J. and Bitton, G. (eds). Toxicity Testing Using Microorganisms, Vol. 1, pp. 75112. CRC Press, Boca Raton, FL. Larson, R.J. and Perry, R.L. (1980) Use of Electrolytic Respirometer to Measure Biodegradation in Natural Waters. Water Res., 15, 697702. Li, K.Y. and Zhang, Y.B. (1996) Oxygen Transfer Limitations in a Respirometer. Water Environ. Res., 68, 3641. Mattman, L.H. (1954) Carbon Dioxide and Routine Cultures in the Diagnostic Laboratory. J. Lab. Clin. Med., 42, 485488. Orhon, D., Yldz, G., Cokgor, E.U. and Sozen, S. (1995) Respirometric Evaluation of Biodegradability of Confectionary Wastewaters. Water Sci. Technol., 32 (12), 1119. Roider, E.M. and Adrian, D.D. (2007) Comparative Evaluation of Three River Water Quality Models. J. Am. Water Res. Assoc., 43 (2), 322333. Siddigi, R.H., Speece, R.E., Engelbrecht, R.S. and Schmidt, J.W. (1967) Elimination of nitrication in the BOD determination with 0.1M ammonia nitrogen. J. Water Pollut. Control Fed., 39 (4), 579589. Suschka, J. and Ferreira, E. (1986) Activated Sludge Respirometric Measurements. Water Res., 20, 137144. Tebbutt, T.H.Y. and Berkun, M. (1976) Respirometric Determination of BOD. Water Res., 10, 613617. Yetis, U., Dilek, F.B. and Gokcay, C.F. (1992) Effects of Heavy Metals on Biological Activity in BOD-Seed Biomass Concentration. In Second International Symposium on Waste Management Problems in Agro-Industries, 2325 September, Istanbul, Turkey. Young, J.C. and Baumann, E.R. (1976) The Electrolytic Respirometer-I. Factors Affecting Oxygen Uptake Measurements. Water Res., 10, 10311040.

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