Bioaugmented sulfur-oxidizing denitrification system with <Emphasis Type="Italic">Alcaligenes defragrans</Emphasis> B21 for high nitrate containing wastewater treatment

The performance of enriched sludge augmented with the B21 strain of Alcaligenes defragrans was compared with that of enriched sludge, as well as with pure Alcaligenes defragrans B21, in the context of a sulfur-oxidizing denitrification (SOD) process. In synthetic wastewater treatment containing 100–1,000 mg NO₃⁻-N/L, the single strain-seeded system exhibited superior performance, featuring higher efficiency and a shorter startup period, provided nitrate loading rate was less than 0.2 kg NO₃⁻-N/m³ per day. At nitrate loading rate of more than 0.5 kg NO₃⁻-N/m³ per day, the bioaugmented sludge system showed higher resistance to shock loading than two other systems. However, no advantage of the bioaugmented system over the enriched sludge system without B21 strain was observed in overall efficiency of denitrification. Both the bioaugmented sludge and enriched sludge systems obtained stable denitrification performance of more than 80% at nitrate loading rate of up to 2 kg NO₃⁻-N/m³ per day.     

Test of Porous Ceramic Material for the Immobilisation of Predominant Nitrifying Bacteria and for the Improvement of the AAO Process

The efficiency of nitrification in a fixed bed reactor was compared to that found in an activated sludge reactor to determine their sensitivity to changing loads and lower temperatures. Two structurally identical lab‐scale systems, using the anaerobic/anoxic/aerobic (AAO) process to remove nitrogen and phosphorus simultaneously, were operated in parallel with secondary clarifiers and sludge return. The first aerobic system was operated as an activated sludge reactor, the second system as a fixed bed reactor. The aerobic fixed bed reactor was filled with porous ceramic materials for the immobilisation of predominantly nitrifying bacteria. The removal efficiencies of 99 % NH₄⁺‐N, 90 % DOC, and 98 % PO₄^3–‐P for normal loads of 0.11 g/L d DOC, 0.06 g/L d NH₄⁺‐N, and 0.0054 g/L d PO₄^3–‐P were achieved for both systems. However, the system with an aerobic fixed bed reactor was characterised by the following advantages over the system with an activated sludge reactor: a shorter time to reach almost complete nitrification, a higher nitrification rate at higher loads of NH₄⁺‐N and a lower sensitivity of nitrification at lower temperatures down to 12 °C.     

EDTA-Facilitated Phytoremediation of Soil with Heavy Metals from Sewage Sludge

The objective of this research was to determine the effect of the chelate EDTA (ethylenediaminetetraacetic acid), which is used in phytoremediation, on plant availability of heavy metals in liquid sewage sludge applied to soil. Sunflower (Helianthus annuus L.) was grown under greenhouse conditions in a commercial potting soil; the tetrasodium salt of EDTA (EDTA Na₄) was added at a rate of 1 g kg^-1 to half the pots. Immediately after seeds were planted, half of the pots with each soil (with or without EDTA) were irrigated with 60 ml sludge, and half were irrigated with 60 ml tap water. For the subsequent five irrigations, plants in soil with EDTA received either sludge or tap water containing 0.5 g EDTA Na₄ per 1000 ml, and plants in soil without EDTA received sludge or tap water without EDTA. Of the four heavy metals whose extractable concentrations in the soil were measured (Cu, Fe, Mn, and Zn), only Zn had a higher concentration in sludge-treated soil with EDTA compared to sludge-treated soil without EDTA. The concentrations of Fe, Cu, and Mn were similar in sludge-treated soil with and without EDTA. Of the three heavy metals whose total concentrations in the soil were measured (Cd, Pb, Cr), Pb (<10 mg kg^-1) and Cd (< 1 mg kg^-1) were below detection limits, and Cr was unaffected by treatment. The concentration of all measured elements in plants (Cd, Cu, Fe, Zn, Pb) was higher than the concentrations measured in the soil. With no EDTA, sludge-treated plants had a higher concentration of the five heavy elements than plants grown without sludge. Cadmium was lower in sludge-treated plants with EDTA than plants with EDTA and no sludge. After treatment with EDTA, the concentrations of Cu, Fe, and Zn were similar in plants with and without sludge. Lead was higher in plants with EDTA than plants without EDTA, showing that EDTA can facilitate phytoremediation of soil with Pb from sewage sludge.     

Biodegradation of coal slurry transport wastewater organics

Summary Activated sludge was successful in reducing the levels of dissolved organic carbon (DOC) in coal slurry wastewaters. DOC removal by the activated sludge ranged from 61% to 97% with a large percentage (21–41%) of this DOC being completely metabolized to CO₂. Second order kinetic constants (k ₂) developed for DOC removal ranged from 1.39·10^–4 to 2.30·10^–1 liter·day^–1·(mg of sludge)^–1, providing evidence that biological treatment was an effective mechanism for reducing the pollution potential of the slurry wastewaters. After treatment with activated sludge a residual DOC remained in the wastewater and data from ultrafiltration studies indicated that this residual carbon was of MW>1000. The activated sludge preferentially removed the lower (MW<1000) molecular weight compounds and the higher molecular weight DOC was more resistant to biological attack. However, extended acclimation (greater than 1 month) enabled the activated sludge to remove the higher molecular weight DOC from the slurry wastewaters.     

Degradation and Fate of Carbon Tetrachloride in Unadapted Methanogenic Granular Sludge

The potential of granular sludge from upflow anaerobic sludge blanket (UASB) reactors for bioremediation of chlorinated pollutants was evaluated by using carbon tetrachloride (CT) as a model compound. Granular sludges cultivated in UASB reactors on methanol, a volatile fatty acid mixture, or sucrose readily degraded CT supplied at a concentration of 1,500 nmol/batch (approximately 10 μM) without any prior exposure to organohalogens. The maximum degradation rate was 1.9 μmol of CT g of volatile suspended solids⁻¹ day⁻¹. The main end products of CT degradation were CO₂ and Cl⁻, and the yields of these end products were 44 and 68%, respectively, of the initial amounts of [¹⁴C]CT and CT-Cl. Lower chlorinated methanes accumulated in minor amounts temporarily. Autoclaved (dead) sludges were capable of degrading CT at rates two- to threefold lower than those for living sludges, indicating that abiotic processes (mediated by cofactors or other sludge components) played an important role in the degradation observed. Reduced components in the autoclaved sludge were vital for CT degradation. A major part (51%) of the CT was converted abiotically to CS₂. The amount of CO₂ produced (23%) was lower and the amount of Cl⁻ produced (86%) was slightly higher with autoclaved sludge than with living sludge. Both living and autoclaved sludges could degrade chloroform. However, only living sludge degraded dichloromethane and methylchloride. These results indicate that reductive dehalogenation, which was mediated better by living sludge than by autoclaved sludge, is only a minor pathway for CT degradation. The main pathway involves substitutive and oxidative dechlorination reactions that lead to the formation of CO₂. Granular sludge, therefore, has outstanding potential for gratuitous dechlorination of CT to safe end products.     

Nickel in soil and maize plants grown on an oxisol treated over a long time with sewage sludge

Abstract

The major limitation for the use of sewage sludge in agriculture is the risk of soil contamination with heavy metals, and their possible transference to man via the food chain. The objective of this study was to evaluate the content of nickel (Ni) in soil by the two methods of digestion (HNO₃ + H₂O₂ + HCl and HClO₄ + HF), and in different parts of maize plants grown on a tropical soil classified as Typic Eutrorthox, that had been treated with sewage sludge for nine consecutive years, and the effects on dry matter and grain production. The experiment was carried out under field conditions in Jaboticabal-SP, using a randomized block design with four treatments and five replicates. Treatments consisted of: 0.0 (control, mineral fertilization), 45.0, 90.0 and 127.5t ha^?1 sewage sludge (dry basis), accumulated during nine years. Sewage sludge was manually applied to the soil and incorporated at 0.1 m depth before sowing the maize. Soil Ni evaluated by Jackson’s method was 76.8% higher than evaluated by the United States Environmental Protection Agency method that digests the samples by heating with concentrated HNO³, H²O² and HCl. Sewage sludge rates did not affect Ni content in the soil. Ni was accumulated in leaf and stem but was not detected in grain. Sewage sludge and mineral fertilization applied to soil for a long time caused similar effects on dry matter and grain production.     

Gratuitous dechlorination of chloroethanes by methanogenic granular sludge

The dechlorinating activity of a methanogenic granular sludge from a methanol-fed upflow anaerobic sludge blanket reactor was investigated with chlorinated ethanes. This unadapted methanogenic consortium degraded all chloroethanes tested. The product formation rates decreased with the number of chlorine substituents. The more highly chlorinated ethanes were also converted, although at a lower rate, in the presence of autoclaved (dead) sludge, indicating the involvement of reduced heat-stable cofactors like vitamin B₁₂ and F₄₃₀. Direct chemical dechlorination of hexa-, penta- and tetrachloroethanes was also observed in medium without sludge, although at a much lower rate. The results show the importance of cometabolic and abiotic (chemical) conversions for the transformation of chlorinated ethanes by the methanogenic consortium. The types of reaction and the products formed were correlated with the Gibbs free-energy change (ΔG ^0′). Reductive hydrogenolysis and dichloroelimination were important dechlorinating mechanisms. Generally, these reactions have a higher ΔG ^0′ value than dehydrochlorination reactions, which occurred less frequently during the transformation of chloroethanes by the methanogenic granular sludge. Received: 8 June 1998 / Received revision: 7 September 1998 / Accepted: 13 September 1998     

Conversion of paper sludge to ethanol. I: Impact of feeding frequency and mixing energy characterization

In this paper, conversion of paper sludge to ethanol was investigated with the objective of optimization of the overall operation costs. Experimental work was undertaken to optimize cellulase loading, and to determine mixing energy requirements. It was found that decreasing feeding frequency (feed additions per residence time) allows the cellulase loading to be decreased at least two fold with no decrease in cellulose conversion but also entails mixing a slurry of higher solids content and lower conversion at the beginning of the operating cycle. The viscosity of paper sludge slurries was found to increase exponentially with decreasing conversion and increasing solid content. In particular, the viscosity (V) was described well by equation V = e^(kX−X ₀^)(S−S ₀^)+C (V viscosity (cp), X conversion, S solid content (g/L), k, X ₀, S ₀, C are empirical parameters). Added costs associated with operating at low feeding frequencies (including higher mixing energy and higher capital costs for the motor and for sludge hold tasks) were found to be small compared to the economic benefits resulting from reduced cellulase loading.     

蚯蚓作用下污泥重金属形态变化及其与化学生物学性质变化的关系

城市污泥处理是一项世界性难题,污泥农业利用是其最简单有效的资源化利用方式之一,但污泥中较高的重金属含量限制了其实际推广应用,利用蚯蚓-超富集植物联合修复污泥重金属的方法已引起国内外研究者的关注。以新鲜城市脱水污泥为研究对象,接种赤子爱胜蚓(Eisenia fetida)进行室内培养试验,系统研究蚯蚓作用下污泥重金属形态的变化,及其与污泥氧化还原条件、化学和微生物性质变化的关系,以期为蚯蚓-超富集植物联合修复技术在污泥重金属处理中的应用提供理论依据。结果表明,试验前期蚯蚓在污泥中能正常生长和存活,前20 d总生物量增加了52%。蚯蚓可以显著促进污泥中的Cu、Zn、Cd、Ni等重金属从残渣态和铁锰态等稳定形态向交换态和水溶态等有效形态转化。还可以显著降低污泥中还原性物质的含量,减缓p H值下降速度,降低总有机碳含量,促进铵态氮向硝态氮转化,减少污泥微生物的数量并增加其种群活性。蚯蚓作用下,污泥中重金属的活化程度与还原性物质的含量呈显著负相关,而与微生物种群的活性呈显著正相关(P0.05)。综上所述,蚯蚓可以促进污泥重金属的活化,并改善污泥的肥力条件,为修复植物在污泥中的正常生长和对重金属离子的快速吸收提供有利条件。     

Acetate treatment in 70°C upflow anaerobic sludge-blanket (UASB) reactors: start-up with thermophilic inocula and the kinetics of the UASB sludges

This study focused on the use of thermophilic anaerobic granulae in the start-up of 70°C acetate-fed upflow anaerobic sludge-blanket (UASB) reactors and the kinetics of granulae grown at 70°C. In the UASB reactors, chemical oxygen demand removal commenced within 48 h of the start-up. The maximum reduction in chemical oxygen demand was 84% with the feed containing yeast and 71% without a yeast supplement. In the bioassays, the yeast-grown sludge converted 98% of the acetate consumed to methane as compared to 92% for the sludge grown without yeast. The highest initial specific methane production rate (µ_{CH 4}) of the UASB sludges grown at 70°C was 0.088 h^–1 at an acetate concentration of 4.6 mM. The higher initial acetate concentration was found to prolong the lag-phase in methane production significantly and to decrease the µ_{CH 4}. The half-saturation constant (K _s), the inhibition constant (K _i), the inhibition response coefficient (n), and the µ_{CH 4} ^max, calculated according to a modified Haldane equation, were 1.5 mM, 2.8 mM, 0.8, and 0.28 h^–1, respectively. The prolonged starvation of the 70°C sludge (15 days) decreased the µ_{CH 4} from about 0.022 h^–1 to 0.011 h^–1 and increased the lag phase in methane production from 6 h to 24 h as compared to non-starved sludge.     

Substrate consumption and excess sludge reduction of activated sludge in the presence of uncouplers: a modeling approach

A mathematical model with a consideration of energy spilling is developed to describe the activated sludge in the presence of different levels of metabolic uncouplers. The consumption of substrate and oxygen via energy spilling process is modeled with a Monod term, which is dependent on substrate and inhibitor. The sensitivity of the developed model is analyzed. Three parameters, maximum specific growth rate (μ _max), energy spilling coefficient (q _max), and sludge yield coefficient (Y _H) are estimated with experimental data of different studies. The values of μ _max, q _max, and Y _H are found to be 6.72 day^-1, 5.52 day^-1, and 0.60 mg COD mg^-1 COD for 2, 4-dinitrophenol and 7.20 day^-1, 1.58 day^-1, and 0.62 mg COD mg^-1 COD for 2, 4-dichlorophenol. Substrate degradation and sludge yield could be predicted with this model. The activated sludge process in the presence of uncouplers that is described more reasonably by the new model with a consideration of energy spilling. The effects of uncouplers on substrate consumption inhibition and excess sludge reduction in activated sludge are quantified with this model.     

Autotrophic denitrification using elemental sulfur

Experimental studies were conducted to determine the feasibility of autotrophic denitrification with Thiobacillus denitrificans as a nitrate removal process for wastewater. S⁰-acclimated activated sludge, which can carry out autotrophic denitrification using S⁰ as electron donors, was prepared by the fill and draw cultivation method. The kinetic constants for the S⁰-denitrification reaction using S⁰-acclimated activated sludge under NO₃-N limiting growth conditions were determined to be Y = 0.33 mg-NO₃-N and b = 0.058 d⁻¹. High percentages of denitrification (over 95%) and rates of denitrification ranging from 0.19 to 0.24 mg-NO₃-N/mg-TOC·d were obtained in the continuous denitrification experiments using S⁰-acclimated activated sludge.     

Enrichment and granulation of Anammox biomass started up with methanogenic granular sludge

A sequencing batch reactor (SBR) seeded with methanogenic granular sludge was started up to enrich Anammox (Anaerobic Ammonium Oxidation) bacteria and to investigate the feasibility of granulation of Anammox biomass. Research results showed that hydraulic retention time (HRT) was an important factor to enrich Anammox bacteria. When the HRT was controlled at 30 days during the initial cultivation, the SBR reactor presented Anammox activity at t = 58 days. Simultaneously, the methanogenic granular sludge changed gradually from dust black to brown colour and its diameter became smaller. At t = 90 days, the Anammox activity was further improved. NH₄⁺-N and NO₂⁻N were removed simultaneously with higher speed and the maximum removal rates reached 14.6 g NH₄⁺-N /(m³ _reactor·day) and 6.67 g NO₂⁻-N /(m³ _reactor·day), respectively. Between t = 110 days and t = 161 days, the nitrogen load was increased to a HRT of 5 days (70 mg/l NH₄⁺ and 70 mg/l NO₂), the removal rates of ammonium and nitrite were 60.6% and 62.5% respectively. The sludge changed to red and formed Anammox granulation with high nitrogen removal activity.     

Increased power generation from primary sludge in microbial fuel cells coupled with prefermentation

Raw primary sludge and the prefermentation liquor (PL) of primary sludge were used to generate electricity in single-chambered air-cathode microbial fuel cells (MFCs). The MFCs treating the primary sludge produced 0.53 V and 370 mW/m² for the maximum potential and power density, respectively. In the primary sludge-fed MFCs, only 5 % of the total energy production was produced from direct electricity generation, whereas 95 % of that resulted from the conversion of methane to electricity. MFCs treating the PL generated the maximum potential of 0.58 V and maximum power density of 885 mW/m², respectively. In the energy production analysis, direct electricity production (1,921 Wh/kg TCOD_rem) in the MFCs treating the PL was much higher than that of the primary sludge-fed MFC (138 Wh/kg TCOD_rem). Volatile suspended solids during 10 days were reduced to 18.3 and 38 % in the primary sludge-fed MFCs and prefermentation reactor, respectively. These findings suggest that a two-stage process including prefermentation and MFCs is of great benefit on sludge reduction and higher electricity generation from primary sludge.     

The impact of titanium dioxide nanoparticles on biological nitrogen removal from wastewater and bacterial community shifts in activated sludge

The potential impact of titanium dioxide nanoparticles (TiO₂ NPs) on nitrogen removal from wastewater in activated sludge was investigated using a sequencing batch reactor. The addition of 2–50 mg L^?1 of TiO₂ NPs did not adversely affect nitrogen removal. However, when the activated sludge was exposed to 100–200 mg L^?1 of TiO₂ NPs, the effluent total nitrogen removal efficiencies were 36.5 % and 20.3 %, respectively, which are markedly lower than the values observed in the control test (80 %). Further studies showed that the decrease in biological nitrogen removal induced by higher concentrations of TiO₂ NPs was due to an inhibitory effect on the de-nitrification process. Denaturing gradient gel electrophoresis profiles showed that 200 mg L^?1 of TiO₂ NPs significantly reduced microbial diversity in the activated sludge. The effect of light on the antibacterial activity of TiO₂ NPs was also investigated, and the results showed that the levels of TiO₂-dependent inhibition of biological nitrogen removal were similar under both dark and light conditions. Additional studies revealed that different TiO₂ concentrations had a significant effect on dehydrogenase activity, and this effect was most likely the result of decreased microbial activity.     

Effect of Mn<Superscript>2+</Superscript> augmentation on reinforcing aerobic sludge granulation in a sequencing batch reactor

Two sequencing batch reactors were synchronously operated to investigate the effect of manganese (II) (Mn²⁺) augmentation on aerobic granulation. Reactor 1 (R1) was added with 10 mg/L Mn²⁺, while there was no Mn²⁺ augmentation in reactor 2 (R2). Results showed that R1 had a faster granulation process than R2 and R1 performed better in chemical oxygen demand (COD) and ammonium nitrogen (NH₄⁺–N) removal efficiencies. Moreover, the mature granules augmented with Mn²⁺ behaved better on their physical characteristics and size distributions, and they also had higher production of extracellular polymeric substances (EPS) content. The result of three-dimensional excitation and emission matrix fluorescence showed that Mn²⁺ had the function of causing organic material diversity (especially proteins diversity) in EPS fraction from granules. Polymerase chain reaction and denaturing gradient gel electrophoresis techniques were employed to analyze the microbial and genetic characteristics in mature granules. The results exhibited that Mn²⁺ augmentation was mainly responsible for the higher microbial diversity of granules from R1 compared with that from R2. Uncultured sludge bacterium A16 (AF234726) and Rhodococcus sp. WTZ-R2 (HM004214) were the major species in R1, while only uncultured sludge bacterium A16 (AF234726) in R2. Moreover, there were eight species of organisms found in both two aerobic granules, and three species were found only in aerobic granules from R1. It could be concluded that Mn²⁺ could enhance the sludge granulation process and have a key effect role on the biological properties during the sludge granulation.     

Monitoring of specific methanogenic activity of granular sludge by confocal laser scanning microscopy during start-up of thermophilic upflow anaerobic sludge blanket reactor

Confocal, laser-scanning microscopy was applied to acquire coenzyme F₄₂₀-based autofluorescence images of middle sections of sludge granules during start-up of a thermophilic reactor that were seeded with mesophilically-grown microorganisms of granular sludge. Digital images were analyzed to calculate weighted averages of autofluorescence. The values were related (r ²=0.97) to specific methanogenic activities of granular sludge as the granules developed to steady state.     

Treatment of old landfill leachate with high ammonium content using aerobic granular sludge

Background

Aerobic granular sludge has become an attractive alternative to the conventional activated sludge due to its high settling velocity, compact structure, and higher tolerance to toxic substances and adverse conditions. Aerobic granular sludge process has been studied intensively in the treatment of municipal and industrial wastewater. However, information on leachate treatment using aerobic granular sludge is very limited.

Methods

This study investigated the treatment performance of old landfill leachate with different levels of ammonium using two aerobic sequencing batch reactors (SBR): an activated sludge SBR (ASBR) and a granular sludge SBR (GSBR). Aerobic granules were successfully developed using old leachate with low ammonium concentration (136 mg L^?1 NH₄ ⁺-N).

Results

The GSBR obtained a stable chemical oxygen demand (COD) removal of 70% after 15 days of operation; while the ASBR required a start-up of at least 30 days and obtained unstable COD removal varying from 38 to 70%. Ammonium concentration was gradually increased in both reactors. Increasing influent ammonium concentration to 225 mg L^?1 N, the GSBR removed 73 ± 8% of COD; while COD removal of the ASBR was 59 ± 9%. The GSBR was also more efficient than the ASBR for nitrogen removal. The granular sludge could adapt to the increasing concentrations of ammonium, achieving 95 ± 7% removal efficiency at a maximum influent concentration of 465 mg L^?1 N. Ammonium removal of 96 ± 5% was obtained by the ASBR when it was fed with a maximum of 217 mg L^?1 NH₄ ⁺-N. However, the ASBR was partially inhibited by free-ammonia and nitrite accumulation rate increased up to 85%. Free-nitrous acid and the low biodegradability of organic carbon were likely the main factors affecting phosphorus removal.

Conclusion

The results from this research suggested that aerobic granular sludge have advantage over activated sludge in leachate treatment.

     

Kinetics of Hydrogen Consumption by Rumen Fluid, Anaerobic Digestor Sludge, and Sediment

Michaelis-Menten kinetic parameters for H₂ consumption by three methanogenic habitats were determined from progress curve and initial velocity experiments. The influences of mass transfer resistance, endogenous H₂ production, and growth on apparent parameter estimates were also investigated. Kinetic parameters could not be determined for undiluted rumen fluid and some digestor sludge from gas-phase measurements of H₂, since mass transfer of H₂ across the gas-liquid interface was rate limiting. However, accurate values were obtained once the samples were diluted. H₂ consumption by digestor sludge with a long retention time and by hypereutrophic lake sediment was not phase transfer limited. The K_m values for H₂ uptake by these habitats were similar, with means of 5.8, 6.0, and 7.1 μM for rumen fluid, digestor sludge, and sediment, respectively. V_max estimates suggested a ratio of activity of approximately 100 (rumen fluid):10 (sludge):1 (sediment); their ranges were as follows: rumen fluid, 14 to 28 mM h⁻¹; Holt sludge, 0.7 to 4.3 mM h⁻¹; and Wintergreen sediment, 0.13 to 0.49 mM h⁻¹. The principles of phase transfer limitation, studied here for H₂, are the same for all gaseous substrates and products. The limitations and errors associated with gas phase determination of kinetic parameters were evaluated with a mathematical model that combined mass transport and Michaelis-Menten kinetics. Three criteria are described which can be used to evaluate the possibility that a phase transfer limitation exists. If it does not exist, (i) substrate consumption curves are Michaelis-Menten and not first order, (ii) the K_m is independent of initial substrate concentration, and (iii) the K_m is independent of biomass (V_max) and remains constant with dilution of sample. Errors in the Michaelis-Menten kinetic parameters are caused by endogenously produced H₂, but they were <15% for rumen fluid and 10% for lake sediment and digestor sludge. Increases in V_max during the course of progress curve experiments were not great enough to produce systematic deviations from Michaelis-Menten kinetics.     

Patterns of carbon flow from glucose to methane in a thermophilic anaerobic bioreactor

[U-¹⁴C]Glucose, added carrier-free to sludge from a thermophilic anaerobic bioreactor being fed a lignocellulose waste, was rapidly turned over with less than one-third of the original radiolabel remaining as glucose after 5 s of incubation. The primary labeled products found were [¹⁴C]acetate and ¹⁴CO₂, which were in a ratio near 2:1. Further incubation resulted in the disappearance of [¹⁴C]acetate and the appearance of an equivalent amount of label as ¹⁴CH₄ and ¹⁴CO₂. No significant production of [¹⁴C]propionate, butyrate, lactate, or ethanol was detected from [¹⁴C]glucose, even if these potential intermediates (unlabeled) were added to the sludge at a concentration of 1 mM to trap any label entering their pools. Addition of 0.8 atm (80 kPa) of H₂ to the headspace over sludge resulted in some accumulation of [¹⁴C]lactate and a corresponding decrease in [¹⁴C]acetate produced from [¹⁴C]glucose. Production of [¹⁴C]propionate, butyrate and ethanol were still not significant in the presence of H₂. Incubation of sludge for 1 h in the presence of hydrogen resulted in increases in the lactate and formate concentrations, but not those of propionate, butyrate, or ethanol. These results demonstrate that glucose was metabolized directly to acetate, CO₂, and H₂ by the microbial populations in the bioreactor with little carbon from glucose flowing through other intermediates, indicating a high degree of coupling between glucose fermentation and hydrogen uptake. The short-term response of these microbial populations to elevated H₂ partial pressures was to increase lactate production.