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The effects of post-treatments and temperature on recovering the methane potential of >2 mm solid fraction of digested cow manure.

The effects of thermal and chemical treatments, mechanical maceration and freezing and thawing on recovering the remaining methane potential of the >2 mm solid fraction of digested cow manure - which accounted for 30% of the original potential of digested cow manure - were studied in laboratory batch assays at 5-20 degrees C and at 35-55 degrees C to evaluate the treatment effects both under long-term (340 d) storage of solids and during active digestion (30 d), respectively. The effects of different treatments on the methane production of the solids varied with incubation temperatures and time. However, in all cases, methane productions at 15 degrees C and lower were slow and low for both untreated and treated solids even after long-term incubation. At 35 and 55 degrees C more methane was recovered from untreated solids producing up to 61-82 ml g(-1) volatile solids (VS)added in 30 d and 179-215 ml g(-1) VSadded in 340 d. Only chemical treatment with or without thermal treatment enhanced the methane yields while some treatments even decreased the yields. An increase in temperature to 35 degrees C of the assays incubated for 6 months at < or =20 degrees C initiated more significant methane production. In conclusion, the methane potential of the digested solids in a farm-scale biogas system can be recovered by active digestion at 35 or 55 degrees C and can be improved to a smaller extent through chemical treatment of separated solids fraction, while methane recovery at lower temperatures and with some of the treatments studied would not be effective.

Animals↗

Have sudden large releases of methane from geological reservoirs occurred since the Last Glacial Maximum, and could such releases occur again?

Methane emissions from geological reservoirs may have played a major role in the sudden events terminating glaciation, both at the start of the Bølling/Allerød and also at the end of the Younger Dryas. These reservoirs include Arctic methane hydrates and also methane hydrate stored in offshore marine sediments in tropical and temperate latitudes. Emissions from hydrate stores may have resonated with tropical wetland emissions, each reinforcing the other. Because methane is such a powerful greenhouse gas, much smaller emissions of methane, compared with carbon dioxide, are required in order to have the same short-term impact by climate forcing. The methane-linked hypothesis has much geological support from sea-floor evidence of emission. However, Greenland ice-core records have been interpreted as showing methane as a consequential factor, rather than the leader, of change. This interpretation can be challenged on the grounds that temperature gradients in Greenland ice record local changes and local timing of a step-like shift in weather fronts, while methane concentrations record changes on a hemispheric and global scale. There are large remaining hydrate reservoirs in the Arctic and in shelf sediments globally, and there is substantial risk of further emissions.

Climate↗

Regulation of methane oxidation in the facultative methanotroph Methylocella silvestris BL2.

The molecular regulation of methane oxidation in the first fully authenticated facultative methanotroph Methylocella silvestris BL2 was assessed during growth on methane and acetate. Problems of poor growth of Methylocella spp. in small-scale batch culture were overcome by growth in fermentor culture. The genes encoding soluble methane monooxygenase were cloned and sequenced, which revealed that the structural genes for soluble methane monooxygenase, mmoXYBZDC, were adjacent to two genes, mmoR and mmoG, encoding a sigma54 transcriptional activator and a putative GroEL-like chaperone, located downstream (3') of mmoC. Transcriptional analysis revealed that the genes were all cotranscribed from a sigma54-dependent promoter located upstream (5') of mmo X. The transcriptional start site was mapped. Transcriptional analysis of soluble methane monooxygenase genes and expression studies on fermentor grown cultures showed that acetate repressed transcription of sMMO in M. silvestris BL2. The possibility of the presence of a particulate, membrane-bound methane monooxygenase enzyme in M. silvestris BL2 and the copper-mediated regulation of soluble methane monooxygenase was investigated. Both were shown to be absent. A promoter probe vector was constructed and used to assay transcription of the promoter of the soluble methane monoxygenase genes of M. silvestris BL2 grown under various conditions and with different substrates. These data represent the first insights into the molecular physiology of a facultative methanotroph.

Beijerinckiaceae↗

Radioactive fingerprinting of microorganisms that oxidize atmospheric methane in different soils.

Microorganisms that oxidize atmospheric methane in soils were characterized by radioactive labelling with (14)CH(4) followed by analysis of radiolabelled phospholipid ester-linked fatty acids ((14)C-PLFAs). The radioactive fingerprinting technique was used to compare active methanotrophs in soil samples from Greenland, Denmark, the United States, and Brazil. The (14)C-PLFA fingerprints indicated that closely related methanotrophic bacteria were responsible for the oxidation of atmospheric methane in the soils. Significant amounts of labelled PLFAs produced by the unknown soil methanotrophs coeluted with a group of fatty acids that included i17:0, a17:0, and 17:1omega8c (up to 9.0% of the total (14)C-PLFAs). These PLFAs are not known to be significant constituents of methanotrophic bacteria. The major PLFAs of the soil methanotrophs (73.5 to 89.0% of the total PLFAs) coeluted with 18:1 and 18:0 fatty acids (e.g., 18:1omega9, 18:1omega7, and 18:0). The (14)C-PLFAs fingerprints of the soil methanotrophs that oxidized atmospheric methane did not change after long-term methane enrichment at 170 ppm CH(4). The (14)C-PLFA fingerprints of the soil methanotrophs were different from the PLFA profiles of type I and type II methanotrophic bacteria described previously. Some similarity at the PLFA level was observed between the unknown soil methanotrophs and the PLFA phenotype of the type II methanotrophs. Methanotrophs in Arctic, temperate, and tropical regions assimilated between 20 and 54% of the atmospheric methane that was metabolized. The lowest relative assimilation (percent) was observed for methanotrophs in agricultural soil, whereas the highest assimilation was observed for methanotrophs in rain forest soil. The results suggest that methanotrophs with relatively high carbon conversion efficiencies and very similar PLFA compositions dominate atmospheric methane metabolism in different soils. The characteristics of the methane metabolism and the (14)C-PLFA fingerprints excluded any significant role of autotrophic ammonia oxidizers in the metabolism of atmospheric methane.

Carbon Radioisotopes↗

Biogeochemical and molecular signatures of anaerobic methane oxidation in a marine sediment.

Anaerobic methane oxidation was investigated in 6-m-long cores of marine sediment from Aarhus Bay, Denmark. Measured concentration profiles for methane and sulfate, as well as in situ rates determined with isotope tracers, indicated that there was a narrow zone of anaerobic methane oxidation about 150 cm below the sediment surface. Methane could account for 52% of the electron donor requirement for the peak sulfate reduction rate detected in the sulfate-methane transition zone. Molecular signatures of organisms present in the transition zone were detected by using selective PCR primers for sulfate-reducing bacteria and for Archaea. One primer pair amplified the dissimilatory sulfite reductase (DSR) gene of sulfate-reducing bacteria, whereas another primer (ANME) was designed to amplify archaeal sequences found in a recent study of sediments from the Eel River Basin, as these bacteria have been suggested to be anaerobic methane oxidizers (K. U. Hinrichs, J. M. Hayes, S. P. Sylva, P. G. Brewer, and E. F. DeLong, Nature 398:802-805, 1999). Amplification with the primer pairs produced more amplificate of both target genes with samples from the sulfate-methane transition zone than with samples from the surrounding sediment. Phylogenetic analysis of the DSR gene sequences retrieved from the transition zone revealed that they all belonged to a novel deeply branching lineage of diverse DSR gene sequences not related to any previously described DSR gene sequence. In contrast, DSR gene sequences found in the top sediment were related to environmental sequences from other estuarine sediments and to sequences of members of the genera Desulfonema, Desulfococcus, and Desulfosarcina. Phylogenetic analysis of 16S rRNA sequences obtained with the primers targeting the archaeal group of possible anaerobic methane oxidizers revealed two clusters of ANME sequences, both of which were affiliated with sequences from the Eel River Basin.

Anaerobiosis↗

Comparative analysis of methane-oxidizing archaea and sulfate-reducing bacteria in anoxic marine sediments.

The oxidation of methane in anoxic marine sediments is thought to be mediated by a consortium of methane-consuming archaea and sulfate-reducing bacteria. In this study, we compared results of rRNA gene (rDNA) surveys and lipid analyses of archaea and bacteria associated with methane seep sediments from several different sites on the Californian continental margin. Two distinct archaeal lineages (ANME-1 and ANME-2), peripherally related to the order Methanosarcinales, were consistently associated with methane seep marine sediments. The same sediments contained abundant (13)C-depleted archaeal lipids, indicating that one or both of these archaeal groups are members of anaerobic methane-oxidizing consortia. (13)C-depleted lipids and the signature 16S rDNAs for these archaeal groups were absent in nearby control sediments. Concurrent surveys of bacterial rDNAs revealed a predominance of delta-proteobacteria, in particular, close relatives of Desulfosarcina variabilis. Biomarker analyses of the same sediments showed bacterial fatty acids with strong (13)C depletion that are likely products of these sulfate-reducing bacteria. Consistent with these observations, whole-cell fluorescent in situ hybridization revealed aggregations of ANME-2 archaea and sulfate-reducing Desulfosarcina and Desulfococcus species. Additionally, the presence of abundant (13)C-depleted ether lipids, presumed to be of bacterial origin but unrelated to ether lipids of members of the order Desulfosarcinales, suggests the participation of additional bacterial groups in the methane-oxidizing process. Although the Desulfosarcinales and ANME-2 consortia appear to participate in the anaerobic oxidation of methane in marine sediments, our data suggest that other bacteria and archaea are also involved in methane oxidation in these environments.

Anaerobiosis↗

[Basic examination on the energization of experimental animal feces by methane fermentation (author's transl)].

Basic experiments of methane fermentation of livestock excreta have been reported by many investigators, yet there has been no report of experiments of methane fermentation of experimental animal feces. We considered that it would be feasible to gasiby experimental animal feces by methane fermentation. In this report, methane gas fermentation of rat feces is studied. The results are summarized as follows: 1. Methane fermentation of rat feces yielded, under the conditions described, 220-4231 of gas per kg of organic matter. 2. Decomposition ratio of BOD removal volume in organic matter was 50-60% when the fermentation condition was optimum. 3. The volume of gas from BOD removal volume in organic matter was 500-600ml/g. 4 Methane gas contents in the gas generated from the feces ranged from about 55-60%. 5. COD removal ratio was above 80%, and BOD removal ratio above 90%. From these data, the authors conclude that under the optimum conditions, described sufficient methane gas can be obtained from rat feces by methane fermentation.

Animals↗

Methane production and oxidation in an anoxic rice soil as influenced by inorganic redox species.

The effects of addition of inorganic redox substances (species of NO3-, Mn4+, Fe3+, and SO4(2-)) on methane production and oxidation in anoxic rice (Oryza sativa L.) soil samples were examined. Sulfate was the most inhibitory for methane production followed by Fe3+, NO3-, and Mn4+, in that order. Addition of rice straw at a rate of 1% (w/w) as a carbon source to increase the electron donor to the electron acceptor ratio did not completely alleviate the inhibitory effects of redox species on methane production. Interestingly, laboratory incubation studies showed that addition of MnO2 and K2SO4 enhanced aerobic methane oxidation in soil samples held at 60% water holding capacity. The suspensions of pretreated soil samples with different redox species, when tested for their ability to oxidize methane in soil solution equivalent medium supplemented with respective redox species under aerobic and anaerobic conditions showed differential effects of redox species. Nitrate and Fe3+ stimulated methane oxidation under anaerobic conditions and retarded it under aerobic conditions. Manganese(IV) ion retarded methane oxidation under anaerobic conditions, but enhanced it under aerobic conditions. However, SO4(2-) stimulated methane oxidation in soil solution equivalent medium under both aerobic and anaerobic conditions.

Agriculture↗

Prediction of methane production from dairy cows using existing mechanistic models and regression equations.

Ruminants may contribute to global warming through the release of methane gas by enteric fermentation. Until now, methane emissions from ruminants were estimated using simple regression equations. The objective of this study was to compare the capacity of dynamic and mechanistic models to that of regression equations to predict methane production from dairy cows. The updated version of the model of Baldwin et al. and a modified version of the model of Dijkstra et al. and the regression equations of Blaxter and Clapperton and Moe and Tyrrell were challenged with 32 experimental diets selected from 13 publications. The predictive capacity of mechanistic models and regression equations was evaluated by comparing predicted and observed methane production using regression analysis. Results of regression showed better prediction of methane production with mechanistic models than with regression equations. The modified model of Dijkstra et al. predicted methane production with the higher R2 (.71) and the smaller error of prediction (19.87% of the observed mean). The model of Baldwin et al. predicted methane production with a similar R2 (.70) but a higher error of prediction (36.93%). However, a large proportion of this error can be eliminated by a correction factor. Predictions using the equations of Moe and Tyrrell and Blaxter and Clapperton were poor (R2 = .42 and .57; error of prediction = 33.72% and 22.93%, respectively). This study demonstrated that from a large variation in diet composition, mechanistic models allow the prediction of methane production more accurately than simple regression equations.

Animal Feed↗

Methane emissions from feedlot cattle fed barley or corn diets.

Methane emitted from the livestock sector contributes to greenhouse gas emissions worldwide. Understanding the variability in enteric methane production related to diet is essential to decreasing uncertainty in greenhouse gas emission inventories and to identifying viable greenhouse gas reduction strategies. Our study focused on measuring methane in growing beef cattle fed corn- or barley-based diets typical of those fed to cattle in North American feedlots. The experiment was designed as a randomized complete block (group) design with two treatments, barley and corn. Angus heifer calves (initial BW = 328 kg) were allocated to two groups (eight per group), with four cattle in each group fed a corn or barley diet. The experiment was conducted over a 42-d backgrounding phase, a 35-d transition phase and a 32-d finishing phase. Backgrounding diets consisted of 70% barley silage or corn silage and 30% concentrate containing steam-rolled barley or dry-rolled corn (DM basis). Finishing diets consisted of 9% barley silage and 91% concentrate containing barley or corn (DM basis). All diets contained monensin (33 mg/kg of DM). Cattle were placed into four large environmental chambers (two heifers per chamber) during each phase to measure enteric methane production for 3 d. During the backgrounding phase, DMI was greater by cattle fed corn than for those fed barley (10.2 vs. 7.6 kg/d, P < 0.01), but during the finishing phase, DMI was similar for both diets (8.3 kg/d). The DMI was decreased to 6.3 kg/d with no effect of diet or phase while the cattle were in the chambers; thus, methane emissions (g/d) reported may underestimate those of the feedlot industry. Methane emissions per kilogram of DMI and as a percentage of GE intake were not affected by grain source during the backgrounding phase (24.6 g/kg of DMI; 7.42% of GE), but were less (P < 0.05) for corn than for barley during the finishing phase (9.2 vs. 13.1 g/kg of DMI; 2.81 vs. 4.03% of GE). The results indicate the need to implement dietary strategies to decrease methane emissions of cattle fed high-forage backgrounding diets and barley-based finishing diets. Mitigating methane losses from cattle will have long-term environmental benefits by decreasing agriculture's contribution to greenhouse gas emissions.

Animal Feed↗

Substrate interactions during aerobic biodegradation of methane, ethene, vinyl chloride and 1,2-dichloroethenes.

Intrinsic biodegradation of trichloroethene and 1,1,1-trichloroethane in groundwater at a Superfund site in California has been observed. An anaerobic zone exists in the area closest to the source location, yielding the expected complement of reductive dechlorination daughter products, including cis-1,2-dichloroethene (cis-DCE) and vinyl chloride (VC). Significant levels of methane and ethene were also generated in the anaerobic zone. The groundwater returns to aerobic conditions downgradient of the source, with methane, ethene, VC, and several other compounds still present. Attenuation of VC in the aerobic zone suggests that it is being biodegraded. In this study microcosms were used to evaluate the role of methane and ethene as primary substrates for aerobic biodegradation of VC. Biodegradation of VC was fastest in the bottles containing ethene, with 40 mumol of VC consumed over a 150 day period, compared to approximately 15-20 mumol with methane or a mixture of methane and ethene. VC did not noticeably inhibit ethene biodegradation but did slow the rate of methane use. Methane inhibited ethene metabolism, which apparently caused a reduction in VC biodegradation when methane was present with ethene. These results suggest that ethene plays an important role during in situ natural attenuation of VC under aerobic conditions. Microcosms were also set up with VC alone. Following a 75 day lag period. VC consumption began and subsequent additions were consumed without a lag, suggesting the presence of organisms capable of using VC as a growth substrate. After providing VC alone for nearly 400 days, aliquots of the enrichment culture were used to evaluate its ability to biodegrade cis- and trans-DCE. Both compounds were readily consumed, although addition of VC as the primary substrate was needed to sustain biodegradation of repeated additions. This result suggests that organisms capable of using VC as a sole substrate may play an active role in aerobic natural attenuation of DCEs.

Bacteria, Aerobic↗

[Biogeochemical cycle of methane in the northwestern shelf of the Black Sea].

Seasonal investigations of methane distribution and rates of its oxidation and generation in the water column and sediments of the Black Sea northwestern shelf were carried out within the framework of the interdisciplinary projects "European River-Ocean Systems" (EROS-2000, EROS-21) and "Biogenic Gases Exchange in the Black Sea" (BigBlack) in August 1995, May 1997, and December 1999. Experiments that involved the addition of 14CH3COONa and 14CO2 to sediment samples showed the main part of methane to be formed from CO2. Maximum values of methane production (up to 559 mumol/(m2 day)) were found in coastal sediments in summer time. In winter and spring, methane production in the same sediments did not exceed 3.6-4.2 mumol/(m2 day). The delta 13C values of methane ranged from -70.7 to -81.8@1000, demonstrating its microbial origin and contradicting the concept of the migration of methane from cold seeps or from the oil fields located at the Black Sea shelf. Experiments that involved the addition of 14CH4 to water and sediment samples showed that a considerable part of methane is oxidized in the upper horizons of bottom sediments and in the water column. Nevertheless, it was found that, in summer, part of methane (from 6.8 to 320 mumol/(m2 day)) arrives in the atmosphere.

Methane↗

The analysis of methods for measurement of methane oxidation in landfills.

Landfills and dumps are important sources of atmospheric methane. There is no generally accepted estimate of the influence of methane oxidation on landfill methane emissions. The present work aimed to analyse different methods for the investigation of methane emission and oxidation in methane-producing environments (wetlands, landfills, sludge checks), and to develop the precise procedure for the landfills. The combination of geochemical and microbiological methods to estimate and monitor the oxidation and emission of methane in landfills during different seasons is proposed. It includes the measurements, both on the surface and at different depths (up to 1 m) of landfill ground of the following parameters: (1) concentrations of methane, carbon dioxide and oxygen; (2) quantity of 13C isotope in gas samples; (3) methane-oxidation activity of landfill grounds assayed with two different methods: (a) in conditions of no moisture or substrate limitations, and (b) in conditions with a minimal deviation to in situ conditions; (4) the density of methanotrophic microbial population.

Carbon Dioxide↗

Methane production and colon cancer.

The cause of the high incidence of methane producers in patients with colorectal cancer is not clear. A total of 270 individuals were studied for methane production, using an end-expiratory breath sampling technique. They were divided into eight groups: 156 healthy controls (group 1); 47 patients with colorectal cancer (group 2); 36 patients (34 of them included in the previous group) after resection of the tumor (group 3); 7 (also included in group 2) with nonresectable tumor (group 4); 29 with nonmalignant diseases of the colon (group 5); 12 with extensive ulcerative colitis (group 6); 12 with ulcerative proctosigmoiditis (group 7); and 12 with colonic polyposis (group 8). Significantly more patients (91.4%) with colorectal cancer in group 2 produced methane than either healthy controls (42.9%) (p less than 0.001) or patients with benign diseases of the colon (41.3%) (p less than 0.001). In 36 patients (group 3) in whom the cancer was resected, the incidence of methane producers fell to 47.2%, similar to the control group, but significantly different from group 2 (p less than 0.001). The percentage of methane producers in patients operated on, but with unresectable cancer, remained very high (87.7%). A significantly higher proportion of patients with extensive ulcerative colitis (group 6) and colonic polyposis (group 7) produced methane than patients with ulcerative proctosigmoiditis (group 7), benign diseases of the colon (group 5), and healthy controls (p less than 0.05). The results suggest that the presence of cancer in the large bowel directly influences methane production. In addition, in the group of diseases with a high risk of malignancy, the prevalence of methane-producing individuals was significantly higher than in the healthy population and in patients with benign diseases of the colon.

Adolescent↗

[Role of exogenous carbon dioxide in the metabolism of methane-oxidizing bacteria].

The object of this work was to study the ability of methane oxidizing bacteria to use CO2 as an acceptor of electrons liberated in methane oxidation and the role of CO2 fixation in the constructive metabolism of the bacteria. All of the studied methane oxidizing bacterial cultures were found to be capable of fixing the 14C of hydrocarbonate. The activity of the process was shown to be similar in different strains. Up to 30% of the carbon in the biomass composition could originate from the carbon of HCO3-. Methane oxidizing bacteria that assimilated C1-compounds via the hexulose phosphate and serine pathways had the same level of HCO3- fixation. No differences were found among strains of the same species, among species, or among genera. The assimilation of HCO3- was catalyzed by PEP-carboxylase (i. e. in a heterotrophous way) or, in some cultures, by ribulose-1,5-diphosphate carboxylase, the key enzyme in the autotrophous pathway of CO2 assimilation. The enzymological mechanisms of HCO3- assimilation are discussed. The biological role of CO2 fixation in the metabolism of methane oxidizing bacteria that use the hexulose phosphate and serine pathways of methane assimilation may be different. The process can either play the role of anapleurotic reactions in the tricarboxylic acid cycle, or be an element of the serine pathway of methane assimilation. Calculations have shown that the extent to which a substrate to be metabolized is reduced seems to determine the activity of exogenous CO2 fixation. The contribution made by HCO3- fixation into the carbon metabolism of methane oxidizing bacteria confirms that they are related to lithotrophous organisms.

Bicarbonates↗

Glycoproteins as substrates for production of hydrogen and methane by colonic bacterial flora.

Hydrogen and methane in human breath derive entirely from bacterial fermentation in the intestinal lumen. The sources of substrates utilized for these reactions have not been completely determined. Basal excretion of both gases occurs in the fasted state, while malabsorbed carbohydrate commonly results in increased hydrogen but not methane production. Using an in vitro fecal incubation system, we have studied hydrogen and methane production from glycoproteins of endogenous as well as dietary origin. All glycoproteins tested yielded hydrogen when incubated with fecal homogenates. Mean hydrogen production from substrates containing less than 3% sugar (human serum albumin, bovine serum albumin, and alpha-casein) averaged 2.2 +/- 0.9% of hydrogen production from equivalent amounts of glucose, while carbohydrate-rich mucin yielded 46.0 +/- 6.7% of hydrogen production from glucose. Glycoproteins of intermediate carbohydrate content, including transferrin and egg white, yielded intermediate values. Methane production from glycoproteins was optimal from carbohydrate-poor protein substrates in fecal homogenates which accumulated hydrogen and became rapidly acidic when incubated with pure carbohydrate. In contrast, methane production was comparable for essentially sugar-free proteins, glycoproteins, and glucose when hydrogen did not accumulate and neutral pH was maintained. We conclude that glycoproteins are substrates for hydrogen and methane production by colonic bacteria from healthy adults. In individuals with bacterial overgrowth syndromes and in protein-losing enteropathy, bacterial catabolism of endogenous glycoproteins may cause increased basal hydrogen and methane excretion. These findings also raise the possibility that measurement of hydrogen or methane after oral administration of dietary glycoproteins may be useful in detection of protein malabsorption.

Bacteria↗

[Formation and utilization of lactate during methane fermentation of cellulose].

The role of lactate was studied in the course of methane formation from cellulose by a mixed bacterial association. Lactate was accumulated in the medium during intensive degradation of cellulose and was assimilated in the period of active gas production. Glucose added to the medium was converted into lactate within 30 min. If lactate served as a substrate for a methane forming association which was previously cultivated in a medium containing cellulose, it was almost completely converted into methane; the lag phase was shorter than in the case of cellulose, while the rate of methane formation was higher. In the presence of sulfate in the medium, sulfate reduction preceded methane production from lactate. The combined culture Desulfovibrio vulgaris+Methanosarcina grew in the medium containing lactate, and completely converted it into methane. The formation of methane from lactate by the mixed microbial association can be attributed to the interaction between sulfate reducing and methane producing bacteria.

Bacteria↗

Use of breath hydrogen and methane as markers of colonic fermentation in epidemiological studies: variability in excretion.

Breath hydrogen and methane are specific end products of colonic fermentation, a process which may play a protective role against colon cancer. To assess the possibility of using these markers in epidemiological studies, we characterized the intra- and intersubject variability of breath hydrogen and methane excretion over 15 consecutive days among 32 men and women of various ethnic backgrounds (16 Asians, 8 Caucasians, 8 Hawaiians). Participants were asked to collect four end-expiratory samples each day, which we had shown previously would optimally characterize daily hydrogen excretion. There was substantial within-subject variation in breath hydrogen over the study, although breath methane levels were more constant over time. We found that about 4 days of measurement of breath hydrogen and 1 day of measurement for breath methane are required to correctly characterize individuals according to their long-term excretion of these gases. This was true for Asians and non-Asians. Although breath methane appears to be more practical to measure, it is a less sensitive marker of colonic fermentation than breath hydrogen. Whereas all subjects excreted hydrogen, only 28% of the subjects excreted methane, and methane excretor status of a few participants varied during the study. Because the breath test is noninvasive and reliable, we tested the multiple day collection protocol among colon cancer patients and controls and found it to be well accepted. We conclude that it is practical to measure breath hydrogen and methane in large epidemiological studies conducted at the individual level. The potential use for these markers is discussed.

Aged↗