Studies On Bio-oxidizing Methane In Coal Mines With Microbial Technology

In this thesis, an efficient methanotrophic system was gained from the rice field soil at Huajiachi campus, Zhejiang University, as enriching source after domestication with methane for a long time and optimization of medium components and culture conditions. A biofilter was designed to control methane concentration in coal mines with the methanotrophic system used to form a biofilm, and fed with methane at the concentration range of 0-20% (v/v) which is far higher than that at explosion range in coal mine gas (5%-16%), for exploring the possibility of bio-oxidation of methane in coal mine gas. The start-up and running process of the biofilter was studied systematically. The running regularity was studied and simulated mathematically during the biofilter was stably operated. The main results of this study are as follows:(1) The effects of methane oxidation of the methanotrophic systems were compared with the rice field soil at Huajiachi campus, Zhejiang University and the soil of Tianziling landfill in Hangzhou as enriching sources. The result indicated that the methane oxidation rate of the methanotrophic system with the rice field soil as enriching source was evidently better than that with the landfill soil. Then, the medium components and the culturing conditions were optimized for the better methanotrophic system. The optimized medium was composed of (g·L^-1): MgSO₄·7H₂O 0.5, KNO₃ 1, Na₂HPO₄·12H₂O 0.36, KH₂PO₄ 0.28, CaCl₂·6H₂O 0.1, NH₄Cl 0.25, and 1 ml trace elements solution. The trace element solution contained (g·L^-1): Na₂EDTA 0.5, FeSO₄·7H₂O 0.4, CuSO₄·5H₂O 0.06. And the optimum conditions were pH 6, the culturing temperature at 30℃, and V_O2:V_CH4 > 2:1. The methanotrophic system could oxidize methane absolutely with concentration in the explosion range of coal mine gas within a short time under the optimum medium and the culturing conditions. It took only 40h for the methanotrophic system to eliminate methane thoroughly with methane concentration of 5%, and 48h when the methane concentration was 10%, and 56h with 15% methane. The methantrophic system always kept increasing of OD₅₆₀ in the process of methane oxidation, indicating that the methanotrophic system had the potential to oxidize methane with a higher concentration.(2) The main influence of poisonous gases on the bio-oxidation of methane in coal mine gas were as the follows: A. as the results of the single factor experiments, there were no distinct changes of CH4 oxidation rate when the content of CO increased from 0 to 250 times, and the same while H₂S and SO₂ increased from 0 to 100 times as the permitted content set by the National Coal Mine Safety Regulations, and no significant inhibition was observed in the CH4 oxidation process, although sometimes there were clear differences in the activity of MMO. The result indicated that the possible concentrations of single CO, H₂S or SO₂ in the coal mine gas could not restrain the bioactivity of methantrophic system. However, the mixture of the three kinds of gas had a inhibitory effect to the CH4 oxidation of the methantrophic system although it was slight and not distinct (p>0.05) . It could be concluded that the co-existence of CO, H₂S, and SO₂ in coal mine gas would not produce obvious inhibition to the methantrophic system as the content of the three kinds of gas tested was far higher than the content limit of that permitted by the National Coal Mine Safety Regulations. No valuable contents of CO H₂S SO₂ in the samples could be detected after incubation for 48 hours, illustrated that some species of bacteria that could exhaust CO, H₂S, and SO₂ survived in the methantroph system. This maybe was one of the possible reasons that CO, H₂S, and SO₂ at the tested concentrations did not produce clear inhibition to the methantroph system. No distinct differences could be observed among the bands of SDS-PAGE of the sample CK, CO-2, CO-5, H₂S-3, H₂S-5, SO₂-3, and SO₂-5, suggesting that the methanotroph system would not produce clear emergency responses to CO, H₂S, and SO₂ at the tested content. However, one band of the sample with all three gases existed was missed compared with the other samples although the methane oxidation rate and the MMO activity didn't have evident variance, which implied that the missed band had no direct relation with the MMO. All the experiment results demonstrated that the possible contents of CO, H₂S, and SO₂ in the coal mine gas in this research did not produce clear inhibition to the CH₄ oxidation activity of methantrophic system. Therefore, the adoption of this methantrophic system would be stable, safe and reliable for treating the coal mine gas.(3) The operation process of the biofilter which was used to control methane concentration under coal mines was systematically studied. Six kinds of packing materials were compared at static and dynamic states. The results indicated that the biofilter filled with large size filtration porcelain balls had the biggest capability for oxidizing methane and standing against the wallop of the inlet gas with its proper specific surface area, grain diameter and density, and was the most available one as the filling material of the biofilter to remove methane in coal mines. The methane elimination capacity went up along with methane inlet loading increasing after the biofilter operated stably. However, it would keep at a stable level when the inlet loading of methane increased to some degree. The methane removal efficiency did not change, but remained nearly 1470 mg /L·h, for different EBRT (empty bed retention time), which was just the optimal removal capacity of this bioreactor. The biofilter had a higher methane removal rate when the inlet methane concentration was lower, and would diminish it with the further increase of inlet methane concentration. The methane removal rate decreased dramatically if the EBRT was shorter. The effect of EBRT on the methane removal rate was not evident when the concentration of methane was lower than 50mg/L. With the concentration increasing, the biofilter had a higher removal rate of methane when the EBRT was longer. Each section of the biofilter had various contributions to the methane removal with different inlet methane concentration. The first section of biofilter had the most contribution to the methane removal with lower inlet methane concentration and longer EBRT (the gas flow rate (GFR) was low). The difference of the contribution rates between each section would diminish when inlet methane concentration increased. More contribution to methane removal efficiency by the second section of the reactor was observed when EBRT was shorter (the inlet GFR was high). The effect of liquid flow rate (LFR) on the operation of biofilter was more significant at higher inlet methane concentrations and GFR. When GFR and the inlet methane concentration were low (30L/h, 35mg/L), the methane removal rate had no evident variance with the liquid flow rate (LFR) from 25ml/min to 75ml/min. That LFR kept at 75ml/min was appropriate when GFR was at 30-90L/h. When GFR was lower than 60L/h, and LFR at 75mL/min, nutrition liquid must be sprayed each 10h again to have enough water for a stable methane oxidation. However, when GFR is higher than 60L/h, the spraying frequency must be enhanced for keeping the biofilm growing regularly. The biofilter had an excellent ability to deal with unexpected events and malfunction. It took about 9h for the biofilter to restore the methane oxidation rate after 3days of running interregnum when GFR was 45L/h, at least 16h if the biofilter intermitted running for 7days, The operation life-span of biofilter biofilm with the large size filtration porcelain balls as packing material had about 160 days when GFR was 45L/h, LFR was 75mL/min, and sprayed 1h every 10h, during which, the biofilter had a good performance to oxidize methane. After that, its capacity to removal methane got worse rapidly as the mould propagated in the biofilter.(4) The microscopic kinetic model for methane's bio-removal and absorption on the surface of the biofilm in the biofilter was constructed according to the material balance and typical Langmuir absorption formula, and showed as follows:The result of contrasting experimental values to the simulation data with the above formula of the outlet methane concentration indicated that the simulation data with the kinetic model were really approaching to the experimental values. All the relevant coefficients were higher than 0.94. The simulation data had good relevance with the experimental data of the bio-removal capacity when GFR and the inlet concentration of methane were lower. Some deviation appeared gradually between the simulated data and the experimental values with GFR and the inlet methane concentration increasing. However, the relevant coefficients of the two surpassed 0.89 when the methane concentrations were within the range of 0-20% tested in this work. Such results demonstrated that the kinetic model constructed based on the "absorption-biofilm" theory was valuably applied to the methane removal process of the biofilter in coal mines. The simulation formula could be realized in the practical process, and could offer some guidance and help for the relative theory study or practical operation.