Anaerobic digestion of swine manure under natural zeolite addition: VFA evolution, cation variation, and related microbial diversity

Batch experiments were carried out on anaerobic digestion of swine manure under 10 % of total solids and 60 g/L of zeolite addition at 35 °C. Four distinctive volatile fatty acid (VFAs) evolution stages were observed during the anaerobic process, i.e., VFA accumulation, acetic acid (HAc) and butyric acid (HBu) utilization, propionic acid (HPr) and valeric acid (HVa) degradation, and VFA depletion. Large decreases in HAc/HBu and HPr/HVa occurred respectively at the first and second biogas peaks. Biogas yield increased by 20 % after zeolite addition, about 356 mL/g VS_added with accelerated soluble chemical oxygen demand degradation and VFA (especially HPr and HBu) consumption in addition to a shortened lag phase between the two biogas peaks. Compared with Ca²⁺ and Mg²⁺ (100–300 mg/L) released from zeolite, simultaneous K⁺ and NH₄ ⁺ (580–600 mg/L) adsorptions onto zeolite particles contributed more to the enhanced biogasification, resulting in alleviated inhibition effects of ammonium on acidogenesis and methanogenesis, respectively. All the identified anaerobes could be grouped into Bacteroidetes and Firmicutes, and zeolite addition had no significant influence on the microbial biodiversity in this study.     

Minimizing nitrous oxide in biological nutrient removal from municipal wastewater by controlling copper ion concentrations

In this study, nitrous oxide (N₂O) production during biological nutrient removal (BNR) from municipal wastewater was reported to be remarkably reduced by controlling copper ion (Cu²⁺) concentration. Firstly, it was observed that the addition of Cu²⁺ (10–100 μg/L) reduced N₂O generation by 54.5–73.2 % and improved total nitrogen removal when synthetic wastewater was treated in an anaerobic–aerobic (with low dissolved oxygen) BNR process. Then, the roles of Cu²⁺ were investigated. The activities of nitrite and nitrous oxide reductases were increased by Cu²⁺ addition, which accelerated the bio-reductions of both nitrite to nitric oxide (NO ₂ ^? ?→?NO) and nitrous oxide to nitrogen gas (N₂O?→?N₂). The quantitative real-time polymerase chain reaction assay indicated that Cu²⁺ addition increased the number of N₂O reducing denitrifiers. Further investigation showed that more polyhydoxyalkanoates were utilized in the Cu²⁺-added system for denitrification. Finally, the feasibility of reducing N₂O generation by controlling Cu²⁺ was examined in two other BNR processes treating real municipal wastewater. As the Cu²⁺ in municipal wastewater is usually below 10 μg/L, according to this study, the supplement of influent Cu²⁺ to a concentration of 10–100 μg/L is beneficial to reduce N₂O emission and improve nitrogen removal when sludge concentration in the BNR system is around 3,200 mg/L.     

Influence of trace elements on biogas production from mango processing waste in 1.5 m3 KVIC digesters

Summary The influence of trace elements (Co²⁺, Ni²⁺ and Fe³⁺) in varying concentrations and combination, was studied in 1.5 m³ Khadi & Village Industries Commission (KVIC) digesters for biogas generation from mangopeel. Addition of these trace metals enhanced the biogas yield and methane content moderately, the maximum being with the iron fed digester. The digesters were always found to be stable without much variation in total volatile fatty acids (VFA), pH, total alkalinity and other parameters. A methane content of 62% and biogas yield of 0.49 m³/kg VS added was obtained with 4000 mg/L FeCl₃ supplemented mangopeel fed digester as compared to control having biogas yield of 0.22 m³/kg VS added with a methane content of about 48–50%.     

Effect of ferrous chloride on biogas production and enzymatic activities during anaerobic fermentation of cow dung and <Emphasis Type="Italic">Phragmites</Emphasis> straw

The effect of ferrous (added as FeCl₂) on the anaerobic co-digestion of Phragmites straw and cow dung was studied by investigating the biogas properties, pH values, organic matter degradation (COD) and enzyme activities (cellulase, protease and dehydrogenase) at different stages of mesophilic fermentation. The results showed that Fe²⁺ addition increased the cumulative biogas yields by 18.1 % by extending the peak period with high daily biogas yields. Meanwhile, the methane (CH₄) contents in the Fe²⁺ added groups were generally higher than the control group before the 15th day. The pH values were not significantly impacted by Fe²⁺ concentrations during the fermentation process. The COD concentrations, cellulase, protease and dehydrogenase activities varied with the added Fe²⁺ concentrations and the stages of the fermentation process. At the beginning stage of fermentation (4th day), Fe²⁺ addition increased the biogas production by improving the cellulase and dehydrogenase activities which caused a decline in COD. At the peak stage of fermentation (8th day), Fe²⁺ addition enhanced the cellulase and protease activities, and resulted in lower COD contents than the control group. When the biogas yields decreased again (13th day), the COD contents varied similar with the protease and dehydrogenase activities, whilst cellulase activities were not sensitive to Fe²⁺ concentrations. At the end of fermentation (26th day), Fe²⁺ addition decreased the cellulase activities, led to lower COD contents and finally resulted the lower biogas yields than the control group. Taking the whole fermentation process into account, the promoting effect of Fe²⁺ addition on biogas yields was mainly attributed to the extension of the gas production peak stage and the improvement of cellulase activities.     

Isolation,selection, and biological characterization research of highly effective electricigens from MFCs for phenol degradation

The microbial fuel cells (MFCs) are recognized to be highly effective for the biodegradation of phenol. For isolating the phenol-degrading bacteria, the sample containing 500 mg/L phenol was collected from the MFCs. The strain (WL027) was identified basing on the 16S rRNA gene analysis and phylogenetic analysis as Bacillus cereus. The effects of pH, temperature, concentrations of phenol, heavy metal ions, and salt on the growth of strain as well as the degradation of phenol have been carefully studied. The WL027-strain exhibited favorable tolerance for the metal cations including Cr²⁺, Co²⁺, Pb²⁺, and Cu²⁺ with the concentration of 0.2 mg/L and NaCl solution with a high concentration of 30 g/L. In 41 h, 86.44% of 500 mg/L phenol has been degraded at the initial pH at 6 and the temperature of 30 °C. The strain was highly active electrogenesis bacteria and the coulombic efficiency reached 64.25%, which showed significant advantage on the efficient energy conversion. Therefore, due to the highly efficient degradation of phenol, WL027-strain could be used in the treatment of phenol-containing wastewater.     

Co-digestion of manure and whey for in situ biogas upgrading by the addition of H2: process performance and microbial insights

In situ biogas upgrading was conducted by introducing H₂ directly to the anaerobic reactor. As H₂ addition is associated with consumption of the CO₂ in the biogas reactor, pH increased to higher than 8.0 when manure alone was used as substrate. By co-digestion of manure with acidic whey, the pH in the anaerobic reactor with the addition of hydrogen could be maintained below 8.0, which did not have inhibition to the anaerobic process. The H₂ distribution systems (diffusers with different pore sizes) and liquid mixing intensities were demonstrated to affect the gas-liquid mass transfer of H₂ and the biogas composition. The best biogas composition (75:6.6:18.4) was obtained at stirring speed 150 rpm and using ceramic diffuser, while the biogas in the control reactor consisted of CH₄ and CO₂ at a ratio of 55:45. The consumed hydrogen was almost completely converted to CH₄, and there was no significant accumulation of VFA in the effluent. The study showed that addition of hydrogen had positive effect on the methanogenesis, but had no obvious effect on the acetogenesis. Both hydrogenotrophic methanogenic activity and the concentration of coenzyme F₄₂₀ involved in methanogenesis were increased. The archaeal community was also altered with the addition of hydrogen, and a Methanothermobacter thermautotrophicus related band appeared in a denaturing gradient gel electrophoresis gel from the sample of the reactor with hydrogen addition. Though the addition of hydrogen increased the dissolved hydrogen concentration, the degradation of propionate was still thermodynamically feasible at the reactor conditions.     

Removal of Contaminating Metals from Soil by Sulfur-Based Bioleaching and Biogenic Sulfide-Based Precipitation

The potential of a hybrid process incorporating sulfur-based bioleaching and sulfide-based precipitation for treatment of metal-contaminated soil was examined in batch-type experiments. The sulfur-based soil bioleaching process with Acidithiobacillus sp. could be initiated at a wide range of initial pH from 4.0 to 6.3. After 15 days, 98% of Zn, 89% of Cu and 79% of Cd was bioleached. The gaseous sulfides recycling from Desulfovibrio sp.-mediated sulfate-reducing reactor via N₂ sparging efficiently treated metal-loaded soil leachate. With a sulfide/metal ratio of 3.0, 88% of Zn, 100% of Cu and 95% of Cd were precipitated, resulting in effluent metal concentrations of 3.5 mg Zn²⁺/L, 0.2 mg Cu²⁺/L and 0.03 mg Cd²⁺/L.

Supplemental materials are available for this article. Go to the publisher's online edition of Geomicrobiology Journal to view the supplemental file.     

Comparative biochemical analysis during the anaerobic digestion of lignocellulosic biomass from six morphological parts of Williams Cavendish banana (Triploid Musa AAA group) plants

We studied banana lignocellulosic biomass (BALICEBIOM) that is abandoned after fruit harvesting, and assessed its biochemical methane potential, because of its potential as an energy source. We monitored biogas production from six morphological parts (MPs) of the “Williams Cavendish” banana cultivar using a modified operating procedure (KOP) using KOH. Volatile fatty acid (VFA) production was measured using high performance liquid chromatography. The bulbs, leaf sheaths, petioles–midribs, leaf blades, rachis stems, and floral stalks gave total biogas production of 256, 205, 198, 126, 253, and 221 ml g^?1 dry matter, respectively, and total biomethane production of 150, 141, 127, 98, 162, and 144 ml g^?1, respectively. The biogas production rates and yields depended on the biochemical composition of the BALICEBIOM and the ability of anaerobic microbes to access fermentable substrates. There were no significant differences between the biogas analysis results produced using KOP and gas chromatography. Acetate was the major VFA in all the MP sample culture media. The bioconversion yields for each MP were below 50 %, showing that these substrates were not fully biodegraded after 188 days. The estimated electricity that could be produced from biogas combustion after fermenting all of the BALICEBIOM produced annually by the Cameroon Development Corporation–Del Monte plantations for 188 days is approximately 10.5 × 10⁶ kW h (which would be worth 0.80–1.58 million euros in the current market). This bioenergy could serve the requirements of about 42,000 people in the region, although CH₄ productivity could be improved.     

Biogas properties and enzymatic analysis during anaerobic fermentation of <Emphasis Type="Italic">Phragmites australis</Emphasis> straw and cow dung: influence of nickel chloride supplement

The importance of nickel (added as NiCl₂) on mesophilic anaerobic fermentation of Phragmites australis straw and cow dung was demonstrated by investigating the biogas properties, pH values, organic matter degradation [chemical oxygen demand (COD)] and enzyme activities (cellulase, protease and dehydrogenase) during the fermentation process. The results showed that Ni²⁺ addition increased the cumulative biogas yields by >18 % by improving the efficiency of first peak stage and bringing forward the second peak stage. The pH values were not significantly influenced by Ni²⁺ addition (p > 0.05). Biogas yields were associated with variations in COD concentrations rather than momentary concentrations. At the start-up stage of fermentation (4th day), the biogas yields increased gradually together with the increase of dehydrogenase activities at elevated Ni²⁺ concentrations when cellulase and protease activities were similar in all test groups. It is suggested that Ni²⁺ addition was mainly dependent on the methanogenic stage. After the start-up stage, the impact of Ni²⁺ addition on biogas production was mainly dependent on its effect on cellulase activities, rather than protease or dehydrogenase activities.     

Influence of cultivation conditions on production of lignocellulolytic enzymes by Ceriporiopsis subvermispora

The aim of this work was to make a survey describing factors that influence the production of extracellular enzymes by white-rot fungus Ceriporiopsis subvermispora responsible for the degradation of lignocellulolytic materials. These factors were: carbon sources (glucose, cellulose, hemicellulose, lignin, maltose and starch), nitrogen sources (ammonium sulphate, potassium nitrate, urea, albumin and peptone), pH, temperature and addition of three different concentrations of Cu²⁺ and Mn²⁺. The cellulase and xylanase activities were similar in medium with different carbon sources and the highest cellulase and xylanase activities were measured in medium with urea and potassium nitrate as nitrogen sources, respectively. The highest laccase activity was observed in medium with lignin and peptone as carbon and nitrogen sources. In other experiments, time course of production of lignocellulolytic enzymes by white-rot fungus C. subvermispora in medium with lignin or glucose as carbon sources was observed.     

Volatile fatty acids (VFAs) and methane from food waste and cow slurry: Comparison of biogas and VFA fermentation processes

The potential of various biomasses for the production of green chemicals is currently one of the key topics in the field of the circular economy. Volatile fatty acids (VFAs) are intermediates in the methane formation pathway of anaerobic digestion and they can be produced in similar reactors as biogas to increase the productivity of a digestion plant, as VFAs have more varying end uses compared to biogas and methane. In this study, the aim was to assess the biogas and VFA production of food waste (FW) and cow slurry (CS) using the anaerobic biogas plant inoculum treating the corresponding substrates. The biogas and VFA production of both biomasses were studied in identical batch scale laboratory conditions while the process performance was assessed with chemical and microbial analyses. As a result, FW and CS were shown to have different chemical performances and microbial dynamics in both VFA and biogas processes. FW as a substrate showed higher yields in both processes (435 ml CH₄/g VS_fed and 434 mg VFA/g VS_fed) due to its characteristics (pH, organic composition, microbial communities), and thus, the vast volume of CS makes it also a relevant substrate for VFA and biogas production. In this study, VFA profiles were highly dependent on the substrate and inoculum characteristics, while orders Clostridiales and Lactobacillales were connected with high VFA and butyric acid production with FW as a substrate. In conclusion, anaerobic digestion supports the implementation of the waste management hierarchy as it enables the production of renewable green chemicals from both urban and rural waste materials.     

A membrane-free baffled microbial fuel cell for cathodic reduction of Cu(II) with electricity generation

A membrane-free baffled microbial fuel cell (MFC) was developed to treat synthetic Cu(II) sulfate containing wastewater in cathode chamber and synthetic glucose-containing wastewater fed to anode chamber. Maximum power density of 314 mW/m³ with columbic efficiency of 5.3% was obtained using initial Cu²⁺ concentration of 6400 mg/L. Higher current density favored the cathodic reduction of Cu²⁺, and removal of Cu²⁺ by 70% was observed within 144 h using initial concentration of 500 mg/L. Powder X-ray diffraction (XRD) analysis indicated that the Cu²⁺ was reduced to Cu₂O or Cu₂O plus Cu which deposited on the cathode, and the deficient cathodic reducibility resulted in the formation of Cu₄(OH)₆SO₄ at high initial Cu²⁺ concentration (500-6400 mg/L). This study suggested a novel low-cost approach to remove and recover Cu(II) from Cu²⁺-containing wastewater using MFC-type reactor.     

Assessment of plant secondary metabolites in oil palm seedlings after being treated with calcium,copper ions and salicylic acid

The effect of calcium, copper ions and salicylic acid (SA) amendment on the incidence of basal stem rot and activity of secondary metabolites in oil palm seedlings were investigated in glasshouse study. Disease incidence (DI) in positive control (T8) was 75% at nine months after inoculation (9 MAI). However, weekly pre-immunisation with Ca^2+?+^?Cu^2+?+^?SA prior to inoculation significantly suppressed DI and delayed disease onset as noted in T7. In the present study, the lowest %DI was observed in T7 (15%) followed by T1, T5, T6, T3, T4 and T2. The Ca²⁺, Cu²⁺ and SA amendments were resulted in earlier and higher accumulation of plant secondary metabolites as noted in leaves, stems and root tissues in response to invasion by Ganoderma boninense. High total phenolic content concentration was detected in T7 (leaf: 233.38 ± 0.12 mg/g; stem: 132.78 ± 0.04 mg/g and root: 86.98 ± 0.28 mg/g). Similar trend was obtained in peroxidase activity, total lignin content and hydrogen peroxide scavenging activity. These results suggested that it could be due to the accumulation of phenolics, peroxidase activities, lignin content and hydrogen peroxide scavenging activities in oil palm seedling tissues which might have collectively contributed to induce resistance against G. boninense.     

Enhancement of erythritol production by Trichosporonoides oedocephalis ATCC 16958 through regulating key enzyme activity and the NADPH/NADP ratio with metal ion supplementation

Erythritol, a well-known natural sweetener, is mainly produced by microbial fermentation. Various metal ions (Al³⁺, Cu²⁺, Mn²⁺, and Ni²⁺) were added to the culture medium of Trichosporonoides oedocephalis ATCC 16958 at 30?mg/L in shake flask cultures. Compared with controls, Cu²⁺ increased the erythritol content by 86% and decreased the glycerol by-product by 31%. After 48 hr of shake flask culture, sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that expression levels of erythrose reductase (ER) in the presence of 30?mg/L CuSO₄?·?5H₂O were higher than those obtained after treatment with other examined metal ions. Furthermore, after 108 hr of batch culture in a 5-L bioreactor, supplementation with 30?mg/L of CuSO₄?·?5H₂O increased the specific erythritol content by 27%. Further studies demonstrated that ER activity under 30?mg/L CuSO₄?·?5H₂O supplementation in a fermentor was overtly increased compared with the control after 60 hr, while glycerol-3-phosphate dehydrogenase activity was clearly reduced in most of the fermentation process. Furthermore, the NADPH/NADP ratio was slightly lower in T. oedocephalis cells treated with Cu²⁺ compared with control cells. These results provide further insights into Cu²⁺ effects on erythritol biosynthesis in T. oedocephalis and should improve the industrial production of erythritol by biological processes.     

Protective effects of pretreatment with Fe2+, Cu2+, and Rb+ on phoxim poisoning in silkworm,Bombyx mori

BackgroundPhoxim is a widely used organophosphorus pesticide in agriculture. People are paying more and more attention to its toxicity. At present, there is no appropriate way to solve the phoxim poisoning of silkworm, which severely affected the development of sericulture. Fe²⁺, Cu²⁺, Rb⁺ exerted their biological effects through various forms in vivo.MethodsTo evaluate the effect of Fe²⁺/Cu²⁺/Rb⁺ on phoxim poisoning in silkworm, Bombyx mori were treated with fresh mulberry leaves soaked in 2.5 mg/L phoxim for 2 min with 50 mg/L FeCl₂, 150 mg/L CuCl₂, or 0.5 mg/L RbCl from 5 days of the fifth-instar silkworm.ResultsFe²⁺, Cu²⁺, and Rb⁺ pretreatments significantly inhibited the phoxim-induced reduction of survival rate and alleviated the phoxim-induced poisoning symptoms. The protective effects of Fe²⁺, Cu²⁺, and Rb⁺ on phoxim poisoning might be due to their enhancement of superoxide dismutase (SOD), catalase (CAT), and carboxylesterase (CarE) in the hemolymph and fat body of silkworm. This enhancement might reduce reactive oxygen species (ROS) accumulation and oxidative stress (OS) caused by phoxim poisoning. Thereby it reduced the damage to silkworm tissues and cells.ConclusionsThese results showed that Fe²⁺, Cu²⁺, and Rb⁺ treatments protected the silkworm from phoxim poisoning by directly enhancing the activity of SOD, CAT, and CarE enzymes and reducing oxidative stress, but not dependent on the high expression of CYP genes. The use of Fe²⁺, Cu²⁺, and Rb⁺ to enhance the activity of SOD, CAT, and CarE enzymes may be an underlying effective way to solve phoxim poisoning in the silkworm industry.     

叶面喷施氨基酸和微量元素对金银花生长发育和质量的影响

以2年生金银花为试验材料,采用叶面喷施法,研究不同浓度的苯丙氨酸(Phe)、酪氨酸(Lyr)以及锌(Zn2+)、铜(Cu2+)对金银花生长发育和质量的影响。结果显示:(1)喷施不同浓度的Phe、Lyr以及Zn2+、Cu2+对叶面积无明显影响;不同处理的叶绿素含量随喷施次数的增加而出现不同程度的下降,喷施浓度适宜则有助于叶绿素的合成;喷施一定浓度的Phe、Lyr以及Zn2+、Cu2+可增加花蕾重量,如经1 000mg/g Phe处理后的花蕾鲜重与干重较对照增加了20.1%和51.4%。(2)不同浓度的Phe、Lyr可显著影响碳代谢,但对氮代谢影响不明显;Zn2+、Cu2+对碳氮代谢产物影响较明显,如喷施10mg/L的CuSO4及ZnSO4可提高可溶性糖及淀粉含量。(3)除Zn2+处理后的花蕾类黄酮含量显著低于对照外,其他处理较CK无显著差异;花蕾总黄酮含量均显著低于对照,但绿原酸含量均高于对照。(4)叶片中离子含量受喷施次数及浓度影响较明显,除30mg/L CuSO4处理外,其它处理的花蕾中Zn2+、Cu2+、Fe2+含量均显著低于对照。研究表明,在金银花的第一茬花抽枝初期喷施适宜浓度的Phe、Lyr(如1 000mg/g Phe、2 000mg/g Lyr)以及Zn2+、Cu2+(如50mg/L ZnSO4、10mg/L CuSO4)可改善金银花的生长发育,并提高产量和质量。     

Copper N2S2 Schiff base macrocycles: The effect of structure on redox potential

A series of bis-salicylidene based N₂S₂ copper macrocycles were prepared, structurally characterised and subjected to electrochemical analysis. The aim was to investigate the effects of length of polymethylene chains between either the imine donors or the sulfur donors on redox state and potential of the metal. The complexes structurally characterised had either distorted square planar or tetrahedral geometries depending on their oxidation state (Cu²⁺ or Cu⁺, respectively), and the N–(CH₂)_n–N bridge was found to be most critical moiety in determining the redox potential and oxidation state of the copper macrocycles, with relatively little change in these properties caused by lengthening the S–(CH₂)_n–S bridge from two to three carbons. In fact, a weakness was observed in the complexes at the sulfur donor, as further lengthening of the S–(CH₂)_n–S methylene bridge to four carbons caused fission of the carbon–sulfur bond to give dimeric rings and supramolecular assemblies. Cu⁺ complexes could be oxidised to Cu²⁺ by tert-butylhydroperoxide, with a corresponding change in the spectrophotometric properties, and likewise Cu²⁺ complexes could be reduced to Cu⁺ by treatment with β-mercaptoethylamine. However, repeated redox cycles appeared to compromise the stability of the macrocycles, most probably by a competing oxidation of the ligand. Thus the copper N₂S₂ macrocycles show potential as redox sensors, but further development is required to improve their performance in a biochemical environment.     

Cu2+-catalyzed oxidative degradation of thyroglobulin

Thyroglobulin (Tg) was subjected to metal-catalyzed oxidation, and the oxidative degradation was analyzed by SDS-polyacrylamide gel electrophoresis under reducing conditions. In contrast to no effect of hydrogen peroxide (H₂O₂) alone on the Tg degradation, the inclusion of Cu²⁺ (30 μM), in combination with 2 mM H₂O₂, caused a remarkable degradation of Tg, time- and concentration-dependent. The action of Cu²⁺ was not mimicked by Fe²⁺, suggesting that Tg may interact selectively with Cu²⁺. A similar degradation of Tg was also observed with Cu²⁺corbate system, and the concentration of Cu²⁺ (5–10 μM), in combination with ascorbate, required for the effective degradation was smaller than that of Cu²⁺ (10–30 μM) in combination with H₂O₂. In support of involvement of H₂O₂ in the Cu²⁺ corbate action, catalase expressed a complete protection. However, hydroxyl radical scavengers such as dimethylsulfoxide or mannitol failed to prevent the oxidation of Tg whereas phenolic compounds, which can interact with Cu²⁺, diminished the oxidative degradation, presumably consistent with the mechanism for Cu²⁺-catalyzed oxidation of protein. Moreover, the amount of carbonyl groups in Tg was increased as the concentration (3–100 μM) of Cu²⁺ was enhanced, while the formation of acid-soluble peptides was not remarkable in the presence of Cu²⁺ up to 200 μM. In further studies, Tg pretreated with heat or trichloroacetic acid seemed to be somewhat resistant to Cu²⁺-catalyzed oxidation, implying a possible involvement of protein conformation in the susceptibility to the oxidation. Based on these observations, it is proposed that Tg could be degraded non-enzymatically by Cu²⁺-catalyzed oxidation.     

Effects of copper on expression of methane monooxygenases,trichloroethylene degradation,and community structure in methanotrophic consortia

Copper plays a key role in regulating the expression of enzymes that promote biodegradation of contaminants in methanotrophic consortia (MC). Here, we utilized MC isolated from landfill cover to investigate cometabolic degradation of trichloroethylene (TCE) at nine different copper (Cu²⁺) concentrations. The results demonstrated that an increase in Cu²⁺ concentration from 0 to 15 μM altered the specific first‐order rate constant k_1,TCE, the expression levels of methane monooxygenase (pmoA and mmoX) genes, and the specific activity of soluble methane monooxygenase (sMMO). High efficiency TCE degradation (95%) and the expression levels of methane monooxygenase (MMO) were detected at a Cu²⁺ concentration of 0.03 μM. Notably, sMMO‐specific activity ranged from 74.41 nmol/(mg_cell h) in 15 μM Cu²⁺ to 654.99 nmol/(mg_cell h) in 0.03 μM Cu²⁺, which contrasts with cultures of pure methanotrophs in which sMMO activity is depressed at high Cu²⁺ concentrations, indicating a special regulatory role for Cu²⁺ in MC. The results of MiSeq pyrosequencing indicated that higher Cu²⁺ concentrations stimulated the growth of methanotrophic microorganisms in MC. These findings have important implications for the elucidation of copper‐mediated regulatory mechanisms in MC.