AbstractMethods for measuring aerobic methane oxidation (MOx) rates in aquatic environments are often based on the incubation of water samples, during which the consumption of methane (CH4) is monitored. Typically, incubation vessels are sealed with butyl rubber because these elastomers are essentially impermeable for gases. We report on the potential toxicity of five different commercially available, lab‐grade butyl stoppers on MOx activity in samples from marine and lacustrine environments. MOx rates in incubations sealed with non‐halogenated butyl were > 50% lower compared to parallel incubations with halogenated butyl rubber stoppers, suggesting toxic effects associated with the use of the non‐halogenated butyl type. Aqueous extracts of non‐halogenated butyl rubber were contaminated with high amounts of various organic compounds including potential bactericides such as benzyltoluenes and phenylalkanes. Comparably small amounts of organic contaminants were liberated from the halogenated butyl rubber stoppers but only two halogenated stopper types were found that did not seem to leach any organics into the incubation medium. Furthermore, the non‐halogenated and two types of the halogenated butyl elastomers additionally leached comparably high amounts of zinc. While the source of the apparent toxicity with the use of the non‐halogenated rubber stoppers remains elusive, our results indicate that leaching of contaminants from some butyl rubber stoppers can severely interfere with the activity of MOx communities, highlighting the importance of testing rubber stoppers for their respective contamination potential. The impact of leachates from butyl rubber on the assessment of biogeochemical reaction rates other than MOx seems likely but needs to be verified.