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Objective To determine the greenhouse gas emissions associated with the energy consumption of Australian ambulance operations, and to identify the predominant energy sources that contribute to those emissions. Methods A two-phase study of operational and financial data from a convenience sample of Australian ambulance operations to inventory their energy consumption and greenhouse gas emissions for 1 year. State- and territory-based ambulance systems serving 58% of Australia's population and performing 59% of Australia's ambulance responses provided data for the study. Results Emissions for the participating systems totalled 67 390 metric tons of carbon dioxide equivalents. For ground ambulance operations, emissions averaged 22 kg of carbon dioxide equivalents per ambulance response, 30 kg of carbon dioxide equivalents per patient transport and 3 kg of carbon dioxide equivalents per capita. Vehicle fuels accounted for 58% of the emissions from ground ambulance operations, with the remainder primarily attributable to electricity consumption. Emissions from air ambulance transport were nearly 200 times those for ground ambulance transport. Conclusion On a national level, emissions from Australian ambulance operations are estimated to be between 110 000 and 120 000 tons of carbon dioxide equivalents each year. Vehicle fuels are the primary source of emissions for ground ambulance operations. Emissions from air ambulance transport are substantially higher than those for ground ambulance transport. [ABSTRACT FROM AUTHOR]