Abstract Background Respiratory quotient (RQ) is an important variable when assessing metabolic status in intensive care patients. However, analysis of RQ requires cumbersome technical equipment. The aim of the current study was to examine a simplified blood gas-based method of RQ assessment, using Douglas bag measurement of RQ (Douglas-RQ) as reference in a laboratory porcine model under metabolic steady state. In addition, we aimed at establishing reference values for RQ in the same population, thereby generating data to facilitate further research. Methods RQ was measured in 11 mechanically ventilated pigs under metabolic steady state using Douglas-RQ and CO-oximetry blood gas analysis of pulmonary artery and systemic carbon dioxide and oxygen content. The CO-oximetry data were used to calculate RQ (blood gas RQ). Paired recordings with both methods were made once in the morning and once in the afternoon and values obtained were analyzed for potential significant differences. Results The average Douglas-RQ, for all data points over the whole day, was 0.97 (95%CI 0.95–0.99). The corresponding blood gas RQ was 0.95 (95%CI 0.87–1.02). There was no statistically significant difference in RQ values obtained using Douglas-RQ or blood gas RQ for all data over the whole day (P = 0.43). Bias was − 0.02 (95% limits of agreement ± 0.3). Douglas-RQ decreased during the day 1.00 (95%CI 0.97–1.03) vs 0.95 (95%CI 0.92–0.98) P < 0.001, whereas the decrease was not significant for blood gas RQ 1.02 (95%CI 0.89–1.16 vs 0.87 (0.80–0.94) P = 0.11. Conclusion RQ values obtained with blood gas analysis did not differ statistically, compared to gold standard Douglas bag RQ measurement, showing low bias but relatively large limits of agreement, when analyzed for the whole day. This indicates that a simplified blood gas-based method for RQ estimations may be used as an alternative to gold standard expired gas analysis on a group level, even if individual values may differ. In addition, RQ estimated with Douglas bag analysis of exhaled air, was 0.97 in anesthetized non-fasted pigs and decreased during prolonged anesthesia.