Insufflation with carbon dioxide (CO2) in colonoscopy has not been widely adopted and, consequently, limited data are available on insufflated gas volume and blood pCO(2). The aim of this study was to compare CO2 and air as an insufflation agent in patients undergoing colonoscopy without sedation in terms of insufflated gas volume, pCO(2), pain and examination time. This was a randomized, double-blind, control trial. Consecutive patients presenting for colonoscopy, excluding those with lung or malignant disease, were randomized into two groups: insufflation with air or with CO2, respectively. Insufflated gas volume, pain, pCO(2) and examination time were assessed. The study cohort comprised 120 patients (66 randomized to CO2 group). No significant difference in insufflated gas volumes was found between the CO2 and air groups. The mean pCO(2) measured before, during (the peak value) and 30 min after colonoscopy were 40, 43 and 40 mmHg, respectively, in both groups. The pain scores in the air group were significantly greater than those in the CO2 group until 3 h after the examination. There was a significant faster cecal intubation time and a trend toward shorter examination time in the CO2 group. The CO2 gas volume used in our study (14.0 L) was much greater than that reported by others (8.3 L), but the pCO(2) values were still within the normal reference range, indicating the safety of CO2 insufflation over a greater range of CO2 gas volume. Among our patients, CO2 insufflation was associated with relatively less pain and a shorter examination time. Based on our results, we recommend that CO2 become the standard gas for insufflation in patients undergoing colonoscopy without sedation.