Background: Sufficient distension of the GI lumen is needed for safe advancement of endoscopes and for careful visualization of the mucosa. Carbon dioxide (CO2) has been proposed as an alternative to room air for insufflation. Aim of the Study: To assess the merits and demerits of the use CO2 insufflation for endoscopy in terms of safety and efficacy. Methods: Electronic and manual searches were combined to search RCTs (Randomized controlled trials). After methodological quality assessment and data extraction, the efficacy and safety of CO2 insufflation were systematically assessed. Results: Ten RCTs met the eligibility criteria and included in the present study; six of which on colonoscopy, two on endoscopic retrograde cholangiopancreatography (ERCP) and two on double-balloon enteroscopy (DBE). Postprocedural pain was assessed. Overall, pain was lower in the CO2 insufflation group compared with the air group. Two RCTs found decreased flatus in the CO2 group compared with the air group, and 3 RCTs showed there was decreased bowel distention on abdominal radiography in the CO2 group compared with the air group. Moreover, CO2 insufflation revealed no consistent advantages in the RCTs of DBE, yet it was still indicated safe as air insufflation in stomach/ oesophagus endoscopic submucosal dissection. PCO2 level showed no significant variation during these procedures. Conclusion: CO2 insufflation is proven to be associated with decreased postprocedural pain, flatus, and bowel distention. CO2 insufflation also appears to be safe in patients without severe underlying pulmonary disease.