Early detection of colorectal anastomotic leakage (AL) may lead to better outcome. AL may be preceded by change in local metabolism and local ischaemia. Microdialysis of the peritoneal cavity is able to measure these changes in real-time and is minimally invasive. The aim of this prospective cohort study was to compare values of intraperitoneal microdialysis in patients with AL to patients without AL after open and laparoscopic colorectal surgery. Twenty-four patients underwent surgery for left-sided, sigmoid and rectal carcinoma with creation of an anastomosis. Intraoperatively a juxta-anastomotical intraperitoneal and subcutaneous microdialysis catheter was placed. The levels of lactate, pyruvate, glucose and glycerol in the dialysate were measured every 4 h during the first 5 post-operative days, and mean values and area under the curve (AUC) were calculated. Mortality was 0 % and morbidity 38 %. In 3 patients (17 %), AL occurred. In patients with AL, post-operative peritoneal lactate level was 3.2 mmol/l (standard deviation (SD) 0.9) for patients without AL, compared to 4.4 mmol/l (SD 1.5) in case of AL (p = 0.03 for AUC). Intraperitoneal glucose levels were 8.1 mmol/l (SD 1.3), compared to 7.8 mmol/l (SD 2.2) in the complicated course (ns for AUC). Mean intraperitoneal lactate/pyruvate-ratio was 19.2 (SD 3) after colorectal surgery without AL compared to 25 (SD 4.7) in case of AL (non-significant (ns) for AUC). No significant differences were observed between patients who underwent laparoscopic resection and those who underwent open resection. Anastomotic leakage was preceded by a significantly higher AUC and mean value of lactate levels during the first 5 post-operative days. To identify cut-off values for clinical use, pooling of data is necessary.