Introduction: In this study, we aimed to evaluate whether fasting plasma citrulline concentration predicts subsequent glucose absorption in critically ill patients. Methods: In a prospective observational study involving 15 healthy and 20 critically ill subjects, fasting plasma citrulline concentrations were assayed in blood samples immediately prior to the administration of a liquid test meal (1 kcal/ml; containing 3 g of 3-O-methylglucose (3-OMG)) that was infused directly into the small intestine. Serum 3-OMG concentrations were measured over the following 4 hours, with the area under the 3-OMG concentration curve (AUC) calculated as an index of glucose absorption. Results: The groups were well matched in terms of age, sex and body mass index (BMI) (healthy subjects versus patients, mean (range) values: age, 47 (18 to 88) versus 49 (21 to 77) years; sex ratio, 60% versus 80% male; BMI, 25.2 (18.8 to 30.0) versus 25.5 (19.4 to 32.2) kg/m2). Compared to the healthy subjects, patients who were critically ill had reduced fasting citrulline concentration (26.5 (13.9 to 43.0) versus 15.2 (5.7 to 28.6) μmol/L; P<0.01) and glucose absorption (3-OMG AUC, 79.7 (28.6 to 117.8) versus 61.0 (4.5 to 97.1) mmol/L/240 min; P=0.05). There was no relationship between fasting citrulline concentration and subsequent glucose absorption (r=0.28; P=0.12). Conclusions: Whereas both plasma citrulline concentrations and glucose absorption were reduced in critical illness, fasting plasma citrulline concentrations were not predictive of subsequent glucose absorption. These data suggest that fasting citrulline concentration does not appear to be a marker of small intestinal absorptive function in patients who are critically ill.