The ectopic deposition of fat in liver and muscle during obesity is well established, however surprisingly little is known about the intestine. We used ob/ob mice and wild type (C57BL6/J) mice fed a high-fat diet (HFD) for 3 weeks, to examine the effects on intestinal mucosal triacylglycerol (TG) accumulation and secretion. Obesity and high-fat feeding resulted in higher levels of mucosal TG and markedly decreased rates of chylomicron secretion, accompanied by alterations in intestinal genes related to anabolic and catabolic lipid metabolism pathways. Overall, the results demonstrate that during obesity and a HFD, the intestinal mucosa exhibits metabolic dysfunction. There is indirect evidence that the lipolytic enzyme monoacylglycerol lipase (MGL) may be involved in the development of obesity. We therefore examined the role of MG metabolism in energy homeostasis using wild type and MGL-/- mice fed low-fat or high-fat diets for 12 weeks. Tissue MG species were profoundly increased, as expected. Notably, weight gain was blunted in all MGL-/- mice. MGL null mice were also leaner, and had increased fat oxidation on the low-fat diet. Circulating lipids levels were decreased in high fat-fed MGL-/- mice, as were the levels of several plasma peptides involved in energy homeostasis. Interestingly, MGL-/- mice had a blunted rate of intestinal TG secretion following an oral fat challenge. The leaner phenotype and improved metabolic serum profile in MGL-/- mice suggested that pharmacological inhibition may be a potential treatment for metabolic disease. To further examine this, C57BL6/J mice were fed low-fat and high-fat diets for 12 weeks, and then given daily oral administration of vehicle or a novel reversible MGL inhibitor for 4 days or 27 days. No changes in food intake were found, nor were adiposity or glucose intolerance substantially altered by inhibitor treatment; this is likely due to the short-term effectiveness of the inhibitor, as the compound was barely detectable 7 hours following administration. Thus, the effects of transient MGL inhibition on energy metabolism are minimal, in contrast to chronic inhibition secondary to genetic ablation.