Factors that influence hepatic oxalate synthesis are poorly defined. Hormones are important regulators of hepatic metabolism and could potentially be involved. The effects of hyperglucagonemia were examined in guinea pigs injected with either saline or pharmacological doses of glucagon for 4 days. Glucagon treatment increased mean urinary oxalate excretion by 77% in male and 34% in female animals. The levels of hepatic peroxisomal enzymes involved in oxalate synthesis declined with glucagon treatment, but experiments with isolated peroxisomes indicated that oxalate synthesis in vitro was unaffected. Glucagon decreased hepatic alanine levels by 66%, lactate by 69%, and pyruvate by 73%, but glycolate and glyoxylate levels were unaffected. This decrease in alanine would substantially lower the activity of alanine-to-glyoxylate aminotransferase activity in vivo and make more glyoxylate available for oxalate synthesis. The decrease in lactate and pyruvate concentrations would stimulate the enzymatic conversion of glyoxylate to oxalate and may account for the increase in oxalate synthesis without an increase in glyoxylate concentration. These results demonstrate that hepatic oxalate synthesis is influenced by metabolic changes and that alterations in hepatic alanine, lactate, and pyruvate concentrations may be important elements.