Benzamidine, an inhibitor of plasmin, was compared with aprotinin (Trasylol) to determine whether it would protect against the degradation of glucagon in human plasma. At concentrations of benzamidine exceeding .05m, insignificant degradation of [125I] glucagon occurred in undiluted plasma up to 72 hr at 4 C. In plasma diluted 1:5 with buffer, glucagon destruction was inhibited by 0.01m benzamidine, which also did not interfere with antigen-antibody affinity using a rabbit antiserum with a high specificity for pancreatic glucagon (Unger antiserum 30K). When added to undiluted plasma in 0.05m benzamidine, recovery of glucagon exceeded 85% as measured by radioimmunoassay in 0.01m benzamidine. Values for immunoreactive glucagon in plasma of subjects after an overnight fast and following arginine stimulation were similar in the presence of benzamidine or Trasylol. At 1% of the cost of Trasylol, benzamidine was found to be as effective as Trasylol in the prevention of glucagon degradation in human plasma.