Plasma concentration of penicillin G was evaluated in beef steers after administration of either a combination of benzathine penicillin G and procaine penicillin G in a 1:1 mixture at a dosage of 9,000 U/kg of body weight, IM (n = 5), 24,000 U/kg, IM (n = 5), or 8,800 U/kg, SC (n = 5), or benzathine penicillin G alone at a dosage of 12,000 U/kg, IM (n = 7). Plasma concentration of penicillin G was measured by use of a high-performance liquid chromatography assay that had a limit of determination of 0.005 microgram/ml. At a dosage for this combination of 9,000 U/kg IM, and 8,800 U/kg, SC, which are approved label recommendations in Canada, and the United States, respectively, mean (+/- SEM) peak plasma concentration was 0.58 (+/- 0.15) and 0.44 (+/- 0.02) microgram/ml, respectively. Although plasma penicillin concentration was quantifiable for 7 days in the steers that received 9,000 U/kg, IM, and for 4 days in the steers that received 8,800 U/kg, SC, the concentration was < 0.1 microgram/ml in both groups after the first 12 hours. After administration of the combination at dosage of 24,000 U/kg, IM, there was an initial peak plasma concentration at approximately 2 hours; thereafter, plasma concentration decreased slowly, with half-life of 58 hours. Although plasma penicillin G concentration was quantifiable for 12 days at this dosage, concentration was < 0.1 microgram/ml after the first 48 hours. After the initial 48 hours, plasma concentration of penicillin was of similar magnitude and decreased at similar rate for the combination at dosage of 24,000 U/kg and for 12,000 U/kg of benzathine penicillin G alone.(ABSTRACT TRUNCATED AT 250 WORDS)