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The aim of the present study was to determine the in-vitro susceptibility of Toxoplasma gondii to dihydrofolate reductase inhibitors and sulphonamides alone, and in combination. It was found that pyrimethamine had the most potent anti-Toxoplasma activity, while sulphadiazine, sulphamethoxazole and sulphametrole were approximately equally effective, but only at a high concentration. The 50% inhibitory concentration of sulphamethoxazole was determined using different fixed concentrations of pyrimethamine or trimethoprim. It was found that a ten-fold increment of the concentration of pyrimethamine, reduced the IC50 of sulphamethoxazole 1000 times. The influence of trimethoprim on the IC50 of sulphamethoxazole was less concentration dependent. Addition of dihydrofolate or tetrahydrofolate did not influence the IC50 of pyrimethamine. In conclusion, the present dose recommendations for sulphonamides, when combined with pyrimethamine, may be unnecessarily high. There may be a justification for dose reduction of sulphonamides in order to prevent side effects.

The following metabolites of sulfadiazine (S) were isolated from monkey urine by preparative HPLC: 5-hydroxysulfadiazine (5OH), 4-hydroxysulfadiazine (4OH) and the glucuronide (5OHgluc) and sulfate conjugate of 5OH (5OHsulf). The compounds were identified by NMR, mass and infrared spectrometry and hydrolysis by beta-glucuronidase. The analysis of S, the hydroxymetabolites (4OH, 5OH) and conjugates N4-acetylsulfadiazine (N4), 5OHgluc and 5OHsulf in human and monkey plasma and urine samples was performed using reversed-phase gradient HPLC with UV detection. In plasma, S and N4 could be detected in high concentrations, whereas the other metabolites were present in only minute concentrations. In urine, S, the metabolites and conjugates were present. The limit of quantification of the compounds in plasma varies between 0.2 and 0.6 microgram/ml (S 0.31, N4 0.40, 4OH 0.20, 5OH 0.37, 5OHgluc 0.33 and 5OHsulf 0.57 microgram/ml). In urine it varies between 0.6 and 1.1 micrograms/ml (S 0.75, N4 0.80, 4OH 0.60, 5OH 0.80, 5OHgluc 0.80 and 5OHsulf 1.1 micrograms/ml). The method was applied to studies with healthy human subjects and Rhesus monkeys. The metabolites 5OH, 5OHgluc and 5OHsulf were present in Rhesus monkey and not in man. Preliminary results of studies of metabolism and pharmacokinetics in Rhesus monkey and man are presented.