Effect of xanthine-related compounds on a theophylline assay using theophylline oxidase

Abstract

Objective: This study investigated a new, nonimmunologic, enzymatic-based theophylline assay for the possible interference of a clinically achievable serum theophylline concentration by four xanthine-related compounds: pentoxifylline, 1-methylxanthine, allopurinol, and oxypurinol.

Rationale: The enzyme theophylline oxidase is used in this assay kit to convert theophylline to 1,3 dimethyluric acid in the presence of cytochrome C. Cytochrome C is reduced to ferrocyte C, which is then measured by ultraviolet spectrophotometry at one minute and again at five minutes. The rate of appearance of ferrocyte C over this time is then related to a one-point 20-micrograms/mL (111-mumol/L) theophylline standard provided with the kit and the concentration of theophylline is calculated. Xanthine-related compounds have the potential to interact with theophylline oxidase via: (1) structural similarity, and (2) inhibition of xanthine oxidase, which increases theophylline metabolite concentrations.

Design: Human serum was spiked with known, therapeutically achievable concentrations of pentoxifylline, 1-methylxanthine, allopurinol, and oxypurinol, both alone and in combination with theophylline in a concentration of 15 micrograms/mL (83.3 mumol/L). Two controls were used: human serum and human serum spiked with theophylline. Data were analyzed statistically with ANOVA, and clinically with +/- 10 percent control criteria.

Results: Means (+/- SEM) for the control samples were 2.0 (0.2) micrograms/mL for serum alone and 15.3 (0.3) micrograms/mL for theophylline-spiked serum. For the test specimens, the results ranged from 2.3 to 2.8 micrograms/mL and from 13.9 to 16.6 micrograms/mL when xanthine-related compounds were added to serum without and with theophylline, respectively.

Conclusions: The results were not statistically or clinically significant for the four xanthine-related compounds at the concentrations studied.