PARAOXONASE HAS A MAJOR ROLE IN THE HYDROLYSIS OF PRULIFLOXACIN (NM441), A PRODRUG OF A NEW ANTIBACTERIAL AGENT

NM441 is a prodrug of the new quinolone carboxylic acid antibacterial agent NM394, A rat serum enzyme (NM441-hydrolase) that catalyzes the h ydrolysis of NM441 to NM394 was purified by ultracentrifugation, hepar in-Sepharose column chromatography, and HPLC with a Mono Q anion excha nge column. The enzyme showed a single protein band after sodium dodec yl sulfate-polyacrylamide gel electrophoresis. Its molecular mass was estimated as 46 kDa, The amino-terminal sequence and two internal amin o acid sequences of the NM441-hydrolase resemble those of mouse, rabbi t, and human serum paraoxonases, Moreover, its enzymatic characteristi cs (optimum pH, calcium requirement, and molecular mass) were similar to those of the paraoxonases. These findings identify the NM441-hydrol ase as rat serum paraoxonase, To determine whether the paraoxonase cat alyzes the hydrolysis of NM441 to NM394 in human serum, we investigate d whether the paraoxonase and NM441-hydrolase activities were correlat ed. There was a positive correlation (r = 0.653, p < 0.005) found in t he sera of 67 healthy volunteers, indicating that paraoxonase is respo nsible for the conversion of NM441 to NM394 in humans, Human paraoxona se shows polymorphism. There was a 9-fold variation in paraoxonase act ivity but only a 2-fold variation in NM441-hydrolase activity. These f indings show that paraoxonase polymorphism does not cause marked inter individual variation in NM441-hydrolase activity and is substrate depe ndent.