ABSTRACT. Urine samples of cats and dogs collected for 24 hr after a subcutaneous injection of orbifloxacin (OBFX) were analyzed. The metabolites were examined using HPLC. In the dog urine, 87% of total was the parent compound and 13% glucuronide compound of OBFX and 96% was parent and 4% metabolite in the cat urine. The metabolite of cat urine was identified as N-hydroxy OBFX, determined by comparison of the extraction of urine with chloroform with the standard compound of N-hydroxy OBFX, using LC/ APCIMS. N-hydroxy OBFX had a weaker antibacterial activity against fluoroquinolone sensitive bacteria than the parent compound. -KEY WORDS: canine, feline, orbifloxacin metabolite.J. Vet. Med. Sci. 60(11): 1259-1261, 1998 pressure, and the residue was dissolved in methanol. A portion (10 µl) of the resulting solution was subjected to LC/APCIMS. A Hitachi Model M-1000 LC/APCIMS spectrometer (Hitachi Co., Ltd., Tokyo, Japan) attached to a Model L-6200 HPLC system (Hitachi Co., Ltd., Tokyo, Japan) was used, with the following measurement conditions: drift voltage 40 V, focus voltage 113 V, vaporization temperature 270°C and desolvation temperature 399°C. The HPLC conditions were almost the same as those described above, except for the composition of mobile phase (methanol: 0.01% trifluoroacetic acid (42: 58 v/v)). The antibacterial activities of metabolites of OBFX were determined by the method of the Japanese Society of Chemotherapy for assay of minimum inhibitory concentration (MIC) [1].The elution patterns of HPLC chromatograms of urine samples were almost the same in the 4 cats. A typical pattern (cat no. 601) is shown in Fig. 1. (a, b). Two peaks (M-2, M-3) were detected at retention times (t R ) of about 10 min and 23 min. When the chloroform extract of the urine sample which including M-2 was analyzed by LC/APCIMS, a protonated molecule (MH + ) appeared at m/z 412 in the mass spectrum of M-2 (Fig. 2), suggesting that the molecular weight of M-2 was 411. The mass spectrum of N-hydroxy OBFX corresponded to that of M-2 (Fig. 2). The mass spectrum of M-3 corresponded to that of standard OBFX. On HPLC analysis, standard OBFX and standard N-hydroxy OBFX were eluted at t R s of 23 min and 10 min, respectively. These findings suggested that M-3 and M-2 were OBFX and N-hydroxy OBFX, respectively. Two peaks (M-1, M-3) were detected in dog urine at t R s of 5 min and 23 min. A typical pattern (dog no. 111 ) is shown in Fig. 1. (A-C). After the treatment of the urine sample of each dog with β-glucuronidase, the peak of M-1 disappeared and the peak of M-3 on the chromatograph increased.The ratio of each metabolite was calculated on the basis of the peak area on HPLC and the molecular absorption coefficient of each metabolite. In the urine of cats, the ratios of OBFX and N-hydroxy OBFX were 0.96 ± 0.004 Orbifloxacin (OBFX: 1-cyclopropyl-5,6,8-trifluoro-1,4-dihydro-7-(cis-3,5-dimethyl-1-piperazinyl)-4-oxoquinoline-3-carboxylic acid), a fluoroquinolone antibacterial agent, has been developed especially for use in veterinary medi...