The tricyclic quinolone antibacterial agent 6,7-dihydro-5,8-dimethyl-9-fluoro-1-oxo-1H,5H-benzo[ij]quinolizine -2-carboxylic acid has an asymmetric center at position 5 of the molecule. The R and S isomers of the compound have been prepared from the corresponding (R)- and (S)-2,5-dimethyl-6-fluoro-1,2,3,4-tetrahydroquinolines, which were separated via their diastereomeric amides of N-tosyl-(S)-proline. The absolute configuration was established by X-ray analysis of one of the diastereomeric amides. The 5-desmethyl analogue was prepared for antibacterial comparison with the isomers and the racemic mixture. It has now been established that the S isomer is much more active than the R isomer. The 5-desmethyl analogue was found to be more active than the R isomer but not as active as the S isomer or the racemic mixture. The importance of stereochemistry at position 5 in this system has been established.

The antimicrobial activity of flumequine (R-802) was characterized by in vitro and in vivo procedures. Assay of the minimal inhibitory concentrations for 321 recent clinical isolates revealed that 88% of the gram-negative bacteria were inhibited by an R-802 concentration of 6.2 ,ug/ml or less. Cross-resistance in laboratory-derived mutants ofProteus vulgaris was essentially complete for R-802, nalidixic acid, and oxolinic acid, although quantitative differences were evident. R-802 was more effective than either of these quinolone antibacterials in preventing the development of experimental murine pyelonephritis (P. vulgaris). R-802 and trimethoprim/sulfamethoxazole (1:5) were equally effective in resolving a P. mirabilis-induced prostatitis of rats.Flumequine (R-802) is a synthetic antibacterial, selected from a series of tricyclic quinolines prepared in Riker Laboratories. The compound is 6,7-dihydro-9-fluoro-5-methyl-1-oxo-1H,5H (U) quinolizine-2-carboxylic acid (Fig. 1) Antimicrobials. Flumequine (R-802) was synthesized in Riker Research Laboratories. Stock solutions of R-802 (1%) for in vitro use were prepared with 0.1 N sodium hydroxide. Nalidixic acid and oxolinic acid were provided by Winthrop Laboratories and Warner-Lambert Research Institute, respectively. Bactrim (sulfamethoxazole and trimethoprim, 5:1), Erythrocin (erythromycin stearate), and Terramycin (oxytetracyline-hydrochloride) were obtained in commercial form.Susceptibility. The antimicrobial spectrum of R-802 was surveyed on tryptone soy agar (TSA) plates with a log,0 dilution series. Microliter aliquots of stock solution were mixed with molten TSA in individual screw-cap vials and dispensed into petri dishes. These preparations were allowed to harden and were surface inoculated with stock culture suspensions. The conditions of incubation were overnight at 37 C in a humidified atmosphere enriched with 10% carbon dioxide. Growth inhibition of those strains that appeared to be most susceptible to R-802 was further examined by a log2 dilution tube technique in nutrient broth (5). Nalidixic acid and oxolinic acid were included in this titration as reference quinolone antibacterials. Bacillus subtilis was deleted in this comparison because of pellicle formation in static broth culture; Proteus vulgaris, which was not tested on TSA because of the "swarming" phenomenon, was included here. The minimal inhibitory concentrations (MICs) of R-802 for clinical isolates of urinary tract infection were similarly determined; the influence ofculture medium pH was examined in selected strains at 0.2-unit intervals over the range 6.0 to 8.0.Cross-resistance of mutant clones selected by growth on oxolinic acid, nalidixic acid, and R-802 was assessed by the replica plating technique (6). Approximately 109 cells were spread over the surface of drug plates containing TSA plus 0.15% bile salts to inhibit swarming ofP. vulgaris 210. The levels of drug permitting development of approximately 10 colonies/plate were as follows: R-802 and oxolinic acid, 10 ,ig/ml; nalidix...

A sensitive and specific high-pressure liquid chromatographic method is described for the determination of the antibacterial drug flumequine and a major metabolite, 7-hydroxyflumequine, in human plasma and urine. The assay was linear over a concentration range of 1 to 120 pLg/ml for both compounds. This method is compared with fluorometric and microbiological assays for flumequine. These latter methods did not differentiate between flumequine and any fluorescent or antimicrobiologically active metabolites. However, because essentially all drug in the plasma was found to be flumequine in radiolabeled studies, levels of unchanged drug in the plasma could be quantitated by either high-pressure liquid chromatography or fluorometry. Although only high-pressure liquid chromatography was able to specifically measure flumequine in the urine, the antimicrobial activity of the urine, which is more therapeutically relevant due to antimicrobially active metabolites, could be quantitated by either the fluorometric or the microbiological assay.Flumequine (I) is a quinolone derivative active against a broad spectrum of gram-negative bacteria, some of which are associated with or responsible for urinary tract infections. It is commercially available in Europe for human and veterinary use. Chemically, I is 9-fluoro-6,7-dihydro-5-methyl-1-oxo-lH,5H-benzo[ii]quinolizine-2-carboxylic acid.Some chemical, microbiological, and metabolic aspects of I have been reported previously (1, 2).In the earlier stages of I development, two microbiological assays and a fluorometric method were employed for analysis of urine and plasma samples. For pharmacokinetic and biopharmaceutical studies, a specific and sensitive highpressure liquid chromatographic (HPLC) method was developed which can simultaneously quantitate I and a 7-hydroxy metabolite, 7-hydroxyflumequine (II) (Fig. 1 hydroxide. The mobile phase was pumped at 1.5 ml/min through a stainless steel column (61 cm by 2.3-mm inside diameter) containing a strong anion-exchange resin (Zipax SAX, Du Pont Co., Wilmington, Del.). The best results were obtained with prepacked columns purchased from the supplier.To 1 ml of plasma were added 100 ,ug of the internal standard, quinaldic acid (100 ,ul of a stock solution of 1 mg of quinaldic acid in 1 ml of methanol), 0.5 ml of 0.6 N HCl, and 5 ml of chloroform. The tubes were shaken on a reciprocal shaker for 5 min (125 cycles/min) and then centrifuged for 10 min at 750 x g. The aqueous phase was discarded. To the chloroform phase was added 0.5 ml of 0.6 N NaOH. The tubes were shaken and centrifuged, and a 10-to 25-,ul sample of the aqueous phase was injected. Urine samples were extracted with 5 ml of chloroform after 300 ,ug of quinaldic acid (300 pAl of the stock solution described above) and 1.0 ml of 0.5 M phosphate buffer, pH 6.0, were added to the original 1 ml of urine sample. A larger amount of internal standard was added to the urine because the larger aqueous volume and higher pH of the urine extraction reduced the recovery of this compound. ...

Resolution of the (R)‐ and (S)‐isomers of the tetrahydroquinoline (I) is achieved by flash chromatography of the diastereomeric amides (III) and (IV).