Potent in vitro interference of fleroxacin in DNA-binding, unwinding and ATPase activities of Bloom helicase.

OBJECTIVE: To study the effect of fleroxacin (FLRX) on biological properties of Bloom (BLM) helicase catalytic core (BLM642-1290 helicase) in vitro and the molecular mechanism of interaction between the two molecules. METHODS: DNA-binding and unwinding activities of BLM642-1290 helicase were assayed by fluorescence polarization and gel retardation assay under conditions that the helicase was subjected to different concentrations of FLRX. Effect of FLRX on helicase ATPase activity was analyzed by phosphorus-free assay based on a colorimetric estimation of ATP hydrolysis-produced inorganic phosphate. Molecular mechanism of interaction between the two molecules was assayed by ultraviolet and fluorescence spectra. RESULTS: The DNA unwinding and ATPase activities of BLM642-1290 helicase were inhibited whereas the DNA-binding activity was promoted in vitro. A BLM-FLRX complex was formed through one binding site, electrostatic and hydrophobic interaction force. Moreover, the intrinsic fluorescence of the helicase was quenched by FLRX as a result of non-radioactive energy transfer. The biological activity of helicase was affected by FLRX, which may be through an allosteric mechanism and stabilization of enzyme conformation in low helicase activity state, disruption of the coupling of ATP hydrolysis to unwinding, and blocking helicase translocation on DNA strands. CONCLUSION: FLRX may affect the biological activities and conformation of BLM642-1290 helicase, and DNA helicase may be used as a promising drug target for some diseases.

Relación estructura-fotoestabilidad u fototoxicidad de fluoroquinolonas / Structure-photostability and phototoxicity relationship in fluoroquinolones

Las quinolonas constituyen una familia de antibióticos de amplio uso en la actualidad, debido a su gran eficacia clínica en el tratamiento de infecciones del tracto respiratorio, urinario, tejidos blandos y enfermedades de transmisión sexual. De acuerdo a su estructura base se pueden clasificar en quinolonas naftiridínicas (enoxacino y ácido nalidíxico), quinolínicas (ciprofloxacino, norfloxacino, ácido oxolínico, rosoxacino) y pirimido-pirimidinicas (ácido pipemídico). Las quinolonas se caracterizan por presentar reacciones de fotosensibilidad y fotolabilidad. El ácido nalidíxico, quinolona de primera generación, presenta reacciones de fototoxicidad, evaluadas en el modelo del glóbulo rojo y en cultivos celulares. El ácido nalidíxico, exento del sustituyente piperazínico presenta una fotolabilidad disminuida. Las fluoroquinolonas, a diferencia del ácido nalidíxico, presentan un anillo piperazínico o metil piperazínico en posición 7, incorporado con el fin de mejorar las propiedades antibacterianas. En este trabajo se investiga la influencia del anillo piperazínico en posición 7 sobre la fotolabilidad y fototoxicidad, demostrándose que las fluoroquinolonas que poseen este sustituyente presentan una fotolabilidad aumentada y una fototoxicidad disminuída, en relación a quinolonas carentes de éste grupo. Es probable que la fotodegradación de quinolonas transcurra mediante un mecanismo radicalario, con la pérdida del grupo carboxílico

Quinolones constitute a large class of synthetic antimicrobial agents that are highly effective in the treatment of respiratory, urinary, sexually transmitted diseases and soft tissue infection. In agreement with his structure the quinolones can be classified into naftaridines (enoxacin and nalidixic acid), quinolines (ciprofloxacin, norfloxacin, oxolinic acid, roxosacin) and pyrid-pyrimidin (pipemidic acid). Some quinolones may undergo photodegradation reactions. On the other hand, quinolones can induce cutaneous photosensitivity reactions.and photolability as well. Nalidixic acid, a first quinolone generation, may undergo phototoxic effects on the red blood cells and in cell culture. Nalidixic acid, which has not the piperazine group in position 7, exhibit a moderated photolability. The fluoroquinolones as oppossed to nalidixic acid have a piperazine ring in position 7. We investigated the influence of the piperazine ring on the phototoxicity and photolability of several quinolones. We demonstrated that the fluoroquinolones with piperazinic group present higher photolability and less phototoxicity .It is possible that the photodegradation of quinolones takes place through a radical pathway, with the loss of a carboxylic acid group

Synthesis and antibacterial activity of 1-(2-fluorovinyl)-7-substituted-4-quinolone-3-carboxylic acid derivatives, conformationally restricted analogues of fleroxacin.

The novel 1-(2-fluorovinyl)-4-quinolone-3-carboxylic acid derivatives Z-15a-c, E-15a-c, Z-16a-c, and E-16a-c, conformationally restricted analogues of fleroxacin (5), were synthesized, and their in vitro antibacterial activity was evaluated. A dehydrosulfenylation of a 2-fluoro-2-[(4-methoxyphenyl)sulfinyl]ethyl group was employed as a key step for the construction of a 2-fluorovinyl group at the N-1 position. It appeared evident that the Z-isomers Z-15a-c and Z-16a-c exhibited 2- to 32-fold more potent in vitro antibacterial activity than the corresponding E-isomers E-15a-c and E-16a-c. Furthermore, since Z-15b showed in vitro antibacterial activity and DNA gyrase inhibition comparable to that of 5, it was hypothesized that the conformation of Z-15b would be equivalent to the active conformer of 5. The results revealed that the antibacterial Z-1-(2-fluorovinyl)quinolone derivatives carry the novel N-1 substituent of the fluoroquinolones.

Inhibition by fluoroquinolones of K(+) currents in rat dissociated hippocampal neurons.

The effects of four fluoroquinolones (sparfloxacin, fleroxacin, ofloxacin and levofloxacin) on K(+) currents were investigated in pyramidal neurons acutely isolated from rat hippocampus, to evaluate their relative potencies for inhibiting these channels. Using patch-clamp electrophysiological techniques, we found that all four compounds inhibited the delayed rectifier K(+) current (I(K)), but with different potencies. Sparfloxacin was the most potent compound, displaying an IC(50) value of 6.44 x 10(-4) M, followed by fleroxacin, ofloxacin and levofloxacin, their IC(50) values being 7.09 x 10(-3), 8.42 x 10(-3) and 1.10 x 10(-2) M, respectively. In contrast, the fast transient K(+) current (I(A)) was blocked only by sparfloxacin (IC(50)=2.86 x 10(-3) M) and fleroxacin (IC(50)=4.38 x 10(-3) M), but not by ofloxacin and levofloxacin even at concentrations up to 1 mM. The K(+) current inhibition was reversible after washout of the compounds. Further study is needed to clarify the possible involvement of this novel action in the adverse effects of fluoroquinolones in the central nervous system (CNS).

Protective effect of fleroxacin against the nephrotoxicity of isepamicin in rats.

The protective effect of fleroxacin on isepamicin-induced nephrotoxicity was investigated. Wistar rats were administered either fleroxacin 100 mg/kg orally, isepamicin 300 mg/kg subcutaneously, or fleroxacin and isepamicin in combination for 14 d. The animals given 300 mg/kg of isepamicin showed a significant increase in urine N-acetyl-beta-D-glucosaminidase (NAG) levels as compared with the control animals which received saline (p<0.01). However, the increase in NAG level was markedly less when isepamicin was administered in combination with fleroxacin (p<0.01). Fleroxacin alone had no effect on urine NAG activity. Serum creatinine and blood urea nitrogen (BUN) levels were significantly higher in animals treated with isepamicin alone than in the control animals (p<0.01) or animals receiving the isepamicin fleroxacin combination (p<0.01). Histopathologically, fleroxacin induced very few cellular alterations, but considerably reduced the manifestation of typical signs of isepamicin nephrotoxicity. This investigation demonstrates that fleroxacin protects animals against isepamicin-induced nephrotoxicity.

[In vitro antibacterial activity of prulifloxacin, a new oral fluoroquinolone].

We compared antibacterial activity of NM394, which is the active metabolite of a prodrug of new fluoroquinolone prulifloxacin (PUFX), against clinical isolates of bacteria with those of ciprofloxacin (CPFX), levofloxacin (LVFX), gatifloxacin (GFLX), tosufloxacin (TFLX) and fleroxacin (FLRX). 1. NM394 showed a broad-spectrum antibacterial activity against both Gram-positive and Gram-negative bacteria. 2. MIC80 of NM394 for methicillin-sensitive Staphylococcus aureus, Streptococcus pneumoniae and Enterococcus faecalis were 0.5 microgram/ml, 2 micrograms/ml and 4 micrograms/ml, respectively. MIC80 of NM394 for Escherichia coli, Klebsiella pneumoniae and Enterobacter cloacae was lower than 0.06 microgram/ml. MIC80 of NM394 for Serratia marcescens and Pseudomonas aeruginosa were 0.25 microgram/ml and 2 micrograms/ml, respectively. 3. Short-time bactericidal activity of NM394 against P. aeruginosa was stronger than those of CPFX, GFLX, LVFX and TFLX. 4. Short-time bactericidal activity of NM394 at Cmax concentration against 12 strains of P. aeruginosa was stronger than those of CPFX, LVFX, GFLX and TFLX.

Effects of fluoroquinolones on the locomotor activity in rats.

During therapy with fluoroquinolones adverse CNS reactions such as dizziness, light-headedness, insomnia or sleepiness are observed in up to 20% of patients. Using a device developed at our institute for the simultaneous registration of the activity of rats housed in single cages, we have investigated the effects of trovafloxacin, fleroxacin or ofloxacin on the locomotor activity of juvenile and adult rats (11 per group) after oral administration of 600 mg/kg for 5 consecutive days. The effects were most pronounced after fleroxacin, which induced a reduction in activity to 36 +/- 9% (mean +/- SD) of the values measured in juvenile rats before treatment and to 60 +/- 21% (mean +/- SD) in adult rats. HPLC analysis of the plasma concentrations in juvenile rats showed that the concentrations of trovafloxacin were considerably lower than those of the other fluoroquinolones that had been studied previously in our laboratory: the peak concentration of trovafloxacin was 14 +/- 2.9 mg/l (mean +/- SD) after a single dose of 600 mg/kg in juvenile rats. Overall, we showed that the locomotor activities of juvenile and adult rats were significantly depressed during treatment with fluoroquinolones. The effects were more pronounced in juveniles. Monitoring of the locomotor activity of rats is a suitable approach to study CNS effects of fluoroquinolones in animals, but pharmacokinetics have to be taken into account.

[Study on the DNA gyrA gene mutation with resistance to fluoroquinolones in Staphylococcus aureus isolated from patients].

This study was aimed at the mechanism of resistance to fluoroquinolones in Staphylococcus aureus isolated from patients in Chengdu. The relationship between the point mutations in the gyrA genes and the resistance of 63 strains (57 fluoroquinone-resistant strains and 6 wild types) isolated clinically in Chengdu were investigated by a combination of restriction fragment length polymorphism analysis. The results revealed that there are 67.27%-92.5% of the fluoroquinolone-resistant strains against norfloxacin, fleroxacin, tosufloxacin, cipofloxacin, ofloxacin and sparfloxacin had a Hinf I site mutation in the gyrA genes, and most of such strains with such mutation in the gyrA genes showed high-level resistance. These indicate that Hinf site mutation in gyrA genes is the mainly cause of the resistance of fluoroquinolone-resistant strains of Staphylococcus aureus in Chengdu region.

Effect of fleroxacin on mouse bladder carcinogenesis with N-butyl-N- (4-hydroxybutyl) nitrosamine.

We recently reported fleroxacin significantly affected cell proliferation in a dose-dependent manner in transitional cell carcinoma cell lines. In this study, we investigated the effect of fleroxacin on mouse bladder carcinogenesis with N-butyl-N-(4-hydroxybutyl) nitrosamine (BHBN). Five-week-old C57BL/6 female mice were divided into three groups. The forced oral administrations of fleroxacin (10 or 50 mg/kg/day) were done for 1 week, then the 0.05% BHBN was given for 8 or 12 weeks. Fleroxacin treatments were continued until sacrifice. The mice were sacrificed at 4 weeks or 8 weeks latent period after BHBN administration, and the histological changes in the bladder were examined. Our study suggested that fleroxacin tended to suppress the development of bladder carcinoma or the malignant changes induced by a shorter period of BHBN administration (8 weeks) in mice. However, in the group of the BHBN administration for 12 weeks, there were no significant effects on the prevention of bladder carcinoma in both of the treatment groups with fleroxacin (10 or 50 mg/kg/day). This study did not make it clear that oral administration of fleroxacin suppressed the development of bladder carcinoma induced by BHBN in mice. However, it is very preliminary data and further experiments need to be done with a much higher dose of the fleroxacin or with other kinds of fluoroquinolone antibiotics.

Mechanisms of action of quinolones against staphylococci and relationship with their in vitro bactericidal activity.

Ciprofloxacin and norfloxacin exhibited mechanism A (requires cell division as well as bacterial protein and RNA synthesis to kill bacteria) and C (active against nondividing bacteria but requires protein and RNA synthesis) against the reference strain Staphylococcus aureus ATCC 25923, yet only mechanism A was exhibited by these fluoroquinolones when tested against three clinical isolates: S. aureus Sa-215, Staphylococcus epidermidis Se-81 and Staphylococcus haemolyticus Sx-1. On the contrary, fleroxacin exerted mechanism A and C against the three clinical isolates but only mechanism A against the reference strain. Ofloxacin displayed mechanism A against S. epidermidis Se-81, mechanism A and C against S. haemolyticus and mechanism A and B (active against nondividing bacteria and does not require protein and RNA synthesis) against the two S. aureus tested. Sparfloxacin showed mechanism A and C against the four Staphylococcus species studied, and temafloxacin was the only fluoroquinolone tested that exhibited mechanism A and B against the four bacterial strains assayed. No correlation was found between the in vitro bactericidal activity (expressed as minimum inhibitory concentration and optimal bactericidal concentration) and the mechanisms of action exhibited by these fluoroquinolones.

[Influence of roxithromycin combined with fleroxacin on bacterial biofilm induced by Pseudomonas aeruginosa].

OBJECTIVE: To investigate the influence of roxithromycin (RXM) on the production of glycocalyx (GLX) and the synergic effect of RXM and fleroxacin (FLRX) on P. Aeruginosa biofilms. METHODS: Bacterial biofilm (BF) was established and was influenced by RXM and FLRX with different concentrations. The samples were detected with scanning electron microscope and by a rapid method after BF was stained with AgNO3 solution. The synergism of antibacterial activities of RXM and FLRX to P. Aeruginosa was studied by computer image analysis and MTT method. RESULTS: GLX production was reduced significantly by both 1/16 MIC and 1/4 MIC RXM (P < 0.01). 1/16 MIC and 1/4 MIC RXM showed no bactericidal activities to P. Aeruginosa in biofilms, while viable counts in biofilms almost had no difference. However, RXM could enhance the bactericidal activity of FLRX to P. Aeruginosa in biofilms. When FLRX of different concentrations was combined with 1/16 MIC and 1/4 MIC RXM respectively, the difference of the stained BF was counted up by computer image analysis system with significance of P < 0.05 and the viable counts were reduced significantly (P < 0.05). The results was in accordance with SEM pictures. CONCLUSION: RXM has no direct bactericidal activities to P. Aeruginosa in biofilms, but it could inhibit GLX production. It can be considered that RXM could enhance antibacterial activities of FLRX by enhancing the permeability of FLRX into biofilms.

[The serum bactericidal activity of domestic fleroxacin and lomefloxacin].

OBJECTIVE: To evaluate antibacterial activities in vivo and in vitro of fleroxacin and lomefloxacin. METHODS: Minimal inhibitory concentration (MIC) and serum bactericidal activity (SBA) domestic fleroxacin and lomefloxacin were determined by agar disk dilution method and microplate method respectively. A clinical observation of fleroxacin and lomefloxacin for treating acute bacillary dysentery was carried out. RESULTS: MIC(90) of fleroxacin against E. coli, K. pneumoniae, S. dysenteriae, S. aureus was 0.25-2 mg/L, being the same as that of ofloxacin. However, MIC(90) of lomefloxacin against the above mentioned species was twice or fourfold as that of ofloxacin and fleroxacin. The percent age of peak SBA > or = 1:8 with ofloxacin, fleroxacin and lomefloxacin against E. coli, K. pneumoniae, S. dysenteriae was 80%-100%, 90%-100% and 50%-80% respectively (P < 0.01). The clinical cure rates of acute bacillary dysentery were 93.3%, 100% and 100% and the bacterial clearance rates were 93.3%, 100% and 100% in the lomefloxacin, fleroxacin and ofloxacin group respectively. CONCLUSION: The antibacterial activity in vivo and in vitro of fleroxacin against E. coli, K. pneumoniae, S. dysenteriae was similar to that of ofloxacin and stronger than that of lomefloxacin.

Photocleavage of DNA by the fluoroquinolone antibacterials.

We have determined the relative efficiencies for the formation of single strand breaks (ssbs) after the UVA irradiation of pBR322 DNA and various fluoroquinolone (fleroxacin, lomefloxacin, norfloxacin) and naphthyridine (nalidixic acid, enoxacin) antibacterials. After correcting for the differences in absorption, the relative order for DNA photocleaving activity under anaerobic conditions is: fleroxacin, lomefloxacin > nalidixic acid >> norfloxacin > enoxacin. In general, fluoroquinolones having fluorine substituents at the C-6 and C-8 positions (lomefloxacin and fleroxacin) are 10-fold more efficient in generating ssbs than those having only a C-6 fluorine atom (norfloxacin). The effect of oxygen on photoinduced DNA damage caused by these antibacterials is complex, but our data imply that active oxygen species are not necessary for DNA scission by these molecules, and indeed, may sometimes inhibit it. Lomefloxacin ethyl ester, which cannot undergo decarboxylation, is as active as lomefloxacin itself. Thus the free radical generated by decarboxylation is unlikely to be the active species involved in photoinduced fluoroquinolone DNA cleavage. For lomefloxacin and fleroxacin, DNA damage probably results from the generation of a carbene at C-8 as a result of photoinduced loss of their F8 atom as fluoride upon UVA irradiation. Fluoroquinolones lacking a C-8 fluorine atom must operate by a different mechanism. While photocleavage of pBR322 DNA does not necessarily mean that duplex DNA will be cleaved under the same conditions, nevertheless lomefloxacin and fleroxacin, the two most photogenotoxic fluoroquinolones, did cause the most damage to the plasmid DNA.

In vitro activity of fleroxacin against multiresistant gram-negative bacilli isolated from patients with nosocomial infections.

In order to evaluate the in vitro activity of fleroxacin against nosocomial gram-negative organisms, 263 multiresistant gram-negative bacilli (203 Enterobacteriaceae and 60 non-fermenting gram-negative bacilli) were isolated from adult patients with nosocomial infections. The different patterns of resistance to eight different antimicrobial agents (ampicillin, carbenicillin, piperacillin, cephalothin, cefamandole, ceftazidime, gentamicin and amikacin) were determined by minimum inhibitory concentration (MIC), using the agar dilution method. The most prevalent multiresistant species isolated were Klebsiella pneumoniae (28.9%), Escherichia coli (24%) and Pseudomonas aeruginosa (12.2%). All these bacterial strains showed three to five resistance patterns to at least three different antibiotics. Resistance to ceftazidime was observed in at least one of the resistance patterns of isolated bacteria. The activity of fleroxacin against multiresistant enteric bacteria was excellent; these strains showed a susceptibility of 79-100%. The susceptibility of P. aeruginosa to antipseudomonal agents was low; however, the activity of fleroxacin against these strains was higher than 60% (MIC < or = 2 microg/ ml), broadly comparable with ciprofloxacin. The resistance to fluoroquinolones detected in this study was no cause for alarm (3%). Consequently, fleroxacin maintains a remarkable activity against Enterobacteriaceae and remains highly active against other gram-negative bacilli. Nevertheless, actions directed at preventing or limiting resistance will be crucial to maintain the viability of fluoroquinolones as important therapeutic agents.

Effects of fluoroquinolones and magnesium deficiency in murine limb bud cultures.

Quinolone-induced arthropathy is probably caused by a lack of functionally available magnesium in immature joint cartilage. We used an in vitro assay to study the effects of fluoroquinolones on cartilage formation in mouse limb buds from 12-day-old mouse embryos in regular and in magnesium-deficient medium. Omission of magnesium from the medium had no adverse effect on the outcome of the culture: limb buds grew and differentiated well in regular and in magnesium-deficient Bigger's medium. Lack of calcium, however, severely impaired the development of the explants; this result was even more enhanced when both minerals (magnesium and calcium) were omitted. Electron microscopy revealed cell necrosis and deposition of electron-dense material in the vicinity of chondrocytes from limb buds after 6 days in a magnesium-free medium. A series of seven fluoroquinolones was tested at 30, 60, and 100 mg/l medium. At a concentration of 30 mg/l sparfloxacin only had a slight effect on limb development. At concentrations of 60 and 100 mg/l sparfloxacin, temafloxacin and ciprofloxacin impaired limb development in vitro concentration-dependently. The effects were enhanced in a magnesium-deficient medium (concentration of magnesium <10 micromol/l). Fleroxacin, lomefloxacin and ofloxacin impaired limb development only slightly; no significant differences were recognizable between the outcome in regular and in magnesium-deficient medium. Pefloxacin did not show any effect on limb development in both media. Using electron microscopy, very similar alterations as described above for the limbs cultured in magnesium-deficient medium were observed with ofloxacin at a concentration of 30 mg/l, which had no effect on the growth of the explants when evaluated macroscopically. The affinity of six fluoroquinolones to magnesium was determined by the use of a fluorescence assay. The affinity to magnesium correlated with the activity of the drugs in the limb bud assay. We conclude that fluoroquinolones have no effect on murine limb development in vitro at concentrations that are achieved under therapeutic conditions (peak concentrations approx. 1-5 mg/l in plasma). Effects at higher concentrations (60 and 100 mg/l) are slightly enhanced (factor 2) if the magnesium concentration in the medium is low. Macroscopically, limbs develop regularly in a magnesium-free medium, but ultrastructurally typical alterations are exhibited (e.g. cell necrosis and pericellular deposition of electron-dense material).

Urinary excretion and bactericidal activities of a single oral dose of 400 milligrams of fleroxacin versus a single oral dose of 800 milligrams of pefloxacin in healthy volunteers.

Twelve healthy volunteers participated in this randomized crossover study to compare the concentrations and recovery levels of fleroxacin and pefloxacin in urine and to assess their bactericidal activities against 12 strains of urinary pathogens with different susceptibilities over a wide range of MICs. The volunteers received a single oral dose of 400 mg of fleroxacin or 800 mg of pefloxacin. The mean cumulative renal excretion of unchanged fleroxacin, N-demethyl-fleroxacin, and N-oxide-fleroxacin accounted for 67, 7, and 6% of the total dose, respectively. The total urinary recovery of pefloxacin and the active metabolite norfloxacin was 34%. In the time-kill and the urinary bactericidal titer (UBT) studies, only the subjects' urine not supplemented with broth was used. With most tested organisms and both quinolones it took more than 8 h to achieve a reduction in CFU of 99.9% (3 log units). Overall, there was a good correlation between UBTs and MICs for the strains. Against Escherichia coli ATCC 25922 the median UBTs were similar for both antibiotics and at least 1:8 for 96 h; against the E. coli strain for which the MIC was 0.5 microgram/ml the UBT was at least 1:4 for 48 h. The UBTs of both drugs against Klebsiella pneumoniae were at least 1:16 for 72 h. The UBTs for Staphylococcus aureus (the MIC for which was 16 micrograms/ml) of both antibiotics were low, and in some of the samples, no bactericidal titers were observed. UBTs for Proteus mirabilis of pefloxacin are significantly higher than those of fleroxacin. For Pseudomonas aeruginosa the median UBTs were present for the 24-to-48-h interval. The same is true for Enterococcus faecalis. Against Staphylococcus saprophyticus, UBTs were present for at least 48 h with both quinolones. Overall, a single oral dose of 400 mg of fleroxacin exhibits UBTs comparable to those of 800 mg of pefloxacin. Therefore, it may be expected that half of the dose of fleroxacin gives comparable results in the treatment of urinary tract infections; this should be substantiated in comparative clinical trials.

In vitro activity of enoxacin versus ciprofloxacin, fleroxacin, lomefloxacin, ofloxacin, pefloxacin, and rufloxacin against uropathogens.

Minimum inhibitory concentrations (MIC) of enoxacin, ciprofloxacin, fleroxacin, lomefloxacin, ofloxacin, pefloxacin and rufloxacin were determined against 400 uropathogens cultured from the urine of patients with complicated and/or hospital-acquired urinary tract infections (UTI) using an agar dilution method. The bacterial spectrum consisted of Entero-bacteriaceae (34.5%), enterococci (31.5%), staphylococci (21.2%) and non-fermenting bacteria (12.8%). Enoxacin inhibited all but one strain (Enterobacter cloacae) of Enterobacteriaceae up to an MIC of 1 mg/l (MIC90 0.25 mg/l). Regarding the total bacterial spectrum, enoxacin inhibited 54.5, 59.5, 76.0 and 83.8% up to an MIC of 1, 2, 4 and 8 mg/l, respectively. If the same breakpoint of resistance for ofloxacin according to DIN 58,940 (NCCLS), i.e. MIC > or = 4 mg/l (> or = 8 mg/l), is also taken for the other fluoroquinolones, and the 126 strains of enterococci are excluded, for which alternative agents, e.g. aminopenicillins, should be considered instead, the following resistance rates were found: ciprofloxacin and enoxacin 15.3% (15.0%), ofloxacin 17.2% (15.3%), pefloxacin 18.2% (15.3%), fleroxacin 19.3% (15.3%), lomefloxacin 19.7% (17.9%) and rufloxacin 31.8% (27.4%). According to their in vitro activity, all fluoroquinolones tested besides rufloxacin show similar rates of resistance against uropathogens and can therefore be considered good alternative agents for the treatment of complicated UTI.

[Synthesis and antibacterial activity of 7-substituted-1-ethyl(2-fluoro-ethyl)-6,8-difluoro-1,4-dihydro-4- oxoquinoline-3-carboxylic acid].

Eighteen 7-substituted-1-ethyl (2-fluoroethyl)-6, 8-difluoro-1, 4-dihydro-4-oxoquinolone-3-carboxylic acid were designed and synthesized. The results of preliminary antibacterial test showed that most of the compounds exhibited definite activities in test against pathogenic bacteria. Especially, compound 17 and 18 showed better activities and surpassed fleroxacin when compared in vitro.

[Inhibitory activity of azithromycin on biofilm synthesis and synergism between azithromycin and fleroxacin on Pseudomonas aeruginosa in biofilms].

OBJECTIVE: The influence of azithromycin (AZT) on the production of glycocalyx (GLX) from P. aeruginosa biofilms and synergism of antibacterial activities between AZT and fleroxacin (FLX) on P. aeruginosa were investigated. METHOD: GLX production was measured by using a L-tryptophan method and viable counts in biofilms was determined by using a methylthiazolyldiphenyltetrazolium(MTT) method. RESULT: GLX production was reduced significantly from 28.0 +/- 8.1 micrograms/ml at 0MIC to 21.8 +/- 8.2 micrograms/ml at 1/16MIC and 16.7 +/- 7.9 micrograms/ml at 1/4MIC respectively (P = 0.0002). 1/16MIC and 1/4MIC AZT showed no bactericidal activities to P. Aeruginosa in biofilms. Viable counts in biofilms were 6.2 +/- 0.61 Lg/cm2 at 0MIC, 6.1 +/- 0.4 Lg/cm2 at 1/16MIC and 6.2 +/- 0.4 Lg CFU/cm2 at 1/4 MIC respectively (P = 0.63). However, AZT could enhance bactericidal activity of FLX on P. aeruginosa in biofilms. When 1/4MIC FLX was combined with 1/16MIC and 1/4 MIC AZT, viable counts were reduced significantly from 6.1 +/- 0.5 Lg CFU/cm2 to 5.9 +/- 0.3 Lg CFU/cm2 and 5.8 +/- 0.4 Lg CFU/cm2 respectively (P = 0.02). When 1/2MIC FLX was combined with 1/16MIC and 1/4MICAZT, viable counts were reduced significantly from 6.3 +/- 0.7 Lg CFU/cm2 to 5.8 +/- 0.5 LgCFU/cm2 and 5.7 +/- 0.6 Lg CFU/cm2 respectively (P = 0.03). CONCLUSION: Because AZT did not have direct bactericidal activities on P. aeruginosa in biofilms but could inhibit GLX production, We considered that AZT could enhance antibacterial activities of FLX by enhancing permeability of FLX into biofilms.