Metabolic and transcriptional activities underlie stationary-phase Pseudomonas aeruginosa sensitivity to Levofloxacin.

IMPORTANCE: The bacterial pathogen Pseudomonas aeruginosa is responsible for a variety of chronic human infections. Even in the absence of identifiable resistance mutations, this pathogen can tolerate lethal antibiotic doses through phenotypic strategies like biofilm formation and metabolic quiescence. In this study, we determined that P. aeruginosa maintains greater metabolic activity in the stationary phase compared to the model organism, Escherichia coli, which has traditionally been used to study fluoroquinolone antibiotic tolerance. We demonstrate that hallmarks of E. coli fluoroquinolone tolerance are not conserved in P. aeruginosa, including the timing of cell death and necessity of the SOS DNA damage response for survival. The heightened sensitivity of stationary-phase P. aeruginosa to fluoroquinolones is attributed to maintained transcriptional and reductase activity. Our data suggest that perturbations that suppress transcription and respiration in P. aeruginosa may actually protect the pathogen against this important class of antibiotics.

Study on the Mechanism of Levofloxacin Combined with Imipenem Against Pseudomonas aeruginosa.

Pseudomonas aeruginosa can develop resistance. Therefore, it is necessary to design proper treatment for it. Pseudomonas aeruginosa can develop resistance against levofloxacin due to the development of efflux pumps. However, the development of these efflux pumps cannot develop resistance against imipenem. Additionally, the MexCDOprJ efflux system which is responsible for the resistance of Pseudomonas aeruginosa to levofloxacin is highly susceptible to imipenem. The objective of the study was to evaluate the emergence of resistance of Pseudomonas aeruginosa against 750 mg levofloxacin, 250 mg imipenem, and a combination of 750 mg levofloxacin and 250 mg imipenem. An in vitro pharmacodynamic model was selected for the evaluation of the emergence of resistance. Pseudomonas aeruginosa strain 236, Pseudomonas aeruginosa strain GB2, and Pseudomonas aeruginosa strain GB65 were selected. Susceptibility testing of both antibiotics was done by agar dilution methodology. A disk diffusion bioassay was performed for antibiotics. RT-PCR measurement was done for the evaluation of expressions of Pseudomonas aeruginosa genes. Samples were tested at 2 h, 4 h, 6 h, 8 h, 12 h, 16 h, 24 h, and 30 h. Levofloxacin and imipenem both individually reported a decrease in colony-forming unit per milliliter of strength in the initial stage but in the later stage both develop resistance individually. Levofloxacin with imipenem had no resistance to Pseudomonas aeruginosa during 30 h. Time after the start of development of resistance or decrease in clinical efficacy was higher for levofloxacin and imipenem combination in all strains. The concentration of Pseudomonas aeruginosa at the time after the start of development of resistance or decrease in clinical efficacy was fewer for levofloxacin and imipenem combination. Levofloxacin with imipenem is recommended for the treatment of infection due to Pseudomonas aeruginosa.

Nephroprotective effect of vitamin D Against Levofloxacin-induced renal injury: an observational study.

The pathogenesis of kidney damage involves complicated interactions between vascular endothelial and tubular cell destruction. Evidence has shown that vitamin D may have anti-inflammatory effects in several models of kidney damage. In this study, we evaluated the effects of synthetic vitamin D on levofloxacin-induced renal injury in rats. Forty-two white Albino rats were divided into six groups, with each group comprising seven rats. Group I served as the control (negative control) and received intraperitoneal injections of normal saline (0.5 ml) once daily for twenty-one days. Group II and Group III were treated with a single intraperitoneal dose of Levofloxacin (50 mg/kg/day) and (100 mg/kg/day), respectively, for 14 days (positive control groups). Group IV served as an additional negative control and received oral administration of vitamin D3 (500 IU/rat/day) for twenty-one days. In Group V, rats were orally administered vitamin D3 (500 IU/rat/day) for twenty-one days, and intraperitoneal injections of Levofloxacin (50 mg/kg/day) were administered on day 8 for 14 days. Group VI received oral vitamin D3 supplementation (500 IU/rat/day) for twenty-one days, followed by intraperitoneal injections of Levofloxacin (100 mg/kg/day) on day 8 for fourteen days. Blood samples were collected to measure creatinine, urea, malondialdehyde, glutathione reductase, and superoxide dismutase levels. Compared to the positive control group, vitamin D supplementation lowered creatinine, urea, and malondialdehyde levels, while increasing glutathione reductase and superoxide dismutase levels. Urea, creatinine, and malondialdehyde levels were significantly (p<0.05) higher in rats administered LFX 50mg and 100mg compared to rats given (LFX + vitamin D). The main findings of this study show that vitamin D reduces renal dysfunction, suggesting that vitamin D has antioxidant properties and may be used to prevent renal injury.

Study effect and mechanism of ofloxacin and levofloxacin on development of Rana nigromaculata tadpoles based on the hypothalamus-pituitary-thyroid axis.

Excessive antibiotics transferred into the aquatic environment may affect the development of amphibians. Previous studies on the aquatic ecological risk of ofloxacin generally ignored its enantiomers. The purpose of this study was to compare the effects and mechanisms of ofloxacin (OFL) and levofloxacin (LEV) on the early development of Rana nigromaculata. After 28-day exposure at environmental levels, we found that LEV exerted more severe inhibitory effects on the development of tadpoles than OFL. According to the enrichment results of differentially expressed genes in the LEV and OFL treatments, LEV and OFL had different effects on the thyroid development of tadpoles. dio2 and trh were affected by the regulation of dexofloxacin instead of LEV. At the protein level, LEV was the main component that affected thyroid development-related protein, while dexofloxacin in OFL had little effect on thyroid development. Furthermore, molecular docking results further confirmed that LEV was a major component affecting thyroid development-related proteins, including DIO and TSH. In summary, OFL and LEV regulated the thyroid axis by differential binding to DIO and TSH proteins, thereby exerting differential effects on the thyroid development of tadpoles. Our research is of great significance for comprehensive assessment of chiral antibiotics aquatic ecological risk.

The Putative Major Facilitator Superfamily (MFS) Protein Named Rv1877 in Mycobacterium tuberculosis Behaves as a Multidrug Efflux Pump.

Efflux pumps are one of the major contributors in the intrinsic multidrug resistance of Mycobacterium tuberculosis. These active transporters, localized in the cytoplasmic membrane, often carry an array of unrelated substances, from toxic substances to metabolites and maintain cellular homeostasis. Rv1877, a putative Major Facilitator Superfamily efflux pump from M. tuberculosis, was investigated in this study. Expression of Rv1877 in Escherichia coli resulted in elevated resistance towards antibiotics of various families. A reversal of this resistance was observed in the presence of sub-inhibitory concentration of the uncoupler carbonyl cyanide-m-chlorophenylhydrazone, indicating its dependence on proton motive force (pmf). Lower intracellular accumulation of the fluoroquinolones ofloxacin and levofloxacin in E. coli cells harbouring Rv1877 implied an active efflux of the drugs. Interestingly, real time, energy-dependent efflux was demonstrated by cells expressing Rv1877 with a lipophilic dye Nile Red. In addition, expression of Rv1877 in trans increased the biofilm formation by the host E. coli cells. Moreover, in silico docking analysis of the molecular interactions between Rv1877 and antibiotics corroborated the experimental observations. Based on the in vivo analyses of Rv1877 in E. coli, it could be designated as a pmf-dependent multidrug transporter with the ability of extruding structurally unrelated antibiotics, preferably some of the fluoroquinolones, and a facilitator of biofilm formation.

Subinhibitory Cefotaxime and Levofloxacin Concentrations Contribute to Selection of Pseudomonas aeruginosa in Coculture with Staphylococcus aureus.

Bacterial species in the polymicrobial community evolve interspecific interaction relationships to adapt to the survival stresses imposed by neighbors or environmental cues. Pseudomonas aeruginosa and Staphylococcus aureus are two common bacterial pathogens frequently coisolated from patients with burns and respiratory disease. Whether the application of commonly used antibiotics influences the interaction dynamics of the two species still remains largely unexplored. By performing a series of on-plate competition assays and RNA sequencing-based transcriptional profiling, we showed that the presence of the cephalosporin antibiotic cefotaxime or the quinolone antibiotic levofloxacin at subinhibitory concentration contributes to selecting P. aeruginosa from the coculture with S. aureus by modulating the quorum-sensing (QS) system of P. aeruginosa. Specifically, a subinhibitory concentration of cefotaxime promotes the growth suppression of S. aureus by P. aeruginosa in coculture. This process may be related to the increased production of the antistaphylococcal molecule pyocyanin and the expression of lasR, which is the central regulatory gene of the P. aeruginosa QS hierarchy. On the other hand, subinhibitory concentrations of levofloxacin decrease the competitive advantage of P. aeruginosa over S. aureus by inhibiting the growth and the las QS system of P. aeruginosa. However, pqs signaling of P. aeruginosa can be activated instead to overcome S. aureus. Therefore, this study contributes to understanding the interaction dynamics of P. aeruginosa and S. aureus during antibiotic treatment and provides an important basis for studying the pathogenesis of polymicrobial infections. IMPORTANCE Increasing evidence has demonstrated the polymicrobial characteristics of most chronic infections, and the frequent communications among bacterial pathogens result in many difficulties for clinical therapy. Exploring bacterial interspecific interaction during antibiotic treatment is an emerging endeavor that may facilitate the understanding of polymicrobial infections and the optimization of clinical therapies. Here, we investigated the interaction of cocultured P. aeruginosa and S. aureus with the intervention of commonly used antibiotics in clinic. We found that the application of subinhibitory concentrations of cefotaxime and levofloxacin can select P. aeruginosa in coculture with S. aureus by modulating P. aeruginosa QS regulation to enhance the production of antistaphylococcal metabolites in different ways. This study emphasizes the role of the QS system in the interaction of P. aeruginosa with other bacterial species and provides an explanation for the persistence and enrichment of P. aeruginosa in patients after antibiotic treatment and a reference for further clinical therapy.

New promising levofloxacin derivatives: Design, synthesis, cytotoxic activity screening, Topo2ß polymerase inhibition assay, cell cycle apoptosis profile analysis.

Newly designed levofloxacin analogues were synthesized to act as topoisomerase II beta inhibitors (Topo2ß). Their cytotoxic activity was screened against breast, liver, and leukemia cancer cell lines. The best activity against liver cancer cell line (Hep3B) was exhibited by the target compounds 3c, 3e, 4a, and 6d (IC50 = 2.33, 1.38, 0.60 and 0.43, respectively). (L-SR) leukemia cancer cell line was pronouncedly affected by compounds 3b, 3g and 4a (IC50 = 1.62, 1.41 and 1.61, sequentially). 3c possessed the best activity against breast cancer cell line (MCF-7) with IC50 = 0.66. Compounds 3c, 3e, 3g, 4a and 4c exhibited Topo2ß inhibition activities exceeding etoposide and levofloxacin as reference drugs and variant cell lines. In DNA-Flow cytometry cell cycle analysis, compound 3c arrested the cell cycle at G2/M phase like etoposide and levofloxacin, while compounds 3e and 4a exhibit its arrest at S phase. In addition, 3c, 3e and 4a showed a significant elevation in active caspase-3 levels (10.01, 8.98 and 10.71 folds, respectively). The effect of the new compounds on normal cells was also investigated including breast (MCF10a), liver (THLE2), and lymphocytic (PCS-800-011) normal cell lines.

Comparing the interaction of the antibiotic levofloxacin with zwitterionic and anionic membranes: Calorimetry, fluorescence, and spin label studies.

The present work compares the interaction of the antibiotic levofloxacin (LVX) with zwitterionic and anionic liposomes composed of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1,2-dipalmitoyl-sn-glycero-3-phospho-(1'-rac-glycerol) (DPPG), respectively. By using differential scanning calorimetry (DSC), and with spin labels incorporated into liposomes at two different depths of the bilayers, we investigated the changes induced on the membrane by increasing concentrations of LVX. Further information was obtained using intrinsic LVX fluorescence. Under the conditions used here, all techniques evinced that LVX has little affinity for DPPC zwitterionic membrane. Opposite to that, LVX exhibits a considerable affinity for anionic bilayers, with membrane partition constants Kp = (3.3 ± 0.5) × 102 and (4.5 ± 0.3) × 102, for gel and fluid DPPG membranes, respectively. On binding to DPPG, LVX seems to give rise to the coexistence of LVX -rich and -poor domains on DPPG membranes, as detected by DSC. At the highest LVX concentration used (20 mol%), DSC trace shows an increase in the cooperativity of DPPG gel-fluid transition, also detected by spin labels as an increase in the bilayer packing. Moreover, LVX does not induce pore formation in either DPPG or POPG vesicles. Considering the possible relevance of LVX-membrane interaction for the biological and toxicological action of the antibiotic, the findings discussed here certainly contribute to a better understanding of its action, and the planning of new drugs.

The Treatment of Tuberculosis.

Tuberculosis (TB) remains a leading cause of infectious death worldwide, and poverty is a major driver. Clinically, TB presents as "latent" TB and active TB disease, and the treatment for each is different. TB drugs can display "early bactericidal activity (EBA)" and / or "sterilizing activity" (clearing persisters). Isoniazid is excellent at the former, and rifampin is excellent at the latter. Pyrazinamide and ethambutol complete the first-line regimen for drug-susceptible TB, each playing a specific role. Drug-resistant TB is an increasing concern, being met, in part, with repurposed drugs (including moxifloxacin, levofloxacin, linezolid, clofazimine, and beta-lactams) and new drugs (including bedaquiline, pretomanid, and delamanid). One challenge is to select drugs without overlapping adverse drug reaction profiles. QTc interval prolongation is one such concern, but to date, it has been manageable. Drug penetration into organism sanctuaries, such as the central nervous system, bone, and pulmonary TB cavities remain important challenges. The pharmacodynamics of most TB drugs can be described by the area under the curve (AUC) divided by the minimal inhibitory concentration (MIC). The hollow fiber infection model (HFIM) and various animal models (especially mouse and macaque) allow for sophisticated pharmacokinetic/pharmacodynamic experiments. These experiments may hasten the selection of the most potent, shortest possible regimens to treat even extremely drug resistant TB. These findings can be translated to humans by optimizing drug exposure in each patient, using therapeutic drug monitoring and dose individualization.

Multiparameter prediction control of in vitro drug delivery into mycobacterium smegmatis induced by microbubble-enhanced sonoporation.

The antibacterial method induced by microbubble-enhanced sonoporation has shown its great potential in facilitating drug delivery into thallus. The enhanced drug delivery induced by microbubble-enhanced sonoporation is a complex event which can be affected by various physical parameters. How to determine the correlation between experimental parameters and the drug delivery efficiency to give the instruction on reasonably choosing the parameters and achieve the control of drug delivery efficiency is impeding further investigations for this complex biophysical process. In the present work, we have explored a number of key parameters affecting the drug delivery efficiency induced by microbubble-enhanced sonoporation using multivariate biological experiments. To achieve the control of the drug delivery efficiency, a multiparameter prediction control method based on modified artificial neural network is presented in this paper. This method is a new modeling method based on combined back-propagation neural network and the multiple model idea to establish quantitative relationship between experimental parameters and drug delivery efficiency. By analyzing the experimental samples, a mapping relationship expression can be deduced to determine the input and output variables of artificial neural network models. Experimental samples were divided into training and test samples. We trained models based on back-propagation neural network to establish their quantitative relationship. In this model, the multiple model idea was introduced into the selection of training samples to modify the traditional back-propagation neural network model to avoid model mismatch caused by poor training sample selection. Numerical experiments results have shown that compared with the traditional model, the identification results obtained by modified model are more closed to experimental results. It is elucidated that an appropriately trained network can act as a good alternative for costly and time-consuming experiments. The findings of this study indicate that this approach can realize the prediction of drug delivery efficiency induced by microbubble-enhanced sonoporation under different experimental parameters, and then achieve the control of drug delivery efficiency through reasonable parameter selection, finally achieve the purpose of efficiently killing bacteria.

Brain uptake and accumulation of new levofloxacin-doxycycline combination through the use of solid lipid nanoparticles: Formulation; Optimization and in-vivo evaluation.

The objective of this study is to investigate the feasibility of delivery of novel levofloxacin/ doxycycline (LEVO/DOX) combination to the brain by intranasal route to achieve a significant local concentration in the brain and a direct nose-to-brain pathway. Solid lipid nanoparticles (SLN) were selected as a drug carrier and employed Box-Behnken design for optimizing LEVO/DOX-SLN to achieve minimum particle size and maximum apparent entrapment efficiency (EE). SLNs were prepared by hot emulsification and characterized. In vitro release of optimized formulations showed prolonged drug release from the optimized formulation. The results of pharmacokinetic study of the optimized SLN-HPMC gel in plasma and brain revealed significant increase in the brain peak concentration (420, 315 ng/g), the AUC 0-360 min (57130 and 48693.13 ng. min/g) in comparison to intranasal LEVO/DOX free solution with the values of (160, 120) ng/g, (36850, 27637.5 ng⋅min/g) for LEVO and DOX, respectively. The optimized LD-SLN-HPMC gel gave a drug-targeting efficiency (DTE %) of 149.815 and 161.969 for LEVO and DOX, respectively, in comparison to the intravenous route. Moreover, the optimized formulation had a direct transport percentage (DTP %) of 33.285 and 40.236 for LEVO and DOX, respectively, which indicates a significant contribution of direct nose-to-brain pathway in brain drug delivery.

Thermosensitive glycol chitosan-based hydrogel as a topical ocular drug delivery system for enhanced ocular bioavailability.

In the present study, we developed and evaluated an in situ gelling system based on hexanoyl glycol chitosan (H-GCS) for enhanced ocular bioavailability. An aqueous solution of H-GCS exhibited a typical sol-gel transition at 32 °C. The formed H-GCS hydrogel was characterized by rheology and scanning electron microscopy (SEM). H-GCS had minimal in vitro cytotoxicity against L-929 and HCEC cells over a concentration range of 0-0.8 mg/mL. Additionally, the H-GCS hydrogel exhibited good ocular tolerance and biocompatibility after a single instillation. Moreover, H-GCS hydrogel significantly prolonged the precorneal retention of fluorescein sodium compared with its aqueous solution. An in vivo pharmacokinetic study demonstrated that the levofloxacin-loaded H-GCS hydrogel could provide a significantly higher Cmax and AUC0-12h compared with the levofloxacin aqueous solution, thus increasing ocular bioavailability. Overall, the proposed H-GCS hydrogel acts as an in situ gelling system that might represent a promising vehicle for topical ocular drug delivery.

New insights into resistance of Helicobacter pylori against third- and fourth-generation fluoroquinolones: A molecular docking study of prevalent GyrA mutations.

BACKGROUND: Fluoroquinolones hinder bacterial DNA replication by inhibiting DNA gyrase. However, mutations, in the QRDR segment of its A subunit (GyrA), cause antibiotic resistance. Here, the interactions of levofloxacin (LVX), gemifloxacin (GXN), and moxifloxacin (MXN) with Helicobacter pylori GyrA, in LVX-resistant vs -sensitive strains, were studied. METHODS: Levoflixacin-sensitive (n = 4) and -resistant (n = 9) H pylori strains, randomly selected from another antibiotic susceptibility study, underwent PCR amplification of gyrA gene, spanning the QRDR segment. The amplified gene fragments were sequenced and aligned. The homology model of H pylori GyrA was built based on that of Escherichia coli, and energy minimization was done. The interaction patterns of LVX, GXN, and MXN with GyrA were analyzed via molecular docking studies. RESULTS: Sequence alignment of the 13 studied strains, created 5 categories of strains: (A) wild type-like (H pylori ATCC26695), (B) N87K-only, (C) D91N-only, (D) N87K + V94L, and (E) D91N + A97V mutations. The minimum inhibitory concentrations (MIC) for LVX-sensitive (category A) and -resistant (categories B-E) strains were <1 mg/L and ≥32 mg/L, respectively. The binding mode of GyrA in category A with LVX identified G35/N87/Y90/D91/V94/G114/S115/I116/D117/G118/D119, as key residues, some residing outside the QRDR segment. Category B strains lost only one interaction (G35), which led to elevated binding free energy (∆G) and full LVX resistance. Categories C-E lost more contacts, with higher ∆G and again full LVX resistance. GXN bound to GyrA of categories A and B via a different set of key residues, while MXN retained the lost contact (G35) in LVX-resistant, category B strains. CONCLUSION: Using molecular docking tools, we identified the key residues responsible for interaction of GyrA with LVX, GXN, and MXN. In the presence of N87K-only mutation, the loss of one of these contacts (ie, G35) led to full LVX resistance. Yet, GXN and MXN overcame this mutation, by retaining all key contacts with GyrA.

Influence of levofloxacin and clarithromycin on the structure of DPPC monolayers.

Research on lipid/drug interactions at the nanoscale underpins the emergence of synergistic mechanisms for topical drug administration. The structural understanding of bio-mimetic systems employing 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) as a lung surfactant model mixed with antibiotics, as well as their biophysical properties, is of critical importance to modulate the effectiveness of therapeutic agents released directly to the airways. In this paper, we investigate the structural details of the interaction between Levofloxacin, 'a respiratory quinolone', and the macrolide Clarithromycin, with DPPC monolayers at the air-water interface, using a combination of Brewster angle microscopy, polarization modulation-infrared reflection-adsorption spectroscopy (PM-IRRAS), surface pressure isotherms and neutron reflectometry (NR) to describe the structural details of this interaction. The results allowed association of changes in the π-A isotherm profile with changes in the molecular organization and the co-localization of the antibiotics within the lipid monolayer by NR measurements. Overall, both antibiotics are able to increase the thickness of the acyl tails in DPPC monolayers with a corresponding reduction in tail tilt as well as to interact with the phospholipid headgroups as shown by PM-IRRAS experiments. The effects on the DPPC monolayers are correlated with the physical-chemical properties of each antibiotic and dependent on its concentration.

Evaluation of Saliva as a Potential Alternative Sampling Matrix for Therapeutic Drug Monitoring of Levofloxacin in Patients with Multidrug-Resistant Tuberculosis.

Saliva may be a useful alternative matrix for monitoring levofloxacin concentrations in multidrug-resistant tuberculosis (MDR-TB) patients. The objectives of this study were (i) to evaluate the correlation between plasma and salivary levofloxacin (Lfx) concentrations in MDR-TB patients and (ii) to gauge the possibility of using saliva as an alternative sampling matrix for therapeutic drug monitoring of Lfx in areas where TB is endemic. This was a prospective pharmacokinetic study that enrolled MDR-TB patients receiving levofloxacin (750- to 1,000-mg once-daily dosing) under standardized treatment regimen in Nepal. Paired blood and saliva samples were collected at steady state. Lfx concentrations were quantified using liquid chromatography-tandem mass spectrometry. Pharmacokinetic parameters were calculated using noncompartmental kinetics. Lfx drug exposures were evaluated in 23 MDR-TB patients. During the first month, the median (interquartile range [IQR]) areas under the concentration-time curve from 0 to 24 h (AUC0-24) were 67.09 (53.93 to 98.37) mg ⋅ h/liter in saliva and 99.91 (76.80 to 129.70) mg ⋅ h/liter in plasma, and the saliva plasma (S/P) ratio was 0.69 (0.53 to 0.99). Similarly, during the second month, the median (IQR) AUC0-24 were 75.63 (61.45 to 125.5) mg ⋅ h/liter in saliva and 102.7 (84.46 to 131.9) mg ⋅ h/liter in plasma, with an S/P ratio of 0.73 (0.66 to 1.18). Furthermore, large inter- and intraindividual variabilities in Lfx concentrations were observed. This study could not demonstrate a strong correlation between plasma and saliva Lfx levels. Despite a good Lfx penetration in saliva, the variability in individual saliva-to-plasma ratios limits the use of saliva as a valid substitute for plasma. Nevertheless, saliva could be useful in semiquantitatively predicting Lfx plasma levels. (This study has been registered at ClinicalTrials.gov under identifier NCT03000517.).

Enantioselective degradation of ofloxacin and levofloxacin by the bacterial strains Labrys portucalensis F11 and Rhodococcus sp. FP1.

Fluoroquinolones are a class of antibiotics widely prescribed in both human and veterinary medicine of high environmental concern and characterized as environmental micropollutants due to their ecotoxicity and persistence and antibacterial resistance potential. Ofloxacin and levofloxacin are chiral fluoroquinolones commercialized as racemate and in enantiomerically pure form, respectively. Since the pharmacological properties and toxicity of the enantiomers may be very different, understanding the stereochemistry of these compounds should be a priority in environmental monitoring. This work presents the biodegradation of racemic ofloxacin and its (S)-enantiomer levofloxacin by the bacterial strains Labrys portucalensis F11 and Rhodococcus sp. FP1 at a laboratory-scale microcosm following the removal and the behavior of the enantiomers. Strain F11 could degrade both antibiotics almost completely when acetate was supplied regularly to the cultures. Enrichment of the (R)-enantiomer was observed in FP1 and F11 cultures supplied with ofloxacin. Racemization was observed in the biodegradation of the pure (S)-ofloxacin (levofloxacin) by strain F11, which was confirmed by liquid chromatography - exact mass spectrometry. Biodegradation of ofloxacin at 450 µg L-1 by both bacterial strains expressed good linear fits (R2 > 0.98) according to the Rayleigh equation. The enantiomeric enrichment factors were comprised between - 22.5% to - 9.1%, and - 18.7% to - 9.0% in the biodegradation of ofloxacin by strains F11 and FP1, respectively, with no significant differences for the two bacteria under the same conditions. This is the first time that enantioselective biodegradation of ofloxacin and levofloxacin by single bacteria is reported.

The study on interactions between levofloxacin and model proteins by using multi-spectroscopic and molecular docking methods.

The interactions of levofloxacin (LEV) with lysozyme (LYZ), trypsin and bovine hemoglobin (BHb) were investigated, respectively, by using multi-spectral techniques and molecular docking in vitro. Fluorescence studies showed that LEV quenched LYZ/trypsin fluorescence in a combined quenching ways and BHb fluorescence in a static quenching with binding constants of .14, .51 and .20 × 105 L mol-1 at 298 K, respectively. The thermodynamic parameters demonstrated that hydrophobic forces, hydrogen bonds, and van der Waals forces played the major role in the binding process. The binding distances between LEV and the inner tryptophan residues of LYZ, trypsin, and BHb were calculated to be 4.04, 3.38, and 4.52 nm, respectively. Furthermore, the results of circular dichroism spectra (CD), UV-vis, and three-dimensional fluorescence spectra indicated that the secondary structures of LYZ, trypsin, and BHb were partially changed by LEV with the α-helix percentage of LYZ-LEV system increased while that of BHb-LEV system was decreased, the ß-sheet percentage of trypsin-LEV system increased from 41.3 to 42.9%. UV-vis spectral results showed that the binding interactions could cause conformational and some micro-environmental changes of LYZ, trypsin, and BHb. The results of molecular docking revealed that in LYZ and trypsin systems, LEV bound to the active sites residues GLU 35 and ASP 52 of LYZ and trypsin at the active site SER 195, and in BHb system, LEV was located in the central cavity, which was consistent with the results of synchronous fluorescence experiment. Besides, LEV made the activity of LYZ decrease while the activity of trypsin increased.

Development of a Prodrug of Levofloxacin to Avoid Chelation with Al3+ and of Pemetrexed Dimedoxomil Esters for Oral Administration.

An ethoxycarbonyl 1-ethyl hemiacetal ester of levofloxacin (LVFX-EHE) avoids insoluble chelate formation with metal-containing drugs in the intestinal tract and is rapidly hydrolyzed to the parent drug. Furthermore, the minimum inhibitory concentration confirms that LVFX-EHE is less likely to cause pseudomembranous colitis because of less susceptibility to normal intestinal bacteria flora. Pemetrexed dimedoxomil, the prodrug of pemetrexed, was synthesized via reaction with medoxomil bromide after modification of L-glutamate with the tert-butyloxycarbonyl protecting group (BOC), followed by hydrolysis of the BOC moiety with trifluoroacetic acid (TFA) in CH2Cl2 at a temperature of 0°C for 2 h. A serum pemetrexed concentration of >2 µg/mL was observed after oral administration of pemetrexed dimedoxomil at a dose of 60 mg/kg to rats.

The complexity of minocycline serum protein binding.

Objectives: Serum protein binding is critical for understanding the pharmacology of antimicrobial agents. Tigecycline and eravacycline were previously reported to have atypical non-linear protein binding; the percentage of free fraction decreased with increasing total concentration. In this study, we extended the investigation to other tetracyclines and examined the factors that might impact protein binding. Methods: Different minocycline concentrations (0.5-50 mg/L) and perfusion media (saline, 0.1 M HEPES buffer and 0.1 and 1 M PBS) were examined by in vitro microdialysis. After equilibration, two dialysate samples were taken from each experiment and the respective antimicrobial agent concentrations were analysed by validated LC-MS/MS methods. For comparison, the serum protein bindings of doxycycline and levofloxacin were also determined. Results: The free fraction of minocycline decreased with increasing total concentration, and the results depended on the perfusion media used. The trends of minocycline protein binding in mouse and human sera were similar. In addition, serum protein binding of doxycycline showed the same concentration-dependent trend as minocycline, while the results of levofloxacin were concentration independent. Conclusions: The serum protein bindings of minocycline and doxycycline are negatively correlated with their total concentrations. It is possible that all tetracyclines share the same pharmacological property. Moreover, the specific perfusion media used could also impact the results of microdialysis. Additional studies are warranted to understand the mechanism(s) and clinical implications of serum protein binding of tetracyclines.

Sugar-based novel niosomal nanocarrier system for enhanced oral bioavailability of levofloxacin.

CONTEXT: Vesicular systems have attracted great attention in drug delivery because of their amphiphilicity, biodegradability, non-toxicity and potential for increasing drug bioavailability. OBJECTIVE: A novel sugar-based double-tailed surfactant containing renewable block was synthesized for preparing niosomal vesicles that could be exploited for Levofloxacin encapsulation, aiming to increase its oral bioavailability. MATERIALS AND METHODS: The surfactant was characterized by 1H NMR, mass spectroscopy and Fourier transform infrared spectroscopy (FT-IR). Its biocompatibility was studied against cell cultures and human blood hemolysis. In vivo acute toxicity was evaluated in mice. The vesicle morphology, size, drug-excipients interaction and entrapment efficiency (EE) were examined using atomic force microscope (AFM), dynamic light scattering (DLS), FT-IR and HPLC. Oral bioavailability studies of Levofloxacin in surfactant-based niosomal formulation were carried out using rabbits and plasma samples were analyzed using HPLC. RESULTS AND DISCUSSION: Vesicles were spherical in shape and the size was 190.31 ± 4.51 nm with a polydispersity index (PDI) of 0.29 ± 0.03. The drug EE in niosomes was 68.28 ± 3.45%. When applied on cell lines, high cell viability was observed even after prolonged exposure at high concentrations. It caused 5.77 ± 1.34% hemolysis at 1000 µg/mL and was found to be safe up to 2000 mg/kg. Elevated Levofloxacin plasma concentration was achieved when delivered with novel vesicles. CONCLUSION: The surfactant was demonstrated to be safe and effective as carrier of Levofloxacin. The study suggests that this sugar-based double-tailed nonionic surfactant could be promising nano-vesicular system for delivery and enhancing oral bioavailability of the hydrophobic Levofloxacin.