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Rationale: Cloxacillin and flucloxacillin are prone to degradation and polymerization in humid and hot environments, and their polymers have long been recognized to trigger allergic manifestations. A series of the degradation and polymerized impurities in cloxacillin and flucloxacillin were separated and characterized to ensure safe use of these drugs by the public., Methods: By studying the chromatographic behavior of the degradation impurities and polymerized impurities in reversed-phase high-performance liquid chromatography (RP-HPLC) gradient elution, the impurities in cloxacillin and flucloxacillin were effectively separated and eluted. RP-HPLC tandem ion trap/time-of-flight mass spectrometry (MS) was applied to characterize the structures of unknown impurities eluted from the RP-HPLC methods for cloxacillin and flucloxacillin. The mechanisms of formation of the impurities in cloxacillin and flucloxacillin were also investigated., Results: The structures of 10 unknown impurities in cloxacillin and 8 unknown impurities in flucloxacillin were elucidated based on the high-resolution MS n data at positive and negative modes, respectively. Six polymerized impurities were found and characterized, of which three were from the polymerization of cloxacillin and three were from the polymerization of flucloxacillin., Conclusions: The study on the impurity profiles of cloxacillin and flucloxacillin provided a scientific basis for improving their production processes and quality control., (© 2024 John Wiley & Sons Ltd.)

The aim of this study was to investigate the transfer of cephalexin, penicillin-G, and ampicillin & cloxacillin from cow's milk to cheese and whey. For this purpose, raw milk was artificially contaminated to different antibiotic levels and then heat-treated to prepare fresh cheese from it. Antibiotic levels of the milk, whey and cheese were measured with LC-MS/MS. The extent of heat degradation was not sufficient to remove the antibiotic residues from milk. Antibiotic concentrations in whey and fresh cheese were in good accordance with the concentration of the same compound in milk suggesting that contamination of the milk will result in contamination of the product. The investigated antibiotics were transferred less into the cheese curd (1.6-12.5% of the original amount), than into the whey (33.2-74.1%). For penicillin-G even 100% (complete removal) was experienced.

Medical devices such as orthopedic and dental implants may get infected by bacteria, which results in treatment using antibiotics. Since antibiotic resistance is increasing in society there is a need of finding alternative strategies for infection control. One potential strategy is the use of antimicrobial peptides, AMPs. In this study, we investigated the antibiofilm effect of the AMP, RRP9W4N, using a local drug-delivery system based on mesoporous titania covered titanium implants. Biofilm formation was studied in vitro using a safranine biofilm assay and LIVE/DEAD staining. Moreover, we investigated what effect the AMP had on osseointegration of commercially available titanium implants in vivo, using a rabbit tibia model. The results showed a sustained release of AMP with equal or even better antibiofilm properties than the traditionally used antibiotic Cloxacillin. In addition, no negative effects on osseointegration in vivo was observed. These combined results demonstrate the potential of using mesoporous titania as an AMP delivery system and the potential use of the AMP RRP9W4N for infection control of osseointegrating implants., (© 2021 The Authors. Journal of Biomedical Materials Research Part B: Applied Biomaterials published by Wiley Periodicals LLC.)

The present work shows the development and evaluation of the veterinary antibiotic cloxacillin benzathine (CLOXB) loaded into poly-ε-caprolactone (PCL) nanocapsules (NC), as a potential new treatment strategy to manage bovine intramammary infections, such as mastitis. Staphylococcus aureus-induced mastitis is often a recurrent disease due to the persistence of bacteria within infected cells. CLOXB-PCL NC were prepared by interfacial deposition of preformed biodegradable polymer followed by solvent displacement method. The mean diameter of NC varied from 241 to 428 nm and from 326 to 375 nm, when determined by dynamic light scattering and by atomic force microscopy, respectively. The zeta potential of NC was negative and varied from -28 to -51 mV. In vitro release studies from the NC were performed in two media under sink conditions: PBS with 1% polyethylene glycol or milk. A reversed-phase HPLC method was developed to determine the NC entrapment efficiency and kinetics of CLOXB release from the NC. Free CLOXB dissolution occurred very fast in both media, while drug release from the NC was slower and incomplete (below 50%) after 9 h. CLOXB release kinetics from polymeric NC was fitted with the Korsmeyer-Peppas model indicating that CLOXB release is governed by diffusion following Fick's law. The fluorescence confocal microscopy images of macrophage-like J774A.1 cells reveal NC uptake and internalization in vitro. In addition, antimicrobial effect of the intramammary administration of CLOXB-PCL NC in cows with mastitis resulted in no clinical signs of toxicity and allowed complete pathogen elimination after treatment. The in vivo results obtained in this work suggest that CLOXB-PCL NC could be a promising formulation for the treatment of intramammary infections in cattle, considering their physicochemical properties, release profiles and effects on bovine mastitis control., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Penicillins are typical of complex ionic samples which likely contain large number of degradation-related impurities (DRIs) with different polarities and charge properties. It is often a challenge to develop selective and robust high performance liquid chromatography (HPLC) methods for the efficient separation of all DRIs. In this study, an analytical quality by design (AQbD) approach was proposed for stability-indicating method development of cloxacillin. The structures, retention and UV characteristics rules of penicillins and their impurities were summarized and served as useful prior knowledge. Through quality risk assessment and screen design, 3 critical process parameters (CPPs) were defined, including 2 mixture variables (MVs) and 1 process variable (PV). A combined mixture-process variable (MPV) design was conducted to evaluate the 3 CPPs simultaneously and a response surface methodology (RSM) was used to achieve the optimal experiment parameters. A dual gradient elution was performed to change buffer pH, mobile-phase type and strength simultaneously. The design spaces (DSs) was evaluated using Monte Carlo simulation to give their possibility of meeting the specifications of CQAs. A Plackett-Burman design was performed to test the robustness around the working points and to decide the normal operating ranges (NORs). Finally, validation was performed following International Conference on Harmonisation (ICH) guidelines. To our knowledge, this is the first study of using MPV design and dual gradient elution to develop HPLC methods and improve separations for complex ionic samples., (Copyright © 2017 Elsevier B.V. All rights reserved.)

This study evaluates the treatment of the antibiotic cloxacillin (CLX) in water by means of electrochemical oxidation, TiO 2 photocatalysis, and the photo-Fenton system. The three treatments completely removed cloxacillin and eliminated the residual antimicrobial activity from synthetic pharmaceutical wastewater containing the antibiotic, commercial excipients, and inorganic ions. However, significant differences in the degradation routes were found. In the photo-Fenton process, the hydroxyl radical was involved in the antibiotic removal, while in the TiO 2 photocatalysis process, the action of both the holes and the adsorbed hydroxyl radicals degraded the pollutant. In the electrochemical treatment (using a Ti/IrO 2 anode in sodium chloride as supporting electrolyte), oxidation via HClO played the main role in the removal of CLX. The analysis of initial by-products showed five different mechanistic pathways: oxidation of the thioether group, opening of the central β-lactam ring, breakdown of the secondary amide, hydroxylation of the aromatic ring, and decarboxylation. All the oxidation processes exhibited the three first pathways. Moreover, the aromatic ring hydroxylation was found in both photochemical treatments, while the decarboxylation of the pollutant was only observed in the TiO 2 photocatalysis process. As a consequence of the degradation routes and mechanistic pathways, the elimination of organic carbon was different. After 480 and 240 min, the TiO 2 photocatalysis and photo-Fenton processes achieved ∼45 and ∼15 % of mineralization, respectively. During the electrochemical treatment, 100 % of the organic carbon remained even after the antibiotic was treated four times the time needed to degrade it. In contrast, in all processes, a natural matrix (mineral water) did not considerably inhibit pollutant elimination. However, the presence of glucose in the water significantly affected the degradation of CLX by means of TiO 2 photocatalysis.

This study was undertaken to investigate the degradation of 6-aminopenicillanic acid (6-APA) and cloxacillin in aqueous solution by the combined effect of subcritical water and the oxidising agents O 2 , H 2 O 2 , and K 2 S 2 O 8 . Nano ZnO was used as a solid catalyst. Response surface methodology was used to determine the optimum experimental parameters (temperature, treatment time, and concentration of oxidising agent). For 6-APA, the maximum organic carbon (TOC) removal rates of 83.54%, 81.11% and 42.42% were obtained using H 2 O 2 , K 2 S 2 O 8 , and O 2 , respectively. For cloxacillin, the maximum TOC removal rates of 67.69%, 76.02% and 14.45% were obtained using H 2 O 2 , K 2 S 2 O 8 , and O 2 , respectively. Additionally, the impact of nano and commercial ZnO on TOC removal rates was determined. Secondary ions produced during the degradation process-such as nitrite, nitrate, sulphate and chloride-were determined using ion chromatography.

Ampicillin exhibited a negative and a positive cotton effects on the circular dichroism (CD) spectra in the wavelength range of 200-280nm. Cloxacillin showed a positive cotton band peaking at 228nm. Three sensitive, precise and accurate CD spectroscopic methods have been developed for the determination of ampicillin and cloxacillin. Method A was used for the determination of ampicillin in presence of cloxacillin by measuring ellipticity at 206nm. Method B and C were employed to determine ampicillin and cloxacillin based on evaluation of ellipticity at 233nm and 228nm, respectively. Methods A, B and C showed linearity in the concentration range of 10-40μgmL(-1), 5-40μgmL(-1) ampicillin and 10-80μgmL(-1) cloxacillin, respectively. The method A was successfully applied to the determination of ampicillin in commercial dosage forms containing equivalent amount of cloxacillin., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Highly selective molecularly imprinted polymers on the surface of silica gels were prepared by a sol-gel process and used as solid-phase extraction adsorbents for the specific recognition, enrichment and detection of cloxacilloic acid in cloxacillin. The obtained polymers were characterized by scanning electron microscopy, FTIR spectroscopy, nitrogen adsorption and desorption, elemental analysis and thermogravimetric analysis. The imprinted polymers not only possessed high adsorption capacity (6.5 μg/mg), but also exhibited fast adsorption kinetics (they adsorb 80% of the maximum amount within 20 min) and excellent selectivity (the imprinted factor was 3.6). A method using the imprinted polymers as solid-phase extraction adsorbents coupled with high-performance liquid chromatography was established with good specificity, linearity (r = 0.9962), precision (ranging from 0.5 to 6.7%), accuracy (ranging from 93.9 to 97.7%) and extraction recoveries (ranging from 78.8 to 89.8%). The limits of detection and quantification were 0.07 and 0.25 mg/g, respectively. This work could provide a promising method in the enrichment, extraction and detection of allergenic impurities in the manufacture, storage and application of cloxacillin., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

A simple, linear gradient, rapid, precise and stability-indicating RP-UPLC method was developed for the determination of Cloxacillin Sodium in its bulk form and formulation. Ultra performance liquid chromatography, a most promising advancement in a world of chromatography, reduces analysis time, increases reliability through higher resolution, sensitivity and selectivity as well as used as an economic method due to reducing solvent consumption. A chromatographic separation of a drug as well as its degradants was achieved using Waters acquity BEH, 2.1 × 100 mm, 1.7 μm C18 column with gradient of mobile phase A: phosphate buffer, pH 6.8 and mobile phase B: methanol:acetonitrile (75:25). The drug and degradants were monitored at a detection wavelength of 225 nm with a flow rate of 0.35 mL/min and an injection volume of 10 µL. The temperature of the column and auto sampler compartments was at 30°C and 25°C ± 1°C, respectively. The retention time of the drug was ∼6.9 min. The resolution of the drug and degradant peak was >1.5 in all cases. Force degradation of CLOX SOD was carried under alkaline, acidic, oxidative, thermal, photo degradation conditions and it was analyzed by the proposed method. The drug degrades under alkaline, acidic and oxidative conditions but was stable in temperature and light. A developed method was validated as per ICH guidelines using validation parameters such as precision, linearity and range, limit of quantification, specificity, assay and robustness., (© The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

In the present study we developed and validated a liquid chromatography/tandem mass spectrometry (LC-MS/MS) assay for the determination of flucloxacillin in human plasma and microdialysis samples and cloxacillin in microdialysis samples, using oxacillin as the internal standard for the assay. The samples were separated on a UPLC BEH C18,1.7 μm column (2.1x50mm) and analyzed by a tandem-quadrupole mass spectrometer in multiple reaction monitoring mode using an electronspray ionization interface. For flucloxacillin the method was demonstrated to be accurate and precise in the linearity range of 1-30 mg/L in plasma and 0.05-5.0 mg/L for microdialysate with a regression coefficient (r) of 0.9986 and 0.9989 in plasma and microdialysate respectively. For cloxacillin it was accurate and precise in the range of 0.1-5.0 mg/L for microdialysate with a regression coefficient of 0.9972. The method presents a high sensitivity for flucloxacillin (lower limit of quantification of 1 mg/L for plasma and 0.05 mg/L for microdialysis samples) combined with a low within- and between-day variation (<5.0% for flucloxacillin and cloxacillin in microdialysis samples and <6.5% for plasma samples of flucloxacillin). The validation experiments for the microdialysis probes showed a relative recovery of 85.5% for flucloxacillin at a flow rate of 1.0 μL/min. The results justify the use of this assay for clinical studies for measuring free unbound tissue concentrations of flucloxacillin in patients with a Staphylococcus aureus bacteremia.

The intrinsic ability of albumin to bind active substances in the physiological fluids has been explored to endow hydrogels with improved capability to regulate drug release. To develop such biomimetic-functional hydrogels, it is critical that albumin conformation is not altered and that the protein remains retained inside the hydrogel keeping its conformational freedom, i.e., it should be not chemically cross-linked. Thus, the hydrogels were prepared with various proportions of albumin by physical cross-linking of anionic polysaccharides (gellan gum and chondroitin sulfate) with the cationic endogen polyamine spermidine under mild conditions in order to prevent albumin denaturation. Texture and swelling properties of hydrogels with various compositions were recorded, and the effect of the preparation variables was evaluated applying neurofuzzy logic tools for hydrogels prepared with and without albumin and associating the antibiotic cloxacillin. Developed hydrogel systems were extensively analyzed by means of nuclear magnetic resonance (NMR) to determine weak-to-medium and strong binding modes and the equilibrium constants of the albumin-cloxacillin association. NMR techniques were also employed to demonstrate the successful modulation of the cloxacillin release from the albumin-containing hydrogels. In vitro microbiological tests carried out with Staphylococcus aureus and Staphylococcus epidermidis confirmed the interest of the albumin-containing hydrogels as efficient platforms for cloxacillin release in its bioactive form.

This work investigates the development, optimization and in vitro evaluation of liquid paraffin-entrapped multiple-unit alginate-based floating system containing cloxacillin by emulsion-gelation method for gastro retentive delivery. The effect of process variables like drug to polymer ratio by weight, and liquid paraffin to water ratio by volume on various physicochemical properties in case of liquid paraffin-entrapped calcium alginate beads containing cloxacillin applicable to drug entrapment efficiency, density and drug release was optimized using 3(2) factorial design and analyzed using response surface methodology. The observed (actual values) responses were coincided well with the predicted values, given by the optimization technique. The optimized beads showed drug entrapment efficiency of 64.63±0.78%, density of 0.90±0.05 g/cm(3), and drug release of 56.72±0.85% in simulated gastric fluid (pH 1.2) after 8h with floating lag time of 8.45 min and floated well over 12h in simulated gastric fluid (pH 1.2). The average size of all dried beads ranged from 1.73±0.04 to 1.97±0.08 mm. The beads were characterized by SEM and FTIR for surface morphology and excipients-drug interaction analysis, respectively. All these beads showed prolonged sustained release of cloxacillin over 8h in simulated gastric fluid (pH 1.2). The cloxacillin release profile from liquid paraffin beads followed Korsmeyer-Peppas model over a period of 8h with anomalous (non-Fickian) diffusion mechanism for drug release., (Copyright © 2011 Elsevier B.V. All rights reserved.)

The study examined the effect of operating conditions (zinc oxide concentration, pH and irradiation time) of the UV/ZnO photocatalytic process on degradation of amoxicillin, ampicillin and cloxacillin in aqueous solution. pH has a great effect on amoxicillin, ampicillin and cloxacillin degradation. The optimum operating conditions for complete degradation of antibiotics in an aqueous solution containing 104, 105 and 103 mg/L amoxicillin, ampicillin and cloxacillin, respectively were: zinc oxide 0.5 g/L, irradiation time 180 min and pH 11. Under optimum operating conditions, complete degradation of amoxicillin, ampicillin and cloxacillin occurred and COD and DOC removal were 23.9 and 9.7%, respectively. The photocatalytic reactions under optimum conditions approximately followed a pseudo-first order kinetics with rate constant (k) 0.018, 0.015 and 0.029 min(-1) for amoxicillin, ampicillin and cloxacillin, respectively. UV/ZnO photocatalysis can be used for amoxicillin, ampicillin and cloxacillin degradation in aqueous solution.

The study examined degradation of the antibiotics amoxicillin, ampicillin and cloxacillin in aqueous solution by the photo-Fenton process. The optimum operating conditions for treatment of an aqueous solution containing 104, 105 and 103 mg/L amoxicillin, ampicillin, and cloxacillin, respectively was observed to be H(2)O(2)/COD molar ratio 1.5, H(2)O(2)/Fe(2+) molar ratio 20 and pH 3. Under optimum operating conditions, complete degradation of amoxicillin, ampicillin and cloxacillin occurred in 2 min. Biodegradability (BOD(5)/COD ratio) improved from approximately 0 to 0.4, and COD and DOC degradation were 80.8 and 58.4%, respectively in 50 min. Photo-Fenton treatment resulted in the release and mineralization of organic carbon and nitrogen in the antibiotic molecule. Increase in ammonia and nitrate concentration, and DOC degradation were observed as a result of organic carbon and nitrogen mineralization. DOC degradation increased to 58.4% and ammonia increased from 8 to 13.5mg/L, and nitrate increased from 0.3 to 14.2mg/L in 50 min.

The study examined the effect of operating conditions of the Fenton process on biodegradability improvement and mineralization of amoxicillin, ampicillin and cloxacillin antibiotics in aqueous solution. In addition, degradation of amoxicillin, ampicillin and cloxacillin under optimum operating conditions were evaluated. The optimum operating conditions for an aqueous solution containing 104, 105 and 103 mg/L amoxicillin, ampicillin, and cloxacillin, respectively were observed to be COD/H2O2/Fe2+ molar ratio 1:3:0.30 and pH 3. Under optimum operating conditions, complete degradation of amoxicillin, ampicillin and cloxacillin occurred in 2 min. In addition, biodegradability improved from approximately 0 to 0.37 in 10 min, and COD and DOC degradation were 81.4% and 54.3%, respectively in 60 min. Maximum biodegradability (BOD5/COD ratio) improvement was achieved in 10, 20 and 40 min at antibiotics concentration 100, 250 and 500 mg/L, respectively for each antibiotic in aqueous solution. Increase in nitrate and ammonia concentration were observed due to mineralization of organic nitrogen, concentration of nitrate increased from 0.3 to 10 mg/L and concentration of ammonia increased from 8 to 13 mg/L in 60 min. The study indicated that Fenton process can be used for pretreatment of amoxicillin, ampicillin and cloxacillin wastewater for biological treatment.

The interaction of cloxacillin sodium with beta-cyclodextrin (beta-CD) has been studied by several analytical techniques, including (1)H NMR, fluorescence spectroscopy, infrared spectroscopy. In this paper, solid inclusion complex of cloxacillin sodium with beta-CD was synthesized by the coprecipitation method. In addition, the characterization of the inclusion complex has been proved by fluorimetry, infrared spectroscopy and 1D, 2D NMR. The experimental results confirmed the existence of 1:1 inclusion complex of cloxacillin sodium with beta-CD. The formation constant of complex was determined by fluorescence method and (1)H NMR. Spacial configuration of complex has been proposed on 2D NMR technique.

Ampicillin and cloxacillin were degraded together under different stress test conditions prescribed by International Conference on Harmonization. The samples so generated were used to develop a stability-indicating high performance liquid chromatographic (HPLC) method for the two drugs. The drugs were well separated from degradation products using a reversed-phase (C-18) column and a mobile phase comprising of acetonitrile:phosphate buffer (pH 5.0), which was delivered initially in the ratio of 15:85 (v/v) for 1 min, then changed to 30:70 (v/v) for next 14 min, and finally equilibrated back to 15:85 (v/v) from 15 to 20 min. Other HPLC parameters were: flow rate, 1 ml/min; detection wavelength, 225 nm; and injection volume, 5 microl. The method was validated for linearity, precision, accuracy, specificity and selectivity. It was also compared with the assay procedures given in British Pharmacopoeia for individual drugs. Similar results were obtained, indicating that the proposed single method allowed selective analysis of both ampicillin and cloxacillin, in the presence of their degradation products formed under a variety of stress conditions. The developed procedure was also applicable to the determination of instability of the drugs in commercial products.

The synthesis and characterization of cloxacillin (clox) complexes with divalent metal ions [Co (II), Cu (II), Ni (II) and Zn (II)] is described. The nature of bonding of the chelated cloxacillin and the structures of the metal complexes have been elucidated on the basis of their physical and spectroscopic data. In all the complexes, the cloxacillin acts as a uninegatively charged bidentate ligand with coordination involving the carboxylate-O and endocyclic-N of the beta-lactam ring. The new compounds have been screened for in-vitro antibacterial activity against Escherichia coli (a), Klebsiella pneumonae (b), Proteus mirabilis (c), Pseudomonas aeruginosa (d), Salmonella typhi (e), Shigella dysentriae (f), Bacillus cereus (g), Corynebacterium diphtheriae (h), Staphylococcus aureus (j) and Streptococcus pyogenes (k) bacterial strains. The brine shrimp bioassay was also carried out to study their in-vitro cytotoxic properties. All compounds, respectively, showed a promising activity (90%) against five bacterial species at 10 microg/ml concentration and a significant activity (52%) against the same test bacteria at 25 microg/ml concentration.

The surface physicochemical properties of two anionic penicillins-cloxacillin and dicloxacillin-in mixed ethanol-water solvent were investigated by surface tension and dynamic light scattering (DLS). The data were analyzed according to the treatment of the Derjaguin-Landau-Verwey-Overbeek (DLVO) theory to study the stability of the systems. The aim of the study is to obtain information about the effects of ethanol on the surface activity, bulk properties, and aggregate stability of these amphiphilic drugs, keeping in mind that both penicillins have the same counterion, and the difference in their structures is only a Cl atom in the phenyl ring that makes dicloxacillin more hydrophobic. The surface tension data show a minimum area per molecule increment with ethanol concentration that is related to the variation of the dielectric constant with the alcohol. Dicloxacillin has lower values of the standard Gibbs energies of adsorption than does cloxacillin, which gives this drug a more marked escaping tendency from the aqueous environment to the air-water monolayer. DLS data was fitted to an exponential function for cloxacillin at any drug or alcohol concentration in the range of concentrations studied that indicates that the system can be modeled as an ergodic system of dilute diffusing monodisperse particles. Dicloxacillin DLS data at an ethanol concentration of 5% (v/v) had to be fitted at a sum of an exponential and a stretched exponential function, which indicates that, besides the drug aggregates, a small population of penicillin clusters with longer relaxation times is present. The stability curves predicted by the DLVO theory, for both penicillins, indicate the predominance of electrostatic repulsion, leading to a stable system over the drug-ethanol concentration range studied, but the height of the reduced pair interaction potential energy barrier decreases with ethanol concentration, thus it is expected to undergo a transition from a stable dispersion to a coagulated one.

Cloxacillin sodium has been shown to form a charge transfer complex of 2:1 stoichiometry with riboflavin (Vitamin B(2)) in aqueous ethanol medium. The enthalpy and entropy of formation of this complex have been determined by estimating the formation constant spectrophotometrically at five different temperatures in pure water medium. Pronounced effect of dielectric constant of the medium on the magnitude of K has been observed by determining K in aqueous ethanol mixtures of varying composition. This has been rationalized in terms of ionic dissociation of the cloxacillin sodium (D(-)Na(+)), hydrolysis of the anion D(-) and complexation of the free acid, DH with riboflavin.

The effect of ethanol on the thermodynamic properties on two anionic amphiphilic penicillins, cloxacillin and dicloxacillin, has been investigated. Cloxacillin and dicloxacillin are two molecules that are similar structurally, differing only by an additional chlorine atom on the phenyl ring of dicloxacillin. The penicillins can be considered as hydrotropes if we considered that the term comprises hydrophilic and hydrophobic moieties that form aggregates by a stacking mechanism as is the case of both penicillins. By means of ultrasound velocities and densities, we have calculated the apparent molar volumes and adiabatic compressibilities. The critical concentrations, cc, and partition coefficients, K, have been determined, the latter using an indirect method based on the pseudophase model with the help of apparent molar data. This method has the advantage of allowing one to calculate the distribution coefficients at concentrations of the solubilizate below the saturation. The standard molar energy of Gibbs change, DeltaG0, on transfer from the aqueous to the micelar phase was calculated from the partition coefficient. The effect of the alcohol involves a slight decrease of the critical concentration because of a headgroup repulsion decrease. The enthalpies of dilution of dicloxacillin in a mixture of water and 15% w/v of ethanol were calculated. The aggregation process is more exothermic in ethanol that in pure water.

A novel drug carrier system was constructed from anionic drug cloxacillin sodium (CLX) and micelle of poly(styrene-b-2-vinyl pyridine-b-ethylene oxide) (PS-PVP-PEO) by incorporating the former into the micelle of the latter. The incorporation of CLX into the micelle of PS-PVP-PEO was confirmed by zeta-potential measurements, dynamic light scattering (DLS), and fluorescence spectroscopy. In the absence of the anionic drug, CLX, the zeta-potential of neat PS-PVP-PEO micelle was +13 mV under aqueous acidic condition due to the presence of positive charges in the PVP unit. The addition of CLX into the micelle of PS-PVP-PEO decreased the zeta-potential of the micelle smoothly and finally led to a minimum zeta-potential around 0 mV. This fact shows that the added CLX is effectively incorporated into the PS-PVP-PEO micelle by electrostatic attraction. In concomitant with the decrease in zeta-potential, a decrease in hydrodynamic diameter from 94 to 69 nm was observed on addition of CLX to the PS-PVP-PEO micellar solution. This fact also indicates the incorporation of CLX into the PS-PVP-PEO micelles because the binding of CLX to the PVP block of the micelles induces a conformational change from an extended to a shrunken form due to the cancellation of the repulsive force in the PVP blocks by CLX. Fluorescence quenching of pyrene by CLX gave additional evidence for the effective bindings of CLX to the PS-PVP-PEO micelles. Further, release of CLX from the nanoaggregates of CLX/PS-PVP-PEO was investigated in vitro. It was found that the release kinetics of the CLX is conformed to a model based on the consecutive chemical kinetics.

Cloxacillin sodium has been shown to form charge transfer (CT) complexes of 1:1 stoichiometry with a number of electron acceptors in 50% (v/v) aqueous ethanol medium. From the trends in the CT absorption bands, the vertical ionization potential of the drug molecule (cloxacillin sodium) has been estimated to be 7.89 eV. The enthalpies and entropies of formation of two such complexes have been determined by estimating the formation constants spectrophotometrically at five different temperatures. The oscillator strengths and transition dipole moments of these complexes have been determined. It has further been noted that the reduction of o-chloranil by aqueous ethanol is completely inhibited by cloxacillin sodium, a phenomenon that makes the present study of formation equilibrium possible.

The interactions and complexation process of the amphiphilic penicillins sodium cloxacillin and sodium dicloxacillin with horse myoglobin in aqueous buffered solutions of pH 4.5 and 7.4 have been examined by equilibrium dialysis, zeta-potential, isothermal titration calorimetry (ITC) and UV-Vis absorbance techniques. A more opened structure of the protein molecules is detected as a consequence of the reduction of pH from 7.4 to 4.5. Binding isotherms and derived Hill coefficients reflect a cooperative binding behavior. Gibbs energies of binding per mole of drug were obtained from equilibrium dialysis data and compared with those derived from the zeta potential taking into account cooperativity. DeltaGads degrees values so obtained are large and negative at low concentrations where binding to the "high-energy" sites occurs and decreases with the drug concentration. The enthalpies of binding have been obtained from ITC and are small and exothermic so that the Gibbs energies of binding are dominated by large increases in entropy consistent with hydrophobic interactions. Other thermodynamic quantities of the binding mechanism, that is, entropy, DeltaSITCi, Gibbs energy, DeltaGITCi, the binding constant, KITCi, and the number of binding sites, ni, were also obtained, confirming the above results. From ITC data and following a theoretical model, the number of bound and free penicillin molecules was calculated, being higher at pH 4.5 than at pH 7.4. The binding of penicillin causes a conformational transition on protein structure as a consequence of the resulting intramolecular repulsion between the penicillin molecules bound to the protein. Thermodynamic quantites (the Gibbs energy of the transition in water, DeltaGw degrees , and in a hydrophobic environment, DeltaGhc degrees) of the denaturation process were calculated, indicating that at pH 4.5 some of the histidine residues are protonated, becoming accessible to solvent and giving rise to a more opened protein structure.

Penicillin sulfones, which structurally incorporate both a 6-position alkylidene substituent and a 2'beta substituent, have been synthesized and evaluated as inhibitors of class C and class A serine beta-lactamases. Incorporation of the 2'beta-substituent generally improves inhibitory activity. Substituents that improve transport across the bacterial cell membrane have also been incorporated.

The G238S substitution found in extended-spectrum natural mutants of TEM-1 beta-lactamase induces a new capacity to hydrolyze cefotaxime and a large loss of activity against the good substrates of TEM-1. To understand this phenomenon at the molecular level, a method to determine the acylation and deacylation elementary rate constants has been developed by using electrospray mass spectrometry combined with UV spectrophotometry. The hydrolysis of penicillins and cefotaxime by TEM-1 and the G238S mutant shows that the behavior of penicillins and cefotaxime is very different. With both enzymes, the limiting step is deacylation for penicillin hydrolysis, but acylation for cefotaxime hydrolysis. Further analyses of the G238S mutant show that the loss of activity against penicillins is due to a large decrease in the deacylation rate and that the increase in catalytic efficiency against cefotaxime is the result of a better Km and an increased acylation rate. These modifications of the elementary rate constants and the hydrolytic capacity in the G238S mutant could be linked to structural effects on the omega-loop conformation in the active site.

Self-association and formation of micellar solutions of cloxacillin sodium salt (CLO-Na) dissolved in heavy water have been investigated by NMR-spectroscopy. Concentration and temperature dependence of proton and carbon chemical shifts of cloxacillin-Na in D2O is presented and certain 1H and 13C NMR line assignments have been substantiated.