A novel high throughput 96-well-based fluorimetric method to measure amikacin in pharmaceutical formulations: development using response surface methodology.

An aminoglycoside antibiotic, amikacin, is used to treat severe and recurring bacterial infections. Due to the absence of a chromophore, however, amikacin must be extensively derivatized before being quantified, both in analytical and bioanalytical samples. In this study, for the first time, we developed a simple and sensitive method for measuring amikacin sulfate using spectrofluorimetry with a 96-well plate reader, based on the design of the experiment's approach. To develop a robust and reproducible spectrofluorimetric method, the influence of essential attributes, namely pH of the buffer, heating temperature, and concentration of reagents, were evaluated using univariate analysis followed by multivariate analysis (central composite design). International Conference of Harmonization guidelines were used to validate the optimized method. The developed technique is linear from 1.9 to 10 µg/ml with a regression coefficient of 0.9991. The detection and quantification limits were 0.649 µg/ml and 1.9 µg/ml, respectively. For the developed method, both intraday and interday precision (%RSD) were less than 5%. Using the method, amikacin concentrations were quantified in prepared amikacin liposomes and commercial formulations of Amicin®. The developed method greatly reduces sample volume and is a rapid, high throughput microplate-based fluorescence approach for the convenient and cost-effective measurement of amikacin in pharmaceutical formulations. In comparison with previously published approaches, the suggested method allowed for quick analysis of a high number of samples in a short amount of time (96 samples in 125 sec), resulting in an average duration of analysis of 1.3 sec per sample.

Concentration of amikacin sulphate in synovial fluid when given in combination with dexamethasone phosphate in intravenous regional limb perfusion in standing horses.

Eight horses underwent IVRLP at two occasions through a 23-gauge 2 cm long butterfly catheter. Regional anaesthesia of the ulnar, median and medial cutaneous antebrachial nerves was performed prior, and an 8 cm rubber tourniquet was placed on the proximal radius for 30 minutes following the infusion. The first infusion consisted of 2 g of amikacin sulphate and 10 mg of dexamethasone phosphate diluted with 0.9% NaCl to a total volume of 100 ml. The second perfusion was performed after a 2-week washout period, the same protocol was used but without dexamethasone phosphate. Synovial fluid samples were collected from the metacarpophalangeal joint at T = 0, 0.5, 2, 12, 24 and 36 h post-infusion. Synovial fluid amikacin sulphate concentrations were determined by use of liquid chromatography/tandem mass-spectrometry. All horses (n = 8) remained healthy throughout the study, and no adverse effects associated with the study were encountered. No statistically significant differences were found in synovial fluid amikacin sulphate concentrations between the treatment and the control group at any of the time points. In conclusion, dexamethasone phosphate can be used in IVRLP concomitantly with amikacin sulphate in cases of distal limb inflammation and pain without decreasing the synovial fluid concentration of amikacin sulphate.

A Status Update on Pharmaceutical Analytical Methods of Aminoglycoside Antibiotic: Amikacin.

Amikacin (AMK) is one of the commonly used aminoglycoside antibiotics, introduced for clinical use in patients suffering from bacterial infections especially life-threatening gram-negative infections. Due to lack of chromophore in the molecule, the detection of AMK during analysis is a challenge. Thus, pre and post-column derivatization techniques are generally used for AMK estimation. This review focuses on different analytical methods used for detection and quantification of AMK in pure or fixed dose combination pharmaceutical formulations and biological samples. Various reported methods described in the literature include high-performance liquid chromatography techniques, pulsed electrochemical detection techniques, Chemiluminescence techniques, Capillary electrophoresis and immunological methods. High-performance-liquid-chromatography based methods with UV/Vis spectrophotometric, fluorescence and mass spectrometric detection are the most prevailing methods employed for the analysis of AMK. This review could be of significant importance in the area of future AMK analytical method development studies.

Charged aerosol detection in early and late-stage pharmaceutical development: selection of regressionmodels at optimum power function value.

In recent years, the use of quantitative liquid chromatography (LC) coupled charged aerosol detection (CAD) for poor UV absorbing analytes in multicomponent mixtures has grown exponentially across academic and industrial sectors. The ballpark of previous LC-CAD reports is focused on practical applications, as well as optimization of critical parameters such as: response dependencies on temperature, nebulization process, analyte volatility, and mobile-phase composition. However, straightforward approaches to deal with the characteristic nonlinear response of CAD still scarce. A highly overlooked parameter is the power function value (PFV), whose optimization enables a detection signal that is more linear with higher signal-to-noise ratio (S/N) and lower relative standard deviation (RSD) of area counts. Herein, a systematic investigation of different regression models (log-log, first-and second-degree polynomial) by both interpolation and extrapolation process in conjunction with PFV optimization throughout the development of LC-CAD assays is reported. The accuracy of the results via interpolation is always good (< 5%) when operating in the vicinity of the optimum PFV regardless the regression model choice. On the contrary, extrapolation process only worked when applying log-log regression at the optimum PFV (accuracy <5%). This outcome indicates that a first-order regression via interpolation can be a safe and simple choice for quantitative LC-CAD in highly regulated laboratories (GLP, GMP, etc.). Whereas a straightforward extrapolation combined with log-log regression can enable the deployment of high-throughput LC-CAD assays, especially but not limited to laboratories where the synthetic process route is undergoing rapid change and optimization (medicinal chemistry, discovery, biocatalysis, process chemistry, etc.). This approach is crucial in developing quantitative LC-CAD assays for poor UV absorbing pharmaceuticals that are sensitive, precise, accurate and robust across early and late-stage pharmaceutical development.

Indirect Determination of Amikacin by Gold Nanoparticles as Redox Probe.

BACKGROUND: Amikacin is an aminoglycoside antibiotic used for many gram-negative bacterial infections like infections in the urinary tract, infections in brain, lungs and abdomen. Electrochemical determination of amikacin is a challenge in electroanalysis because it shows no voltammetric peak at the surface of bare electrodes. OBJECTIVE: In this approach, a very simple and easy method for indirect voltammetric determination of amikacin presented in real samples. Gold nanoparticles were electrodeposited at the surface of glassy carbon electrode in constant potential. METHODS: The effect of several parameters such as time and potential of deposition, pH and scan rates on signal were studied. The cathodic peak current of Au3+ decreased with increasing amikacin concentration. Quantitative analysis of amikacin was performed using differential pulse voltammetry by following cathodic peak current of gold ions. RESULTS: Two dynamic linear ranges of 1.0 × 10-8-1.0 × 10-7 M and 5.0 × 10-7-1.0 × 10-3 M were obtained and limit of detection was estimated 3.0× 10-9 M. CONCLUSION: The method was successfully determined amikacin in pharmaceutical preparation and human serum. The effect of several interference in determination of amikacin was also studied.

Analytical validation of a quantitative method for therapeutic drug monitoring on the Alinity®c Abbott.

OBJECTIVE: The aim of this study was to evaluate the analytical performance of the Alinity®c Abbott compared to the Architect® immunoassay system for the determination of drugs having a narrow therapeutic index. METHODS: Valproic acid, amikacin, gentamicin, phenobarbital and vancomycin were analyzed using Particle-Enhanced Turbidimetric Inhibitor Immunoassay (Petinia), phenytoin and theophylline were analyzed using an immunoenzymatic method and a colorimetric method was performed to quantify lithium. The methods were validated according to the total error approach. Seven validation standards were analyzed in quintuplet during four days to establish the limits of the methods. Dilution integrity and interferences (hemolysis and high concentrations of bilirubin and lipids) were also tested. Depending on the analyte, the results obtained for twenty to forty patients on the Alinity® were compared to those obtained on the Architect®. RESULTS: The bias and the coefficients of variation for repeatability and for intermediate precision were lower than 15% for all drugs. Accuracy profiles were acceptable (acceptance limits fixed at 30%) in the validated ranges. The lower limits of quantification (LLOQ) were similar to those determined by Abbott except for gentamicin for which we determined a LLOQ at 1.22 mg/L while Abbott determined it at 0.5 mg/L. All assays diluted linear and analyte concentrations were not affected by interferences. Concentrations obtained for real samples on the Alinity®c are comparable to those obtained on the Architect®ci. CONCLUSIONS: The analytical validation of a method suitable for therapeutic drug monitoring of drugs on the Alinity®c meets the requirements of European Medicines Agency.

Predictors of insufficient peak amikacin concentration in critically ill patients on extracorporeal membrane oxygenation.

BACKGROUND: Amikacin infusion requires targeting a peak serum concentration (Cmax) 8-10 times the minimal inhibitory concentration, corresponding to a Cmax of 60-80 mg/L for the least susceptible bacteria to theoretically prevent therapeutic failure. Because drug pharmacokinetics on extracorporeal membrane oxygenation (ECMO) are challenging, we undertook this study to assess the frequency of insufficient amikacin Cmax in critically ill patients on ECMO and to identify relative risk factors. METHODS: This was a prospective, observational, monocentric study in a university hospital. Patients on ECMO who received an amikacin loading dose for suspected Gram-negative infections were included. The amikacin loading dose of 25 mg/kg total body weight was administered intravenously and Cmax was measured 30 min after the end of the infusion. Independent predicators of Cmax < 60 mg/L after the first amikacin infusion were identified with mixed-model multivariable analyses. Various dosing simulations were performed to assess the probability of reaching 60 mg/L < Cmax < 80 mg/L. RESULTS: A total of 106 patients on venoarterial ECMO (VA-ECMO) (68%) or venovenous-ECMO (32%) were included. At inclusion, their median (1st; 3rd quartile) Sequential Organ-Failure Assessment score was 15 (12; 18) and 54 patients (51%) were on renal replacement therapy. Overall ICU mortality was 54%. Cmax was < 60 mg/L in 41 patients (39%). Independent risk factors for amikacin under-dosing were body mass index (BMI) < 22 kg/m2 and a positive 24-h fluid balance. Using dosing simulation, increasing the amikacin dosing regimen to 30 mg/kg and 35 mg/kg of body weight when the 24-h fluid balance is positive and the BMI is ≥ 22 kg/m2 or < 22 kg/m2 (Table 3), respectively, would have potentially led to the therapeutic target being reached in 42% of patients while reducing under-dosing to 23% of patients. CONCLUSIONS: ECMO-treated patients were under-dosed for amikacin in one third of cases. Increasing the dose to 35 mg/kg of body weight in low-BMI patients and those with positive 24-h fluid balance on ECMO to reach adequate targeted concentrations should be investigated.

Stability indicating RP-HPLC method development and validation of cefepime and amikacin in pure and pharmaceutical dosage forms

A simple, accurate, isocratic stability indicating RP-HPLC method was developed for the determination of cefepime and amikacin in Pure and its pharmaceutical formulations. The method consists of methanol: acetonitrile:acetate buffer 75:20:05 (v/v) mobile phase at pH 5.1 with C18 column as stationary phase. The flow rate and detection wave length were 1.0 mL/min and 212 nm respectively. The linearity range for the method was found to be 2.5-25 µg/mL for amikacin and 10-100 µg/mL cefepime respectively. The developed method was validated as per ICH guidelines and the results of all the validation parameters were well within their acceptance values. Also the forced degradation studies were conducted with standard drugs. Degradation products formed during the different stress conditions were separated from both drugs. This validated method was applied for the simultaneous estimation of cefepime and amikacin in commercially available formulation sample.

Evaluation of dynamics of derivatization and development of RP-HPLC method for the determination of amikacin sulphate.

The absence of chromophore and/or conjugated system, prerequisite for UV and florescent light detection, or absorbance at very low wavelength necessitates the development of simple and reliable methods for the determination of amikacin sulphate. Therefore, the present study describes for the first time dynamics of the drug derivatization using ninhydrin reagent and development and validation of a simple RP-HPLC method, using diode array detector (DAD). The variables such as heating time, heating type, drug-reagent ratio, reagent composition and storage temperature of the derivative were optimized. The analyte and aqueous ninhydrin solution upon heating for 2.00-5.00 min produced the colored drug-derivative which was stable for one month at refrigeration. The derivatized drug (20.00µL) was eluted through a column - Eclipse DB-C18 (5.00 µm, 4.60×150.00 mm), maintained at 25°C- using isocratic mobile phase comprising water and acetonitrile (70:30, v/v) at a flow rate of 1.00 mL/min, and detected at 400 nm. The method was found to be reliable (98.08-100.72% recovery), repeatable (98.02-100.72% intraday accuracy) and reproducible (98.47-101.27% inter day accuracy) with relative standard deviation less than 5%. The results of the present study indicate that the method is easy to perform, specific and sensitive, and suitable to be used for the determination of amikacin sulphate in bulk and pharmaceutical preparations using less expensive/laborious derivatization.

Susceptibility of Malassezia pachydermatis to aminoglycosides.

Previous studies have evaluated the action of gentamicin against Malassezia pachydermatis. The aim of this study was to evaluate in vitro susceptibility of M. pachydermatis to the aminoglycosides- gentamicin, tobramycin, netilmicin and framycetin. The minimum inhibitory concentration (MIC) of gentamicin was determined following methods M27-A3 microdilution and Etest® . The Etest® was used to determine the minimum inhibitory concentration (MIC) of the tobramycin and netilmicin. The Kirby-Bauer test was used to determine the antibiotic susceptibility to the framycetin. The MIC50 and MIC90 were 8.12 µg/mL and 32.5 µg/mL by microdilution method for gentamicin. The MIC50, determined by the Etest® , was 8 µg/mL for gentamicin and netilmicin and 64 µg/mL for tobramycin. The MIC90 was 16 and 32 µg/mL for gentamicin and netilmicin respectively. The MIC90 was outside of the detectable limits for tobramycin. To framycetin, 28 strains (40%) of the 70 M. pachydermatis isolates tested showed a diameter of 22 mm, 22 strains (31.42%) showed a diameter of 20 mm, 16 strains showed a diameter of ≤ 18 mm, and only 5.71% of the isolates showed a diameter of ≥ 22 mm. This study provides evidence of high in vitro activity of the aminoglycosides-gentamicin, tobramycin, netilmicin and framycetin against M. pachydermatis. For gentamicin Etest® showed similar values of MIC50 y MIC90 that the obtained by microdilution method. We considered Etest® method could be a good method for these calculations with aminoglycosides.

Antibacterial activity of human simulated epithelial lining fluid concentrations of amikacin inhale alone and in combination with meropenem against Acinetobacter baumannii.

BACKGROUND: Acinetobacter baumannii(ACBN) is a MDR organism causing pneumonia in ventilated patients. High MICs often result in insufficient lung exposures, thus poor outcomes have been observed with parenteral antimicrobials. Amikacin Inhale(AMK-I), is a drug-device combination of amikacin and a Pulmonary Drug Delivery System device. We aimed to describe the pharmacodynamic profile of human simulated epithelial lining fluid(ELF) exposures of AMK-I and intravenous meropenem alone and in combination against ACBN with variable susceptibility profiles. METHODS: AMK-I ELF exposures and the ELF profile of meropenem achieved after intravenous administration were evaluated in an in vitro pharmacodynamic model. Nine ACBN with amikacin/meropenem MICs of 2-512/2 to >64 mg/L were utilized. MICs were repeated post exposure to assess the development of resistance. RESULTS: AMK-I monotherapy rapidly achieved and sustained bactericidal activity for isolates with amikacin MIC ≤128 mg/L. For isolates with MICs of 256 and 512 mg/L initial reductions in bacterial density were observed followed by regrowth. The combination produced similar bactericidal activity against ACBN with amikacin MICs of ≤128. While the combination regimen produced initial reductions and prolonged the duration of activity against organisms with MICs of 256 and 512 mg/L, regrowth and MIC elevations were noted during the 72-h exposure period. CONCLUSION: The combination achieved rapid and sustained efficacy when amikacin MICs were ≤128 mg/L and prolonged the duration of activity compared to monotherapy for organisms with MICs 256 mg/L and 512 mg/L. These data support the utility of AMK-I as an adjunct for the treatment of pneumonia caused by A. baumannii with MICs above current susceptibility break-points.

A simplified guide for charged aerosol detection of non-chromophoric compounds-Analytical method development and validation for the HPLC assay of aerosol particle size distribution for amikacin.

Amikacin, an aminoglycoside antibiotic lacking a UV chromophore, was developed into a drug product for delivery by inhalation. A robust method for amikacin assay analysis and aerosol particle size distribution (aPSD) determination, with comparable performance to the conventional UV detector was developed using a charged aerosol detector (CAD). The CAD approach involved more parameters for optimization than UV detection due to its sensitivity to trace impurities, non-linear response and narrow dynamic range of signal versus concentration. Through careful selection of the power transformation function value and evaporation temperature, a wider linear dynamic range, improved signal-to-noise ratio and high repeatability were obtained. The influences of mobile phase grade and glassware binding of amikacin during sample preparation were addressed. A weighed (1/X2) least square regression was used for the calibration curve. The limit of quantitation (LOQ) and limit of detection (LOD) for this method were determined to be 5µg/mL and 2µg/mL, respectively. The method was validated over a concentration range of 0.05-2mg/mL. The correlation coefficient for the peak area versus concentration was 1.00 and the y-intercept was 0.2%. The recovery accuracies of triplicate preparations at 0.05, 1.0, and 2.0mg/mL were in the range of 100-101%. The relative standard deviation (Srel) of six replicates at 1.0mg/mL was 1%, and Srel of five injections at the limit of quantitation was 4%. A robust HPLC-CAD method was developed and validated for the determination of the aPSD for amikacin. The CAD method development produced a simplified procedure with minimal variability in results during: routine operation, transfer from one instrument to another, and between different analysts.

Fecal microbiota transplantation: is there a role in the eradication of carbapenem-resistant Klebsiella pneumoniae intestinal carriage?

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Evaluation of 10-minute versus 30-minute tourniquet time for intravenous regional limb perfusion with amikacin sulfate in standing sedated horses.

The objective of the article is to determine the synovial fluid concentrations of amikacin sulfate (AS) after performing an intravenous regional limb perfusion (IVRLP) with the tourniquet in place for 10 minutes and compare this with 30 minutes. This is a randomised crossover experimental design. Animals were seven healthy adult horses. Horses underwent two IVRLP procedures in a randomised, crossover design. The horses received IVRLP with 2 g AS diluted to 60 ml using 0.9% saline in the cephalic vein of alternate limbs with the tourniquet in place for either 10 or 30 minutes, with a washout period between procedures. Movement of the horses was recorded. Synovial fluid was collected from the radiocarpal and metacarpophalangeal joints at five minutes and 24 hours after IVRLP, and amikacin concentration was compared between groups. Amikacin was quantified by a fluorescence polarisation immunoassay. There was no significant difference between the synovial amikacin concentrations noted between 10 minute and 30 minute IVRLP. Horses in the 30 minute tourniquet application group demonstrated more movement than the 10 minute tourniquet group. There was a significant difference between groups with regard to the systemic venous blood concentration of amikacin one minute after tourniquet release with mean systemic venous blood concentrations of AS being higher (P=0.001) in the 10 minute group. Ten minute application of a wide elastic tourniquet may be sufficient when performing IVRLP in standing sedated horses.

Design of a surface plasmon resonance immunoassay for therapeutic drug monitoring of amikacin.

The therapeutic drug monitoring (TDM) of pharmaceutical drugs with narrow therapeutic ranges is of great importance in the clinical setting. It provides useful information towards the enhancement of drug therapies, aiding in dosage control and toxicity risk management. Amikacin is an aminoglycoside antibiotic commonly used in neonatal therapies that is indicated for TDM due to the toxicity risks inherent in its use. Current techniques for TDM such as high performance liquid chromatography (HPLC) and gas chromatography-mass spectrometry (GC-MS) are costly, time consuming, and cannot be performed at the site of action. Over the last decades, surface plasmon resonance (SPR) biosensors have become increasingly popular in clinical diagnostics due to their ability to detect biomolecular interactions in real-time. We present an SPR-based competitive immunoassay for the detection of the antibiotic amikacin, suitable for TDM in both adults and neonates. We have obtained high specificity and sensitivity levels with an IC50 value of 1.4ng/mL and a limit of detection of 0.13ng/mL, which comfortably comply with the drug's therapeutic range. Simple dilution of serum can therefore be sufficient to analyze low-volume real samples from neonates, increasing the potential of the methodology for TDM. Compared to current TDM conventional methods, this SPR-based immunoassay can provide advantages such as simplicity, potential portability, and label-free measurements with the possibility of high throughput. This work is the foundation towards the development of an integrated, simple use, highly sensitive, fast, and point-of-care sensing platform for the opportune TDM of antibiotics and other drugs in a clinical setting.

Development and validation of an HPLC method for determination of Amikacin in water samples by solid phase extraction and pre-column derivatization.

This work presents a rapid and sensitive high performance liquid chromatography method for the determination of amikacin in water samples with solid phase extraction and pre-column derivatization. Amikacin residue was extracted from water samples with solid phase extraction cartridge. Then the extraction solution was derivatized with 4-chloro-3,5-dinitrobenzotrifluoride in the presence of triethylamine at 70°C in 20 min. The amikacin derivative was separated on a C18 column and detected by application of UV detection at 238 nm. The limit of detection is 0.2 µg/L with a signal-to-noise ratio of 3 and linearity is established over the concentration range from 0 to 500.0 µg/L. Recoveries of the amikacin in three types of water samples are from 87.5 % to 99.6 % and RSDs are 2.1 %-4.5 %. This method can be used for the quantification of amikacin residues in water samples.

Development of a Fourier transform infrared spectroscopy coupled to UV-Visible analysis technique for aminosides and glycopeptides quantitation in antibiotic locks.

Antibiotic Lock technique maintains catheters' sterility in high-risk patients with long-term parenteral nutrition. In our institution, vancomycin, teicoplanin, amikacin and gentamicin locks are prepared in the pharmaceutical department. In order to insure patient safety and to comply to regulatory requirements, antibiotic locks are submitted to qualitative and quantitative assays prior to their release. The aim of this study was to develop an alternative quantitation technique for each of these 4 antibiotics, using a Fourier transform infrared (FTIR) coupled to UV-Visible spectroscopy and to compare results to HPLC or Immunochemistry assays. Prevalidation studies permitted to assess spectroscopic conditions used for antibiotic locks quantitation: FTIR/UV combinations were used for amikacin (1091-1115cm(-1) and 208-224nm), vancomycin (1222-1240cm(-1) and 276-280nm), and teicoplanin (1226-1230cm(-1) and 278-282nm). Gentamicin was quantified with FTIR only (1045-1169cm(-1) and 2715-2850cm(-1)) due to interferences in UV domain of parabens, preservatives present in the commercial brand used to prepare locks. For all AL, the method was linear (R(2)=0.996 to 0.999), accurate, repeatable (intraday RSD%: from 2.9 to 7.1% and inter-days RSD%: 2.9 to 5.1%) and precise. Compared to the reference methods, the FTIR/UV method appeared tightly correlated (Pearson factor: 97.4 to 99.9%) and did not show significant difference in recovery determinations. We developed a new simple reliable analysis technique for antibiotics quantitation in locks using an original association of FTIR and UV analysis, allowing a short time analysis to identify and quantify the studied antibiotics.

Liquid chromatographic determination of amikacin sulphate after pre-column derivatization.

A novel high performance liquid chromatographic (HPLC) method with a pre-column derivatization reaction has been developed and validated. The method was used for the determination of the aminoglycoside antibiotic amikacin sulphate (AMK) in the presence of its synthetic precursor kanamycin sulphate in pure form and in different pharmaceutical preparations. The pre-column derivatization was based on Hantzsch condensation reaction and the obtained coloured products were separated using an isocratic reversed-phase high performance liquid chromatographic method. The separation was achieved on a Spherisorb C18 ODS2 (250 × 4.6 mm, 5 µm) column using a mobile phase composed of acetonitrile-0.1 M sodium acetate buffer (pH 5.0; 25:75, v/v). The column temperature was adjusted at 35 °C and the flow rate at 2 mL min(-1). The detection was carried out at 330 nm by using photo-diode array detector. Different conditions for the optimization of the derivatization reaction as well as for the HPLC measurement were studied. Moreover, AMK was subjected to forced degradation by oxidation, hydrolysis, photolysis and dry heat. Degradation products did not interfere with the assay, which can thus be considered selective and specific. The proposed method was validated for linearity, precision, accuracy, specificity and robustness. Also, it was used to check the purity of AMK in the presence of KAN (related impurity) at the pharmacopoeial limit (0.5%).

Amikacin reverse iontophoresis: optimization of in vitro extraction.

The aim of this work was to optimize amikacin reverse iontophoretic extraction across the skin in vitro, for non-invasive drug monitoring. Reverse iontophoresis experiments were performed using vertical diffusion cells. The lower chamber, simulating body fluids, contained amikacin bisulphate and acetaminophen, as marker for electroosmosis, while the upper chamber was filled with the appropriate extraction solution. The effect of concentration of amikacin in the dermal bathing solution and the effect of extraction solution composition and pH were studied. The results show that the extraction of amikacin was independent of pH and always in the anode-to-cathode direction, in agreement with the positive charge of the drug. The presence of amikacin in the bathing solution did not modify acetaminophen extraction at pH 4.0, while the extraction was reduced at pH 8.0. In conclusion, amikacin can be extracted across the skin in vitro by reverse iontophoresis. Owing to the charge of the molecule, extraction takes place at the cathode. Using acetaminophen as neutral marker, it was shown that amikacin can interact with the skin and alter its permselectivity at pH 8.0.

Development and validation of a capillary electrophoresis method with capacitively coupled contactless conductivity detection (CE-C(4) D) for the analysis of amikacin and its related substances.

Amikacin is a semisynthetic aminoglycoside antibiotic derived from kanamycin A that lacks a strong UV absorbing chromophore or fluorophore. Due to the physicochemical properties of amikacin and its related substances, CE in combination with capacitively coupled contactless conductivity detection (CE-C(4) D) was chosen. The optimized separation method uses a BGE composed of 20 mM MES adjusted to pH 6.6 by l-histidine and 0.3 mM CTAB that was added as flow modifier in a concentration below the CMC. Ammonium acetate 20 mg.L(-1) was used as internal standard. 30 kV was applied in reverse polarity on a fused silica capillary (73/48 cm; 75 µm id). The optimized separation was obtained in less than 6 min with good linearity (R(2) = 0.9996) for amikacin base. It shows a good precision expressed as RSD on relative peak areas equal to 0.1 and 0.7% for intraday and interday, respectively. The LOD and LOQ are 0.5 mg.L(-1) and 1.7 mg.L(-1) , respectively.