Quantification Of Amoxicillin Research Articles

A HPLC-MS/MS method for screening of selected antibiotic adulterants in herbal drugs.

The use of herbal products adulterated with conventional drugs increases the risk of developing microbial resistance and causes herb-to-drug interaction, leading to severe clinical consequences. The complex nature of herbal products has been a challenge for the unambiguous identification of adulterants. The improved analytical selectivity and sensitivity of hyphenated techniques such as high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) enable the confirmatory screening of adulterants in herbal products. Simultaneous screening of adulterants is necessary and efficient because it has been established that more than one chemical adulterant may be present in one herbal product. An HPLC-MS/MS method for the simultaneous detection and quantification of amoxicillin, ampicillin, metronidazole, ciprofloxacin, sulfamethoxazole, and trimethoprim in powdered herbal drugs was developed. Deuterated metronidazole-d3, trimethoprim-d3, ciprofloxacin-d8, and sulfamethoxazole-d4 were used as internal standards (ISs). For each analyte, two transitions were monitored using protonated molecules as precursor ions. The extraction of analytes from herbal products was performed using a simple methanol : water : formic acid (90 : 10 : 0.05, v/v) extraction solvent. Chromatographic separation was done in a gradient of 0.01% formic acid in methanol and 0.01% formic acid in MilliQ water. The calibration curves were linear (r2 ≥ 0.996) over the range of 0.005-2.5 μg mL-1 for all compounds except metronidazole, whose range was 0.005-1 μg mL-1. The limit of detection (LOD) ranged from 0.012 to 0.046 μg mL-1, while the limit of quantification (LOQ) ranged from 0.066 to 0.153 μg mL-1. The accuracy, expressed as the recovery of spiked herbal products, ranged from 45% to 114%. The precision, expressed as relative standard deviation (RSD) at two concentration levels, ranged from 1.6% to 15.9%. The matrix effect expressed as the matrix factor (MF) ranged from 0.79 to 0.92. The developed method was applied to powder herbal products purchased in Tanzania. Amoxicillin, ampicillin, trimethoprim, sulfamethoxazole, and ciprofloxacin were not detected in all samples. Metronidazole was detected in eight samples with the highest concentration of 1.38 μg g-1. The developed method is suitable for the detection of all the studied antibiotic adulterants in herbal products. Quantification can be performed for all the compounds except ciprofloxacin due to its lower recovery.

Development of a New Electrochemical Sensor Based on Mag-MIP Selective Toward Amoxicillin in Different Samples

This work describes an electrochemical sensor for the selective recognition and quantification of amoxicillin and a β-lactam antibiotic in real samples. This sensor consists of a carbon paste electrode (CPE) modified with mag-MIP (magnetic molecularly imprinted polymer), which was prepared by precipitation method via free radical using acrylamide (AAm) as functional monomer, N,N′-methylenebisacrylamide (MBAA) as a crosslinker, and potassium persulfate (KPS) as initiator, to functionalized magnetic nanoparticles. The magnetic non-imprinted polymers (mag-NIP) were prepared using the same experimental procedure without analyte and used for the preparation of a CPE for comparative studies. The morphological, structural, and electrochemical characteristics of the nanostructured material were evaluated using Field emission gun scanning electron microscopy (FEG-SEM), Transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FTIR), Vibrating sample magnetometry (VSM), X-ray diffraction (XRD), and voltammetric technique. Electrochemical experiments performed by square wave voltammetry show that the mag-MIP/CPE sensor had a better signal response compared to the non-imprinted polymer-modified electrode (mag-NIP/CPE). The sensor showed a linear range from 2.5 to 57 μmol L−1 of amoxicillin (r2 = 0.9964), with a limit of detection and a limit of quantification of 0.75 and 2.48 μmol L−1, respectively. No significant interference in the electrochemical signal of amoxicillin was observed during the testing experiments in real samples (skimmed milk and river water). The proposed mag-MIP/CPE sensor could be used as a good alternative method to confront other techniques to determine amoxicillin in different samples.

Diffuse reflectance near infrared-chemometric methods development and validation of amoxicillin capsule formulations.

Objective:The aim of present study was to establish near infrared-chemometric methods that could be effectively used for quality profiling through identification and quantification of amoxicillin (AMOX) in formulated capsule which were similar to commercial products. In order to evaluate a large number of market products easily and quickly, these methods were modeled.Materials and Methods:Thermo Scientific Antaris II near infrared analyzer with TQ Analyst Chemometric Software were used for the development and validation of the identification and quantification models. Several AMOX formulations were composed with four excipients microcrystalline cellulose, magnesium stearate, croscarmellose sodium and colloidal silicon dioxide. Development includes quadratic mixture formulation design, near infrared spectrum acquisition, spectral pretreatment and outlier detection. According to prescribed guidelines by International Conference on Harmonization (ICH) and European Medicine Agency (EMA) developed methods were validated in terms of specificity, accuracy, precision, linearity, and robustness.Results:On diffuse reflectance mode, an identification model based on discriminant analysis was successfully processed with 76 formulations; and same samples were also used for quantitative analysis using partial least square algorithm with four latent variables and 0.9937 correlation of coefficient followed by 2.17% root mean square error of calibration (RMSEC), 2.38% root mean square error of prediction (RMSEP), 2.43% root mean square error of cross-validation (RMSECV).Conclusion:Proposed model established a good relationship between the spectral information and AMOX identity as well as content. Resulted values show the performance of the proposed models which offers alternate choice for AMOX capsule evaluation, relative to that of well-established high-performance liquid chromatography method. Ultimately three commercial products were successfully evaluated using developed methods.

Development of an Liquid Chromatography–Tandem Mass Spectrometry Method for the Determination of Amoxicillin in Broth Medium and its Application to anIn VitroPharmacokinetic and Pharmacodynamic Model

A simple, rapid and highly sensitive liquid chromatographic-tandem mass spectrometry (LC-MS-MS) method has been developed and validated for the quantification of amoxicillin in broth-a liquid bacterial culture medium. After appropriate dilution with ultrapure water, broth samples containing amoxicillin and an internal standard (IS) were extracted by acetonitrile and dichloromethane. The extract was injected into the system. The analyte and the IS were separated by a prepacked Atlantis C18 column using acetonitrile-0.1% formic acid as a mobile phase and detected by selected reaction monitoring in electrospray ionization positive ion mode. The calibration curve of amoxicillin was linear over the concentration range of 0.05-20.00 µg/mL. The mean recovery of amoxicillin from broth was 71.7%, and the intra- and interday precision and accuracies of the assay were within 10%. Amoxicillin was stable in broth for 12 h at room temperature (24°C), for 6.5 months at -80°C and for 24 h after preparation in an autosampler at room temperature. It has been successfully applied to an in vitro pharmacokinetic (PK) and pharmacodynamic (PD) model in which the broth is used for bacterial growth. The method provides high-throughput biological analysis to facilitate the in vitro PK and PD model of amoxicillin.

Highly sensitive determination and selective immobilization of amoxicillin using carbon ionic liquid electrode

Carbon ionic liquid electrode (CILE) was used as a substrate for immobilization and determination of amoxicillin (AMX) in phosphate buffer solution. The electrochemical response of AMX showed an irreversible oxidation peak in the first scan and redox peaks corresponding to the electrooxidation of product in the subsequent scans which gradually grew upon repetitive scanning. Effect of experimental conditions on electrooxidation of AMX and its adsorbed intermediates was investigated, and possible mechanism for strong immobilization is suggested. The electrode response was linear in the concentration range of 5.0 to 400.0 μmol L−1 AMX with a detection limit of 0.8 μmol L−1 AMX. There was no serious interference from other β-lactam family of antibiotics such as ampicillin and penicillin. The electrode showed good selectivity as well as high sensitivity toward quantification of AMX in pharmaceuticals and urine samples.

Validation of a chromatographic method for amoxicillin determination in wastewaters after its degradation by advanced oxidation process

A rapid analytical procedure based on high performance liquid chromatography for identification and quantification of amoxicillin in wastewaters after its degradation by advanced oxidation process (AOP) was developed and validated. The proposed methodology was validated concerning to linearity, precision, limit of quantification, and limit of detection according to the required INMETRO and ANVISA (regulatory agencies in Brazil) standards. The method was applied to the determination of amoxicillin in wastewaters from wastewater treatment plant of a Brazilian pharmaceutical industry spiked with amoxicillin after AOP treatment process on a pilot scale reactor. The method allows the determination of the referred drug in a concentration range of 10–100 mg L−1 with a detection limit of 2.32 mg L−1, a quantification limit of 7.04 mg L−1, and enabling control of contaminant removal.

Estimation of Amoxicillin in Presence of High Concentration of Uric Acid and Other Urinary Metabolites Using an Unmodified Pyrolytic Graphite Sensor

Voltammetric determination of amoxicillin (AMX) has been carried out using an unmodified edge plane pyrolytic graphite (EPPG) sensor. The peak corresponding to the oxidation of AMX was found close to ∼560 mV, both in cyclic as well as in square wave voltammetry. AMX has been quantified using square wave voltammetry in the linear concentration range of 5–500 μM at pH 7.2. Sensitivity and detection limit of 0.0742 μA μM−1 and 0.84 μM was achieved respectively. The well separated peak of AMX, from the common metabolites present in urine, such as UA, xanthine and hypoxanthine, demonstrates the selective nature of the developed sensor. Also, no change in the peak current of AMX, in the presence of clavulanic acid and ampicillin (used commercially in combination with AMX), indicated the practical utility of the developed sensor in pharmaceutical industries. The analytical applicability of the sensor was checked by analyzing different pharmaceutical formulations and by quantification of AMX in the urine sample of the patients undergoing treatment with AMX. The results were further validated using HPLC and a comparison demonstrated the successful application of the proposed sensor for the fast, simple and selective analysis of AMX at pharmaceutical and medical fronts.

Sequential injection chromatography against HPLC and CE: Application to separation and quantification of amoxicillin and clavulanic acid

Sequential injection chromatography (SIC) has been recently proposed as an alternative separation technique. SIC has the advantages of inexpensiveness, rapid operation procedure, small dimension, short stabilization time, friendly maintenance process and less reagent consumption. In contrast, SIC has had suffered from some limitations. In the current study, a higher pressure resistant selection valve with additional ports than that available in commercially SIC systems was installed. This development allows achieving solution propulsion without showing leakage and linking more standard/sample solutions. In addition, a new method for the separation and quantification of amoxicillin and clavulanic acid in pharmaceutical formulations has been optimized and validated. A comparative study on the efficiency of the SIC method with previous HPLC and CE methods was conducted as well. Besides the benefits of the instrumentation of SIC, the proposed method has shown some advantages over previous HPLC and CE methods with respect to reagent consumption and sample frequency. Other such analytical characters of the SIC method as resolution, peak symmetry, number of theoretical plates, linearity range, accuracy, precision and limits of detection and quantification, recorded comparable results with those obtained from previous HPLC and CE methods.

Development and validation of a method to determine amoxicillin in physiological fluids using micellar liquid chromatography

A simple and robust method was developed for the routine identification and quantification of amoxicillin by micellar LC. Amoxicillin, a beta-lactamase inhibitor, is one of the most commonly prescribed drugs in the treatment of urine and skin structure infections. In this work, amoxicillin was determined in urine samples without any pretreatment step in a phenyl column using a micellar mobile phase of 0.10 M SDS and 4% butanol at pH 3. A UV detection set at 210 nm was used. Amoxicillin was eluted at 5.1 min with no interference by the protein band or endogenous compounds. Linearities (r >0.9998), intra- and interday precisions were determined (RSD (%) 0.4-2.7% and 0.3-5%, respectively, in micellar media, and 0.14-2.6% and 0.13-6%, respectively, in urine), and robustness was studied in the method validation. LOD and LOQ were 0.04 and 0.1 microg/mL in micellar media and 0.14 and 0.34 microg/mL in urine, respectively. Recoveries in the urine matrix were in the range of 95-110%. The validated method proved to be reliable and sensitive for the determination of amoxicillin in urine samples.

Study on the formation of an amoxicillin adduct with methanol using electrospray ion trap tandem mass spectrometry

The possibility of using the protonated methanol-adduct of antimicrobial amoxicillin for its identification and quantification at residue levels has been investigated, since it is impossible to completely suppress the formation of these adducts when methanol is present in the solvent system. This process has been monitored over time and as a function of concentration. It was determined that adducts were instantly formed and that the abundance of the protonated methanol-adduct at m/z 398 increased at the expense of the protonated molecule m/z 366 with storage time. The effect of several common solvents and mobile-phase additives on the ionization efficiency of amoxicillin and the formation of the methanol adduct has also been investigated. It was shown that the mass spectra of amoxicillin were strongly influenced by the solvent in which the analyte is dissolved and by the analyte concentration, as well as by the composition of mobile phase. Methanol was determined to be the best spray solvent, as it provided spectra with the lowest abundance of dimer ions. It was also determined that acetic acid as the mobile-phase additive provided the highest signal intensities, while ammonium acetate should not be used as an additive for the determination of amoxicillin at residue levels. Using high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC/ESI-MS/MS), fragmentation of the protonated molecules and the protonated methanol-adduct ions, in both positive and negative ion mode, has been performed. The fragmentation was stable and strong product ion spectra were obtained. The linearity of the MS detector response, and that of the chromatographic method, was tested. Due to the linear behaviour it was concluded that the protonated methanol-adduct ion can be used for analytical purposes, i.e. for identification and quantification of amoxicillin at trace levels.

Rapid determination of amoxicillin in premixes by HPLC

A rapid analytical procedure for routine identification and quantification of amoxicillin in premixes by high performance liquid chromatography was developed and tested. The ground premix samples were extracted for 10 min using 100 ml extraction mixture water–methanol (800:200, v/v). The extract was analyzed by reversed-phase on Agilent Zorbax SB-C18 column (4.6 mm × 150 mm, i.d., 5 μm particle size) with water–methanol–phosphoric acid–triethylamine (842:150:4:4) containing 10 mM hexane-1-sulfonic acid sodium salt (pH 3.5) as mobile phase. UV detection was carried out at 230 nm. The method was validated for specificity, linearity, solution stability, accuracy, precision, limit of detection, and limit of determination. The detector response for amoxicillin was linear over the selected concentration range from 2.0 to 40.0 mg ml −1 with a correlation coefficient 0.9999. The mean accuracy was 100.1% with a standard deviation of 0.6%. The limit of detection and the limit of determination are 0.1 and 0.3 mg ml −1, respectively, which corresponds to 10 and 30 mg kg −1, respectively, in real premix sample. The sample and standard solutions were stable for 4 h. The method is selective and can be used in routine analysis.