Current Pharmaceutical Analysis - Volume 9, Issue 2, 2013

The extrapyramidal side effect parameters of typical and atypical antypsychotics were correlated with affinity chromatographic data determined on the melanin-based column. The chromatographic study was performed according to the hypothesis that extrapyramidal symptoms (EPS) as side effects of the use of antipsychotic drugs at clinically effective doses are correlated to the affinity of these drugs to neuromelanin. For that aim the polymerization product of L-DOPA (melanin) was immobilized onto aminopropyl silica and the binding efficiency of melanin towards antipsychotics has been determined. The results indicate that melanin based-column can be used to evaluate the risk of EPS of drug candidates to antipsychotic drug therapy.

Tablet splitting has been a practice widely used. The aim of this work is to investigate the influence of splitting on content uniformity of 25 mg captopril tablets, a common dosage form. Split tablets were obtained by using the hands, a knife or a tablet-splitter. Captopril reference product and two different brands codified as X and Y were used in this study. The method used for quantification was that described in The United States Pharmacopoeia, 30th ed. The results showed that the quality of the analyzed products was affected by splitting processes (by either using the hands, a knife or a tabletsplitter). Nevertheless, the use of tablet splitter produced halves more uniform than the other division processes considered. Moreover, similar products codified as X and Y had a greater discrepancy in content uniformity when knife was used to split the tablets, while splitting the tablets by hands produced the highest variation in the reference product. Considering that lack of information about the split tablets can result in an inadequate therapy, problems related to captopril tablet splitting could be minimized by the distribution of splitting devices.

Column switching LC/MS methods were developed for rapid identification of impurities in cefdinir. Based on the mechanism by which cephalosporins are degraded, stress tests were designed and performed. It was found that eight main impurities were degradation products and five impurities originated from the synthesis process. Cefdinir bulk material was eluted as gradient on a C18 column, with 0.25% tetramethyl ammonium hydroxide solution (pH5.5)-acetonitrilemethanol as mobile phase. According to the retention time of different impurities, the target impurities were separated and enriched in another chromatographic column using column switching technology. Then the target impurity was desalted using an elution of 0.5% formic acid solution. Mass analysis was performed by elution with a mobile phase consisting of 1.0% formic acid-acetonitrile (3:7). The structures of ten related impurities were characterized on the bases of MS/MS data, general mass fragmentation pathway of cefdinir, and UV spectra. This study provides the material basis for the related substances of cefdinir in the Chinese Pharmacopeia 2010 edition. It also provides an effective means for the rapid identification of impurities in chromatographic systems containing non-volatile salt.

A specific ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was established and validated for the simultaneous quantification of paclitaxel prodrug (PP) and paclitaxel (PT) in human plasma. PP, PT and an internal standard 13C6-labelled PT (13C6-PT) were extracted from plasma by tert-butyl methyl ether and separated on an ACQUITY UPLC BEH C18 column (50 mm × 2.1 mm, 1.7 μm) using a mobile phase of methanolacetonitrile 50:50 (v/v) at a flow rate of 0.4 mL/min. At positive electrospray ionization mode, multiple reaction monitoring of the precursor-product ion transitions of m/z 882.2→313.9 for 13C6-PT, 876.2→307.9 for PT and 1216.5→647.8 for PP was used for the quantification. The linear calibration curve was obtained in a concentration range of 1-1000 ng/mL with a lower limit of quantification of 1 ng/mL. For both analytes, the values of intra- and inter-day precision were within 12.5% and accuracy fell in the ranges of 91.5-103.4%. The recovery ranged from 88.1% to 94.2% and the matrix effects from 89.3% to 95.4%. PP and PT were stable under short-term temperature and post-preparative conditions. The method was applied to stability of PP in human plasma and released PT from PP.

The purpose of this study was to develop a sensitive and rapid method for the determination of venlafaxine in human saliva. Blank saliva samples were obtained from healthy volunteers. Their extraction was carried out on SPE columns. Cartridges of the columns were washed with water and the adsorbate was eluted with methanol. The eluate was evaporated and the residue was dissolved in a mixture of acetonitrile and water (50:50, v/v). Venlafaxine was analyzed on a HPLC system with UV detection at 226 nm. The mobile phase was a mixture of acetonitrile and a phosphate buffer (30:70 v/v) at a flow rate of 1 mL min–1. Linearity of the calibration curve was obtained in the range of 1–1000 ng mL–1 of venlafaxine in saliva. The limit of detection was 1 ng mL–1 (S/N = 3) and the limit of quantification was 3 ng mL–1 (S/N = 10). The RSD values for intra–day study varied between 0.21 and 1.67% and those for inter–day study did not exceed 8.14%. The developed method was applied to the analysis of saliva samples without interference peaks and enabled determination of the venlafaxine concentration in depressed persons. The method can be useful for controlling intake of the drug by patients.

The aim of this work was to develop a microbiological assay of vancomycin employing microplates and kinetic reading mode, and to validate the developed method by assessing of specificity and selectivity, linearity, linear range, accuracy and precision parameters. Established conditions comprised a vancomycin standard-curve in concentrations from 0.6 to 1.0 μg mL-1, and soybean-casein broth inoculated with 5% of Bacillus subtilis (ATCC 6633) suspension. Satisfactory results were obtained after a 5 hour incubation. The developed method showed appropriate selectivity and linearity in the range from 0.6 to 1.0 μg mL-1, respectively, satisfactory accuracy (recovery = 97.0%) and precision (RSD = 7.8%).

Two different strategies were used to encapsulate oleanolic acid into nanoparticles, including nanoprecipitation and liposome technique. In nanoprecipitation method, the detailed formulations were investigated by varing the parameters of total good solvent/poor solvent volume ratio, surfactant mass ratio. The effects of different physical situations were considered in the experiment. The formulation which exhibits the most satisfactory colloidal stability and particles' formation was identified. The average diameter of nanoparticles is about 150 nm shown by transmission electron microscopy (TEM). Secondly, proliposome and nano-sized liposomes were prepared. The liposomes contain a hydrophobic oleanolic acid core, an amphiphilic soybean lecithin monolayer and a hydrophilic PEG protective coating. They are dispersed individually and distributed around 110-140 nm in diameters. Encapsulation enficiencies (EE) of the two methods were calculated by high performance liquid chromatograpy. The EE of nanoparticles obtained are 86.7% and 92.6%, respectively. Furthermore, the stability of nanoparticles was explored in different physicochemical situations. The results demonstrate that nanoparticles can possess the higher stability at 4°C.

Resolution of ternary and quaternary mixtures of paracetamol (PAR), desxtromethorphan (DEX), phenylephrine (PHE), and chlorpheniramine (CHL) was successfully achieved with minimum sample pre-treatment and without analyte separation using a rapid method based on partial least squares (PLS) and UV spectral data. Multivariate PLS calibration was performed using 35 ternary and quaternary mixtures and the model was employed for prediction of analyte in 15 external test samples. Simultaneous determination of the analytes was possible by analyzing absorbances between 210 and 300 nm. Q2 values which are a measure of the predictivity of the PLS models were between 0.9696 and 0.9880 in ternary mixtures and between 0.9230 and 0.9757 in quaternary mixtures. The PLS models were successfully applied to determine the analytes in real pharmaceutical preparations. The mean recoveries for the analytes in the real samples were between 95.25 and 102.4% in ternary and between 93.52 and 103.55% in quaternary mixtures.

A simple, sensitive and rapid isocratic reversed phase (RP) liquid chromatographic (LC) method was developed for the estimation of milnacipran in bulk and its formulations. Separations were carried out using the mobile phase consisting of 25mM phosphate buffer, acetonitrile and methanol on LiChrospher® 100 RP-18 analytical column at a flow rate of 1.0 mL/min and detection wavelength of 220 nm. The method developed as per the ICH guidelines, exhibited linearity over the analytical range of 25-3000 ng/mL with regression equation y = 801.1 x - 10767 [mean peak area = 801.1 concentration (ng/mL) - 10767] and regression coefficient, r2 = 0.999. The method demonstrated selectivity with no interfering peaks eluting in the vicinity of the drug peak and was consistent with good recovery. Forced degradation studies were also performed on the drug samples to demonstrate the stability indicating power of the method. The developed method being sensitive, precise and accurate in the studied range, can be employed for the routine estimation of in bulk and its formulations.

Quantification of pegylated drugs in biological matrices is often complex and challenging. This paper describes a sensitive HPLC method with fluorescence detection for the analysis of pegylated resveratrol, resveratrol and its metabolites. Solvent mediated fluorescence enhancement and quenching effects were explored to develop a highly sensitive HPLC method for analysis of resveratrol-PEG conjugates. The effect of solvent composition on fluorescence intensity of resveratrol-PEG conjugate was evaluated by varying the concentration of methanol in the solvent mixture. The fluorescence intensity of resveratrol-PEG was found to be dependent on the methanol concentration and was at maximum at 100% methanol. The HPLC assay method developed, with a linear gradient, allowed the maximum detection of the resveratrol- PEG conjugate with methanol at 95% of the mobile phase without affecting sepertion of resveratrol and its metabolites. The LOQ for resveratrol-PEG was 300 ng/mL (equivalent to 30 ng/mL resveratrol), nearly ten times more sensitive than HPLC with UV detection (3 μg/mL). The peaks detected for resveratrol-PEG, resveratrol and its metabolites in HPLC were identified qualitatively by LC/MS and LC-MS/MS (metabolites) and found to correlate with their respective molecular masses. The results of this study demonstrate that the developed HPLC assay method can accurately and selectively quantify resveratrol-PEG conjugates in the presence of resveratrol metabolites without affecting the sensitivity of resveratrol analysis. The application of solvent mediated fluorescence enhancement in HPLC potentially permits the analysis of, not only resveratrol-PEG conjugates, but also other resveratrol-polymer conjugates in samples obtained from in vitro or in vivo biological studies.

The antioxidant capacity of capsules containing blueberry based products which are included among the group of integrators owing to their antioxidant capacity and produced by various drug firms was investigated. The results of the investigation are compared to rank these products in order to their antioxidant capacity. In order to measure antioxidant capacity, our laboratory has recently developed a special electrochemical method based on a superoxide dismutase (SOD) biosensor to determine the superoxide radical. The results obtained by applying the SOD biosensor method to various blueberry based integrators were compared with the results obtained with the spectrophotometric (FRAP) method based on N,N-dimethyl-p-phenylenediamine (DMPD-FeCl3) and with those obtained also using the ORAC fluorimetric (TRAP) method. One of the more interesting aspects of the article is the good agreement it evidences of the results of the three methods for measuring antioxidant capacity. The three methods differ among themselves: an Electron Transfer (ET) method, a Hydrogen Atom Transfer Method (HAT) and an electrochemical based biosensor method of the Monitoring Superoxide Radical (MSR) type. It is also shown how the antioxidant capacity of the fresh vegetable is in any case always greater than that of any food supplement obtained from the same type of vegetable.

A comprehensive approach using multiple qualitative and quantitative methods coupled with multivariate statistical analysis was developed for the quality evaluation of Arnebiae Radix currently marketed in China. A simple, rapid and accurate liquid chromatographic method by optimization of the current ones was established for determination of β, β'-dimethylacrylalkannin. Moreover, hierarchical clustering analysis (HCA) and principle component analysis (PCA) were applied to classify and differentiate the different samples. In the qualitative analyses, two known markers (l-shikonin and β, β'-dimethylacrylalkannin) were detected in all samples. The quantitative analyses showed that these samples were qualified with the high contents of both total hydroxynaphthoquinones and β, β'-dimethylacrylalkannin. However, their chemical profiles from either TLC or HPLC analysis as well as the pharmacognostic features showed some differences from each other. By HCA and PCA, all samples especially imported ones could be clearly separated and classified into two major groups, of which four samples including two indigenous herbs and two imported ones from Pakistan and the authentic herb of Arnebiae euchromae Radix were grouped together but could be further differentiated by PCA study. Some potential markers besides l-shikonin and β, β'-dimethylacrylalkannin were found accountable for such discrimination but need to be further elucidated. This study demonstrated that a combination of qualitative and quantitative analyses with multivariate statistical analysis could be an efficient approach for the comprehensive quality evaluation of Arnebiae Radix.

Although there are many different methods for measuring the carbon monoxide and carbon dioxide content under biological conditions, not all of them are appropriate for detecting the very low concentrations. This review discusses the principles of the most commonly used methods for the detection and quantitative determination of these species, as well as the advantages and limitations of the methods described. The considered analytical methods are discussed in relation to their accuracy, precision and sensitivity together with the extent to which any interference can occur. The described methods for measuring carbon monoxide content under biological conditions fall into six main groups, while in the case of the quantitative determination of carbon dioxide the use of ten techniques is discussed.