Capillary electrophoresis-mass spectrometry in impurity profiling of pharmaceutical products

Generally reversed-phase high-performance liquid chromatography (RP-HPLC) methods are extensively applied during quality control of pharmaceutical products. Since capillary electrophoresis (CE) is based on a different separation principle and consequently results in a unique selectivity compared to RP-HPLC, it can advantageously be used as an orthogonal technique. CE equipped with a mass spectrometer detector provides even more information that can be helpful for identification and structural elucidation purposes. CE-MS was recently implemented in the method development approach to support impurity profiling of pharmaceutical products. In this paper the application of CE-electrospray ionization (ESI)-MS/MS to the impurity profiling of galantamine hydrobromide in stressed Reminyl Extended Release (ER) capsules is discussed. Reminyl ER samples were stressed at different storing conditions. The impurity profile of these samples was compared with the current RP-HPLC and chiral CE method, but also with CE-ESI-MS/MS. The combination of these three methods provided valuable data that allowed understanding comprehensively the impurity profile of these samples. Two impurities were detected at concentrations lower than 0.05%, which did not occur in nonstressed samples. Chromatographic data and the fragmentation patterns of galantamine and related compounds were also examined for identification of these two degradation products.     

Determination of captopril in pharmaceutical preparation and biological fluids using two- and three-way chemometrics methods

Spectrophotometric method has been developed for the direct quantitative determination of captopril in pharmaceuticalpreparation and biological fluids(human plasma and urine)samples.The method was accomplished based on parallel factoranalysis(PARAFAC)and partial least squares(PLS).The study was carried out in the pH range from 2.0 to 12.8 and with aconcentration from 0.70 to 61.50 μg mL~(-1)of captopril.Multivariate calibration models such as PLS at various pH and PARAFACwere elaborated from ultraviolet spectra deconvolution and captopril determination.The best models for this system were obtainedwith PARAFAC and PLS at pH 2.0.The applications of the method for determination of real samples were evaluated by analysis ofcaptopril in pharmaceutical preparations and biological fluids with satisfactory results.The accuracy of the method,evaluatedthrough the RMSEP,was 0.5801 for captopril with best calibration curve by PARAFAC and 0.6168 for captopril with PLS at pH 2.0model.     

Improvement of the decision efficiency of the accuracy profile by means of a desirability function for analytical methods validation. Application to a diacetyl-monoxime colorimetric assay used for the determination of urea in transdermal iontophoretic extracts

Validation of analytical methods is a widely used and regulated step for each analytical method. However, the classical approaches to demonstrate the ability to quantify of a method do not necessarily fulfill this objective. For this reason an innovative methodology was recently introduced by using the tolerance interval and accuracy profile, which guarantee that a pre-defined proportion of future measurements obtained with the method will be included within the acceptance limits. Accuracy profile is an effective decision tool to assess the validity of analytical methods. The methodology to build such a profile is detailed here. However, as for any visual tool it has a part of subjectivity. It was then necessary to make the decision process objective in order to quantify the degree of adequacy of an accuracy profile and to allow a thorough comparison between such profiles. To achieve this, we developed a global desirability index based on the three most important validation criteria: the trueness, the precision and the range. The global index allows the classification of the different accuracy profiles obtained according to their respective response functions. A diacetyl-monoxime colorimetric assay for the determination of urea in transdermal iontophoretic extracts was used to illustrate these improvements.     

Assay of cisapride in pharmaceutical formulations by extraction spectrophotometry

Four simple and sensitive spectrophotometric methods (A-D) for the assay of cisapride in pure and dosage forms based on the formation of chloroform soluble ion-associates under specified experimental conditions are described. Four acidic dyes, namely, Suprachen Violet 3B (SV 3B, method A), Erioglaucine A (EG-A, method B), Naphthalene Blue 12 BR (NB-12BR, method C) and Tropaeolin 000 (TP 000, method D) are utilized. The extracts of the ion-associates exhibit absorption maxima at 595, 640, 620 and 500 nm for methods A, B, C and D, respectively. Beer's law and the precision and accuracy of the methods are checked by the UV reference method. The results are reproducible with an accuracy of +/-1.0%. The methods are found to be suitable for the determination of cisapride in the presence of the other ingredients that are usually present in dosage forms.     

Validation of UV spectrophotometric and nonaqueous titration methods for the determination of carvedilol in pharmaceutical formulations

Ultraviolet (UV) spectrophotometric and nonaqueous volumetric methods are described for the determination of carvedilol in pharmaceutical formulations. Linearity, precision, and accuracy were evaluated according to the validation guidelines of the International Conference on Harmonization and the United States Pharmacopeia for both methods. The UV spectrophotometric procedure was performed in ethanol at 244 nm. Good linearity was obtained between 2 and 7 microg/mL with a correlation coefficient of 0.9999. The intra- and interday precision values were <2% for all samples analyzed. The accuracy, determined from recovery studies, was between 97.5 and 102.2%. The other procedure was based on the volumetric quantitation of carvedilol in a nonaqueous medium with 0.01 M perchloric acid and 1% violet crystal as the indicator. The validation of the volumetric method yielded good results that included linearity (r of > 0.999), precision (relative standard deviations of <2% for intra- and interday precision), and accuracy (96.4-102.4%). The methods were applied to tablets and compounded capsules. Statistical analysis by analysis of variance showed no significant difference between the results obtained by the proposed methods.     

Place of DSC purity analysis in pharmaceutical development

DSC purity analysis is based on thermodynamic phase diagrams for substances (purity ≥98%) which undergo a melting point. Impurities which have eutectic behaviour with the analyte are determined together. DSC purity analysis obtained from a single melting event of a 1–2 mg sample is, therefore, extremely attractive for the global assessment of eutectic impurities. The main advantages in early development lie in the very small amount of material necessary and the very fast analysis time. However, the DSC purity analysis cannot replace chromatographic methods which deliver specific individual levels of impurities. Furthermore, a complete validation of a DSC purity method is difficult and time consuming. Despite these limitations, DSC is the best support for the development of chromatographic methods, for purity profile and stability assessment during pharmaceutical development. Parameters of purity determination and validation aspects are discussed. Examples of use in pharmaceutical development are given.     

Active content determination of non-coated pharmaceutical pellets by near infrared spectroscopy: Method development, validation and reliability evaluation

A robust near infrared (NIR) method able to quantify the active content of pilot non-coated pharmaceutical pellets was developed. A protocol of calibration was followed, involving 2 operators, independent pilot batches of non-coated pharmaceutical pellets and two different NIR acquisition temperatures. Prediction models based on Partial Least Squares (PLS) regression were then carried out. Afterwards, the NIR method was fully validated for an active content ranging from 80 to 120% of the usual active content using new independent pilot batches to evaluate the adequacy of the method to its final purpose. Conventional criteria such as the R², the Root Mean Square Error of Calibration (RMSEC), the Root Mean Square Error of Prediction (RMSEP) and the number of PLS factors enabled the selection of models with good predictive potential. However, such criteria sometimes fail to choose the most fitted for purpose model. Therefore, a novel approach based on accuracy profiles of the validation results was used, providing a visual representation of the actual and future performances of the models. Following this approach, the prediction model using signal pre-treatment Multiplicative Scatter Correction (MSC) was chosen as it showed the best ability to quantify accurately the active content over the 80-120% active content range. The reliability of the NIR method was tested with new pilot batches of non-coated pharmaceutical pellets containing 90 and 110% of the usual active content, with blends of validation batches and industrial batches. All those batches were also analyzed by the HPLC reference method and relative errors were calculated: the results showed low relative errors in full accordance with the results obtained during the validation of the method, indicating the reliability of the NIR method and its interchangeability with the HPLC reference method.     

Flow-through chloroquine sensor and its applications in pharmaceutical analysis.

Poly (vinyl chloride) membrane electrodes that responded selectively towards the antimalarial drug chloroquine are described. The electrodes were based on the use of the lipophilic potassium tetrakis(4-chlorophenyl)borate as ion-exchanger and bis(2-ethylhexyl)adipate (BEHA), or trioctylphosphate (TOP) or dioctylphenylphosphonate (DOPP) as plasticizing solvent mediator. All electrodes produced good quality characteristics such as Nernstian- and rapid responses, and are minimally interfered with by the alkali and alkaline earth metal ions tested. The membranes were next applied to a flow-through device, enabling it to function as flow-injection analysis (FIA) detector. The performance of the sensor after undergoing the FIA optimization was further evaluated for its selectivity characteristics and lifetime. Results for the determination of chloroquine in synthetic samples that contained common tablet excipients such as glucose, starch, and cellulose, and other foreign species such as cations, citric acid or lactic acid were generally satisfactory. The sensor was also successfully used for the determination of the active ingredients in mock tablets, synthetic fluids and biological fluids. The sensor was applied for the determination of active ingredients and the dissolution profile of commercial tablets was also established.     

Spectrophotometric determination of glimepiride in pharmaceutical preparations based on the formation of charge-transfer and ion-pair complexes

Two rapid, simple, accurate and sensitive spectrophotometric methods were developed for the determination of glimepiride in pharmaceutical preparations. The first method was based on the formation of a charge-transfer complex of the drug, as n-electron donor, with 7,7,8,8-tetracyanoquinodimethane (TCNQ), as π-acceptor. The second method was based on the formation of ion-pair complexes between the examined drug and bromothymol blue (BTB). The proposed methods were validated for linearity, limit of detection, limit of quantification, precision, accuracy, robustness and specificity. The calibration was linear over the concentration range of 10–80 and 20–120 μg/mL for methods I and II, respectively. The limits of detection were 2.6 and 2.8 μg/mL. The proposed methods were applied to the determination of the drug in pharmaceutical preparations. The results obtained were in good agreement with those obtained using the reference method (HPLC). There was no significant difference in the accuracy and precision as revealed by the accepted values of t- and F-tests, respectively.     

Sublimation measurements of pharmaceutical compounds by isothermal thermogravivletry

Procedures for measuring sublimation rates of pharmaceutical compounds by isothermal thermogravimetry are discussed. Experimental data was obtained using the Mettler TA4000 thermogravimetric system. The sublimation rate is measured directly from the mean weight loss per unit time in the linear region of the monitored TG profile at a set isothermal temperature. This data when fitted to the Arrhenius equation yields the sublimation enthalpy. For the benzoic acid reference, the enthalpy so calculated is 99% of the value obtained from direct vacuum TG measurements. Thermal degradation in the solid state or pre-melting can effect a departure from the characteristic linear mass loss-time sublimative profile. Data pertaining to several established Merck drugs is discussed. Examples where loss of residual solvent, onset of thermal degradation and pre-melting phenomena affect the measurement, are presented.     

Highly sensitive spectrofluorimetric determination of ephedrine hydrochloride in pharmaceutical preparations

A sensitive and specific spectrofluorimetry method was developed and validated for the quantification of ephedrine (EP) in pharmaceutical preparations. The method is based on the fluorescent enhancing reaction of EP with 7-chloro-4-nitrobenzofurazan (NBD-C1; derivatization reagent), in borate buffer of pH 9 to yield a yellow, fluorescent product. Under these experimental conditions, the derivatized product of EP had excitation and emission wavelength maxima at 458 and 516 nm, respectively. The linear range of this method was 20-2500 ng/mL. The detection limit was 7.3 ng/mL EP. Intra- and interday precisions of the assay at 3 concentrations within this range were 0.037-1.77%. The low relative standard deviation values indicate good precision, and high recovery values indicate excellent accuracy of the method. The proposed method was applied to the determination of the examined drugs in pharmaceutical formulations, and the results indicate that the method is equally as accurate, precise, and reproducible as the official method.     

Wavelength selection framework for classifying food and pharmaceutical samples into multiple classes

Near infrared (NIR) spectroscopy is an efficient, low‐cost analytical technique widely applied to identify the origin of food and pharmaceutical products. NIR spectra‐based classification strategies typically use thousands of equally spaced wavelengths as input information, some of which may not carry relevant information for product classification. When that is the case, the performance of predictive and exploratory multivariate techniques may be undermined by such noisy information. In this paper, we propose an iterative framework for selecting subsets of NIR wavelengths aimed at classifying samples into categories. For that matter, we integrate Principal Components Analysis (PCA) and three classification techniques: k‐Nearest Neighbor (KNN), Probabilistic Neural Network (PNN) and Linear Discriminant Analysis (LDA). PCA is first applied to NIR data, and a wavelength importance index is derived based on the PCA loadings. Samples are then categorized using the wavelength with the highest index and the classification accuracy is calculated; next, the wavelength with the second highest index is inserted into the dataset and a new classification is performed. This forward‐based iterative procedure is carried out until all original wavelengths are inserted into the dataset used for classification. The subset of wavelengths leading to the maximum accuracy is chosen as the recommended subset. Our propositions performed remarkably well when applied to four datasets related to food and pharmaceutical products. Copyright © 2016 John Wiley & Sons, Ltd.     

Parafac and PLS applied to determination of captopril in pharmaceutical preparation and biological fluids by ultraviolet spectrophotometry

A new ultraviolet spectrophotometric method has been developed for the direct qualitative determination of captopril in pharmaceutical preparation and biological fluids such as human plasma and urine samples. The method was accomplished based on parallel factor analysis (PARAFAC) and partial least squares (PLS). The study was carried out in the pH range from 2.0 to 12.8 and with a concentration from 0.70 to 61.50 microg ml(-1) of captopril. Multivariate calibration models PLS at various pH and PARAFAC were elaborated from ultraviolet spectra deconvolution and captopril determination. The best models for this system were obtained with PARAFAC and PLS at pH = 2.04 (PLS-PH2). The applications of the method for the determination of real samples were evaluated by analysis of captopril in pharmaceutical preparations and biological (human plasma and urine) fluids with satisfactory results. The accuracy of the method, evaluated through the root mean square error of prediction (RMSEP), was 0.58 for captopril with PARAFAC and 0.67 for captopril with PLS-PH2 model. Acidity constant of captopril at 25 degrees C and ionic strength of 0.1 M have also been determined spectrophotometrically. The obtained pKa values of captopril are 3.90 +/- 0.05 and 10.03 +/- 0.08 for pKa. and pKa2, respectively.     

PARAFAC Decomposition of Three‐Way Kinetic‐Spectrophotometric Spectral Matrices Based on Phosphomolymbdenum Blue Complex Chemistry for Nitrite Determination in Water and Meat Samples

Abstract

A three‐way analytical methodology experimentally based on kinetic‐spectrophotometric and parallel factor analysis (PARAFAC) chemometrics analysis was assessed for the quantification of nitrite in water and meat samples. This method is based on the reduction of phosphomolybdic acid to phosphomolymbdenum blue complex by sodium sulfide. The obtained phosphomolymbdenum blue complex is oxidized by the addition of nitrite and this causes a reduction in intensity of the blue color. Three‐way data matrices were generated by acquisition of ultraviolet‐visible (UV‐Vis) spectra (600–900 nm) as a function of the time and of different relative concentration of the nitrite (0.10–2.10 µg mL^?1). The PARAFAC trilinear model, without restrictions, was used in the data analysis. A full decomposition of the data matrices was obtained (spectra, concentration, and time profile). It was shown that kinetic methods coupled to three‐way chemometrics analytical methods can be used for the development of robust sensors for the analysis of nitrite in water and meat samples. The accuracy of the method, evaluated through the root mean square error of prediction (RMSEP), was 0.0515 and 0.1181 for nitrite by PARAFAC and partise least squares (PLS) models respectively. The results with the PARAFAC model are better than those of the PLS model, according to results, it being possible to recover the spectra and kinetic profiles, as well as the initial concentration of nitrite with good accuracy.