Current Pharmaceutical Analysis - Volume 18, Issue 6, 2022

Background: A pharmaceutical will be clinically accepted if it is impurity-free and its dose is accurately maintained. For this, the contribution of analytical techniques for developing and validating a new pharmaceutical dosage form cannot be overlooked. Introduction: Tolterodine tartrate is a potent competitive muscarinic receptor antagonist. It binds to the muscarinic M3 receptors in the bladder selectively and competitively. It is used to treat urinary incontinence and overactive bladder syndrome. The 5-hydroxymethyl derivative is the pharmacologically active metabolite of Tolterodine tartrate, which is as potent as the main drug. It is available with α-adrenergic blocker Tamsulosin in combined pharmaceutical formulations, treating benign prostatic hyperplasia in men. This review article presents several analytical methods, including HPLC, HPTLC, UV-Visible spectrophotometry, spectrofluorimetry, electroanalytical, and various hyphenated techniques like GC-MS, LC-MS, LC-MS-MS, etc., for estimation of Tolterodine tartrate is present as a single material or in combined form in bulk materials, different pharmaceutical formulations, and biological matrices. Conclusion: HPLC and spectrophotometry are the most widely used methods for determining the drug in the bulk and pharmaceutical dosage form among all these methods. LC-MS and LC-MSMS are widely used for the estimation of Tolterodine tartrate from plasma and other biological fluids. All the methods included in this article are accurate, sensitive, and cost-effective.

Background: High-Performance Liquid Chromatography (HPLC)-Ultraviolet (UV) and Liquid Chromatography (LC)-Mass Spectrometry (MS)/MS methods have been used to analyse abiraterone (ART); however, a single-quadrupole mass spectrometer with LC-MS systems has never been used to analyse ART. Objective: The study aimed to establish a novel, simple assay of quantitating ART in rat plasma through LC-MS. Methods: The analytical procedure involved the extraction of ART and D4-ART (internal standard, IS) from rat plasma through simple protein precipitation. Chromatographic separation was achieved using an isocratic mobile phase (acetonitrile: 5 mM ammonium formate with 0.1% formic acid, 50:50 v/v) at a flow rate of 0.30 mL/min on a Waters XBridge® C18 column with a total run time of 5 min. LC-MS ion transitions monitored were 350.1 and 354.1 for ART and IS, respectively. The method was validated, and the results met acceptance criteria. Results: The lower limit of quantitation achieved was 1 ng/mL, and linearity was 1-8000 ng/mL. The intra- and inter-day precisions were 1.26%-14.20% and 5.49%-13.08%, respectively, in rat plasma. Conclusion: LC-MS offers a novel, specific, sensitive, and accurate method for quantifying ART and it was successfully applied to pharmacokinetic studies of ART in rats.

Background: To optimize therapy for patients with human immunodeficiency virus-tuberculosis (HIV-TB) coinfection, we developed an ultra-high-performance liquid chromatography/- tandem mass spectrometry (UHPLC-MS) method to monitor seven second-line anti-tuberculosis drugs. Methods: Blood samples (n = 70) were collected from 35 patients with HIV-TB coinfection; the plasma sample was protein-precipitated and diluted with a solution containing heptafluorobutyric acid. The plasma concentrations of rifabutin (RBT), clofazimine (CLO), moxifloxacin (MFX), prothionamide (PTH), levofloxacin (LFX), amikacin (AMK), and para-aminosalicylic acid (PAS) were detected by UHPLC-MS/MS method. Results: In these 70 samples, the mean concentrations of RBT, CLO, MFX, PTH, LFX, and AMK were 173.8 (10.0–550.0), 61.1 (54.4–67.7), 646.6 (25.0–2480.0), 120.5 (50.0–597.0), 1565.9 (100.0–3480.0), and 10753.0 (400.0–76 700.0) μg/L, respectively. Only one sample was detected to have PAS with a concentration less than the lower limit of quantification. Most of the drug concentrations detected in these patients were lower than the targeted concentrations in TB patients. Conclusion: We created a simple UHPLC-MS method for simultaneously quantifying anti-TB drugs. The plasma concentrations in HIV-TB co-infected patients were lower than the targeted concentrations. It is important to monitor anti-TB drugs in the future. This method will facilitate the monitoring of anti-TB drugs in the future.

Background: Various illegal phosphodiesterase-5 (PDE5) inhibitors have been indiscriminately added to food and dietary supplements since the early 2000s without evaluating their side effects or pharmaceutical effects. Objective: The metabolic pathway of benzamidenafil was evaluated using liquid chromatography quadrupole time-of-flight mass spectrometry (LC-Q-ToF-MS). Methods: In vitro metabolic samples were prepared using human liver microsomes, and in vivo metabolic samples were obtained from drug-administered rats. The metabolic samples were extracted to remove biological interferences and injected into the LC-Q-ToF-MS system for analysis. Their mass spectra and chromatograms were compared and interpreted. Results: A total of nine metabolites were evaluated in the benzamidenafil-treated samples. All error values were within 5 ppm with high accuracy. Conclusion: These newly identified metabolites of benzamidenafil could be used to construct relevant legal databases and applied to related forensic cases.

Background: Lenvatinib is a potent drug utilized in the medication of thyroid cancer and it acts as a tyrosine kinase inhibitor. Thus, the development and validation of Lenvatinib and allied impurities in rat plasma, and its pharmacokinetic study, are one of the most significant areas of modern pharmaceutical analysis. Objective: The current study conducts bioanalytical system validation and pharmacokinetic analysis of Lenvatinib and associated impurities in rat plasma with LC-MS/MS. Methods: The current study involves bioanalytical system validation and pharmacokinetic analysis of Lenvatinib and associated impurities in rat plasma using LC-MS/MS. Gradient elution of Lenvatinib with a flow rate of 1 mL/min and an X-Bridge phenyl column (150x4.6 mm, 3.5μ) was used in the optimized process. In this method, buffer (1 mL formic acid in 1 liter of water) and acetonitrile mixture was used as the mobile phase. Results: By using Carfilzomib as the internal norm and impurity-4 as the active metabolite and 30 minute run time, Lenvatinib and its associated impurities were separated. The linearity was in the range of 10 percent to 200 percent of rat plasma, and each analyte R² value was found to be 0.999. Conclusion: This work indicates that all parameters, such as precision, recovery, accuracy, and stability, were achieved as per USFDA guidelines. This approach can be used to investigate Lenvatinib impurities and conduct pharmacokinetic studies involving rat plasma.

Objective: A simple, sensitive, and accurate in-vitro dissolution method has been developed for Olmesartan Medoxomil (OLM), Chlorthalidone (CHLR) & Cilnidipine (CIL) drug combination according to USP dissolution testing methodologies with different discriminating mediums and validated as per ICH guidelines. Methods: The in-vitro dissolution profile was obtained using 900 ml of phosphate buffer pH 6.8 with 1.0% SLS at 37 °C ± 0.5 °C as dissolution medium and USP II (paddle) at 75 rpm. The average % in-vitro drug release was above 80% within 45 minutes for the above drug combination. The drug release profile was evaluated by RP-HPLC method. Chromatographic separation was done on Hypersil-BDS C-18 (12.5cm x 4.6mm x 5μm) column using gradient program with initial mobile phase ratio of 55:45 (v/v) mixture of ammonium acetate buffer (pH 5.0) and acetonitrile at a flow rate of 1.0 ml/min with detection wavelength 260 nm. Results: The method was validated with respect to specificity, linearity, precision, accuracy, and robustness. The method was found to be linear in the range of 7.0-21.0 μg/ml for CHLR (R² = 0.9982), 22.5-67.5 μg/ml for OLM (R² = 0.9999) and 5.5 -16.5 μg/ml for CIL (R² = 0.9995) respectively. The % recovery data were found between 98.3 % to 104.1%. The % RSD for method and intermediate precision of method did not exceed more than 2%. Conclusion: The proposed in-vitro method can be applied successfully for routine quality control analysis to check the quality of above drug combination.

Background: (S)-oxiracetam has shown better efficacy than racemic oxiracetam in the trial. However, there is no report on the method used to detect the genotoxic impurity of S-(+)- epichlorohydrin(S-(+)-ECH) that may be contained in (S)-oxiracetam. Objective: Here, we have proposed and verified a detailed analysis method for the (S-(+)-ECH) impurity in (S)-oxiracetam using a gas chromatography(GC) system. Methods: The chromatography system includes a Supelco DB-624 capillary column. The initial column temperature was 50°C, kept for 4 minutes, and then increased to 220°C at a rate of 10°C/min for 4 minutes. Nitrogen was used as the carrier gas. The constant flow rate was 3.8 mL/min, the split ratio was 10:1, the injection port temperature was maintained at 200°C, and a hydrogen flame ionization detector was used. S-(+)-ECH was analyzed in detail by this analysis method, and three different batches of samples were measured. The GC system was used for verification. Results: The method verification results included the system suitability, specificity, sensitivity, accuracy, precision, and the good correlation coefficient between peak responses and concentrations were over 0.990. Conclusion: The method proposed in this study is the most detailed GC method currently used for the analysis of S-(+)-ECH impurities in (S)-oxiracetam. The proposed method has the advantages of fast, convenience, low cost, etc. It provides a reference method for quality control of genotoxic impurity of (S)-oxiracetam.

Background: Hybrid polymeric materials have been in research focus owing to their outstanding progression in drug targeting. A new Quality by Design approach by RP-HPLC was developed and validated for the estimation of hybrid polymeric nanoparticles of Ritonavir in rat plasma. Objective: The main objective of the present study was to develop and validate a simple, robust, and accurate method by QbD approach for the detection of hybrid polymeric nanoparticles of Ritonavir (RTV) in plasma. Methods: The mobile phase consisting of a mixture of Acetonitrile: HPLC grade water (60:40 v/v), 1.0 ml/min flow rate and UV detection at 240 nm. Critical Analytical Attributes (CAAs) were screened and selected by Taguchi orthogonal array model. Box-Behnken's three-level, the 3- factorial design, was employed to create and analyze a "Design Space" (DoE). This design was statistically analyzed by ANOVA, contour-plot, and 3D response surfaces plots, which demonstrated that the model was statistically significant. The developed method was validated as per the ICH guidelines Q2 (R1). Results: The developed method showed excellent linearity between 100 and 600 ng/mL with good regression (R2>0.998), LOD (10 ng/mL) and LOQ (30 ng/mL). The validation results of the tested parameters were found within the acceptable limit. Conclusion: From the study, it was concluded that QbD driven bioanalytical method is suitable for the in-vitro and in-vivo estimation of RTV from bulk as well as from hybrid polymeric nanoparticles formulation.