Journal of Pharmaceutical and Biomedical Analysis Open最新文献

Metronidazole (MTZ) is a broad-spectrum antibiotic with numerous routes of administration, including topical. Topical application of MTZ gel or cream results in very low systemic absorption, resulting in the need for a sensitive extraction method to quantify plasma concentrations. Currently published methods are not suitable for analysis of plasma concentrations after topical application, as undetectable MTZ concentrations commonly occur. We validated a simple extraction method for MTZ recovery from plasma and quantified it using an LC-MS/MS analytical method. Methods: Plasma samples were spiked with MTZ (0.5 – 5 ng/mL) and internal standard (tinidazole, 2 ng/mL). MTZ was extracted by liquid-liquid extraction using ethyl acetate and acetonitrile mixture (4:1) as the extraction solvent. A quadrupole mass spectrometer interfaced with an Acquity H-Class HPLC was used to quantify MTZ concentrations in positive ion mode. A Kinetix C18 analytical column (150 mm × 4.6 mm i.d., 5 µm particle size) was used for separation. The plasma extraction method was validated for various parameters, including % recovery, precision, accuracy, and stability. Results: The extraction method demonstrated high MTZ recovery, ranging from 93.7 – 97.5%. The calibration curve prepared using MTZ samples extracted from plasma (0.5 – 5 ng/mL) had excellent linearity with R² = 0.999. The extracted samples also showed higher autosampler and freeze-thaw stability over a 72-hr period. The mean intra- and inter-day accuracy and precision of the extraction assay ranged from 97 to 101.6% and 2.7 – 4.8% RSD, respectively. The assay was highly efficient, with a limit of quantification (0.53 ± 0.04 ng/mL) lower than previously published methods (≥5 ng/mL). The extraction method was successfully validated using LC-MS/MS and can be used to extract and detect trace amounts of MTZ in plasma after topical application.

A biosimilar considered as a biomolecule medicinal product that is comparable to a reference medicinal product in terms of the structural, functional, biological, and clinical attributes. Glurazyme® was developed as a biosimilar to Cerezyme® (imiglucerase) and was approved in CIS countries (Russian Federation, Belarus, Kazakhstan) and recently Algeria for the treatment of type 1 and type 3 Gaucher disease. The quality assessment of Glurazyme® was performed in accordance with the Rules for the Study of Biological Medicines of the Eurasian Economic Union harmonized with the ICH comparability guideline and the biosimilar guidelines of the European Medicines Agency and Food and Drug Administration. Extensive side-by-side comparison was employed with state-of-the-art and orthogonal assays designed to interrogate all expected physicochemical and biological activities, including those known to affect the mechanisms of action for imiglucerase. Similarity evaluation was performed on the basis of tolerance intervals determined from about 10 lots of commercial Cerezyme®. Mainly three discrepancies of quality attributes were established concerning oxidized and deamidated forms as well as phosphorylated oligomannose N-glycans reflecting the difference between cultivation and down-stream processes of both medicinal products. Nevertheless, all of them possess a little or no influence on safety and efficacy.

Polysaccharide activation by periodate oxidation to form aldehydes, followed by conjugation with proteins via reductive amination is a general procedure to make polysaccharide-protein conjugate vaccines. Controlling the level of residue aldehyde content after conjugation is critical to ensure vaccine product stability. Herein, to support conjugation process optimization, we developed a 3-methyl-2-benzothiazolone hydrazone (MBTH) chemistry-based colorimetric method, which can measure low-level residual aldehyde polysaccharide-protein conjugate in high throughput 96-well plate format. This mechanism of detection works in two steps. First, MBTH reacts with the aldehyde group to form azine. Then the excess MBTH was oxidized by ferric ammonium sulfate in sulfamic acid to form a reactive cation, which reacts further with azine to form formazan, a characteristic blue chromophore at 610 nm, for colorimetric detection. The method performance, including detection limit, linearity range, matrix effect, kinetics, and color stability was systematically studied. For samples with severe matrix interference, a supplemental size exclusion chromatography (SEC) method was also developed as an alternative method.

Diabetic kidney disease (DKD) onset and progression is a major cause of end-stage renal failure in diabetic patients, however, no practical method has been reported to predict the progression rate of renal function decline. Nine serum compounds are reported to associate with prognosis in type 1 diabetes patients; however, quantitative analytical methods for these compounds lacks. Herein, we developed a simultaneous quantitative method for 15 compounds, including Niewczas’s nine biomarker candidates, associated with renal function and its prognosis in kidney disease patients to achieve a prognostic method of renal function decline in DKD patients. This report describes the development and validation of a LC–MS/MS analytical method for 15 compounds of biomarker candidates using human plasma, serum, and urine as sample matrices. The analytes are N-acetyl-L-alanine, N6-acetyl-L-lysine, N-acetyl-L-serine, N-acetyl-L-threonine, phenyl sulfate, pseudouridine, N6-threonylcarbamoyladenosine, tryptophan 2-C-mannoside, tyrosine O-sulfate, creatinine, p-cresol sulfate, 4-ethylphenyl sulfate, indoxyl sulfate, N1-methyladenosine, and trimethylamine N-oxide. The Capcell Pak ADME-HR column was compared to several general columns and selected as the most suitable column for the simultaneous analysis of all 15 compounds. The proposed method was validated for selectivity, accuracy, precision, stability, dilution integrity, and parallelism. This report describes the suitability of the calibration ranges established and the actual sample concentrations of serum and urine from type 2 diabetic patients, as well as new findings on the unknown analyte levels of several compounds in these samples. The proposed method can be used to aid the development of prognostic methods for renal function decline in patients with DKD.

There has been a growing concern over the contamination of pharmaceutical products with nitrosamines (NAs) such as N-nitrosodimethylamine (NDMA). To quantify NA levels in drugs using reversed-phase liquid chromatography (LC), the sample solution should achieve a high drug concentration to detect trace NAs, and an appropriate amount of hydrophilic NAs should be retained. However, these are difficult to achieve, and no suitable method has yet been developed. The present study was the first to develop a sample preparation method to achieve this by combining drugs with formic acid (FA), followed by the removal of active pharmaceutical ingredients (APIs) from samples via crystallization. This method was successfully applied for the sensitive quantification of eight NAs in poorly water-soluble acidic atorvastatin (ATS) and basic itraconazole (ITC) via LC–ultraviolet (LC-UV) detection. The removal rate of ITC via recrystallization exceeded 99.96 %, whereas most NAs remained as solutes. Assuming that the enhancement in ITC solubility directly translates to heightened analytical sensitivity, a > 100-fold increase in sensitivity was attained compared to conventional methodologies. This sample preparation method would be applicable to other poorly water-soluble drugs, contributing to the control of NA content in various formulations to realize the safe delivery of pharmaceuticals to patients.

Over the last two decades, surface plasmon resonance (SPR) sensors have advanced significantly, becoming an important tool in disciplines such as biosensing, chemical sensing, and material characterization. SPR has gained popularity in biosensing because of its great sensitivity and specificity in detecting biomolecular interactions. This review provides an overview of the recent developments of the SPR biosensor technology and its applications in medical diagnostics. To provide an up-to-date overview of the area, the review includes the most recent works from the last decade. Furthermore, it explores various configurations (prism, grating, fiber optic, waveguide modulated) and wave properties (angle, wavelength, phase) being tracked for sensing together with strategies for enhancing sensitivity and selectivity through mechanisms such as surface coatings, sensing mediums, and immobilization techniques.

Tinospora cordifolia (TC) is known for its immense therapeutic applications in 'Ayurveda', which has created considerable scientific interest in pharmacology. Thus, a targeted and rapid bioanalytical UHPLC-ESI-MS/MS method was developed and validated for the extraction of its bioactive markers from diverse classes of diterpenes as tinosporide (TC1) and phytosterol-20-β-hydroxyecdysone (TC2) and isoquinoline alkaloids-jatrorrhizine (TC3), tetrahydropalmatine (TC4) from the TC stem extract (TCE) in rat plasma by solid phase extraction technique (SPE). The optimum recovery (≥ 90 %) was achieved for TC1–4 and internal standard fluoxymesterone (IS) with the SPE method on the C18 phase. The analytes were subjected to chromatographic analysis on the Agilent C18 Zorbax Eclipse Plus column (4.6 × 100 mm, 3.5 µ) with a gradient program using 0.1 % acetic acid in water (% v/v) and acetonitrile as mobile phase at a flow rate of 0.500 mL/min. The MS/MS quantification and validation were performed on the Shimadzu 8045 tandem mass spectrometer associated with the heated-ESI probe in SRM mode. The precursor to product ion transitions m/z 416.20→375.10 (TC1), 481.40→445.20 (TC2), 339.15→323.05 (TC3), 356.25→192.10 (TC4) and 337.20→91.00 (IS) were used for quantification. Also, in silico ADMET prediction and in vivo pharmacokinetics revealed that TC1–4 was well absorbed from the GI tract and could act as a P-GP substrate. In the pharmacokinetic study, TC1–4 could be detected by this validated bioanalytical method. The TC1 was found bioavailable, having an optimum half-life of > 9.0 h to exhibit therapeutic activity with other TC bioactive markers in vivo. This research is the first comprehensive study on in silico and in vivo pharmacokinetics of TC biomarkers, which may aid in further pre-clinical and clinical trial investigations.

Inflammatory long non-coding RNAs (lncRNAs) including PACER (p50-associated COX-2 extragenic RNA) and NKILA (NF-Kappa B Interacting long non-coding RNA) and have recently emerged as essential regulators in immune and inflammation pathways in patients with multiple sclerosis (MS), which have possible worthiness as diagnostic, prognostic, and therapeutic targets. In the current study, we aimed to evaluate the expressions of PACER, NKILA lncRNAs, CTCF (CCCTC-binding factor), and NF-κB in the PBMC (peripheral blood mononuclear cells) from MS patients and control subjects. We detected the expression levels of PACER, CTCF, NKILA, and NF-kB using real-time PCR in 39 MS patients and 37 healthy controls. The findings show that the expression levels of PACER lncRNA and CTCF, but not NKILA lncRNA and NF-κB, have significantly decreased in MS patients compared to the controls, so they are probably involved in MS pathogenesis. Notably, the area under the ROC curve for PACER and CTCF was up to 0.80 and 0.68, respectively. Based on these observations, the PACER and CTCF had been shown to have the best efficiency in the discrimination of disease status between MS patients and healthy controls. A low expression level of CTCF was correlated with PACER lncRNA and NF-KB expression levels as well as some factors including disease duration, age, and EDSS. Altogether, these results demonstrate that PACER lncRNA and CTCF were potentially related to the MS risk. However, further functional investigations are required to confirm the roles of these genes in the etiology of MS.

The study aimed to develop a supercritical fluid chromatography-tandem mass spectrometry (SFC-MS/MS) method for the enantioselective separation and quantification of pantoprazole in rat plasma. The method utilized protein precipitation with acetonitrile to prepare plasma samples. SFC was performed on an Acquity ultra-performance convergence chromatography (UPCC) system. An Acquity UPCC Trefoil™ CEL2 column was employed for enantioseparation, and isocratic elution was achieved using a mobile phase consisting of CO₂-methanol (81:19, v/v). The detection of pantoprazole enantiomers and the internal standard phenacetin was carried out using a Xevo TQD triple quadrupole mass spectrometer in selected reaction monitoring mode with positive electrospray ionization. The developed method successfully achieved full separation of S- and R-pantoprazole enantiomers. Calibration curves were linear in the concentration range of 10–5000 ng/mL for both S- and R-pantoprazole. The method met the acceptance criteria for carry-over, accuracy, precision, extraction recovery, matrix effect, stability, and dilution integrity, demonstrating its stereoselectivity, sensitivity, accuracy, precision, and reliability. The validated method was then applied to investigate the toxicokinetics of racemic pantoprazole and evaluate the potential of chiral inversion between S- and R-pantoprazole in rats. Following oral administration of 200 mg/kg racemic pantoprazole once daily for 90 consecutive days, there was no significant difference in toxicokinetic parameters between the two enantiomers on both day 1 (single-dose experiment) and day 90 (multiple-dose experiment). This suggests that there is no enantioselectivity in the toxicokinetic behaviors of racemic pantoprazole in rat plasma. Additionally, bidirectional chiral inversion between S- and R-pantoprazole in plasma was observed after single-dose and multiple-dose oral administrations of S-pantoprazole (20, 100, and 200 mg/kg) or R-pantoprazole (200 mg/kg) to rats. Compared to single and low doses, there was no accumulation of each enantiomer in rat plasma following repeated and escalated dosing. Overall, this study provided the first report on the toxicokinetics of pantoprazole in rat plasma and presented the first SFC-MS/MS method for the simultaneous quantification of pantoprazole enantiomers in rat plasma.

Cocoa shell is a by-product from cocoa industry which contains bioactive compounds of high and attractive value for food, pharmaceutical and cosmetics industry. However, cocoa shell can be contaminated by undesirable materials that, even in small amounts, would not change the color, aroma, and taste characteristics of the final product. Identification and prediction of this impurity are performed using expensive methods that require chemicals and produce residues. Thus, this work aims to investigate the performances of benchtop (867–2535 nm) and portable (900–1700 nm) near-infrared (NIR) spectrometer for fast prediction of cocoa shell powder impurities. Mixtures (n = 432) of cocoa shell powders with leaves, pods, stem fragments and nibs at several proportions (0–20 % w/w), were analyzed. Multivariate calibration models were developed using partial least-squares regression (PLSR) with raw spectra and various preprocessing approaches applied to the spectra. The most informative spectral variables were selected by variable importance in projection (VIP) method. Results obtained for the benchtop (R²_P> 0.99 and RMSEP<0.71) and low-cost portable (R²_P> 0.92 and RMSEP<1.70) devices are promising, and portable spectrometer could be used to certify cocoa shell purity.