Liquid Chromatography-tandem Mass Spectrometry Method Research Articles

Simultaneous determination and pharmacokinetics study of three triterpenoid saponins in rat plasma by ultra-high-performance liquid chromatography tandem mass-spectrometry after oral administration of Astragalus Membranaceus leaf extract.

A selective and sensitive ultra-high-performance liquid chromatography-tandem mass spectrometry method was developed and validated for the determination of three triterpenoid saponins isolated from Astragalus membranaceus leaf extract. In this article, a method for simultaneous determination of Huangqiyenin A, Huangqiyenin E, and Huangqiyenin K was established for the first time. The method was successfully applied to the pharmacokinetic study of Astragalus membranaceus leaf extract after oral administration. Liquid-liquid extraction was applied to plasma sample preparation. Multiple reaction monitoring mode with an electrospray ion source in positive electrospray ionization was chosen to quantify the analytes. Chromatographic separation was performed on a Waters HSS T3 column, using gradient elution with a mobile phase composed of acetonitrile and 5mM ammonium acetate/water. The pharmacokinetic results showed that all three compounds had the characteristics of rapid absorption-slow metabolism trend. The time of maximum plasma concentration of Huangqiyenin A is higher than Huangqiyenin E and Huangqiyenin K. And the maximum plasma concentration of Huangqiyenin A is higher as well. The pharmacokinetic results revealed the pharmacokinetic characteristics of the three analytes in rat plasma, which could provide a helpful reference for the further study of Astragalus membranaceus leaf extract.

An improved ultra-high performance liquid chromatography-tandem mass spectrometry method determining hispidulin and homoplantaginin in rat plasma and associated pharmacokinetic studies

AbstractFlavonoids are the most abundant components in Salvia plebeia, with significant pharmacological antioxidant and hepatoprotective properties. Hispidulin and homoplantaginin are the main flavonoid components in S. Plebeia. In this study, we established an ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) to determine hispidulin and homoplantaginin in rat plasma samples, which were precipitated using acetonitrile-methanol (9:1, v/v). We used a UPLC HSS T3 (100 mm × 2.1 mm, 1.7 μm diameter) chromatographic column, an acetonitrile-water (containing 0.1% formic acid) mobile phase, and a gradient elution flow rate of 0.4 mL min−1 in an elution time of 4 min. We used electrospray ionization (ESI) detection in positive ion mode, and multiple reaction monitoring mode (MRM) for quantitative analysis: m/z 301 → 286 for hispidulin, m/z 463 → 301 for homoplantaginin, and m/z 465 → 303 for internal standard (IS). In pharmacokinetic studies, 24 rats were orally administered hispidulin and homoplantaginin (5 mg kg−1) and received sublingual intravenous injections (1 mg kg−1) at two different doses, four groups with six rats/group. Differences in hispidulin and homoplantaginin pharmacokinetics in rat plasma were evaluated. The calibration curve showed good linearity in the 0.5–1,000 ng mL−1 range, with r > 0.99. Precision, accuracy, recovery, matrix effects, and stability results all met standard biological sample detection requirements. Our pharmacokinetic studies showed hispidulin bioavailability was much higher than homoplantaginin at 17.8% and 0.1%, respectively.

Simultaneous determination of seven bile acids to study the effect of ivermectin on their plasma levels in rat by UHPLC–MS/MS

Bile acids (BAs) are considered to be important diagnostic biomarkers to understand the pathophysiology of hepatobiliary and metabolic diseases. BAs regulate lipid and glucose metabolism by binding to farnesoid X receptor (FXR). To date, there were no reports on the effect of an exogenous FXR modulator, ivermectin (IVM), on the plasma BA profiles in rats. To explore the effect of IVM on plasma BA levels in rat, an ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC–MS/MS) method was developed and validated for simultaneous determination of seven major BAs in rat plasma. The developed method was selective, specific, accurate and precise for the quantification of plasma BAs. Sprague–Dawley rats were orally administered with IVM at a dose of 5 mg/kg once a day for 14 days and the plasma BAs were determined before and after IVM exposure using developed UHPLC–MS/MS method. Once-daily administration of IVM for 14 days resulted in significant reduction in cholic acid and deoxycholic acid levels while glycodeoxycholic acid and taurodeoxycholic acid levels were not affected. Interestingly, tauro-α-muricholic acid and tauro-β-muricholic acid levels were significantly increased. This study revealed that IVM has an important effect on plasma BA profiles in rats. This report provides an analytical methodology that can be applied to investigate the effect of drugs or pathophysiological factors on plasma BA levels.

Development and validation of an LC-MS/MS method for the determination of AZD7648 in rat plasma: Application to a pharmacokinetic study.

AZD7648 is a potent DNA-PK inhibitor that is being developed for the treatment of ovarian cancer. The study aimed to develop a simple and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to determine the concentration of AZD7648 in rat. AZD7648 was extracted from plasma by acetonitrile-mediated protein precipitation. The quantification was performed on a Thermo Vantage TSQ mass spectrometer with ibrutinib as an internal standard. A Waters Acquity UPLC BEH C18 column combined with 0.1% aqueous formic acid and acetonitrile was employed for chromatographic separation. The precursor-to-product ion transitions were m/z 421.2 > 337.2 and m/z 441.2 > 138.1 for AZD7648 and internal standard, respectively. This method was successfully validated according to the US Food and Drug Administration guidance. The calibration curve was linear over the concentration range of 0.5-1,000 ng/ml with correlation coefficient >0.999. The precision expressed as the coefficient of variation was <8.09%, while the accuracy expressed as relative error ranged from -10.00 to 9.08%. The mean recovery was >94.49%. AZD7648 was stable in rat plasma after storage under certain conditions. The validated method was demonstrated to be selective, sensitive and reliable, and has been successfully applied to the pharmacokinetic study of AZD7648 in rat plasma after oral and intravenous administration (1 mg/kg).

Development and validation of an ultra-high performance liquid chromatography-tandem mass spectrometry method for quantifying lenacapavir plasma concentrations: Application to therapeutic monitoring

Although current antiretroviral therapy (ART) effectively suppresses HIV in the blood, regimens may fail due to suboptimal treatment history and non-adherence to ART. In these scenarios, accumulation of viral resistance mutations to ART drug classes may occur. For these treatment-experienced people living with HIV (PLWH), activity against resistant viral strains is required; lack of therapeutic efficacy will result in continued viral replication and progression to acquired immunodeficiency syndrome. New treatment options have emerged. Lenacapavir is a first-in-class long-acting HIV-1 capsid inhibitor approved for the treatment of HIV in treatment-experienced patients. Lenacapavir is approved with an initiation regimen of oral and subcutaneous injection dosing followed by subcutaneous self-injection every 6 months. With infrequent dosing, therapeutic drug monitoring may be necessary to ensure adequate concentrations are consistently achieved in the plasma to assure treatment adherence and prevent further HIV resistance formation. To this end, we developed and validated a highly selective ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method to quantify lenacapavir concentrations in human plasma. A simple protein precipitation with acetonitrile followed by supernatant dilution was performed. Lenacapavir and its stable labeled internal standard were separated at 1.90 min using a multi-step UPLC gradient. The assay for lenacapavir quantification was extensively validated according to the United States Food and Drug Administration Bioanalytical Guidelines over a clinically relevant range of 0.1 to 500 ng/mL with excellent linearity (R2 ≥ 0.9960). This analytical method achieves acceptable performance of trueness (89.7–104.1 %), repeatability, and precision (CV < 15 %). We applied this method to quantify a clinical sample and to determine the percent protein-unbound. This method can be utilized for the therapeutic monitoring of lenacapavir in human plasma for monitoring HIV treatment efficacy.

An accurate and precise liquid chromatography–tandem mass spectrometry method for the determination of six phosphatidylethanol homologues in whole blood with phospholipid interferences minimized

Alcohol consumption is associated with a wide risk of different diseases, injury and death, and has significant social and economic consequences worldwide. Phosphatidylethanol (PEth) is a group of promising direct alcohol biomarkers, with a significantly longer half-life in blood than ethanol, which can be measured to predict different drinking patterns, such as heavy- and social drinking. This study aimed to develop and validate an accurate and precise LC-MS/MS method for the determination of six PEth homologues in whole blood with minimal interference from unwanted phospholipids. Different organic solvent mixtures for liquid-liquid extraction were investigated to obtain satisfactory recovery of PEth homologues and removal of the lyso-phospholipids and other early eluting phospholipids. The mixture of heptane/2-propanol (80:20, v:v) gave lower phospholipid background and better signal/noise values for the PEth peaks. An LC-MS/MS TQ-S system from Waters was used for the instrumental analysis. The main part of unwanted phospholipids were separated from the PEth homologues on an Acquity BEH C18 column (50 × 2.1 mm ID, 1.7 µm particles) using a buffer-free mobile phase of 0.025 % ammonia in Type 1 water, pH 10.7, as solvent A and methanol as solvent B. Validation and quantification of 22 authentic blood samples showed that the developed LC-MS/MS method is sensitive, precise and accurate for the determination of the six PEth homologues in whole blood. Lower limit of quantification was 10 nM for all compounds. No matrix effects were observed, possibly due to the successful strategies incorporated to avoid the influence of unwanted phospholipids.

Determination of restricted dyes in textile raw material solid wastes by ultra-high performance liquid chromatography-tandem mass spectrometry

A rapid and highly sensitive method for the quantification of 34 restricted dyes (including acid, basic, disperse, direct, and azo dyes) in solid textile raw material wastes was developed by employing ultrasonic extraction coupled with ultra-high performance liquid chromatography-tandem mass spectrometry(UHPLC-MS/MS). More specifically, the proposed method employed methanol as the extraction solvent, while the mobile phases consisted of acetonitrile and 10 mmol/L ammonium acetate + 0.05% ammonia. A good linearity was achieved over the concentration range of 0.01–200 ng/mL with correlation coefficients (R) between 0.991–0.999, limits of detection (LODs) of 0.25–40.0 µg/kg (S/N = 3) and limits of quantification (LOQs) of 0.84–133.4 µg/kg (S/N = 10). 34 dyes were recovered at three levels ranging from 84.5 to 106.9% with relative standard deviation (RSDs) ranging from 0.59% to 10.61%. Further, the method was applied for the accurate analysis of 32 counts of cotton yarn, waste cotton, and printed fabrics within 15 min. The dyestuffs accurately quantified by this rapid chromatographic procedure covered a wide range of carcinogenic and allergenic dyestuffs listed in the Oeko-Tex Standard 100 (version 02.2023) colourants. The ultrasound technique combined with the ultra-high performance liquid chromatography-tandem mass spectrometry method proposed in this work is thus suitable for the rapid screening, confirmation, and quantitative detection of industrial synthetic dyes within solid waste originating from textile raw materials.

Diverse associations observed between pregnancy complications and RBC or plasma folates determined by an in-house developed LC-MS/MS method

Background As folates are essential for embryonic development and growth, it is necessary to accurately determine the levels of folates in plasma and red blood cells (RBCs) for clinical intervention. The aims of this study were to develop and validate a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for quantitation of folates in plasma and RBCs and to examine the association between plasma and RBC folate concentrations and gestational diabetes mellitus (GDM), gestational hypertension (GH) and preeclampsia (PE). Methods With the in-house developed LC-MS/MS, a retrospective cross-sectional study was conducted. The healthy pregnant women of first- (n = 147), second- (n = 84) and third-trimester (n = 141) or the women diagnosed with GDM (n = 84), GH (n = 58) or PE (n = 23), that were aged between 22 and 46 years old and registered at our institute, were subjected for measurement of folic acid (FA) and 5-methyltetrahydrofolate (5-MTHF), followed by appropriate statistical association analysis. Results The assay for simultaneous quantitation of FA and 5-MTHF in plasma and RBCs was linear, stable, with imprecision less than 15% and recoveries within ±10%. The lower limits of quantification for FA and 5-MTHF measurement in whole blood were 0.57 and 1.09 nmol/L, and in plasma were 0.5 and 1 nmol/L, respectively. In the association analysis, the patients with lower RBC folate level (<906 nmol/L) presented higher risks of PE development (OR 4.861 [95% CI 1.411–16.505]) by logistic regression and restricted cubic spline (RCS) regression in a nonlinear fashion. In addition, higher level of plasma folates in pregnancy was significantly associated with GH risk but may be protective for the development of GDM. Conclusions The in-house developed LC-MS/MS method for folates and metabolites in plasma or RBC showed satisfactory analytical performance for clinical application. Further, the levels of folates and metabolites were diversely associated with GDM, GH and PE development.

Optimization of a ultra-high-pressure liquid chromatography-tandem multiple reaction monitoring mass spectrometry method for monitoring haloacetic acids as chlorinated disinfection by-products in the fresh-cut industry

Haloacetic acids (HAAs) are one of the most important chlorinated disinfection by-products generated during water disinfection in the fresh-cut industry, and they can remain in the product, resulting in a consumer health risk. In this study, ultra-high-pressure liquid chromatography-tandem multiple reaction monitoring mass spectrometry (UHPLC-MRM) analysis used for drinking water was optimized and applied for the quantification of nine HAAs (HAA9) in fresh-cut lettuce and process water samples, with the complex matrix interferences for separation, and quantification problems. The method showed good selectivity, specificity and linearity, satisfactory values for trueness (recoveries of 80–116 %), precision (<22 %), and uncertainty (<55 %). Quantification limits varied from 1 to 5 µg L−1 or µg kg−1. The matrix effect for tribromoacetic, bromochloroacetic and chlorodibromoacetic acid was corrected by matrix-matched calibration and standard addition. After storage at -20 °C, only monobromoacetic acid was the HAA which loss happened after 7 days. The application of the methodology in lettuce and process water samples from the industry was successfully implemented. Therefore, this method could be employed for the quality control and regulatory analysis of HAAs in fresh products and process water from the fruit and vegetable industry.

New sensitive LC-MS/MS method for the simultaneous determination of 13 phenolic and carboxylic acid pesticide biomarkers in human urine, including dicamba

The commercialization in 2016 of genetically engineered seeds tolerant to dicamba and/or 2,4-dichlorophenoxyacetic acid (2,4-D) has caused a rapid increase in the use of these herbicides. New questions about the reproductive and chronic health effects of long-term exposure to these herbicides have been raised. To assess exposure to dicamba and other pesticides of interest in the Heartland Study, a birth cohort study based in the United States, a new analytical method was needed. The present study describes the development and validation of this new solid phase extraction and liquid chromatography-tandem mass spectrometry method that detects simultaneously 13 pesticides or their metabolites in 250 μL of urine. More specifically, the method allows the analysis of dicamba, 2,4-D and 2,4,5-trichlorophenoxyacetic acid (2,4,5-T), which are herbicides, of malathion dicarboxylic acid (MDA), para-nitrophenol (PNP), 3,5,6-trichloro-2-pyridinol (TCPy), 2-diethylamino-6-methylpyrimidin-4-ol (DEAMPY) and 2-isopropyl-6-methyl-4-pyrimidinol (IMPY), which are metabolites of organophosphate insecticides, and finally of cis-3-(2,2-Dichlorovinyl)-2,2-dimethylcyclopropane carboxylic acid (cis-DCCA), trans-3-(2,2-Dichlorovinyl)-2,2-dimethylcyclopropane carboxylic acid (trans-DCCA), 3-Phenoxybenzoic acid (3-PBA), 4-Fluoro-3-phenoxybenzoic acid (4-F-3-PBA) and cis-3-(2,2-Dibromovinyl)-2,2-dimethylcyclopropane carboxylic acid (cis-DBCA), which are metabolites of synthetic pyrethroids insecticides. The method was validated under ISO/IEC 17025 guidance. The limit of detection (LOD) in urine samples was 0.10 μg/L for dicamba, while the LOD for other analytes ranged between 0.0038 μg/L and 0.091 μg/L. Accuracy was evaluated by analyzing samples from two External Quality Assessment Schemes, namely G-EQUAS and OSEQAS. Preliminary results obtained following the analysis of 91 urine samples taken from pregnant women enrolled in the Heartland Study are presented here. This method is suitable for human biomonitoring studies.

Development, validation, and implementation of an ultratrace analysis method for the determination of moenomycin A, in aquatic animal products.

Moenomycin A, an antimicrobial growth promoter widely used as an additive in aquaculture feedstuffs, has been restricted for use in the European Union and China due to its potential risk of promoting resistant strains of pathogenic bacteria and causing residues in aquatic animal products. Although methods for analyzing moenomycin A in feedstuffs have been developed, no established method exists for aquatic matrices. In this study, we present, for the first time, a sensitive and validated high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method for the determination of moenomycin A in aquatic animal products. Samples were extracted using methanol and purified with the QuEChERS method employing C18 sorbent. The aliquot was dried under a nitrogen stream, reconstituted with methanol-water solvent, and analyzed by HPLC-MS/MS. The developed method exhibited good linearity (r2 > 0.995) over a wide concentration range (1-100μg/L) and a low limit of detection (1µg/kg). Average recoveries ranged between 70 and 110% at spiked concentrations of 1, 50, and 100μg/kg, with associated intra- and inter-day relative standard deviations of 1.25 to 7.32% (n = 6) and 2.91 to 10.08% (n = 3), for different representative aquatic animal production, respectively. To the best of our knowledge, this is the first reported HPLC-MS/MS method for the quantification of moenomycin A in aquatic animal products. The new approach was effectively employed in the analysis of moenomycin A across various aquatic samples.

Correlations among the plasma concentrations of first-line anti-tuberculosis drugs and the physiological parameters influencing concentrations.

Background: The plasma concentrations of the four most commonly used first-line anti-tuberculosis (TB) drugs, isoniazid (INH), rifampicin (RMP), ethambutol (EMB), and pyrazinamide (PZA), are often not within the therapeutic range. Insufficient drug exposure could lead to drug resistance and treatment failure, while excessive drug levels may lead to adverse reactions. The purpose of this study was to identify the physiological parameters influencing anti-TB drug concentrations. Methods: A retrospective cohort study was conducted. The 2-h plasma concentrations of the four drugs were measured by using the high-performance liquid chromatography-tandem mass spectrometry method. Results: A total of 317 patients were included in the study. The proportions of patients with INH, RMP, EMB, and PZA concentrations within the therapeutic range were 24.3%, 31.5%, 27.8%, and 18.6%, respectively. There were positive associations between the concentrations of INH and PZA and RMP and EMB, but negative associations were observed between the concentrations of INH and RMP, INH and EMB, RMP and PZA, and EMB and PZA. In the multivariate analysis, the influencing factors of the INH concentration were the PZA concentration, total bile acid (TBA), serum potassium, dose, direct bilirubin, prealbumin (PA), and albumin; those of the RMP concentration were PZA and EMB concentrations, weight, α-l-fucosidase (AFU), drinking, and dose; those of the EMB concentration were the RMP and PZA concentrations, creatinine, TBA and indirect bilirubin; and those of the PZA concentration were INH, RMP and EMB concentrations, sex, weight, uric acid and drinking. Conclusion: The complex correlations between the concentrations of the four first-line anti-TB drugs lead to a major challenge in dose adjustment to maintain all drugs within the therapeutic window. Levels of TBA, PA, AFU, and serum potassium should also be considered when adjusting the dose of the four drugs.

Development of enantioselective high-performance liquid chromatography-tandem mass spectrometry method for the quantitative determination of 3,4-methylenedioxy-methamphetamine (MDMA) and its phase-1 metabolites in human biological fluids

In the present study enantioselective high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods were developed for the quantitative determination of 3,4-methylenedioxy-methamphetamine (MDMA) and its major phase-1 metabolites 4-hydroxy-3-methoxyamphetamine (HMA), 4-hydroxy-3-methoxymethamphetamine (HMMA) and 3,4-methylenedioxyamphetamine (MDA) in human plasma, sweat, oral fluid (OF) and urine. The simultaneous separation of all these compounds and their respective enantioseparation was accomplished on two polysaccharide-based chiral columns. The Lux AMP column with a proprietary chiral selector enabled baseline separation of the enantiomers of MDMA, HMA and HMMA while MDA enantiomers could not be separated with this column under the experimental conditions used in this study. The Lux i-Amylose-3 column based on amylose tris(5-chloro-3-methylphenylcarbamate) as chiral selector baseline-separated the enantiomers of MDMA, HMMA and MDA while the enantiomers of HMA could not be separated. Thus, the various samples were analyzed by using both columns alternatively in combinations with acetonitrile containing 25% (v/v) 5 mM ammonium bicarbonate buffer at pH 11.0 as mobile phase. Analysis time was less than 4 min with the Lux AMP column and less than 6 min with the Lux i-Amylose-3 column. Both methods were validated and applied to the enantioselective determination of MDMA and its phase-I metabolites in human biological fluids, and enantioselective metabolism of MDMA was confirmed.

SAT628 Effect Of Hepatic Impairment On The Pharmacokinetics, Safety, And Tolerability Of Oral Paltusotine, A Non-peptide, Selective Somatostatin Receptor Subtype 2 Agonist

Abstract Disclosure: R. Luo: Employee; Self; Crinetics Pharmaceuticals Inc. A. Casagrande: Employee; Self; Crinetics Pharmaceuticals Inc. S. Oun: Employee; Self; Crinetics Pharmaceuticals Inc. Y. Wang: Employee; Self; Crinetics Pharmaceuticals Inc. R. Struthers: Employee; Self; Crinetics Pharmaceuticals Inc. A. Krasner: Employee; Self; Crinetics Pharmaceuticals Inc. Paltusotine is an investigational oral, once-daily, nonpeptide, small molecule somatostatin receptor type 2 agonist in clinical development for the treatment of acromegaly and neuroendocrine tumors. This multicenter, open-label, phase 1 study assessed the influence of varying degrees of hepatic impairment on the pharmacokinetics (PK), safety, and tolerability of paltusotine. Participants with mild, moderate, and severe hepatic impairment based on Child-Pugh classification and matched healthy participants received a single 20 mg oral dose of paltusotine after an overnight fast of at least 10 hours. Analysis of paltusotine in plasma was performed using a validated liquid chromatography-tandem mass spectrometry method. Geometric mean ratios of PK parameters were calculated from an analysis of variance (primary analysis) with hepatic condition (normal, mild, moderate, severe) as a fixed effect and the log transformed PK parameter as the dependent variable. A sensitivity analysis (matched pairs mixed models for repeated measures) accounted for participant age and BMI. Overall, 36 participants were included (mild [n=8], moderate [n=8], and severe [n=6] hepatic impairment; matched healthy [n=14]). Most participants were male (72.2%); mean age was 57.3 years, and mean BMI was 30.7 kg/m2. Following single oral administration of 20 mg paltusotine, the mean plasma concentration of paltusotine peaked rapidly, with the median time to maximum concentration ranging from 1.6 to 3.0 hours postdose across all groups. The maximum plasma concentration (Cmax) and total plasma exposure (area under the curve from time 0 extrapolated to infinity [AUC0-inf]) of paltusotine were similar across all hepatic impairment groups when compared with healthy participants as indicated by the geometric mean ratio (90% CI) values (participants with mild, moderate, and severe hepatic impairment: Cmax: 1.35 [0.79-2.31], 0.76 [0.45-1.31], and 1.05 [0.58-1.90], respectively; AUC0-inf: 1.00 [0.60-1.66], 0.75 [0.45-1.24], and 0.90 [0.51-1.57], respectively). Results of the sensitivity analysis were consistent with those from the primary analysis. Overall, the most common (≥5% of participants) treatment-emergent adverse events (TEAEs) were diarrhea (16.7%) and headache (5.6%). The incidence of TEAEs was similar across all 4 groups. No participant discontinued the study because of a TEAE, and no serious drug-related TEAEs were reported. In conclusion, the peak and total plasma exposures and safety and tolerability profile of paltusotine were similar across participants with varying degrees of hepatic impairment compared with participants with normal hepatic function. There were no changes in paltusotine plasma exposure that would be considered clinically meaningful or sufficient to warrant dose adjustment for mild, moderate, or severe hepatic impairment. Support: Crinetics Pharmaceuticals. Presentation: Saturday, June 17, 2023

Development of a novel enantioselective high-performance liquid chromatography–mass spectrometry method for the differentiation of dextro- and levo-methorphan and their O-demethylated metabolites in human blood and its application to post-mortem samples

Recently we proposed an isocratic enantioselective high-performance liquid chromatography-tandem mass spectrometry (LC–MS/MS) method for the separation and quantitative determination of dextro- (DXM) and levo-methorphan (LVM) and their pharmacologically relevant metabolites, dextrorphan and levorphanol, respectively, in human blood samples. This method was based on the polysaccharide-based chiral column Lux AMP, a specialty column characterized with high stability in mobile phases of pH 11.0 and above. The use of a single-source column is a limitation for any analytical method. Therefore, the major goal of the present study was to develop an enantioselective method for the differentiation of dextro- and levo-methorphan, as well as their metabolites dextrorphan and levorphanol, using Lux Cellulose-3 as alternative chiral column with methanol containing 0.1 % diethylamine mobile phase. A newly developed method uses a chiral selector part of HPLC columns available from multiple manufacturers and a fairly common mobile phase. The method was validated and applied to post-mortem blood samples. Out of 50 analyzed new samples, dextromethorphan (DXM) was detected in 17 samples. Of these 17 cases DXM was accompanied with LVM in 7 samples. The proposed analytical method is relatively simple, accurate and fast and can be adopted for routine use in forensic and clinical toxicology laboratories.

Pharmacokinetics of a New Neuroprotector - Indenoquinoxalinone Derivative after Intravenous Administration in Rabbits and Rats.

The specific JNK inhibitor and NO donor 11H-indeno[1,2-b]quinoxalin-11-one oxime (IQ-1) demonstrated pronounced neuroprotective properties in an in vivo model of ischemic stroke in rats. The pharmacokinetic behavior of IQ-1 was studied in two animal species (rats, rabbits) after intravenous administration in a dose of 1 mg/kg. IQ-1 concentrations in venous blood plasma were measured by the liquid chromatography-tandem mass spectrometry method. The pharmacokinetics of IQ-1 was adequately described by the two-compartmental model. The calculated C0 for IQ-1 in rabbit and rat plasma were 2239.83±1229.55 and 1552.50±182.23 ng/ml, respectively. Two animal species are characterized by extensive tissue distribution of IQ-1 (Vss exceeded the total body water in rabbits and rats by 3.6 and 5.6 times, respectively) and high clearance values (88-94% of hepatic blood flow).

Correlation of drug exposure and bacterial susceptibility with treatment response for Mycobacterium avium complex lung disease: protocol for a prospective observational cohort study

IntroductionThe burden of Mycobacterium avium complex (MAC) lung disease is increasing globally and treatment outcome is in general poor. Therapeutic drug monitoring has the potential to improve treatment outcome by...

INVESTIGATION OF RECENTLY ABUSED DRUGS IN DRIED BLOOD SPOTS USING FTA CARDS BY LIQUID CHROMATOGRAPHY TANDEM MASS SPECTROMETRY: VALIDATION AND APPLICATION TO REAL SAMPLES

A robust and selective liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was established for the analysis of 19 illicit drugs and metabolites in whole blood dried blood spots (DBS) using FTA cards. Samples prepared using Ahlstrom Munksjö Gensaver™ Colorcards were extracted with methanol. Analytical separation of the analytes was maintained using an Agilent Poroshell column with mobile phase A (0.1% formic acid in water) and B (methanol). Multiple Reaction Monitoring (MRM) with positive ionisation on LC-MS/MS was used to develop the method. The method was validated and met the acceptance criteria with acceptable results for carry-over, linearity, specificity, sensitivity, accuracy, precision, matrix effect and recovery. The method was applied to positively reported whole blood samples from patients suspected of drug abuse. Good quantitative agreement was obtained between the DBS and whole blood methods. Application to real DBS samples showed that this method is a good alternative and useful technique for the detection of drugs of abuse in forensic toxicology and appears to provide a good alternative storage condition.

Simultaneous determination of spiropidion and its five major metabolites in sweet orange fruit and various processing by-products using ultra-high performance liquid chromatography-tandem mass spectrometry

The present work reported the application of an ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method for simultaneous analysis of spiropidion and its five major metabolites in sweet orange fruit and by-products throughout the whole industrial juicing process of the orange fruit. The reversed-dispersive solid phase extraction (r-DSPE) with multi-walled carbon nanotubes (MWCNTs) was employed for the extraction and purification. The established method was validated and satisfactory parameters (linearity, trueness, precision, sensitivity, matrix effect and stability) were obtained. And then, the field trial of spiropidion on sweet oranges has been conducted and the effect of commercial juicing processing on the residue of spiropidion and its metabolites was further investigated. The various processing factors (PFs) for washing, juicing, sterilization, concentrating and essential oil collecting were also determined. The final results indicated that washing processing reduced residues by 18.4%; the juicing step allowed a significant decrease of the spiropidion residue by 34.2–70.8%, with PFs value in the range of 0.290–0.658. However, high level of residual spiropidion (ranging from 4.016 to 4.205 mg/kg) was detected in orange essential oil, with PFs value of 17.157. All the above results demonstrated the efficiency of the established method in the routine control analysis of spiropidion residues in sweet orange fruits and their by-products, and will facilitate the further intensive research on its spatial distribution, transfer and degradation during the different processing procedures of the sweet orange fruits.

Determination of Caffeine in Human Plasma by Using Liquid Chromatography-Tandem Mass Spectrometry

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated to determine caffeine (CAF) in human plasma. The plasma samples were extracted by protein precipitation using CAF-D3 as an internal standard (IS). The chromatographic separation was performed on GL Sciences InertSustain C18 Column (4.6 x 50 mm, 5 µm) maintained at 40 °C with a mobile phase consisting of formic acid, water, and methanol at a 1 mL/min of flow rate using two separate lines. CAF was detected and identified by mass spectrometry with electrospray ionization (ESI) in positive ions and multiple-reaction monitoring (MRM) mode. The MRM transitions of m/z 195.10 &amp;gt; 138.00 for CAF and 198.10 &amp;gt; 141.10 for IS were used for quantification. The standard curve was linear in the range of 10 - 10000 ng/mL for CAF. The within-batch precision and accuracy were evaluated by analyzing QC samples at five different concentration levels with six replicates in a batch. The between-batch precision and accuracy were determined by analyzing three different batches. The within-batch accuracy and precision was -8.76% - 9.61% and 0.95% - 7.22%, respectively. The between-batch accuracy and precision was -7.47% -1.42% and 1.83% - 8.66%, respectively. The results of the intra- and inter-day precision and accuracy studies were within the limits. The validated method applied to a pharmacokinetic study and the test product containing 60 mg CAF administered to total of 12 subjects. The mean ± SD of maximum plasma concentration (Cmax) was found to be 147.94 ± 139.39 ng/mL and the mean ± SD of area under the plasma concentration-time curve from zero to last measurable concentration (AUC0-tlast) was found to be 1119.59 ± 1468.30 h.ng/mL for the fasting conditions. The median time to reach peak plasma concentration (Tmax) was found to be 12.00 (6.50 - 12.00). The developed and validated method can be used for bioavailability and bioequivalence studies in human plasma samples.