Development and Application of a LC-MS/MS Method for Simultaneous Quantification of Four First-Line Antituberculosis Drugs in Human Serum

A simple, rapid, and sensitive liquid chromatography (LC)/mass spectrometry (MS) method was established and validated for simultaneous quantitation of pyrazinamide, isoniazid, rifampicin, and ethambutol in human blood sample. Samples were pretreated by a single-step precipitation with acetonitrile. Chromatographic separation was achieved on XSelecT HSS T3 column by gradient elution with a total run time of 5.0 min. MS detection was performed by a triple quadrupole tandem mass spectrometer in the multiple reaction monitoring mode with a positive electrospray ionization source. Isotope-labeled internal standard, especially rifampicin-D8, was applied to adjust for the loss during sample treatment. The established LC-MS/MS method showed a wide analytical range (pyrazinamide: 1.02∼60.0 μg/mL, isoniazid: 0.152∼10.0 μg/mL, rifampicin: 0.500∼30.0 μg/mL, and ethambutol: 0.0998∼5.99 μg/mL) and a good linearity (r > 0.99 for the four analytes) with acceptable accuracy and precision (90.15%∼104.62% and 94.00%∼104.02% for intra- and interaccuracy, respectively; RSD%: <12.46% and <6.43% for intra- and interprecision, respectively). It also showed excellent recoveries (79.24%∼94.16% for all analytes) and absence of significant matrix effect. This method was successfully applied to the quantification of four first-line antituberculosis (anti-TB) drugs, suggesting its suitability for therapeutic drug monitoring in the clinical practices.

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A UPLC-MS/MS Assay for Simultaneous Determination of Two Antipsychotics and Two Antidepressants in Human Plasma and Its Application in Clinic

Current Pharmaceutical Biotechnology ◽

10.2174/1389201020666190830150549 ◽

2020 ◽

Vol 21 (1) ◽

pp. 60-69

Author(s):

Houli Li ◽

Xiaoliang Cheng ◽

Di Zhang ◽

Maoyi Wang ◽

Weihua Dong ◽

...

Keyword(s):

Simultaneous Determination ◽

Human Plasma ◽

High Performance ◽

Medication Compliance ◽

Multiple Reaction Monitoring ◽

Gradient Elution ◽

Individual Therapy ◽

Therapeutic Drug ◽

Ionization Source

Background: Antidepressants and antipsychotics are widely prescribed drugs for the treatment of mental diseases. Therapeutic drug monitoring (TDM) is recommended for patients taking these drugs to ensure pharmaceutical efficacy, medication compliance and prevent toxicity. Objective: An ultra-high performance liquid chromatography/tandem-mass spectrometry (UPLC-MS/ MS) method was developed for simultaneous determination of two Antidepressants-Fluoxetine (FLU) and Escitalopram (ESC), and two antipsychotics-risperidone (RIS) and aripiprazole (ARI), in human plasma. Methods: The sample was processed by simple protein precipitation and the targeted analytes were separated on a C18 column by gradient elution with a mobile phase containing 0.1% formic acid (v/v) and acetonitrile. All the analytes were qualitative and quantitative measured by electrospray ionization source with Multiple Reaction Monitoring (MRM) in positive ion mode. A total of 56 plasma samples were obtained from out- or in-patients who were taking the cited four drugs for further analysis. Results: The calibration curves for FLU, ESC, RIS and ARI were linear in the range of 45-1800, 4-320, 2-200 and 50-1800 ng/mL, respectively. The entire analytical time for the analytes was 7.0 min for each run and the extraction efficiency was more than 90%. The sample was stable within various storage conditions. The trough concentrations in patients were measured with the validated method. Conclusions: The developed method was successfully used for simultaneous determination of FLU, ESC, RIS and ARI in the plasma of the patients, which provides effective technical support for routine TDM of these four drugs and is of great clinic value for individual therapy.

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Development of a liquid chromatography tandem mass spectrometry method for the simultaneous measurement of voriconazole, posaconazole and itraconazole

Annals of Clinical Biochemistry International Journal of Laboratory Medicine ◽

10.1177/0004563216686378 ◽

2017 ◽

Vol 54 (6) ◽

pp. 686-695 ◽

Cited By ~ 2

Author(s):

John M Wadsworth ◽

Anna M Milan ◽

James Anson ◽

Andrew S Davison

Keyword(s):

Therapeutic Drug Monitoring ◽

Drug Monitoring ◽

Matrix Effects ◽

Multiple Reaction Monitoring ◽

Internal Standard ◽

Gradient Elution ◽

Turnaround Time ◽

Therapeutic Drug ◽

Tandem Mass ◽

Lower Limits

Background Azole-based antifungals are the first-line therapy for some of the most common mycoses and are now also being used prophylactically to protect immunocompromised patients. However, due to variability in both their metabolism and bioavailability, therapeutic drug monitoring is essential to avoid toxicity but still gain maximum efficacy. Methods Following protein precipitation of serum with acetonitrile, 20 µL of extract was injected onto a 2.1 × 50 mm Waters Atlantis dC18 3 µm column. Detection was via a Waters Quattro Premier XE tandem mass spectrometer operating in ESI-positive mode. Multiple reaction monitoring (MRM) detected two product ions for each compound and one for each isotopically labelled internal standard. Ion suppression, linearity, stability, matrix effects, recovery, imprecision, lower limits of measuring interval and detection were all assessed. Results Optimal chromatographic separation was achieved using gradient elution over 8 minutes. Voriconazole, posaconazole and itraconazole eluted at 1.71, 2.73 and 3.41 min, respectively. The lower limits of measuring interval for all three compounds was 0.1 mg/L. The assay was linear to 10 mg/L for voriconazole (R2 = 0.995) and 5 mg/L for posaconazole (R2 = 0.990) and itraconazole (R2 = 0.991). The assay was both highly accurate and precise with % bias of voriconazole, posaconazole and itraconazole, respectively, when compared with previous NEQAS samples. The intra-assay precision (CV%) was 1.6%, 2.5% and 1.9% for voriconazole, posaconazole and itraconazole, respectively, across the linear range. Conclusion A simple and robust method has been validated for azole antifungal therapeutic drug monitoring. This new assay will result in a greatly improved sample turnaround time and will therefore vastly increase the clinical utility of azole antifungal drug monitoring.

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Development and Validation of an Up-to-Date Highly Sensitive UHPLC-MS/MS Method for the Simultaneous Quantification of Current Anti-HIV Nucleoside Analogues in Human Plasma

Pharmaceuticals ◽

10.3390/ph14050460 ◽

2021 ◽

Vol 14 (5) ◽

pp. 460

Author(s):

Amedeo De Nicolò ◽

Alessandra Manca ◽

Alice Ianniello ◽

Alice Palermiti ◽

Andrea Calcagno ◽

...

Keyword(s):

Human Plasma ◽

Multiple Reaction Monitoring ◽

Internal Standard ◽

Reversed Phase ◽

Therapeutic Drug ◽

People Living With Hiv ◽

Simultaneous Quantification ◽

Working Solution ◽

Tenofovir Alafenamide ◽

Combination Regimens

Therapeutic options to treat HIV infection have widened in the past years, improving both effectiveness and tolerability, but nucleoside reverse transcriptase inhibitors (NRTIs) are still considered the standard backbone of the combination regimens. Therapeutic drug monitoring (TDM) can be useful for these drugs, due to concentration–effect relationship, with risk of ineffectiveness, toxicity or adherence concerns: in this scenario, robust and multiplexed methods are needed for an effective TDM activity. In this work, the first validated ultra-high spectrometry liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS) method is described for the high-sensitive simultaneous quantification of all the currently used NRTIs in human plasma, including tenofovir alafenamide (TAF), following FDA and EMA guidelines. The automated sample preparation consisted in the addition of an internal standard (IS) working solution, containing stable-isotope-linked drugs, protein precipitation and drying. Dry extracts were reconstituted with water, then, these underwent reversed phase chromatographic separation: compounds were detected through electrospray ionization and multiple reaction monitoring. Accuracy, precision, recovery and IS-normalized matrix effect fulfilled guidelines’ requirements. The application of this method on samples from people living with HIV (PLWH) showed satisfactory performance, being capable of quantifying the very low concentrations of tenofovir (TFV) in patients treated with TAF.

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Determination of sarecycline by UPLC-MS/MS and its application to pharmacokinetic study in rats

Acta Chromatographica ◽

10.1556/1326.2020.00796 ◽

2020 ◽

Author(s):

Yonghui Shen ◽

Deru Meng ◽

Feifei Chen ◽

Hui Jiang ◽

Liming Hu ◽

...

Keyword(s):

Pharmacokinetic Study ◽

Multiple Reaction Monitoring ◽

Internal Standard ◽

Gradient Elution ◽

Rat Plasma ◽

Chronic Inflammatory Disease ◽

Linear Calibration ◽

Limit Of Quantification ◽

Acquity Uplc ◽

Intravenous And Oral Administration

AbstractSarecycline is a narrow-spectrum antibiotic for the treatment of acne, which is a chronic inflammatory disease of the hair follicle sebaceous glands. In the study, UPLC-MS/MS was used to establish a rapid and accurate analytical method. The sarecycline was determined with poziotinib as internal standard (IS) in rat plasma. An ACQUITY UPLC HSS T3 column (2.1 × 100 mm, 1.8 μm) could performe chromatographic separation with the mobile phase (methanol: water of 0.1% formic acid) with gradient elution. The ions of target fragment were m/z 488.19→410.14 for sarecycline and m/z 492.06→354.55 for poziotinib, which could quantify the electrospray ionization of positive multiple reaction monitoring (MRM) mode. The linear calibration curve of the concentration range was 1–1,000 ng/mL for sarecycline with a lower limit of quantification (LLOQ) of 1 ng/mL. The mean recovery was between 82.46 and 95.85% for sarecycline and poziotinib in rat plasma. RSD for precision of inter-day and intra-day were between 3.24 and 13.36%, and the accuracy ranged from 105.26 to 109.75%. The developed and validated method was perfectly used in the pharmacokinetic study and bioavailability of sarecycline after intravenous and oral administration in rats.

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Development and Validation of a Sensitive UHPLC-MS/MS Method for the Measurement of Gardneramine in Rat Plasma and Tissues and its Application to Pharmacokinetics and Tissue Distribution Study

Molecules ◽

10.3390/molecules24213953 ◽

2019 ◽

Vol 24 (21) ◽

pp. 3953 ◽

Cited By ~ 1

Author(s):

Zhao ◽

Tan ◽

Chen ◽

Sun ◽

Wang ◽

...

Keyword(s):

Multiple Reaction Monitoring ◽

Indole Alkaloid ◽

Internal Standard ◽

Gradient Elution ◽

Rat Plasma ◽

Limit Of Quantification ◽

Tissue Samples ◽

Distribution Study

As a novel monoterpenoid indole alkaloid, gardneramine has been confirmed to possess excellent nervous depressive effects. However, there have been no reports about the measurement of gardneramine in vitro and in vivo. The motivation of this study was to establish and validate a specific, sensitive, and robust analytical method based on UHPLC-MS/MS for quantification of gardneramine in rat plasma and various tissues after intravenous administration. The analyte was extracted from plasma and tissue samples by protein precipitation with methanol using theophylline as an internal standard (I.S.). The analytes were separated on an Agilent ZORBAX Eclipse Plus C18 column using a gradient elution of acetonitrile and 0.1% formic acid in water at a flow rate of 0.3 mL/min. Gardneramine and I.S. were detected and quantified using positive electrospray ionization in multiple reaction monitoring (MRM) mode with transitions of m/z 413.1→217.9 for gardneramine and m/z 181.2→124.1 for I.S.. Perfect linearity range was 1–2000 ng/mL with a correlation coefficient (r2) of ≥0.990. The lower limit of quantification (LLOQ) of 1.0 ng/mL was adequate for application to different preclinical studies. The method was successfully applied for determination of gardneramine in bio-samples.

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P63 UPLC/MS/MS assay for the simultaneous determination of seven antibiotics in human serum – application to pediatric studies

Archives of Disease in Childhood ◽

10.1136/archdischild-2019-esdppp.101 ◽

2019 ◽

Vol 104 (6) ◽

pp. e43.2-e43

Author(s):

S Magreault ◽

O Chaussenery-Lorentz ◽

T Storme ◽

E Jacqz-Aigrain

Keyword(s):

Multiple Reaction Monitoring ◽

Internal Standard ◽

Ultra Performance Liquid Chromatography ◽

Routine Activity ◽

Therapeutic Drug ◽

Pharmacokinetic Studies ◽

Pediatric Dose ◽

Liquid Chromatography Tandem Mass ◽

Sampling Volume ◽

Acquity Uplc

BackgroundAntimicrobials are widely used in children but pediatric dose regimens are not always validated, and PK studies, required to validate dosage, are difficult to conduct in children. Low sampling volume limits the number of PK samples drawn per patient and analytical methods adapted to small volumes are not always available. Due to the wide inter-patient pharmacokinetic (PK) variability in children, particularly neonates, therapeutic drug monitoring is required to adapt dosage to individual patients. In such clinical and analytical context, our aim was to develop a unique, rapid and highly sensitive ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) assay to quantify 7 antibiotics (amoxicillin, azithromycin, cefotaxime, ciprofloxacin, meropenem, metronidazole and piperacillin) in low sample volumes (50 µL) for both routine monitoring and pharmacokinetic studies.MethodsAfter protein precipitation by acetonitrile, the antibiotics and their associated deuterated internal standard were separated on a Waters Acquity UPLC HSS T3 (100 mm x 2.1 mm; 1.8 µm). The mobile phases consisted of a gradient of ammonium acetate (pH 2.4; 5mM) and acetonitrile acidified with 0.1% (v/v) formic acid (started ratio of 93:7, v/v), run at 0.5 mL/min flow rate (total run time: 2.75 min). Ions were detected in the turbo-ion-spray-positive and multiple-reaction-monitoring modes.ResultsThis method was linear from 0.1–50 µg/mL. Accuracy and precision were evaluated using Quality Control (2, 10, 35 µg/mL). Validation of the method proved that precision, selectivity and stability were all within the recommended limits.ConclusionThis method has the advantage of a unique, efficient and standardized analytical tool for rapid measurement of 7 antibiotics in low blood volume. It has been successfully applied for routine activity and pharmacokinetic studies in children and neonates.Disclosure(s)Nothing to disclose.

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Development and Validation of UPLC-MS/MS Method for Determination of Enasidenib in Rat Plasma and Its Pharmacokinetic Application

International Journal of Analytical Chemistry ◽

10.1155/2020/5084127 ◽

2020 ◽

Vol 2020 ◽

pp. 1-7

Author(s):

Shuang-long Li ◽

Yong-liang Zhu ◽

Yi Zhang ◽

Shu-han Liu ◽

Xiang-die Wang ◽

...

Keyword(s):

Mass Transfer ◽

Multiple Reaction Monitoring ◽

Internal Standard ◽

Gradient Elution ◽

Rat Plasma ◽

Bioanalytical Method Validation ◽

Fda Approval ◽

Elution Method ◽

Acquity Uplc ◽

Development And Validation

In our research, a straightforward UPLC-MS/MS method, with diazepam as the internal standard (IS), was proposed and acknowledged to determine the concentrations of enasidenib in rat plasma. When preparing the sample, we used acetonitrile for protein precipitation. The gradient elution method was used, and the mobile phase was acetonitrile and 0.1% formic acid. Diazepam was used as the IS. We used the Acquity UPLC BEH C18 column to separate enasidenib and IS. Under the positive ion electrospray ionization (ESI) source conditions, the mass transfer pairs of enasidenib were monitored by multiple reaction monitoring (MRM) to be m/z 474.2 ⟶ 456.1 and m/z 474.2 ⟶ 267.0, and the IS mass transfer pairs were m/z 285.0 ⟶ 154.0. Enasidenib had good linearity (r2 = 0.9985) in the concentration range of 1.0–1000 ng/mL. Besides, the values of intraday and interday precision were 2.25–8.40% and 3.94–5.46%, respectively, and the range of the accuracy values varied from −1.44 to 2.34%. Matrix effect, extraction recovery, and stability were compliant with FDA approval guidelines in terms of bioanalytical method validation. We had established a new method that had been applied to the pharmacokinetic study of enasidenib in rats.

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Validated RP-HPLC Method for Simultaneous Determination of Ribavirin, Sofosbuvir and Daclatasvir in Human Plasma: A Treatment Protocol Administered to HCV Patients in Egypt

Journal of Chromatographic Science ◽

10.1093/chromsci/bmz038 ◽

2019 ◽

Vol 57 (7) ◽

pp. 636-643 ◽

Cited By ~ 2

Author(s):

Aya A Youssef ◽

N Magdy ◽

Lobna A Hussein ◽

A M El-Kosasy

Keyword(s):

Human Plasma ◽

Method Validation ◽

Treatment Protocol ◽

Internal Standard ◽

Gradient Elution ◽

Hplc Method ◽

Array Detector ◽

Therapeutic Drug ◽

Bioanalytical Method Validation ◽

Rp Hplc

Abstract Egypt has the highest prevalence of hepatitis C virus (HCV) in the world thus it launched a national program for eliminating HCV aiming to treat 300,000 HCV patients per year. Three anti-HCV co-administered drugs; ribavirin (RBV), sofosbuvir (SF) daclatasvir (DAC) were simultaneously determined in human plasma by a validated, simple and sensitive RP-HPLC method using propyl paraben as an internal standard. Liquid–liquid extraction using ethyl acetate was used for samples extraction. Chromatographic separation was achieved on Scharlau® C18 column (250 × 4.6 mm2, 5 μm). Gradient elution was employed with a mobile phase mixture of water and acetonitrile at a flow rate 1 mL/min. UV detection using photodiode array detector was carried out at 207, 260 and 312 nm for RBV, SF and DAC, respectively. Method validation was performed according to the FDA guidelines for bioanalytical method validation. The calibration curves were linear over the ranges (0.5–80, 0.1–40 and 0.5–80 μg/mL) with average recoveries (100.64–108.28%, 98.48–105.91% and 97.68–101.38%) for RBV, SF and DAC, respectively. The intra-day and inter-day precision and accuracy results were within the acceptable limits. Stability assays revealed that the three studied analytes were stable during sample storage, preparation and injection. The method can be successfully applied in routine analysis of plasma of HCV patients treated with this combination therapy which aids in therapeutic drug monitoring and patients’ follow-up especially in Egypt and other developing countries fighting HCV.

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Development of a Validated UPLC-MS/MS Method for Analyzing Major Ginseng Saponins from Various Ginseng Species

Molecules ◽

10.3390/molecules24224065 ◽

2019 ◽

Vol 24 (22) ◽

pp. 4065 ◽

Cited By ~ 1

Author(s):

Ling Yang ◽

Chi-Lin Li ◽

Yung-Yi Cheng ◽

Tung-Hu Tsai

Keyword(s):

Partition Coefficient ◽

Multiple Reaction Monitoring ◽

Internal Standard ◽

Gradient Elution ◽

Ultra Performance Liquid Chromatography ◽

Reversed Phase ◽

Hydroxyl Groups ◽

Ginsenoside Rb1 ◽

Ginsenoside Re ◽

Ginsenoside Rd

Ginsenosides, which contain one triterpene and one or more sugar moieties, are the major bioactive compounds of ginseng. The aim of this study was to develop and optimize a specific and reliable ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for the analysis of twelve different resources of ginseng. The six marker compounds of ginsenoside Rb1, ginsenoside Rb2, ginsenoside Rc, ginsenoside Rd, ginsenoside Re, and ginsenoside Rg1, as well as an internal standard, were separated by a reversed-phase C-18 column with a gradient elution of water and methanol-acetonitrile. The multiple-reaction monitoring (MRM) mode was used to quantify and identify twelve market products. The results demonstrated that not only is the logarithm of its partition coefficient (cLog P; octanol-water partition coefficient) one of the factors, but also the number of sugars, position of sugars, and position of the hydroxyl groups are involved in the complicated separation factors for the analytes in the analytical system. If the amount of ginsenoside Rb1 was higher than 40 mg/g, then the species might be Panax quinquefolius, based on the results of the marker ginsenoside contents of various varieties. In summary, this study provides a rapid and precise analytical method for identifying the various ginsenosides from different species, geographic environments, and cultivation cultures.

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Pharmacokinetic Comparison of Isoalantolactone and Alantolactone in Rats after Administration Separately by Optimization of an UPLC-MS2Method

Journal of Chemistry ◽

10.1155/2014/354618 ◽

2014 ◽

Vol 2014 ◽

pp. 1-8 ◽

Cited By ~ 3

Author(s):

Renjie Xu ◽

Mengyue Wang ◽

Ying Peng ◽

Xiaobo Li

Keyword(s):

Multiple Reaction Monitoring ◽

Internal Standard ◽

Rat Plasma ◽

Sprague Dawley ◽

Ionization Source ◽

Fragment Ions ◽

Sprague Dawley Rats ◽

Liquid Chromatography Tandem Mass ◽

Positive Ion

Isoalantolactone and alantolactone are two major active ingredients that are present in many medicinal plants. In this study, a sensitive and rapid ultraperformance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method was developed for determination of the two compounds in rat plasma, separately. In this method, an electrospray ionization source was applied and operated in positive ion mode; multiple reaction monitoring (MRM) was selected for quantification using target fragment ions 233.2→187.1 for isoalantolactone (alantolactone) and 245.1→189.1 for internal standard (IS). Retention time of the lactones and IS was within 3.0 min. Further calibration suggested a linear regression can be calculated within 2.5–500 ng/mL for isoalantolactone and 4–500 ng/mL for alantolactone. This method was used to compare the pharmacokinetic characteristics of isoalantolactone and alantolactone at a single dose of 5 mg/kg into male Sprague-Dawley rats by intravenous administration separately. The levels oft1/2, Kel, CL,Cmax, and AUC were significantly increased in the alantolactone group compared to isoalantolactone. These results suggested that isoalantolactone was distributed and eliminated more rapidly than alantolactone in rats when administered, respectively.

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