Mass Fragmentography of Cortisol and Cortisone: Preliminary Studies on the Development of a Reference Method

2002 ◽  
Vol 67 (1) ◽  
pp. 1-9 ◽  
Author(s):  
Hugh L. J. Makin ◽  
David J. H. Trafford ◽  
Norman F. Taylor
Keyword(s):  
Human Plasma ◽  
Trimethylsilyl Ether ◽  
Spectral Characteristics ◽  
Internal Standard ◽  
Reference Method ◽  
High Specificity ◽  
Mass Fragmentography ◽  
Mass Spectral ◽  
Target Values ◽  
Two Peaks

Cortisol and cortisone are the two main glucocorticoids found in human plasma. Cortisol is commonly measured as a means of assessing adrenal function, usually by immunoassay, but such procedures must be subject to proper quality control which ideally uses analyte target values measured by isotope dilution mass fragmentography (GC-MS). Three derivatives of these steroids have been investigated for their chromatographic and mass spectral characteristics in order to develop a simple and rapid GC-MS method for this purpose. Trimethylsilyl ether-O-methyloxime derivatives gave poor sensitivity and thesyn- andanti-isomers were resolved into two peaks. Cyclic methylboronate-O-methyloxime derivatives gave excellent sensitivity but were difficult to prepare, occasionally giving rise, by some unknown mechanism, to 11-oxidation of cortisol. The underivatised 11-hydroxyl of this derivative of cortisol also gave rise to chromatographic problems. Enol-trimethylsilyl ethers however were easy to prepare and gave excellent sensitivity (10-17 times that achieved with the trimethylsilyl ether-O-methyloxime derivatives). Straight line calibration graphs were obtained and the method gave clean traces allowing high specificity and sensitive measurement of cortisol and cortisone in human plasma using tri-deuterated cortisol as an internal standard.

scholarly journals Validated ICP-MS method for measurement of plasma and intracellular antimony concentrations applied to pharmacokinetics of meglumine antimoniate

2020 ◽  
Author(s):  
Diana J. Garay-Baquero ◽  
David E. Rebellon-Sanchez ◽  
Miguel D. Prieto ◽  
Lina Giraldo-Parra ◽  
Adriana Navas ◽  
...  
Keyword(s):  
Human Plasma ◽  
Inductively Coupled Plasma ◽  
Mononuclear Cells ◽  
Internal Standard ◽  
Reference Method ◽  
Clinical Samples ◽  
Peripheral Blood Mononuclear ◽  
Meglumine Antimoniate ◽  
Icp Ms ◽  
Limit Of Quantitation

Aim A high-throughput method using inductively coupled plasma mass spectrometry (ICP-MS) was developed and validated for the quantitative analysis of antimony in human plasma and peripheral blood mononuclear cells (PBMCs) from patients with cutaneous leishmaniasis undergoing treatment with meglumine antimoniate. Methods For this study, antimony was digested in clinical samples with 1% TMAH / 1% EDTA and indium was used as internal standard. Calibration curves for antimony, over the range of 25 to 10000 ng/mL were fitted to a linear model using a weighting of 1/concentration2. Accuracy, precision and stability were evaluated. Results Taking the lower limit of quantitation (LLOQ) to be the lowest validation concentration with precision and accuracy within 20% (25% at the LLOQ), the current assay was successfully validated from 25 to 10000 ng/mL for antimony in human plasma and PBMCs. Dilution studies demonstrated that concentrations up to 100000 ng/mL of antimony in plasma were reliably analyzed when diluted into the calibration range. Conclusion This protocol will serve as a baseline for future analytical designs, aiming to provide a reference method to allow inter-study comparisons.

scholarly journals Stability-Indicating Methods for Determination of Tiapride in Pure Form, Pharmaceutical Preparation, and Human Plasma

2007 ◽  
Vol 90 (6) ◽  
pp. 1554-1565 ◽  
Author(s):  
Fadia H Metwally ◽  
Mohammad Abdelkawy ◽  
Nada S Abdelwahab
Keyword(s):  
Human Plasma ◽  
Pharmaceutical Preparation ◽  
Internal Standard ◽  
Reference Method ◽  
Pure Form ◽  
Stability Indicating ◽  
First Derivative ◽  
Beer's Law ◽  
Beer’S Law

Abstract Four different stability-indicating procedures are described for determination of tiapride in pure form, dosage form, and human plasma. Second derivative (D2), first derivative of ratio spectra (1DD), spectrofluorimetric, and high-performance column liquid chromatographic (LC) methods are proposed for determination of tiapride in presence of its acid-induced degradation products, namely 2-methoxy-5-(methylsulfonyl) benzoic acid and 2-diethylaminoethylamine. These approaches were successfully applied to quantify tiapride using the information included in the absorption, excitation, and emission spectra of the appropriate solutions. In the D2 method, Beer's law was obeyed in the concentration range of 1.59 g/mL with a mean recovery of 99.94 1.38% at 253.4 nm using absolute ethanol as a solvent. In 1DD, which is based on the simultaneous use of the first derivative of ratio spectra and measurement at 245 nm in absolute ethanolic solution, Beer's law was obeyed over a concentration range of 1.59 g/mL with mean recovery 99.64 1.08%. The spectrofluorimetric method is based on the determination of tiapride native fluorescence at 339 nm emission wavelength and 230 nm excitation wavelength using watermethanol (8 + 2, v/v). The calibration curve was linear over the range of 0.23 g/mL with mean recovery of 99.66 1.46%. This method was also applied for determination of tiapride in human plasma. A reversed-phase LC method performed at ambient temperature was validated for determination of tiapride using methanoldeionized watertriethylamine (107 + 93 + 0.16, v/v/v) as the mobile phase. Sulpiride was used as an internal standard at a flow rate of 1 mL/min with ultraviolet detection at 214 nm. A linear relation was obtained over a concentration range of 230 g/mL with mean recovery of 99.66 0.9%. Results were statistically analyzed and compared with those obtained by applying the reference method. They proved both accuracy and precision.

Toward absolute methods in clinical chemistry: application of mass fragmentography to high-accuracy analyses.

1976 ◽  
Vol 22 (11) ◽  
pp. 1789-1801 ◽  
Author(s):  
I Björkhem ◽  
R Blomstrand ◽  
O Lantto ◽  
L Svensson ◽  
G Ohman
Keyword(s):  
Clinical Chemistry ◽  
Internal Standard ◽  
High Specificity ◽  
Mass Fragmentography ◽  
Reference Methods ◽  
Relative Standard ◽  
High Degree ◽  
Definitive Methods ◽  
Very High

Abstract We review recently developed reference methods based on mass fragmentography (specific ion monitoring) with use of isotope-labeled internal standard, for determination of cholesterol, triglycerides, urea, glucose, cortisol, progesterone, and testosterone. With an optimal ratio between standard and material to be determined, the relative standard deviation of these methods, based on duplicate determinations, is between 1.3 and 2.7%. The very high specificity of the methods, in combination with the fact that the ratio between labeled and unlabeled molecules is determined with a high degree of accuracy, makes it likely that the most significant errors in the methods are related to errors in pipetting. The possibility is discussed that these methods might be developed into "absolute" or "definitive" methods by subjecting each individual step in the determination to a detailed analysis with respect to possible error. We also discuss some of the consequences of the different reference methods on the choice of routine methods.

Development and Validation of a New Method for the Determination of Anti-hepatitis C Agent Simeprevir in Human Plasma using HPLC with Fluorescence Detection

2020 ◽  
Vol 16 (4) ◽  
pp. 428-435
Author(s):  
Ahmed F.A. Youssef ◽  
Yousry M. Issa ◽  
Kareem M. Nabil
Keyword(s):  
Hepatitis C ◽  
Human Plasma ◽  
Fluorescence Detection ◽  
Internal Standard ◽  
Protein Precipitation ◽  
Assay Method ◽  
Fluorescence Detector ◽  
Limit Of Quantification ◽  
Relative Standard

Background: Simeprevir is one of the recently discovered drugs for treating hepatitis C which is one of the major diseases across the globe. Objective: The present study involves the development of a new and unique High-Performance Liquid Chromatography (HPLC) method using fluorescence detection for the determination of simeprevir (SIM) in human plasma. Methods: Two methods of extractions were tested, protein precipitation using acetonitrile and liquidliquid extraction. A 25 mM dipotassium hydrogen orthophosphate (pH 7.0)/ACN (50/50; v/v), was used as mobile phase and C18 reversed phase column as the stationary phase. The chromatographic conditions were optimized and the concentration of simeprevir was determined by using the fluorescence detector. Cyclobenzaprine was used as an internal standard. Results: Recovery of the assay method based on protein precipitation was up to 100%. Intra-day and inter-day accuracies range from 92.30 to 107.80%, with Relative Standard Deviation (RSD) range 1.65-8.02%. The present method was successfully applied to a pharmacokinetic study where SIM was administered as a single dose of 150 mg SIM/capsule (Olysio®) to healthy individuals. Conclusion: This method exhibits high sensitivity with a low limit of quantification 10 ng mL-1, good selectivity using fluorescence detection, wide linear application range 10-3000 ng mL-1, good recovery and highly precise and validation results. The developed method can be applied in routine analysis for real samples.

Bioanalytical method development and validation of simultaneous analysis of telmisartan and hydrochlorthiazide in human plasma using LC-MS/MS

2016 ◽  
Vol 5 (03) ◽  
pp. 4862 ◽  
Author(s):  
Mathew George* ◽  
Lincy Joseph ◽  
Arpit Kumar Jain ◽  
Anju V.
Keyword(s):  
Human Plasma ◽  
Retention Time ◽  
High Performance ◽  
Method Development ◽  
Matrix Effects ◽  
Solid Phase ◽  
Internal Standard ◽  
Assay Method ◽  
Buffer System ◽  
Internal Standards

A simple, sensitive, rapid and economic high performance thin layer chromatographic method and a mass spectroscopic assay method has been developed for the quantification of telmisartan and hydrochlorthiazide combination in human plasma. The internal standards and analytes were extracted from human plasma by solid-phase extraction with HLB Oasis1cc (30mg) catridges. The scanning and optimization for the samples are done using methanol: water (50:50). The samples were chromatographed using reverse phase chromatography with C-18 column of different manufacturers like Ascentis C18 (150×4. 6, 5µ) using the buffer system Acetonitrile: Buffer (80:20%v/v) which consist of 2±0. 1Mm ammonium format at a flow rate of 0. 7ml/min at a column oven temperature 35±10c. The internal standard used was hydrochlorthiazide13c1, d2 and telmisartand3. The extraction techniques include conditioning, loading, washing and elution, drying followed by reconstitution of the dried samples. The volume injected was 10µl with the retention time of 3-4 min for telmisartan, 1-2 min for hydrochlorthiazide and for the internal standards the retention time was 3-4 min for telmisartand3 and 1-2 min for hydrochlorthiazide c13d2. The rinsing solution was Acetonitrile: HPLC grade water in the ratio (50:50). The above developed method was validated using various parameters like selectivity and sensitivity, accuracy and precision, matrix effects, % recovery and various stability studies. The method was proved to be sensitive, accurate, precise and reproducible. The preparation showed high recovery for the quantitative determination of telmisartan and hydrochlorthiazide in human plasma.

scholarly journals Development and Validation of a Method for Quantification of Favipiravir as COVID-19 Management in Spiked Human Plasma

Molecules ◽  
2021 ◽  
Vol 26 (13) ◽  
pp. 3789
Author(s):  
Mohammad Hailat ◽  
Israa Al-Ani ◽  
Mohammed Hamad ◽  
Zainab Zakareia ◽  
Wael Abu Dayyih
Keyword(s):  
Human Plasma ◽  
Internal Standard ◽  
Weighting Factor ◽  
Hplc Method ◽  
Bioanalytical Method Validation ◽  
Spiked Human Plasma ◽  
Fda Guidance ◽  
Significant Difference ◽  
Us Fda ◽  
Carry Over

In the current work, a simple, economical, accurate, and precise HPLC method with UV detection was developed to quantify Favipiravir (FVIR) in spiked human plasma using acyclovir (ACVR) as an internal standard in the COVID-19 pandemic time. Both FVIR and ACVR were well separated and resolved on the C18 column using the mobile phase blend of methanol:acetonitrile:20 mM phosphate buffer (pH 3.1) in an isocratic mode flow rate of 1 mL/min with a proportion of 30:10:60 %, v/v/v. The detector wavelength was set at 242 nm. Maximum recovery of FVIR and ACVR from plasma was obtained with dichloromethane (DCM) as extracting solvent. The calibration curve was found to be linear in the range of 3.1–60.0 µg/mL with regression coefficient (r2) = 0.9976. However, with acceptable r2, the calibration data’s heteroscedasticity was observed, which was further reduced using weighted linear regression with weighting factor 1/x. Finally, the method was validated concerning sensitivity, accuracy (Inter and Intraday’s % RE and RSD were 0.28, 0.65 and 1.00, 0.12 respectively), precision, recovery (89.99%, 89.09%, and 90.81% for LQC, MQC, and HQC, respectively), stability (% RSD for 30-day were 3.04 and 1.71 for LQC and HQC, respectively at −20 °C), and carry-over US-FDA guidance for Bioanalytical Method Validation for researchers in the COVID-19 pandemic crisis. Furthermore, there was no significant difference for selectivity when evaluated at LLOQ concentration of 3 µg/mL of FVIR and relative to the blank.

scholarly journals Analysis of Pembrolizumab in Human Plasma by LC-MS/HRMS. Method Validation and Comparison with Elisa

Biomedicines ◽  
2021 ◽  
Vol 9 (6) ◽  
pp. 621
Author(s):  
Aurélien Millet ◽  
Nihel Khoudour ◽  
Jérôme Guitton ◽  
Dorothée Lebert ◽  
François Goldwasser ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Human Plasma ◽  
High Resolution Mass Spectrometry ◽  
Internal Standard ◽  
Therapeutic Drug ◽  
Limit Of Quantification ◽  
Immunoglobulin G4 ◽  
Positive Mode ◽  
Surrogate Peptide ◽  
First Time

Pembrolizumab is a humanized immunoglobulin G4-kappa anti-PD1 antibody used in the treatment of different solid tumors or haematological malignancies. A liquid chromatography coupled with a high resolution mass spectrometry (orbitrap technology) method was fully developed, optimized, and validated for quantitative analysis of pembrolizumab in human plasma. A mass spectrometry assay was used for the first time a full-length stable isotope-labelled pembrolizumab-like (Arginine 13C6-15N4 and Lysine 13C6-15N2) as an internal standard; the sample preparation was based on albumin depletion and trypsin digestion and, finally, one surrogate peptide was quantified in positive mode. The assay showed good linearity over the range of 1–100 μg/mL, a limit of quantification at 1 μg/mL, excellent accuracy from 4.4% to 5.1%, and also a between-day precision below 20% at the limit of quantification. In parallel, an in-house ELISA was developed with a linearity range from 2.5 to 50 µg/mL. Then, results were obtained from 70 plasma samples of cancer patients that were treated with pembrolizumab and quantified with both methods were compared using the Passing-Bablok regression analysis and Bland-Altman plotting. The LC-MS/HRMS method is easy to implement in the laboratory for use in the context of PK/PD studies, clinical trials, or therapeutic drug monitoring.

scholarly journals Development and validation of a liquid chromatography-tandem mass spectrometric method for the determination of lenalidomide in human plasma and its application on bioequivalence studies

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
R. Gopinath ◽  
S. T. Narenderan ◽  
M. Kumar ◽  
B. Babu
Keyword(s):  
Liquid Chromatography ◽  
Human Plasma ◽  
Internal Standard ◽  
Spectrometric Method ◽  
Tandem Mass ◽  
Mass Spectrometric Method ◽  
Mass Detection ◽  
Triple Quadrupole Mass Spectrometer ◽  
Liquid Chromatography Tandem Mass ◽  
Tandem Mass Spectrometric

AbstractA simple, sensitive, and specific liquid chromatography-tandem mass spectrophotometry (LC-MS/MS) method was developed and validated for the quantification of lenalidomide in human plasma. The separation was carried out on a symmetry, C18, 5-μm (50 × 4.6 mm) column as stationary phase and with an isocratic mobile phase of 0.1% formic acid in water-methanol in the ratio of (15:85, v/v) at a flow rate of 0.5 mL/min. Protonated ions formed by electrospray ionization in the positive mode were used to detect analyte and fluconazole (internal standard). The mass detection was made by monitoring the fragmentation of m/z 260.1/148.8 for lenalidomide and m/z 307.1/238.0 for internal standard on a triple quadrupole mass spectrometer. The developed method was validated over the concentration range of 10–1000 ng/mL for lenalidomide in human plasma with a correlation coefficient (r2) was 0.9930. The accuracy and precision values obtained from six different sets of quality control samples analyzed on separate occasions ranged from 99.41 to 106.97% and 2.88 to 4.22%, respectively. Mean extraction recoveries were 98.06% and 88.78% for the analyte and IS, respectively. The developed method was successfully applied for analyzing lenalidomide in human plasma samples.

scholarly journals 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

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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