Comparison between external and internal standard calibration in the validation of an analytical method for 1‐hydroxypyrene in human urine by high‐performance liquid chromatography/tandem mass spectrometry
44
Citation
10
Reference
10
Related Paper
Citation Trend
Abstract:
Abstract 1‐Hydroxypyrene is a metabolite of pyrene, a member of the class of polycyclic aromatic hydrocarbons (PAHs) whose toxic properties in some cases include carcinogenicity. The determination of 1‐hydroxypyrene in human urine is used as a biological indicator for exposure to PAHs, which is related to the combustion of organic materials, like smoking, living in urban environments, and eating grilled or smoked food. The determination of 1‐hydroxypyrene by high‐performance liquid chromatography (HPLC) with fluorescence detection has very good sensitivity but it is not highly specific: this can reduce accuracy in the quantitative determination of low levels of analyte in a complex matrix like urine. An HPLC method that uses triple quadrupole mass detection has been validated with the objective both to improve the signal‐to‐noise (S/N) ratio and to achieve the maximum specificity for the analyte in those urine samples that are richer in possible inteferents. The calibration range for 1‐hydroxypyrene is from 0.005–0.1 µg/L in the urine of non‐smoking healthy volunteers. After solid‐phase extraction, samples were analyzed by HPLC/tandem mass spectrometry (MS/MS) in the multiple reaction monitoring (MRM) mode. In order to obtain reliable results quantitative analysis must be performed by means of the internal standard method (we used deuterium‐labelled 1‐hydroxypyrene): the method accuracy is not less than 85%. The S/N ratio at a concentration of 0.1 µg/L is about 10, and therefore this can be considered the lowest limit of quantitation. The method performance does not change if urine samples are measured using a calibration curve prepared in methanol, thus reducing the time of analysis and costs. Copyright © 2006 John Wiley & Sons, Ltd.Keywords:
Internal standard
Matrix (chemical analysis)
Cite
LC-MS and LC-MS/MS methods were established to identify the main components of teicoplanin.By using electrospray ionization and positive ion monitoring,mass spectrometric detection were operated on a sigle quadrupole mass spectrometer and a triple quadrupole tandem mass spectrometer.Six components of teicoplanin were identified and determined.The established methods can be used to analyze teicoplanin qualitatively and quantitatively in the new drug development for quality control and stability study.
Teicoplanin
Hybrid mass spectrometer
Quadrupole mass analyzer
Cite
Citations (0)
Internal Standard (ISTD) is a well-known chromatographic technique, aimed to compensate sample size variations, where known amount of a component, called internal standard is added to both standard and unknown samples. The classic Internal Standard quantification method plots the response ratio (analyte to internal standard) versus amount ratio (again analyte to internal standard). Internal standard component itself does not have any calibration curve. Quantification procedure uses this plot to get concentration ratio from response ratio. We demonstrate here, that this approach may cause systematic errors in the case of non-(linear through origin) detector response to concentration of analyte or internal standard. We also offer an alternative calculation scheme allowing wide variations of standard and analyte concentrations and non-(linear through origin) calibrations of components. In the case of non-directly proportional calibrations it requires that External standard dependencies of both internal standard component and analyte are measured. Offered scheme of Internal standard calculations splits into two independent parts: 1. Calculation of Relative concentration, i.e. concentration of analyte, provided concentration of Internal Standard is known, using External Standard calibration curves. 2. Constuction of improved calibration curves (Universal calibration) that can be simplified for the case of linear through origin dependencies. We provide a proof, that the classic response ratio scheme for linear through origin calibration is a particular case of the presented approach. The described calculation scheme is successfully used for Internal Standard calculations in Chrom&Spec chromatographic software for more than 15 years. Calibration Predictive relationship between input and detector response • Input: calibration samples – concentrations of components • Output: peak area or height • Prediction: Predict unknown input looking at response ESTD Calibration • Response (Area or Height) versus Quantity • Quantity is provided without error false • Response is measured with random normally distributed error – sometimes true External Standard Calibration Curve • Axes: Q – Quantity (NOT Concentration), R – Response (Area or Height) • Independent variable: Typically Q, sometimes R • Calibration curve: polynomial interpolation • Prediction: either solution of polynomial equation (independent Q) or value of polynomial (independent R) – we denote either of them W(R) Quantification: External Standard (raw) Concentration • Quantity of injected substance Qx = W(Rx) • Concentration of initial sample Cx = Qx/V = W(Rx)/(Vinj) • Vinj – injection volume ISTD Targets Reason Axis • Sample-size variations Q • Effect of sample preparations Q • Instrument drift R All reasons are always acting together
Internal standard
Standard addition
Cite
Citations (0)
Internal standard is an external compound which is mixed with targeted analytical solution and matrix as a constant concentration and use for preparing calibration standard curve by using ratio of analyte area and internal standard area with analyte concentration and internal standard concentration. This calibration curve used for quantification of unknown concentration of anlayte of interest. This article provide necessary information about internal standard like its selection procedure, characterization, types and response factor , to all analyst who are connected with drug analysis. This article is more important and I think first article which focuses a clear idea about internal standard use in drug analysis.
Internal standard
Standard curve
Standard addition
Matrix (chemical analysis)
Cite
Citations (2)
Internal standard
Bioanalysis
Isotope dilution
Sample Preparation
Cite
Citations (376)
Internal standard
Standard solution
Standard addition
Cite
Citations (26)
Protein precipitation
Scutellarin
Bioequivalence
Cite
Citations (4)
Ceramides play an important role in a variety of cellular functions including cell differentiation and apoptosis, responses to DNA damage and stress, and transcriptional events. Detection of ceramides in mammalian cells is required for many biological studies. Here, we report a validated method using LC-MS/MS-MRM on an Agilent 6410 triple quadrupole mass spectrometer to simultaneously quantify six ceramides extracted from mammalian cell lysates following a 5 min HPLC gradient. This method demonstrated outstanding sensitivity, accuracy, reproducibility, and speed of analysis.
Cite
Citations (9)
A method for introducing internal standards by direct infusion into the LC effluent for the quantitative LC−MS analysis of environmental samples is described. This postcolumn introduction method was found to be effective in correcting quantitative errors associated with matrix signal suppression. However, unlike surrogate or volumetric internal standards, the performance of the postcolumn method does not depend on the selection of an internal standard that shares identical elution time with the target analyte. As a result, either structural analogues (target analyte derivatives) or isotopically labeled compounds may be applied as effective internal standards. Furthermore, the postcolumn introduction method allows the application of one internal standard to address signal suppression effects for several analytes in a single LC−MS run. In contrast, volumetric and surrogate introduction methods require an isotopically labeled internal standard for each analyte to be quantified.
Internal standard
Quantitative Analysis
Analytical procedures
Matrix (chemical analysis)
Standard addition
Cite
Citations (72)
This laboratory experiment employs an innovative approach to introduce undergraduates to liquid chromatography–tandem mass spectrometry (LC–MS/MS) by developing a method based on multiple reaction monitoring (MRM). During this qualitative investigation, students operated a triple quadrupole mass spectrometer in the MS scan and product ion scan modes to determine precursor and product ions, respectively, for a vanillin, ethyl vanillin, and coumarin standard mixture. After identifying precursor → product ion transitions, students conducted a 2 min liquid chromatography run in MRM mode. Students were astonished by the speed, selectivity, and sensitivity of the LC–MS/MS method compared to that of a liquid chromatograph with ultraviolet detection.
Cite
Citations (19)
Stable isotope-labeled internal standards are of great utility in providing accurate quantitation in mass spectrometry (MS). An implicit assumption has been that there is no "cross talk" between signals of the internal standard and the target analyte. In some cases, however, naturally occurring isotopes of the analyte do contribute to the signal of the internal standard. This phenomenon becomes more pronounced for isotopically rich compounds, such as those containing sulfur, chlorine, or bromine, higher molecular weight compounds, and those at high analyte/internal standard concentration ratio. This can create nonlinear calibration behavior that may bias quantitative results. Here, we propose the use of a nonlinear but more accurate fitting of data for these situations that incorporates one or two constants determined experimentally for each analyte/internal standard combination and an adjustable calibration parameter. This fitting provides more accurate quantitation in MS-based assays where contributions from analyte to stable labeled internal standard signal exist. It can also correct for the reverse situation where an analyte is present in the internal standard as an impurity. The practical utility of this approach is described, and by using experimental data, the approach is compared to alternative fits.
Internal standard
Cite
Citations (33)