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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
Author(s) -
Pigini D.,
Cialdella A. M.,
Faranda P.,
Tranfo G.
Publication year - 2006
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/rcm.2407
Subject(s) - chemistry , chromatography , analyte , mass spectrometry , detection limit , urine , high performance liquid chromatography , tandem mass spectrometry , selected reaction monitoring , internal standard , triple quadrupole mass spectrometer , matrix (chemical analysis) , biochemistry
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.