Suitability evaluation of new endogenous biomarkers for the identification of nitrite‐based urine adulteration in mass spectrometry methods

This paper reviews various pitfalls observed during developing, validation, application, and interpretation of drug testing approaches using GC-MS and low-and high-resolution LC-MS. They include sampling and storage of body samples, sample adulteration and contamination, analyte stability, sample preparation without or with cleavage of conjugates, extraction, derivatization, internal standardization, false negative and positive results by GC-MS or LC-MS screening and/or confirmation procedures including artifact formation, ion suppression or enhancement by electrospray ionization, and finally pitfalls in data interpretation. Conclusions and prospects close the Tutorial. K E Y W O R D S

Section: Samplingmentioning

confidence: 99%

Pitfalls in drug testing by hyphenated low‐ and high‐resolution mass spectrometry

Maurer

2020

“…Experiments on KNO 2 were described elsewhere. 13 ROC curves, potential cut-offs, sensitivity and specificity were calculated using both MetaboAnalyst 4.0 using the method closest to left corner combining best specificity and best sensitivity 22 and GraphPad Prism 7.0 (GraphPad Software, La Jolla, CA, USA). 13 ROC curves, potential cut-offs, sensitivity and specificity were calculated using both MetaboAnalyst 4.0 using the method closest to left corner combining best specificity and best sensitivity 22 and GraphPad Prism 7.0 (GraphPad Software, La Jolla, CA, USA).…”

Section: Data Analysis For Concentration and Ph Influence And Biomamentioning

confidence: 99%

“…4,9,10 With recent intentions in several US states to ban artificial urine products, chemical adulteration attempts might become of higher interest. 13 To unambiguously identify urine adulteration attempts, a suitable biomarker needs to comprise the ability to reliably differentiate a However, for a broad applicability of these markers, more data are needed considering the large variety of different chemical adulterants. Kluge et al recently evaluated the potential of co-measurement of selected endogenous compounds in an untargeted screening approach for identification of artificial urine products.…”

Section: Introductionmentioning

confidence: 99%

“…Experiments with KNO 2 are described in detail in previous work 12,13. RT, room temperature; ROC, receiver operating characteristics.…”

mentioning

confidence: 99%

“…Experimental conditions for each experiment and adulterant. Experiments indicated as H 2 O 2 include addition of H 2 O 2 and peroxidaseTable S1) for ROC analysis of PCC, H 2 O 2 , I 2 , and NaOCl were treated as described in Section 2.3 and quantified against a six-point calibration curve prepared in synthetic urine 21 in MultiQuant 2.1 and given as μmol concentration/mmol creatinine using a validated method 13. Experiments with KNO 2 are described in detail in previous work 12,13.…”

mentioning

confidence: 99%

See 2 more Smart Citations

Evaluation of endogenous urinary biomarkers for indirect detection of urine adulteration attempts by five different chemical adulterants in mass spectrometry methods

Steuer

Kamber

Kræmer

2018

Self Cite

Reliable detection of urine adulteration attempts to circumvent positive drug testing represents a critical step for laboratories in abstinence control settings. An ideal workflow for high-throughput testing would involve simultaneous detection of adulteration attempts in the same run with drug detection. Monitoring of degraded or oxidized endogenous urinary compounds as indirect markers has been previously evaluated for that purpose exemplified for the adulterant potassium nitrite (KNO 2 ).Fifteen, previously identified endogenous markers should now be evaluated for their general applicability to detect adulteration attempts for the adulterants hypochloritebased bleach (NaOCl), peroxidase and peroxide (H 2 O 2 ), pyridinium chlorochromate (PCC), and iodine (I 2 ). Initial experiments revealed similar results for the tested adulterants regarding degradation of indolylacryloylglycine (IAG), uric acid (UA), or UA derivatives. 5-Hydroxyisourate (HIU), the oxidation product of UA, was however only formed by KNO 2 , PCC, and H 2 O 2 . Amino acids showed larger adulterantdependent differences. All reactions were shown to be influenced by the adulterant concentration and the urinary pH with large variances depending on compound and adulterant. Except for HIU/PCC, all markers were stable within +/− 30% variation for all adulterants at −20°C. Receiver operating characteristics indicated best sensitivity and specificity over all adulterants for IAG (specificity 0.9, sensitivity 1.0) and UA (specificity 1.0, sensitivity 0.9). HIU gave best results for KNO 2 , PCC, and H 2 O 2 and N-acetylneuraminic acid for PCC and H 2 O 2 , respectively. When integrating a limited number of targets into existing screening methods, monitoring of UA, IAG, N-acetylneuraminic acid, and HIU is recommended.

Identification of new urinary gamma‐hydroxybutyric acid markers applying untargeted metabolomics analysis following placebo‐controlled administration to humans

Steuer

Raeber

Boxler

et al. 2019

Self Cite

Gamma‐hydroxybutyrate (GHB) is a short‐chain fatty acid that occurs naturally in the mammalian brain and is prescribed as a medication against narcolepsy or used as a drug of abuse. Particularly, its use as a knock‐out drug in cases of drug‐facilitated crimes is of major importance in forensic toxicology. Because of its rapid metabolism and resulting narrow detection windows (<12 hours in urine), detection of GHB remains challenging. Thus, there is an urgent call for new markers to improve the reliable detection of GHB use. In the framework of a randomized, placebo‐controlled, crossover study in 20 healthy male volunteers, urine samples obtained 4.5 hours post‐administration were submitted to untargeted mass spectrometry [MS, quadrupole time of flight (QTOF)] analysis to identify possible new markers of GHB intake. MS data from four different analytical methods (reversed phase and hydrophilic interaction liquid chromatography; positive and negative electrospray ionization) were filtered for significantly changed features applying univariate and multivariate statistics. From the resulting 42 compounds of interest, 8 were finally identified including conjugates of GHB with carnitine, glutamate, and glycine as well as the endogenous compounds glycolate and succinylcarnitine. While GHB conjugates were only detectable in the GHB, but not in the placebo group, glycolate and succinylcarnitine were present in both groups albeit significantly increased through GHB intake. Untargeted metabolomics proved as a suitable tool for the non‐hypothesis driven identification of new GHB markers. However, more studies on actual concentrations, detection windows, and stability will be necessary to assess the suitability of these markers for routine application.