Kara Lynch (Presenter)
University of California San Francisco
Bio: Dr. Kara Lynch is an Associate Professor of Laboratory Medicine at the University of California San Francisco and the Associate Chief of the Clinical Chemistry and Toxicology Laboratory at San Francisco General Hospital and Trauma Center. She received her PhD in Biochemistry from the University of Wisconsin and did a post-doctoral fellowship in Clinical Chemistry in the COMACC approved training program at UCSF. Her primary area of research is in Clinical Toxicology. Her laboratory conducts studies aimed at identifying and quantifying toxins, drugs of abuse and adulterants in biological specimens using mass spectrometry and correlating the findings with clinical pathologies.
Authorship: Kara L. Lynch (1) and Jennifer M. Colby (2)
(1) Department of Laboratory Medicine, University of California San Francisco and San Francisco General Hospital (2) Department of Pathology, Microbiology, and Immunology, Vanderbilt University
The use of LC-HRMS is becoming more common for small molecules identification in a variety of analytical fields. With LC-HRMS, data acquisition is untargeted and the resulting data can be analyzed using targeted, suspect, and untargeted data analysis. The continued use of novel psychoactive substances (NPS) presents a significant analytical challenge for toxicology laboratories trying to keep their analytical methods up-to-date. Purchasing all possible NPS analytical standards is not practical for most laboratories. The objective of this study was to evaluate the utility of suspect screening with and without library matching to identify emerging amphetamine-type NPS in our patient population.
Introduction: The use of liquid chromatography high resolution mass spectrometry (LC-HRMS) is becoming more common for the detection of small molecules in a variety of analytical fields. One advantage of LC-HRMS over traditional techniques is the ability to perform untargeted data collection. This theoretically allows for acquisition of data (accurate mass, retention time, isotope and fragmentation pattern) for all analytes within the method’s specified mass range. The data can then be analyzed using targeted, suspect, and untargeted data analysis. For targeted data analysis, the acquired data is analyzed for the presence of a set list of compounds for which accurate mass, retention time, isotope and fragmentation pattern are known and have been analytically validated. With suspect analysis, the acquired data is searched for a list of compounds for which only the exact mass and theoretical isotope pattern are known. A mass spectrum for some of these compounds may also be available in the library used by the laboratory. Therefore, suspect analysis allows for the preliminary identification of a wider range of compounds compared to targeted data analysis. Suspect analysis is routinely used in the analysis of waste water and veterinary residue analysis, however, the utility of suspect analysis for clinical toxicology has yet to be evaluated or realized.
The use of novel psychoactive substances (NPS) continues to be a growing problem causing significant morbidity among drug users. When one NPS is regulated at the state or federal level, often a new class emerges. This presents a significant challenge for toxicology laboratories trying to keep their analytical methods up-to-date and relevant. Purchasing all possible NPS analytical standards is not possible for most clinical laboratories. The objective of this study was to evaluate the utility of suspect screening with and without library matching to identify new and emerging amphetamine-type NPS in our patient population.
Methods: All urine samples sent to the San Francisco General Hospital Clinical Laboratory for routine drug screening, for patients under the age of thirty (excluding neonates), were included in the study. Samples were collected for the month of October in 2014 and 2015. Each sample was diluted 1:5 in mobile phase and tested using a HRMS method. Briefly, HRMS data was acquired with an ABSciex TripleTOF®5600 system operating in positive ion mode, collecting full scan data with IDA triggered acquisition of product ion spectra. Chromatographic separations were performed on a Phenomenex Kinetex 3.5 um C18 column (50 x 3.00 mm, 2.6 µm). Mobile phase A consisted of 0.05% formic acid in 5 mM ammonium formate. Mobile phase B consisted of 0.05% formic acid in 50% methanol 50% acetonitrile. The elution gradient was ramped linearly from 2% to 100% B over 10 minutes. The method was previously validated and is in routine use with targeted data analysis for ~200 drugs and metabolites. Data analysis was done using PeakView® and MasterView® software (version 2.0, AB Sciex). For suspect analysis, a list of 749 NPS drugs and metabolites and their corresponding chemical formulas was generated and used for chromatogram extraction. An in-house spectral library, a vendor-supplied library and the Wiley library were used for data analysis.
Results: One-hundred and forty samples (64 female, 76 male, average age 24) were collected and analyzed during the month of October 2014. Half of these samples were sent from the emergency department and the other half from outpatient clinics. Using targeted data analysis, 188 drugs and metabolites were identified in these samples. Thirty-two were positive for methamphetamine. Using NPS suspect analysis, three preliminary positives were identified in one of the samples. Suspect analysis identified the presence of N-methyl-2AI (mass, isotope and library match), 2-AI (mass and isotope match, no available spectrum in the libraries), and methoxyamphetamine (mass, isotope and library match). Analytical standards were purchased and used to confirm the presence of these compounds. This sample came from a patient that was on an involuntary psychiatric hold in the psychiatric emergency department and the clinicians suspected the patient was under the influence of an amphetamine-like stimulant; however, the immunoassay result for amphetamines was negative. Analysis of the data collected in October 2015 is underway.
Conclusions: NPS are not encountered as frequently as methamphetamine in our patient population. This may be unique to patients in San Francisco or hospital patients at large and may not apply to synthetic cannabinoids which were not investigated in this study. Suspect screening is a useful technique for NPS screening, with and without the availability of library spectra. It saves the laboratory from having to purchase reference standards for many compounds that may never be identified and provides for the detection of preliminary positives immediately while waiting for a reference standard for confirmation. It also allows screening for compounds that are new and for which no reference materials are available.
References & Acknowledgements:
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