Restek at MSACL 2024

17-22 March–Monterey, CA, U.S.

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TECHNICAL POSTERS:

Analysis of Δ-8-THC, Δ-9-THC, and Isomer Metabolites in Whole Blood by LC-MS/MS

Wednesday, March 20, 2:30-3:45 p.m.
Haley Berkland (presenter), Jamie York
Restek Corporation

Abstract
The testing of whole blood samples for tetrahydrocannabinol (Δ-9-THC) consumption is routine and has been around for many decades. As more isomers of Δ-9-THC become available on the market, testing becomes more complicated, and novel methods are needed to achieve isomeric resolution. One such isomer, Δ-8-THC, is federally unregulated in the United States and readily available for purchase in many stores. This compound forms its own hydroxylated and carboxylated metabolites, (11-OH-Δ-8-THC and Δ-8-THC-COOH), that must be chromatographically resolved from their isomeric metabolites.

An LC-MS/MS method was successfully developed on a Raptor FluoroPhenyl 100 x 3.0 mm, 2.7 μm column for reliable and accurate testing of Δ-8/9-THC isomers and isomer metabolites in whole blood. The method was determined to be quick, rugged, and sensitive enough to meet reporting guidelines for clinical and forensic toxicology laboratories.

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Exploring Different HPLC Column Chemistries for Optimal Separation of 13 Bile Acids by LC-MS/M

Wednesday, March 20, 12:15 -1:30 p.m.
Haley Berkland¹ (presenter), Paul Connolly¹, Justin Steimling¹, Katharina Schramm²
1. Restek Corporation, 2. ARUP Institute for Clinical and Experimental Pathology

Abstract
The analysis of bile acids in human plasma is an important diagnostic tool for the detection of liver disease and can also be used as indicators of potentially harmful side effects of new drugs. There are two main types of bile acids based upon their functional groups: free (or unconjugated) bile acids and conjugated bile acids, primarily glycine- or taurine-bound. Quantitation of bile acids in matrix can be very challenging due to a number of factors. These include structural similarities, varying polarity and stereochemistry, the presence of isomers, limited fragmentation of unconjugated bile acids in a mass spectrometer, high endogenous levels, and matrix effects caused by phospholipids or triglycerides.

Three column chemistries were explored in this study for the analysis of 13 bile acids in plasma in order to resolve a matrix interference. Biphenyl, FluoroPhenyl, and ARC-18 stationary phases were tested on a 100 x 2.1 mm, 2.7 μm column dimension. The ARC-18 stationary phase showed good selectivity for the target analytes. This phase can also resolve all three isomers sets with adequate resolution, and successfully separated the matrix interference from UDCA-D4, which is necessary for accurate quantitation of these analytes.

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The Benefits of 2.1 mm Internal Diameter Analytical Columns for the Analysis of Drugs of Abuse by LC-MS/MS

Wednesday, March 20, 2:30-3:45 p.m.
Samantha Herbick (presenter), Jamie York
Restek Corporation

Abstract
The biphenyl stationary phase offers advantageous selectivity compared to a C18 column for drugs of abuse panels, but choosing the right column dimension is key to obtaining robust and accurate data. Every column dimension can be advantageous in different scenarios, but generally clinical labs are all working towards the same goals: high throughput, low sample volume, good sensitivity, and low cost. In this work, the advantage of 2.1 mm internal diameter (ID) columns is discussed and demonstrated for the analysis of drugs of abuse by LC-MS/MS.

Two methods were developed to analyze common isobars in drugs of abuse panels on Raptor Biphenyl columns: one method used a 50 x 2.1 mm, 2.7 μm column with a flow rate of 0.6 mL/min and the other used a 50 x 4.6 mm, 2.7 μm column with a flow rate of 0.9 mL/min. The two methods were compared for efficiency, sensitivity, resolution (Rs), consumption of mobile phase, and robustness. Buprenorphine was used to demonstrate sensitivity differences between the two different column dimensions. It was found that the 2.1 mm ID column produced twice the signal response of the 4.6mm internal diameter column when injection volume is held constant. Twice the amount of sample had to be injected on the larger-bore column to produce the same signal response, thus introducing the chromatography to greater matrix effects with more impact to instrument cleanliness.

In summary, the 2.1 mm ID column was able to demonstrate superior sensitivity while consuming less resources and minimizing impact to the MS while still providing adequate resolution of isobars and excellent column robustness.

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Using a Virtual Liquid Chromatography Tool to Develop Methods for Novel Psychoactive Substances

Thursday, March 21, 9:15-10:30 a.m.
Jared Burkhart (presenter), Haley Berkland, Melinda Urich, Justin Steimling, Jamie York, Chris Nelson, Tim Yosca, John Garrett
Restek Corporation

Abstract
Novel psychoactive substances (NPS) have created a challenge for toxicology laboratories. New NPS are constantly disappearing as fast as they emerge, making it difficult to stay on top of which compounds are necessary to add to laboratory testing scopes. The development and optimization of liquid chromatography (LC) separations is time-consuming and costly, often requiring several steps, including literature research, column selection, method scouting, method development, and method optimization.

To alleviate the burden of sacrificing instrument uptime, labor, and materials, an instrument-free software modeling tool was developed to include a comprehensive drugs of abuse (DoA) library. This online tool allows users to obtain optimized separations while maintaining critical pair resolution by adjusting parameters, such as column dimension, mobile phase, gradient programs, and more for almost 300 compounds, including the 38 newly added NPS drugs.

To verify the ability of the modeler to develop methods for NPS, three methods were developed and optimized using the chromatogram modeler for the following NPS subclasses: 1) synthetic opioids and toxic adulterants, 2) designer benzodiazepines, and 3) stimulants and synthetic cannabinoids. All methods utilized a Raptor Biphenyl 100 x 2.1, 2.7 μm column, and based on the defined acceptance criteria, each NPS method was successfully transferred from the virtual model to an LC-MS/MS instrument.

As NPS continue to proliferate the illicit drug market, the burden of adding these compounds to laboratory testing scopes becomes the obligation of LC method developers. Utilizing tools such as a virtual chromatography modeler can help method developers deal with the challenges these emerging compounds present.

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