20 Buyers’ Guide 2021
Evaluation of Mixed-Mode Ion Exchange SPE for the Extraction of Opioids from Human Urine by UHPLC-MS/MS
Author: James Edwards, Chromatography Applications Scientist, Porvair Sciences Ltd -
james.edwards@porvairsciences.com Background
The goal of any solid phase extraction (SPE) or sample preparation method development is to obtain the best analyte recovery while minimising the concentration of contaminating compounds reaching the final analysis sample. Traditional loose-filled SPE products are susceptible to a wide range of technical issues arising from poor packing of the resins into cartridges. Inconsistent packing can be problematic during method development resulting in voiding and channelling thereby increasing the risk of obtaining poor analyte recoveries.
The MicroluteTM CP SPE range eliminates
loose packing in the manufacturing process and therefore, overcomes these technical issues often experienced during sample preparation. Instead it consists of a unique, solid hybrid polymer structure composed of an interconnected network of evenly distributed pores immobilised with the retentive media. This design enhances the flow-through of samples to maximise interactions between analytes and the solid phase to deliver a highly reproducible SPE method.
This application describes the SPE LC- MS/MS methodology using a strong cation exchange (SCX) SPE 30 mg 96 well microplate for the determination of 12 opioids from human urine. It also examines the performance benefits of a hybrid polymer product versus a loose packed plate.
Introduction The Opioid Crisis
Opioids have been viewed as some of the most effective drugs for pain relief and classed as an essential part for reducing suffering of patients [1]. Due to their effectiveness in pain relief, this class of drugs has caused a lot of controversy due to their administration for recreational purposes.
This has led to a normalisation of their use, leading to widespread addiction and abuse of their use all over the world. Europe has been estimated to have had 1.3 million high risk users in 2017 [2] and the United States had an estimated 10.3 million people aged 12 or older misusing opioids in 2018 [3].
With this increase in abuse of opioids, comes a greater importance in laboratory testing for the presence of opioids to help detect patients misusing opioids while still allowing other patients to get the pain relief they need. Laboratory testing has a key part to play to help with solving the opioid crisis in the United States. Quest diagnostics (a large national clinical U.S laboratory) analysed their drug testing data from 2011 - 2017 which contained 3.9 million de-identified drug monitoring tests. This reported findings that in every year from 2011 - 2017, the majority of tests performed were classed as inconsistent with the expected result for their prescribed amount [4]. Therefore it is possible to conclude from these results that testing is needed to confirm if a patient is abusing their prescriptions or using other illicit drugs with their prescription. Chromatographic urine drug testing is there to allow a definitive verification if a patient is using their opioids as prescribed or disregarding their plan and leading towards abuse [5].
Detection of Opioids
Two tests are currently used for detection of opioids in urine: an immunoassay screening test and a chromatographic test. The standard opioid immunoassays are typically designed to detect the natural morphine- like molecules (morphine and codeine) but do not detect synthetic opioids like fentanyl and methadone [6]. The advantages of using chromatographic techniques is that it is possible to identify all the different opioids in one method to target different classes. It also provides a quantifiable result on each opioid present in the sample instead of the qualitative immunoassay result.
Other techniques for determining opioids in urine include a chromatographic ‘dilute and shoot’ method [7, 8], this involves diluting a sample with an internal standard solution and injecting straight onto LC-MS system. However, compared to a solid phase extraction (an SPE) method, it does not allow concentration of a sample. With low levels of opioids often present in urine samples this could result in false negatives, especially so on opioids that do not give a strong response on a mass spectrometer. SPE also removes any matrix components from the urine sample which are present in a dilute and shoot method. These can affect ionisation of the opioids and build up in the MS source resulting in unreliable
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