Joshua Cooper, IsilDilek, Uma Sreenivasan
Cerilliant Corporation, 811 Paloma Drive, Suite A, Round Rock, TX 78665
LC-MS/MS is a powerful tool that brings numerous benefits to the clinical sample analysis arena. However, due to the complexity of the instrumentation there are some unique challenges that also accompany these benefits. Even following sample extraction and cleanup, matrix effects from the samples can cause interferences or impact ionization efficiency. Deuterium-labeled internal standards are the most common and prevalent labeled internal standards used to compensate for matrix effects. Some deuterium labeled compounds may exhibit hydrogen-deuterium scrambling/exchange in the collision cell which can impact MS/MS transition selection.
In this study we investigated numerous variables that potentially contribute to scrambling in order to ascertain reproducibility and impact on scrambling ratios: influencesof different LC-MS systems (tandem quadrupolevs. quadrupoletime-of-flight), matrix selection, concentration, with and without HPLC, collision energies, and deuterium placement in the internal standard. Numerous small molecules of clinical importance were investigate including: hydroxyvitaminD, testosterone, immunosuppressants, bath salts, and spice cannabinoids.
Labeled 25-Hydroxyvitamin D2 and D3 Scrambling in Serum
Transitions Comparisons for Native and Labeled 25-Hydroxyvitamin D2 and D3 in EtOH on 6410
Notes: 25-Hydroxy D2-d6 water loss→2 water loss has same transition as 25-Hydroxyvitamin D3 parent→waterloss. Can be problem if compounds are not well resolved chromatographically.
Selection of Transitions Greatly Impacts Observed Scrambling
Testosterone Scrambling at Minor Transitions
* or Scrambling %13Cn-1/13Cn
Figure 1. Testosterone Chromatograms on XevoG2
Figure 2. (±)-11-nor-9-Carboxy-Δ9-THC Scrambling at m/z 327
Figure 3. JWH-073 3-Hydroxybutyl Metabolite Scrambling at m/z 216
Figure 4. Everolimus Scrambling at m/z 389
Spice Cannabinoids, Bath Salts, and Immunosuppressants Scrambling Comparison usinf Xevo G2
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