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Fentanyl (Figure 1) is a very potent opioid analgesic and can be detected in the sub – ppb to low ppb range following chromatographic separation through a C18column using reversed

phase (RP) HPLC conditions. When fentanyl and the internal standard D5-fentanyl were separated using RP-HPLC the retention times of the two components were5.0 minutes and 7.0 minutes respectively. The retention time of the mobile phase was 1.5 minutes. If the resolution (Rs) was 1.5, using a suitable expression, calculate the number of theoretical plates required to facilitate this separation. Fentanyl can be routinely analysed using a RP HPLC system linked toa MS (ESI) detector. What characteristics must the interface possess ifit is to adequately link the instrumentation to allow for detection of the analyte? What are the guidelines for method validation that would enable the determination of fentanyl (and its derivatives) using RP HPLC?

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