| Summary: | The intracellular drug concentration is needed for determination of target exposure at the site of action regarding its pharmacological action and adverse effects. Sesamol is an antiproliferative molecule from <i>Sesamum indicum</i> with promising health benefits. We present a method for measuring the intracellular sesamol content using reverse-phase HPLC with a UV diode array in melanoma cells. Sesamol was completely resolved by isocratic elution (4.152 ± 0.008 min) with methanol/water (70%, <i>v</i>/<i>v</i>) through a 30 °C, 5-µm C-18 column and detection at 297 nm. The present assay offers high sensitivity, fast elution, and an accurate and linear nominal concentration range of 10−1000 ng/mL (<i>R</i><sup>2</sup> = 0.9972). The % accuracy of the sesamol quality control sample was −3.36% to 1.50% (bias) with a 0.84% to 5.28% relative standard deviation (RSD), representing high repeatability and high reproducibility. The % recovery was 94.80% to 99.29%, which determined that there was no loss of sesamol content during the sample preparation. The validated method was applied to monitor intracellular sesamol concentration after treatment from 5 min to 24 h. The remaining intracellular sesamol content was correlated with its antiproliferative effect (<i>R</i><sup>2</sup> = 0.9483). In conclusion, this assay demonstrated low manipulation, quick elution, and high sensitivity, precision, accuracy, and recovery, and it was successfully applied to the quantification of sesamol in target cells.
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