Optimization of dispersive liquid liquid microextraction method for determination of trace salivary melatonin using high performance liquid chromatography

Due to low concentration of melatonin and also the existence of other compounds in body fluids, measuring the amount of melatonin is a serious challenge for the analysts. Before measurement of melatonin amounts using chromatographic method, sample preparation is unavoidable. The aim of current study is develop a new sample preparation method that not only have high accuracy and validity, but also can using by conventional equipment available in the laboratory to determine trace levels of salivary melatonin. For this purpose, the dispersive liquid-liquid microextraction (DLLME) method, that has not been used so far to preparation of salivary melatonin samples, were optimized.
Optimization procedure involved evaluation of eight factors affecting DLLME method. From these factors, seven were examined in four levels, and just pH was done in five levels. Accuracy and precision of the optimized method was evaluated at three concentrations of 50, 100, 250 pg.ml-1 by achieving CV% for day-to-day and within-day reproducibility.
Optimum values were determined. Accuracy and precision of the optimized method was investigated at three concentrations of 50, 100, and 250 pg.ml-1 by achieving CV% of 6.29, 2.39, and 1.82 for the "day-to-day" and 4.49, 2.68, and 1.83 for "within-day" reproducibility.
Due to the results of this study for Coefficient of variations, the optimized method was provided with proper accuracy and precision. Although optimized extraction factors must be fit analyte, but these factors can be used for similar chemical compounds with a slight modification.