Abstract
Spinach (Spinacia oleracea) is one of the most famous vegetables worldwide, rich in essential metabolites for various health benefits. It is a valuable plant source that has the potential to be a nutraceutical. In the process of developing functional foods, it is crucial to establish a standard compound and quantitative analysis methods to ensure standardization. Thus, the analytical method validation was established using 3',4',5-trihydroxy-3-methoxy-6,7-methylenedioxyflavone 4'-glucuronide (TMG) as a marker compound. The specificity, accuracy, linearity, precision, repeatability, limit of detection (LOD), and limit of quantification (LOQ) was proved according to validation guideline. The specificity was confirmed from the BPI and UV chromatogram of the standard compound (TMG) as the predominant metabolite in the spinach sample. The calibration curve of TMG (15.625 ~ 500 µg/ml) had reasonable linearity (R2 = 0.999) considering with LOD and LOQ values, respectively. Recovery rate of TMG was 93.31-100.98% for FDS and 90.37–95.04% for SEC. The precision was less than 3 and 6% in the intra-day and inter-day. The spinach samples (FDS and SEC) were validated using TMC as a standard compound to comply with AOAC and KFDA regulations for approving functional ingredients in health functional food.