Quantitative Assessment of an Antimalarial Flavonoid in the Citrus maxima(Burm.) Merr. Hydroalcoholic Root Extract using HPTLC Densitometric Analysis

Purpose: The main purpose of the study was to standardize the hydroalcoholic (1:1) root extract (CM-HA) of Citrus maxima(Burm.) Merr using luteolin, a moderately powerful antimalarial flavonoid. Methods: The quantitative estimation of phenolic and flavonoid contents were done using standard methods. The qualitative and quantitative characterization of CM-HA was also carried out using High Performance Thin Layer Chromatography (HPTLC) based densitometric analysis using a validated method. Results: The quantities of phenolic and flavonoid contents were found to be 68.83 ± 1.83 mg gallic acid equivalent per gm and 40.04 ± 0.97 mg quercetin equivalent per gm of CM-HA. In HPTLC analysis, the marker compound (luteolin) was separated using a solvent system made up of toluene, ethyl acetate, and formic acid (10:8:1 v/v/v). Luteolin was quantified using densitometric scanning under a deuterium lamp at a 350 nm wavelength in absorbance mode. With the ideal solvent system, this validated approach produced a compact band of luteolin at an R_fvalue of 0.532 ± 0.029. Before quantitative estimation, the created method’s precision, accuracy and reproducibility were verified as per the guideline of the International Council on Harmonization (ICH). For luteolin, linear results were seen graphically with a correlation coefficient of 0.990322 in the 400–1400 ng/band range. It was discovered that luteolin had detection and quantification limits of 30 and 80 ng/band, respectively. Eventually, it was discovered that there were 7.203 ± 0.557 mg of luteolin per 1 g of CM-HA. Conclusion: The study confirmed the presence of the antimalarial flavonoid luteolin in sufficient amount in the root of the plant.