Simultaneous quantification of diarylheptanoids in Alnus nepalensis using a validated HPTLC method

Area of Science:

• Phytochemistry
• Analytical Chemistry
• Pharmacognosy

Background:

• Alnus nepalensis contains bioactive metabolites like platyphylloside, oregonin, and hirsutanonol 5-O-β-D-glucopyranoside.
• Accurate quantification of these marker compounds is essential for quality control and understanding their biological roles.

Purpose of the Study:

• To develop and validate a sensitive, selective, and robust high-performance thin-layer chromatographic (HPTLC) method.
• To enable simultaneous qualitative and quantitative determination of platyphylloside, oregonin, and hirsutanonol 5-O-β-D-glucopyranoside in Alnus nepalensis leaves.

Main Methods:

• Isolation and simultaneous quantification of three marker compounds using thin-layer chromatography densitometry.
• Development of an HPTLC method using silica gel 60F254 plates and a chloroform:methanol:formic acid mobile phase.
• Quantification via densitometric reflection/absorption at 610 nm after derivatization with vanillin-sulfuric acid reagent.

Main Results:

• A validated HPTLC method achieved precise and accurate quantification within a linear range of 333-3330 ng/spot (r² = 0.999).
• The method demonstrated good correlation, peak purity, precision, robustness, and appropriate limits of detection and quantitation as per ICH guidelines.
• Specificity was confirmed through retention factor and spectra correlation between standard and sample markers.

Conclusions:

• A reliable HPTLC-densitometric method was successfully developed and validated for the simultaneous determination of key bioactive diarylheptanoids in Alnus nepalensis.
• This method provides a robust tool for the quality assessment and standardization of Alnus nepalensis-derived materials.