Novel chromatographic methods for fingerprinting the bioactive constituents of herbal medicines used as hepatoprotective and adaptogenic in herbo-mineral regimen

Abstract

Shilajit, along with Triphala, has been one of the most widely used herbal drug combinations for the symptomatic treatments of hyperthyroidism. However, the high polarity difference among active constituents proposes difficulties while analyzing its quality. In this study, two high-performance thin-layer chromatography methods were developed for chromatographic fingerprinting of the proposed herbal mixture of Triphala and Shilajit. A reliable HPTLC method effectively analyzed bioactive phytoconstituents such as quercetin, myricetin, gallic acid, ellagic acid, tannic acid, ascorbic acid, and humic acid. Method 1 showed good separation of quercetin, myricetin, and tannic acid, but separating gallic and ellagic acid proved difficult. To address this issue, the 2D-TLC approach involves a second mobile phase. Although humic acid and ascorbic acid were not detected due to their highly polar and non-polar nature, so a different HPTLC method 2 was developed. The successful method of development using the Quality by Design approach and validated. The developed method successfully analyzed the bioactive constituents of Shilajit, in combination with Triphala, and validation parameters such as linearity, accuracy, precision, range, specificity, robustness, the limit of detection (LOD), and limit of quantification (LOQ) agreed with given values.