Development and Validation of Thin Layer Chromatography-Densitometric Method for Quantification of Kaempferol and Chlorogenic Acid in Methanolic Extract of Dragea Volubilis
Keywords:Dragea volubilis, Kaempferol and chlorogenic acid, High-performance thin layer chromatography, Validation
The present study aims to prepare a methanolic extract of Dragea volubilis and to quantitatively estimate kaempferol and chlorogenic acid using newly developed and validated method of high-performance thin layer chromatography (HPTLC) using TLC densitometry technique. Dried leaves obtained from D. volubilis were defatted with petroleum ether and extracted in a Soxhlet extractor with methanol. The solvent was evaporated using a Rota evaporator. Quantitative analysis of the extract was done using TLC and HPTLC. A mixture of toluene: ethyl acetate: glacial acetic acid (5.5:4:0.5) for kaempferol and ethyl acetate: acetic acid: water (7:1.5:1.5) for chlorogenic acid was used as a mobile phase. The chromatography was performed on a TLC plate precoated with silica gel GF254 and the developed TLC plates were visualized and quantified at 367 nm and 327 nm for kaempferol and chlorogenic acid, respectively. The percentage yield was found to be 0.0397 ± 0.0001 and 0.0755 ± 0.0000 for kaempferol and 0.0755 ± 0.0000 for chlorogenic acid. Rf. values 0.95 and 0.61 were observed for kaempferol and chlorogenic acid, respectively. The method was validated for instrumental precision, repeatability, coefficient of determination (r2), linearity range (ng), LOD (ng), LOQ (ng), and intra-day and inter-day precision for both kaempferol and chlorogenic acid, and all the parameters were found to be within the acceptable range as per the ICH guidelines. The newly developed HPTLC method was found to be prompt, cost-effective, precisely accurate, and reproducible for the qualitative as well as quantitative analysis of kaempferol and chlorogenic acid in methanolic extract of D. volubilis.
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Copyright (c) 2022 Amit Sharma, Naresh Gill, Rajesh Kumar
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