Validation of HPTLC method for the analysis of luteolin in Cardiospermum halicacabum Linn.

Mona Kukkar, Rajiv Kukkar, Ajay Saluja


Background: Luteolin is one of the constituent of Cardiospermum halicacabum, which is traditionally used as an important
component of plant‑based medicine. Aim and Objective: This study presents the first report of thin layer chromatography (TLC)
densitometric method, which has been developed and validated for quantification of luteolin from aerial parts of C. halicacabum.
Materials and Methods: Chromatographic separation was achieved on silica gel 60 F254 plates with using the solvent system of Toluene:Ethyl acetate: Formic acid (10:9:1). Detection of luteolin was carried out in the absorption‑reflection mode at 254 nm. TLC plates were dried under stream of hot air and then subjected to densitometric scanning using a Camag TLC scanner III (Camag, Switzerland) with win CATS software in the absorbance‑reflectance scan mode. The accuracy of the method was checked by conducting various validation parameters according to ICH (International Conference on Harmonizaation) guidelines. Results: The system was found to give compact spots for luteolin (retention factor, Rf = 0.55). The calibration plot was linear in the range of 500-3000 ng of luteolin. The correlation coefficient of 0.997 was indicative of good linear dependence of peak area on concentration. The concentration of luteolin was found to be 0.50% w/w in aerial parts of C. halicacabum. The limit of detection (LOD) and limit of quantification value
for luteolin were found to be 22.45 ng and 68.03 ng, respectively. Recovery values from 99.37 and 99.80 to 100.58 showed excellent reliability and reproducibility of the method. Conclusion: The proposed HPTLC method for quantitative monitoring of luteolin in C. halicacabum can be used for routine quality testing of C. halicacabum extract used in formulations.
Key words: Cardiospermum halicacabum, high performance thin layer chromatography, luteolin

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