Simultaneous quantification of vasicine and vasicinone in different parts of Justicia adhatoda using high-performance thin-layer chromatography‒densitometry: comparison of different extraction techniques and solvent systems

The present study was carried out for screening an efficient extraction method and extracting solvent in different plant parts of Justicia adhatoda L. for the development of a rapid and simultaneous determination of vasicine and vasicinone using a validated high-performance thin-layer chromatography (HPTLC) method. Four different extraction techniques, viz., maceration extraction (ME), ultrasonic-assisted extraction (UAE), microwave-assisted extraction (MAE), and heat reflux extraction (HRE), as well as extracting solvents (water, methanol, acetone, and acetonitrile) were used. Chromatographic separation was carried out on pre-coated silica gel plate G60 F254 using ethyl acetate‒methanol‒ammonia (8:2:0.2, V/V) as the mobile phase, and densitometric analysis was carried out in absorbance mode at 270 nm for vasicine and vasicinone, respectively. The optimized mobile phase gave well-defined peaks at RF of 0.37 and 0.57 for vasicine and vasicinone, respectively. The calibration curve area versus concentration was found to be linear with regression coefficient (r) values of 0.9956 and 0.9919 in the range of 100‒400 ng band‒1 for vasicine and vasicinone, respectively. The HPTLC method was validated in terms of precision, specificity, recovery, and accuracy. The best quantitative estimation result for vasicine and vasicinone was obtained by microwave-assisted extraction in leaf parts using methanol as solvent. Under these conditions, it was found that this plant contained approximately 6.38 mg g‒1 vasicine and 3.59 mg g‒1 vasicinone, respectively. The present HPTLC method was found to be precise, reliable, and can be used for the quality control of J. adhatoda L. samples and different J. adhatoda extracted samples.