An ultrasound-assisted extraction and chromolithic LC method was developed for simultaneous determination of glycyrrhizic acid (GA) and glycyrrhetinic acid (GL) from the root extract of Glycyrrhizza glabra using RPLC-PDA. The developed method was validated according to the International Conference on Harmonisation. The method exhibited good linearity (r2 > 0.9989) with high precision and achieved good accuracies between 97.5 to 101.3% of quantitative results. The method is more sensitive and faster (resolved within ten minutes) than the earlier developed methods using normal LC columns.