During the last few decades, chemo preventive and chemotherapeutic compounds against various types of cancer have been isolated from various plants. Glycyrrhezic acid (2-20%), Glycyrrhizin, is present as a major chemical constituent of natural anticancer compound Glycyrrhiza glabra L. (Fabaceae). The present work is aimed to investigate the in-vitro cytotoxic screening of standard 18 β-glycyrrhetic acid and also for natural anticancer drug Glycyrrhiza glabra L. (Fabaceae) using three different extracts (chloroform, methanol and water) of the drug through MTT method. Cell viability of previously identified 18 β-glycyrrhetic acid in three different extracts of Glycyrrhiza glabra L. (Fabaceae) was determined by two fold trypan blue method using two different cell lines MCF7-cancerous and Vero-normal cell line and it was quantified through HPTLC method. The results of the HPTLC study indicated that the amount of 18 β-glycyrrhetic acid into three different extracts (chloroform, methanol and water extract) of Glycyrrhiza glabra L. (Fabaceae) was 26.6, 12.5 and 5.6 μg/g, respectively. The percentage viability of two different cell lines was 45.71% for Vero-normal cell line and 78.78% for MCF7-cancerous cell line. IC 50 values of standard 18 β-glycyrrhetic acid was 0.412±0.004 μM and those for the three different extracts (chloroform, methanol and water)of Glycyrrhiza glabra L. (Fabaceae) on MCF7 cell line were 0.4485±0.001, 0.9906±0.001 and 1.288±0.005 μM, respectively. From the above result, it can be said that 18 β-glycyrrhetic acid could be considered as a potential source of natural anticancer component and the percentage of which was higher in the chloroform extract.