Abstract: In the present investigation, alkaloid profiling of Rauwolfia serpentina plant samples, including roots, leaves and callus, were performed which were collected from different localities. The CAF of samples suggest that roots are rich in alkaloid content as compared to leaves and callus. The maximum CAF was obtained from the in vitro regenerated roots (496 mg g-1). The TLC analysis of all the samples, including roots, leaves and callus, showed that roots are rich in reserpine and they also contain other alkaloids besides reserpine. Callus tissues also contain reserpine, but in the leave samples alkaloids could not be estimated. However, reserpine was detected in all the root and leaf samples by HP-TLC. The alkaloid profiling through HPLC detected a number of alkaloids from all the plant samples. The highest reserpine content was obtained from the in vitro regenerated roots (33 mg g-1) and the least from the leaves.