This work was conducted on Antigonon leptopus to optimize in vitro propagation conditions for shootlets multiplication and callus culture under effect of some factors such as types of cytokinins (BA and 2ip) at different concentrations (0.2, 0.4 and 0.6 mg/l) plus 0.1 mg/l of IBA that were examined for proliferated shoots with high rooting ability. Also, different concentrations of growth regulators (2,4-D, TDZ and NAA) under both light and darkness incubation were evaluated for callus tissues development. The secondary metabolites for both shootlets and callus tissues were estimated. For obtaining shootlet culture development with more proliferation and elongation of shootlets with the highest number of leaves, rooting percent and roots length, MS medium was added with 0.2 mg/l 2ip plus 0.1 mg/l IBA. While, MS supplemented with 0.4 2iP plus 0.1 IBA mg/l resulted in the highest values of all estimated pigments (Chl. a, b, carotenoid and anthocyanin) in shootlets. Using high concentration of 2ip (0.6 mg/1) enhanced the production of phenolic, flavonoids compounds, and antioxidant activity in shootlets. Low concentration of both NAA (2.5 mg/l) and TDZ (1.0 mg/l) under darkness condition produced the highest callus fresh weight. However, the same treatment (NAA at 2.5 mg/l+ TDZ at1mg/l) under light condition caused significant increment of the above-mentioned active compounds in callus tissues. The successfully acclimatized plantlets showed variations in morphological characters due to transferring them to different growth media (peat moss, perlite, clay and sand) in the greenhouse.