The present work has been undertaken with the aim to formulate hair growth gel formulation containing extracts of Hibiscus rosa sinensis flower 1%, Eclipta alba whole plant 1% and Solanum nigrum plant berries 0.5% which are preferably used in case of Alopecia, i.e., baldness pattern as an effective herbal therapy showing 5 α-reductase inhibitory activity. The formulated gel was evaluated for parameters such as pH which was found to be 6.68, viscosity 4731 cps, spreadability 11.05 (g-cm/sec) whereas consistent homogeneity was found with no skin irritation. Simultaneous quantification of bioactive markers was done by HPTLC. Quercetin, β-sitosterol and linoleic acid were selected as bioactive markers for quantification of Hibiscus rosa sinensis flower, Eclipta alba whole plant and Solanum nigrum plant berries extract respectively in the formulation. The aforementioned markers have 5 α-reductase inhibitory activity. β-sitosterol, quercetin and linoleic acid was found to be 0.1377, 0.120 and 0.379% w/w respectively in the formulated gel.