The medical plant research has received more attention among researchers especially after the Covid-19 pandemic. This research performed to evaluate the antifungal, anti-lung cancer (A549), and anti-hyperglycemic activities of aqueous extract of Andrographis paniculata flower. Interestingly, A. paniculata flower aqueous extract contains pharmaceutically valuable phytochemicals such as alkaloid, phenolics, terpenoids, tannins, flavonoids, and protein. It also showed fine antifungal activity against test fungal pathogens in the following order as: Aspergillus niger > Fusarium solani > Trichoderma harzianum > A. parasiticus > P. expansum > Penicillium janthinellum with lowest MIC values as ranged from 100 to 300 mug mL(-1). Interestingly, this aqueous extract also showed considerable anti-lung cancer activity, evidenced by dose and time dependent lung cancer cell line (A549) growth/proliferation inhibition/cytotoxicity activity (65%) at 300 mug mL(-1) concentration. This can be achieved by plant extract through inducing the secretion of apoptosis related proteins such as TNF alpha, IFN-gamma, and interleukin 2 leads to apoptosis in A549 cells. It also showed fine anti-diabetic activity by inhibiting alpha -amylase (58.41%) than alpha-glucosidase (54.74%) at 200 mug mL(-1) concentration. The UV as well as FTIR results demonstrated that the aqueous extract of A. paniculata flower contains pharmaceutically valuable bioactive compounds, which may be responsible for the wide range of biomedical applications. CI - Copyright (c) 2023 Elsevier Inc. All rights reserved.