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Objective. The red rhizome of Angiopteris helferiana has been used widely to treat muscle fatigue, bone pain, and skin infection in Nepal. However, scientific evidence for its bioactive compounds and their bioactivities was lacking till January, 2024. Therefore, we investigated to validate and advance the further use and development. Methods. Column chromatography, including MCI gel CHP20P, Sephadex LH‐20, ODS, and Silica gel, was used for the isolation of compounds from 70% methanol extract of A. helferiana rhizomes. The isolated compounds were evaluated for their TLC‐based antioxidant and antibacterial activity against Staphylococcus aureus, Klebsiella pneumonia, Escherichia coli, and Pseudomonas aeruginosa by adopting the agar well diffusion protocol. Furthermore, in silico molecular docking against the penicillin binding protein of E. coli (PBP1b, PBP2, and PBP3 TPd) and ADME toxicity of the isolated compounds was predicted. Results and Discussion. Angiopteroside (1) and osmundalactone (2) were isolated for the first time from the red rhizomes of A. helferiana. The structure of the isolated compounds was elucidated on the basis of spectroscopic (1H‐, 13C‐, and 13C/DEPT‐NMR) and spectrometric (LC MS/ESI and IR) analyses and comparison with reported literature. Both compounds were inactive towards DPPH scavenging activity. Compound 1 showed poor inhibitory activity against E. coli, with inhibition zone range of 2–7 mm. However, both isolated compounds were found to be resistant against S. aureus, K. pneumonia, and P. aeruginosa. Based on in silico molecular docking forecasting, compound 1 revealed the good binding affinity with PBP1b (−6.5 kcal/mol), PBP2 (−5.8 kcal/mol) and PBP3 TPd (−6.1 kcal/mol) compared to positive control meropenem with PBP1b (−6.6 kcal/mol), PBP2 (−6.9 kcal/mol), and PBP3 TPd (−6.6 kcal/mol). Similarly, compound 2 showed weak preference for PBP1b (−4.3 kcal/mol), PBP2 (−4.4 kcal/mol), and PBP TPd (−4.5 kcal/mol). Compounds 1 and 2 were predicted to be safe in terms of hepatotoxicity, cytotoxicity, mutagenicity, and carcinogenicity, with a potential to induce nephrotoxicity similar to that of meropenem. Conclusion. This study successfully isolates two lactones with antibacterial properties and inspires the researcher to further research, development, and formulation. [ABSTRACT FROM AUTHOR]

Ethnomedicinally, more than 2000 plants were found to be used in Nepal. Among them, the red colored rhizome of Angiopteris helferiana and the bark of Saurauia fasciculata have been used widely to treat muscle fatigue, bone pain, fever, postpartum hemorrhage, and thirst by healers in Kaski and Tanahun districts, Nepal. However, scientific evidence towards their traditional uses is lacking till December, 2023. Therefore, we report the phytochemicals, total phenolic content (TPC), total flavonoid content (TFC), total carbohydrate content (TCC), antioxidant and antibacterial activities of A. helferiana and S. fasciculata extracts. Phytochemical analysis indicated that A. helferiana and S. fasciculata extracts were potential sources of chemicals such as phenols, flavonoids, tannins, terpenoids, saponins, and carbohydrates. The TPC, TFC, and TCC of extracts were determined by using an ultraviolet visible spectrophotometer. Among the extracts tested, A. helferiana extracts showed the highest phenolic and carbohydrate contents of 208.33 ± 12.96 mg of gallic acid equivalent/g and 564.16 ± 2.92 mg of D‐glucose equivalent/g of dry extract, respectively. Similarly, S. fasciculata revealed the highest flavonoid content of 30.35 ± 0.1 mg quercetin equivalent/g of dry extract. The extract of A. helferiana and S. fasciculata exhibited potent antioxidant activity by scavenging 2,2‐diphenyl‐1‐picrylhydrazyl radicals with an IC50 of 25.9 µg/ml and 31.07 µg/ml, respectively. The antibacterial activity of the A. helferiana and S. fasciculata extract against Staphylococcus aureus, Pseudomonas aeruginosa, and Escherichia coli was determined using an agar‐well diffusion protocol that revealed the potential antibacterial activity of A. helferiana against E. coli. The present study will help validate the traditional uses of A. helferiana rhizomes and S. fasciculata barks as a healing medicine and inspire the researcher towards further research, development, and formulation. [ABSTRACT FROM AUTHOR]