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Two populations of Physalis patula Miller collected in different localities (Querétaro and Teotihuacán) of Central México were analyzed. Eight new compounds including three withanolides, physapatolides A-C (1-3), two labdanes, 12-epi-physacoztomatin (10) and physapatulone (12), besides three 12-O-glucosyl labdanes, patulosides A-C (13-15), were isolated from these collections. A series of known withanolides, flavonoids, labdanes, sucrose esters, and sterols were also isolated. Withanolides isolated of each population exhibited different substitution patterns of ring D, thus while 15,16-dioxygenated withanolides were found in the Querétaro population, oxygenation occurs at C-14 and C-15 in those isolated from Teotihuacán population. The structures of all the isolated compounds were determined by analysis of their spectroscopic and spectrometric data. The absolute configuration assigned to physapatolide A (1) was confirmed by X-ray diffraction analysis of its acetyl derivative 1a, while those of labdane diterpenoids were based on chemical correlations. The antibacterial activity of 16 of the isolated compounds (1-3, 5-9, 13, 14, and 16-21) and two derivatives (13a, 14a) was evaluated against five ATCC bacterial strains (Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Staphylococcus aureus and Bacillus subtilis). Compounds 1-3 inhibited around 100 % the growth of S. aureus at 250 μg/mL (w/v). The minimum inhibitory concentrations (MIC) of compounds 1, 2, and 3, against S. aureus were 62.5, 125 and 62.5 μg/mL, respectively, whereas their MIC's against a Methicillin-resistant S. aureus (MRSA) clinical isolate were 125, 62.5 and 62.5 μg/mL, respectively., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)