The objective of the present work was to optimize the extraction of phytochemicals from Hamelia patens Jacq. by ultrasound-assisted extraction. Taguchi L9 orthogonal array was used to evaluate the factors solid/liquid ratio (1:8, 1:12, and 1:16), extraction time (10, 20, and 30 min), and ethanol concentration (0, 35, and 70%). Total polyphenols were the response variable. Chromatographic fractionation using Amberlite XAD-16 was carried out and the total polyphenols, flavonoids, and condensed tannins were quantified. The redox potential, the reduction of the 2,2-diphenyl-1-picrylhydrazyl (DPPH), and the lipid oxidation inhibition were determined. Anti-bacterial activity was evaluated. The phytochemicals were identified by liquid chromatography coupled to mass spectrometry. Optimal extraction conditions were a solid/liquid ratio of 1:16, ethanol of 35%, and 10 min of ultrasound-assisted extraction. Maximum polyphenol content in the crude extract was 1689.976 ± 86.430 mg of gallic acid equivalents (GAE)/100 g of dried plant material. The purified fraction showed a total polyphenols content of 3552.84 ± 7.25 mg of GAE, flavonoids 1316.17 ± 0.27 mg of catechin equivalents, and condensed tannins 1694.87 ± 22.21 mg of procyanidin B1 equivalents, all per 100 g of purified fraction. Its redox potential was 553.93 ± 1.22 mV, reducing 63.08 ± 0.42% of DPPH radical and inhibiting 77.78 ± 2.78% of lipid oxidation. The polyphenols demonstrated antibacterial activity against Escherichia coli, Klebsiella pneumonia, and Enterococcus faecalis. The HPLC-ESI-MS analysis revealed the presence of coumarins, hydroxycinnamic acids, and flavonoids.