| Summary: | The prevalence of multidrug-resistant (MDR) bacteria has recently increased dramatically, seriously endangering human health. Herein, amoxicillin (Amoxi)-conjugated gold nanoparticles (AuNPs) were created as a novel drug delivery system to overcome MDR bacteria. MDR bacteria were isolated from a variety of infection sources. Phenotype, biotype, and 16S rRNA gene analyses were used for isolate identification. Additionally, <i>Juniperus excelsa</i> was used for the production of AuNPs. The conjugation of AuNPs with Amoxi using sodium tri-polyphosphate (TPP) as a linker to produce Amoxi-TPP-AuNPs was studied. The AuNP and Amoxi-TPP-AuNP diameters ranged from 15.99 to 24.71 nm, with spherical and hexagonal shapes. A total of 83% of amoxicillin was released from Amoxi-TPP-AuNPs after 12 h, and after 3 days, 90% of the medication was released. The Amoxi-TPP-AuNPs exhibited superior antibacterial effectiveness against MRSA and MDR <i>E. coli</i> strains. Amoxi-TPP-AuNPs had MICs of 3.6–8 µg mL<sup>−1</sup> against the tested bacteria. This is 37.5–83 fold higher compared to values reported in the literature. Amoxi-TPP-AuNPs exhibit a remarkable ability against MRSA and <i>E. coli</i> strains. These results demonstrate the applicability of Amoxi-TPP-AuNPs as a drug delivery system to improve therapeutic action.
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