The extensive and improper use of antibiotics has escalated antimicrobial resistance (AMR) into a global crisis. This issue not only affects human health but also increases mortality rates and negatively impacts the environment worldwide. ESBL-PE strains, which include representative Enterobacterales such as Escherichia coli and Klebsiella pneumoniae, produce ESBL enzymes that counteract commercially available drugs through hydrolysis. These strains can cause severe and potentially fatal infections, including pneumonia, urinary tract infections, and meningitis. As the efficacy of drugs against AMR bacteria dwindles, the development of new antibacterial agents becomes crucial. Chinese medicinal herbs, as natural products, present promising sources for future antimicrobial therapies. This project aims to identify potential antimicrobial agents, evaluate the antibacterial activities of different Chinese medicinal herbs, and compare the method efficacy against different ESBL-PE strains using disc diffusion and broth microdilution methods. The results revealed that six Chinese medicinal herbs, including Arnebiae Radix, Sarcandrae Herba, Scutellariae Radix, Coptidis Rhizoma, Prunellae Spica, and Magnoliae Cortex, demonstrated effective antimicrobial activity. The minimum inhibitory concentration (MIC) for these herbs against one of the ESBL-producing E. coli strains and K. pneumoniae strains was then determined. The MIC of different plant extracts ranged from 1.5625 to 25 mg/mL for the ESBL-producing E. coli strain, and 0.78125 to 3.125 mg/mL for the ESBL-producing K. pneumoniae strain. It was concluded that the broth microdilution method was more useful, accurate and precise in comparing two antimicrobial susceptibility testing methods. Therefore, these six Chinese medicinal herbs could potentially serve as novel and effective antibacterial agents to combat the growing issue of antimicrobial resistance, and the broth microdilution method should be prioritized for determining antimicrobial effects.