Background. Epidermal growth factor receptor inhibitors (EGFRIs) and tyrosine kinase inhibitors (TKIs) are key drugs in targeted cancer therapy. However, they may cause skin toxicity. We previously prepared a modified Huang-Lian-Jie-Du (mHLJD) decoction cream using 10 herbs, which effectively alleviated EGFRI/TKI-induced skin toxicity. In the present study, we identified the reference markers of the mHLJD decoction and investigated the anti-inflammatory and antibacterial effects of the mHLJD decoction extract. Methods. We performed high-performance liquid chromatography (HPLC) to determine the composition of the mHLJD decoction. Human epidermoid A431 cells were treated with tumor necrosis factor (TNF)-α to induce inflammation; then, the effects of the mHLJD decoction extract on the cytokine expression were determined using a cytokine array and by performing real-time quantitative polymerase chain reaction (qPCR). The antibacterial effects of the extract were examined using disk diffusion and microdilution assays. Results. HPLC results revealed that the mHLJD decoction primarily consisted of geniposide, berberine chloride, baicalin, coptisine, and palmatine. TNF-α treatment increased the expression of certain cytokines, including IL-1β, IL-8, M-CSF, and TGF-β2; however, pretreatment with the mHLJD decoction extract reduced their expression. The qPCR results demonstrated a decreased mRNA expression of IL-8, M-CSF, and TGF-β2. The antibacterial assay revealed that the extract exerted inhibitory effects on Staphylococcus aureus, forming an inhibition zone at the minimum inhibitory concentrations of 3.125 and 6.25 mg/mL; however, the extract exerted no effects on Escherichia coli and Pseudomonas aeruginosa. Conclusions. We developed an HPLC method to quantify the reference markers of the mHLJD decoction. The bioactivity analysis provided the potential mechanisms underlying the effects of the mHLJD decoction on EGFRI/TKI-induced skin toxicity.