Periodontal disease consists of chronic gingival inflammation characterized by both degradation of the periodontal connective tissue and alveolar bone loss. Drug therapy is used as an auxiliary treatment method in severe chronic periodontitis, aggressive periodontitis, and periodontitis-associated systemic disease. Nal-P-113, a modified antimicrobial peptide, specifically replaces the histidine residues of P-113 with the bulky amino acid β-naphthylalanine, and our previous studies have verified that this novel peptide is not toxic to the human body within a certain concentration range. The objective of the present study was to evaluate the effect of Nal-P-113 on periodontal pathogens and periodontal status in clinical studies. In a split-mouth clinical trial, the pocket depth and bleeding index values tended to decrease in the experimental group compared with those in the control group. SEM results verified that Nal-P-113 restrained the maturation of plaque. Based on real-time polymerase chain reaction, the levels of Fusobacterium nucleatum, Streptococcus gordonii, Treponema denticola, and Porphyromonas gingivalis in subgingival plaque were decreased when the subjects were given Nal-P-113. Bacterial growth curve analysis and a biofilm susceptibility assay verified that Nal-P-113 at a concentration of 20 μg/mL restrained the growth of S. gordonii, F. nucleatum, and P. gingivalis and biofilm formation. Therefore, Nal-P-113 effectively reduces periodontal pathogens and ameliorates periodontal status.