The effectiveness of an erbium-doped: yttrium, aluminum and garnet (Er: YAG) laser (EYL) for the treatment of peri-implant disease (PID) remains unclear. The aim of this study was to compare non-surgical EYL therapy for PID with locally delivered minocycline hydrochloride (MC) ointment therapy by evaluating clinical, microbiological, and biochemical markers. Thirty-seven patients with PID were randomly assigned to either the EYL group (n = 18) or the MC group (n = 19). The clinical, microbiological, and biochemical markers at baseline and at 1 and 3 months after treatment were compared between the two groups. Subgingival plaque and peri-implant crevicular fluid (PICF) were collected from the diseased pockets. In the EYL group, probing pocket depth (PPD) was significantly decreased after treatment when compared with baseline. On the other hand, in the MC group, there was no significant decrease in PPD after treatment. Specific bacteria associated with PID were not determined. The counts of both Gram-positive and -negative species did not significantly decrease in the EYL group at 3 months after treatment. In the MC group, the counts of almost all bacterial species were significantly decreased after treatment. Biochemical marker analysis of PICF revealed significantly lower levels of metalloproteinase (MMP)-9 in the EYL group, as compared with the MC group at 3 months after treatment (p= 0.009). Non-surgical therapy with an EYL for PID was clinically effective, with decreased MMP-9 levels in PICF, which may lead to reduced peri-implant tissue destruction.