Objective To evaluate the efficacy and safety of three treatment approaches to applying Botulinum Toxin Type A (BoNTA) for crow's Feet. Methods Thirty female subjects with moderate-to-severe crow's feet were included in this comparative in vivo study. They were randomly divided into three groups, including the local intramuscular, intradermal microdroplet injection, and nanomicroneedle delivered with BoNTA therapy group. After one session, evaluations were done at the time points of weeks one, four, and twelve after the treatment. The assessments included subjective satisfaction, blinded clinical assessment, and the biophysical parameters (skin collagen content, elasticity, hydration, and sebum contents). Results For dynamic wrinkles, intramuscular injection and intradermal microdroplets injection were more effective than nanomicroneedles. For static wrinkles, nanomicroneedles and intradermal microdroplets injection were more effective. However, the intramuscular injection had no significant effect on static wrinkles. At one week and four weeks after the treatment, the skin elasticity, collagen content, and hydration of nanomicroneedle group and intradermal microdroplet group increased more significantly than those of the intramuscular injection group; at twelve weeks after the treatment, the skin elasticity, collagen content, and hydration of intradermal microdroplet group were higher than those of other two groups. However we observed no statistically significant difference in sebum content between the three groups before and after the treatment. Conclusion BoNTA delivered through nanomicroneedles and intradermal microdroplets injection can effectively treat crow's feet. This trial is registered with [2016]KY018-01, registered 16 Feb 2016.