Background: The use of performance-enhancing substances (PES), food supplements (FS), and traditional herbs (TH) has become increasingly common among athletes worldwide, often leading to inadvertent doping violations and health risks. In Kenya, mixed martial arts (MMA) disciplines such as boxing, wrestling, Taekwondo, karate, weightlifting, and bodybuilding have seen rapid growth. However, limited data exists regarding athletes’ knowledge of PES, FS, and TH and the methods employed in their application. Objective: To investigate the levels of knowledge on performance-enhancing substances, food supplements, traditional herbs, and methods of administration among mixed martial arts athletes in Kenya. Methodology: A cross-sectional survey was conducted among 771 registered MMA athletes across 23 counties in Kenya. Stratified proportionate random sampling was used to select participants from six disciplines. Data were collected using a structured questionnaire with 16 knowledge-related items. Descriptive statistics were computed, and one-way ANOVA and independent samples t-tests were used to examine differences in knowledge levels across sports disciplines and gender. Results: Knowledge scores varied significantly across disciplines (F(5, 765) = 9.87, p < .001). Wrestlers had the highest mean knowledge score (M = 14.05, SD = 2.41), while bodybuilders and Taekwondo athletes recorded the lowest scores (M = 10.93 and 11.10, respectively). Male and female athletes did not differ significantly in knowledge scores. Misconceptions were common, with more than 60% of respondents incorrectly believing that banned substances would be listed on supplement labels and only 37.2% aware that the Anti-Doping Agency of Kenya (ADAK) does not approve nutritional supplements. Conclusion: The study reveals substantial knowledge gaps regarding PES, FS, and TH among Kenyan MMA athletes. While some disciplines demonstrate higher awareness, others remain vulnerable to misinformation and potential doping violations due to limited access to accurate information. Recommendation: Targeted anti-doping education programs should be implemented with an emphasis on disciplines showing low knowledge levels. ADAK and sports federations should develop sport-specific and accessible learning resources and increase the frequency of workshops to improve athletes’ knowledge and safeguard their health and careers.