Knowledge of physical activity (PA) can be considered a predictor of public health for society. Thus, this study aims to show content validity of the “Knowledge Questionnaire on World Health Organization (WHO) Recommendations on PA and Health” (CUAFYS-A) and reference values on adults’ knowledge of the WHO recommendations on PA. This is a quantitative, non-experimental, descriptive, and cross-sectional study, in which 579 adults completed an online questionnaire with demographic data. The questionnaire was made up of 9 items to measure PA related to knowledge. For the elaboration of the items of the questionnaire a disciplinary team formed it and for the analysis of results, a descriptive analysis of these was applied. Then an inferential analysis was performed, content validity, construct validity, and reliability were analyzed. The CUAFYS-A after its content analysis, obtained appropriate results in terms of pertinence and relevance; additionally, it showed Cronbach’s alpha coefficient of 0.62. Thereby, it was concluded the CUAFYS-A questionnaire proved to be a valid and reliable instrument to show reference values and to evaluate the knowledge of adults of PA and health according to the WHO recommendations.