Improving the sexual and reproductive health (SRH) of adolescent girls is one of the primary aims of the Sustainable Development Goals (SDGs). Adequate and accurate knowledge, a favorable attitude, safe behavior, and regular practice contribute to adolescent girls’ SRH, maternal health, and child health. Considering this, this study aims to explore the level of knowledge, attitudes, and practices (KAP) of SRH among college-going older adolescent girls in Chittagong district, Bangladesh. An institution-based cross-sectional study was conducted in four colleges among the older adolescent girl age group of 16–17 years old (N = 792) attending a higher secondary grade in Chittagong district. Data were collected using a structured and self-administered questionnaire. Descriptive statistics and multiple linear regression analyses were used to summarize the SRH-related KAP and identify the associated factors, respectively. The level of knowledge about puberty, family planning, maternal health, and HIV/AIDS was not satisfactory among the older adolescent girls. Different myths are common in the rural area with regards to menstruation, which impose several restrictions on adolescent girls and adult women. Standardized coefficients of beta (β) and p value < 0.05 in linear regression analyses demonstrated that being a student of the science group (β = 0.29, p < 0.001) and reading about or watching SRH issues on media (β = 0.21, p < 0.001) were significantly associated with older adolescent girls’ high level of knowledge in this regard. Furthermore, being a student of the science group (β = 0.17, p < 0.001), urban residence (β = 0.20, p < 0.001), regular SRH communication (at least once a month) with a mother/sister/friend (β = 0.10, p = 0.003), and reading or watching any SRH content on media (β = 0.22, p < 0.001) appeared as predictors of adolescent girls’ positive attitude towards SRH issues. Moreover, being a student of the science group (β = 0.07, p = 0.048), urban residence (β = 0.22, p < 0.001), regular SRH discussions with a mother/sister/friend (β = 0.09, p = 0.005), pre-knowledge on periods before menarche (β = 0.12, p < 0.001), and reading or watching any SRH content on media (β = 0.18, p < 0.001) are the most important factors influencing a regular hygienic practice of SRH. This study suggests strengthening SRH-related comprehensive education programs incorporated into the curriculum, the effective use of mass media, and supplying behavioral change communication materials.