The world all over, Nigeria inclusive, is faced with a lot of environmental problems as a result of unlawful refuse disposal, technological innovations, oil spillage, release of greenhouse gases, pollution and many more. This calls for immediate and pragmatic approaches in Chemistry Education most especially, Green Chemistry, which emphasizes the need to design chemical processes and products by using eco-friendly and non-toxic feed stock while reducing or eliminating the generation of hazardous substances. Therefore, there is the need for adequate knowledge of Green Chemistry for every Chemistry teacher. This study therefore, investigated the role of gender on Chemistry teachers’ knowledge of Green Chemistry in Lagos State, Nigeria. Descriptive survey research design was adopted in this study. Multi-stage sampling technique was used for the selection of schools while 160 (93 males and 67females) teachers participated in the study using convenient sampling technique. A research instrument titled “Teachers’ Knowledge of Green Chemistry Questionnaire”(TKGCQ) was used to elicit information about the perceived and measured knowledge of Chemistry teachers. The instrument which was made up of three Sections (A, B, &C) was validated by experts in items development in Chemistry Education. The reliability of the perceived knowledge (Section B) was established using Cronbach’s alpha which gave a reliability index of 0.71; while that of the measured knowledge (Section C) was determined using Split-half method with a reliability index of 0.68. Two hypotheses were formulated and tested using independent sample t-test statistic at 0.05 level of significance. The findings showed that both male and female teachers had low and similar perceived knowledge of Green Chemistry. But gender influenced their actual content knowledge; the male teachers had more knowledge of Green Chemistry than their female counterparts. It was therefore, recommended, among others, that, policy makers should embrace the need to integrate Green Chemistry principles into teacher education curriculum, while relevant capacity building workshops/seminars and in-service training programmes should be made available for Chemistry teachers at all levels, with more opportunities for female teachers to acquire adequate knowledge in Green Chemistry.