Study aim: To determine the effectiveness of a newly designed series of fitness segments that can be used to provide healthy behaviour knowledge (HBK) for 5th grade Physical Education classes.Material and methods: 641 pupils from six intervention (n = 401) and five control (n = 240) schools were used. The intervention schools received a five-week intervention of 12-minute fitness segments for each class period. A cognitive test that has been shown to produce valid and reliable scores (28-questions) was used to assess pupils’ HBK at pre – and post – examination. Pedometers were also used to examine physical activity levels differences. Hierarchical Linear Modeling using a two-level multilevel model was used to investigate mean changes in HBK between intervention and control groups.Results: Pupils in the intervention classes had 0.8-unit greater mean improvements in HBK than did control pupils (p < 0.05). Girls also had greater mean changes in HBK than did boys (p < 0.05). There was no statistical difference between intervention and control groups in physical activity levels.Conclusion: This type of fitness segment can be one strategy used by teachers to aid the instruction of HBK without decreasing physical activity participation in classes.