Bicycling posture leads to considerable discomfort and a variety of chronic injuries. This necessitates a proper bicycle design to avoid injuries and thereby enhance rider comfort. The objective of this study was to investigate the muscle activity during cycling on three different bicycle designs, i.e., rigid frame (RF), suspension (SU) and sports (SP) using surface electromyography (sEMG). Twelve male volunteers participated in this study. sEMG signals were acquired bilaterally from extensor carpi radialis (ECR), trapezius medial (TM), latissimus dorsi medial (LDM) and erector spinae (ES), during 30min of cycling on each bicycle and after cycling. Time domain (RMS) and frequency domain (MPF) parameters were extracted from acquired sEMG signals. From the sEMG study, it was found that the fatigue in right LDM and ES were significantly (p<0.05) higher in SP bicycle. This was corroborated by a psychophysical assessment based on RBG pain scale. The study also showed that there was a significantly lesser fatigue with the SU bicycle than the RF and SP bicycles. © 2013 Elsevier Ltd and The Ergonomics Society.