Introduction One of the major cues that help in auditory stream segregation is spectral profiling. Musicians are trained to perceive a fine structural variation in the acoustic stimuli and have enhanced temporal perception and speech perception in noise. Objective To analyze the differences in spectral profile thresholds in musicians and nonmusicians. Methods The spectral profile analysis threshold was compared between 2 groups (musicians and nonmusicians) in the age range between 15 and 30 years old. The stimuli had 5 harmonics, all at the same amplitude (f0 = 330 Hz, mi4). The third (variable tone) has a similar harmonic structure; however, the amplitude of the third harmonic component was higher, producing a different timbre in comparison with the standards. The subject had to identify the odd timbre tone. The testing was performed at 60 dB HL in a sound-treated room. Results The results of the study showed that the profile analysis thresholds were significantly better in musicians compared with nonmusicians. The result of the study also showed that the profile analysis thresholds were better with an increase in the duration of music training. Thus, improved auditory processing in musicians could have resulted in a better profile analysis threshold. Conclusions Auditory stream segregation was found to be better in musicians compared with nonmusicians, and the performance improved with an increase in several years of training. However, further studies are essential on a larger group with more variables for validation of the results.