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There are players which have hardware and software enhancements of which have obvious differences, for example iAudio X5 with BBE, To me it is something I prefer for music listening, BBE gives an Audio spatial enhancement a 3D effect for audio of which you cant do with equalizer settings, some player have Bass enhancements such as many of the Sony players do. If you set all the Equalizer settings flat on all players and use the same headphones for testing purposes you can audibly tell differences that are small to large from player to player, some players have more distortion than others, and some have a greater audio response time to the frequencies they are playing.
All players will have differences just because there hardware is different, for instance they have different “wattage”, voltage and current outputs of which will give perceptible audio differences. The software enhancements such as “BBE from iAudio creates difference in “time” of how the waveforms are reproduced on the player.
In answer to your question; Yes there will always be differences small or big that can be audibly perceived.
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