If you import the file into the iTunes library then it will make a back up of it, so for example if the original file was on your desktop and then you import it, you can then delete the file on your desktop and not worry about it being gone.
Just give it a try, but back up the file first in another location and then see if it makes a difference when it's deleted, but as I said I'm pretty sure the file will be duplicated once added to iTunes.
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