Don't worry about performance - two disks are only better when you're working with them both at the same time, such as when copying from one to the other.
Or if you have two VMs active and working at the same time, having each on its own disk would improve disk throughput, on condition of having a multi-core (or multi-thread) CPU. But if both active VMs are on the same hard disk, then it's totally unimportant on which disk they are.
That said, here's some mathematics: 2 x 320 = 640 > 500. So you gain 140 GB.
If you go for the 2 disk solution, for me 320GB is too large for a system disk. I would go even further and partition the first disk into, say, 40 GB system disk with Windows and all the applications. This make it easier to backup an image of the system disk, which I do systematically just-in-case before I let Microsoft's Windows Update destroy my running system.
In that case I would have:
C = disk 1 partition 1 (40 GB)
D = disk2 (320 GB)
E = disk1 partition 2 (280 GB)
Please note that repartitioning the C drive will probably involve reinstallation of Windows (or restore from the OEM restore partition after re-partitioning).