You really should try Windows 7 32bit. Think about it, DOS and Windows 3.1 applications shouldn't run on XP either. It's all running in a virtual machine included with XP (and 2K, NT4 before) and still included in Windows 7. The CPU goes into "Virtual 8086 mode" (used in Windows 3.x already to multitask DOS applications), and the main reason the virtual machine is not included in 64bit editions of Windows is that when the CPU is in 64bit mode, Virtual 8086 is inaccessible (that's the way AMD made the architecture).
Win 7 works well on old hardware too, I've seen it on a PC from 2002, it can actually use XP drivers. I put a 2000/XP driver for the network card, and an old Catalyst for the Radeon 9200. The installer for the driver will most probably fail but you can look at the extracted files (or extract yourself with a program that can deal with .exe, .cab etc.) and pick the .inf manually from the device manager, using a GUI unchanged since Windows 95.
The one software issue is you can't seem to be able to run graphical DOS apps (as well as not being able to fullscreen a text console), text mode DOS apps do work.
About serial ports I don't think there's a shortage of PCI and PCIe cards with them (plus RS232 still built-in on most motherboards, either at the back or on header). PCIe was designed to be software compatible with PCI by the way. Look at modern full ATX mobos with COM and LPT on the back (or no LPT if you don't need it) and at least two PCI slots. No shortage of them. Even DOS can still be run.
The only problem I can see is if you need ISA slots. Even then some Pentium 3 or Athlon hardware could be tried, and more recent stuff with ISA can be ordered it's just rare and more expensive.