Reference #
The Reference # field is used to enter the Invoice Number received from your supplier. In many cases when your suppliers send you invoices, you'd like to be able to reference to them while creating bills. You can add their invoice numbers for reference in this field.
Purchase Order #
The Purchase Order # field is the number generated by your internal purchase order system that you share with your supplier prior to the service being delivered.
You can learn more about the fields on the Bill layout here, Getting Started with Bills.