Almost two-thirds of the Dutch senate voted for ratification, with opposition coming mostly from far-left and far-right parties.
The country's EU allies were counting on the Netherlands to ratify the treaty signed in 2014 as it was the only EU country that had not yet done so.
With the senate's vote, the ratification process has been completed for the entire EU, although the treaty had already been applied provisionally.
With the war Ukraine's east showing no signs of abating, European Commission President Jean-Claude Juncker said, "The vote sends an important signal from the Netherlands and the entire EU to our Ukrainian friends: Ukraine's place is in Europe. Ukraine's future lies with Europe."
Juncker thanked the Dutch government and the leadership of other parties "for their efforts in bringing this process to a positive conclusion."
He added, "We are nearly there. Our association agreement, including the deep and comprehensive free trade area component, is now one step closer to being ratified. I would like to see the process now being finalised swiftly, in time for the EU-Ukraine Summit in July."
Juncker said the EU was fully committed to its partnership with Ukraine, describing it as "one of our closest and most valued" allies.
He said, "The association agreement has already increased trade between us, has brought increased prosperity for entrepreneurs, has helped to initiate and consolidate a number of reforms in Ukraine, and has brought new opportunities to EU and Ukrainian citizens alike.
"Let us harness the positive momentum generated by the Dutch vote to further strengthen our partnership."