Seeking some leverage over a newly-assertive Moscow, European Union leaders agreed sanctions against top Russian politicians and stepped up efforts to cut the bloc's energy dependence on Russia.
Signing the Association Accord "symbolises the importance both sides attach to this relationship... and the joint will to take it further," EU president Hermann Van Rompuy told Ukraine interim premier Arseniy Yatsenyuk.
The EU was offering Ukraine its "steadfast support," Van Rompuy said, promising help to get the country's struggling economy back on track.
"We are sure that together we will succeed," Yatsenyuk said after the document was signed.
At a very different ceremony, in Moscow, President Vladimir Putin signed documents sealing Crimea's absorbtion into Russia.
"Today we have a serious, momentous event," Putin said.
"I want to congratulate you, all the inhabitants of the country, Russian citizens, the inhabitants of Crimea and Sevastopol on this landmark -- without any exaggeration -- event."
The annexation of Crimea, after a referendum on Sunday internationally condemned as illegal, prompted both the US and EU to impose sanctions on Russia.
US President Barack Obama blacklisted 20 Russian lawmakers, senior government officials and businessmen, in addition to 11 already targeted, and warned Moscow it faced international isolation if it did not reverse course.
Moscow responded in kind, announcing sanctions of its own against nine US officials, including top political figures and presidential aides.
After long talks in Brussels, EU leaders hit 12 more Russians with travel bans and asset freezes, bringing their list to 33.
Among them was Russian Deputy Premier Dmitry Rogozin, also targeted by Washington and who vehemently dismissed the measures.
"All these sanctions aren't worth a grain of sand of the Crimean land that returned to Russia," Rogozin said in a tweet.
But in Ukraine, former prime minister Yulia Tymoshenko, who was freed last month after spending three years in jail on what she said were trumped up charges, declared Putin had "lost Ukraine forever after declaring war on us".