The dispute over the EU ban on beef from animals administered certain growth-promoting hormones dates back to 1996, with the new EU challenge coming after a WTO appeal body recommended that an earlier ruling against Brussels be re-considered.
"We are convinced that our legislation on hormones is fully in line with WTO law: the restrictions on hormone-treated beef are based on solid scientific evidence showing risks for human health," said EU spokesman Peter Power.
The World Trade Organisation in 1998 ruled that the EU had violated trade rules by banning the hormone-treated beef, thereby allowing the United States and Canada to impose trade sanctions on the bloc.
But the EU argued in 2003 that it had scientific grounding for the ban making the restriction valid under trade rules. However, the argument was rejected by the US and Canada and the two countries maintained trade sanctions against the bloc.
The EU, its earlier challenge to the sanctions having been rejected, brought the case again to the WTO on Monday after a WTO appeal body recommended that the question of the US and Canadian sanctions be reconsidered.