Valencia, Champions League finalists in 2000 and 2001, are one of a number of strong Spanish contenders, and Rafael Benitez's side have already enjoyed success in England this term -- beating Liverpool 1-0 in the first phase.
Arsenal, the English champions, looked out of form as they succumbed 2-0 at United in the premier league on Saturday.
But Wenger told reporters on Monday: "We will make it right tomorrow, don't worry.
"This game is the biggest test we've faced so far this season in Europe...Valencia defend well and are very clever on the break.
"But I don't like it when people question our spirit.
"The game (against United) on Saturday was one we could have won as well. We were not given a footballing lesson.
"We were disappointed but we are not worried."
Wenger said he was planning to call French compatriot Gerard Houllier, the Liverpool manager, to talk about Valencia's strengths and weaknesses.
After beating AS Roma 3-1 away in the first game of the second phase last month, a victory at Highbury on Tuesday would be a huge step towards qualifying for the knockout stages.
Arsenal head group B with three points, followed by Valencia and Ajax Amsterdam on one point apiece with Roma yet to get off the mark.
Wenger also refuted suggestions from United's Ole Gunnar Solskjaer -- made in British newspapers -- that Arsenal players had begun to 'squabble' with each other towards the end of their Old Trafford defeat.
"Our spirit is one of this club's assets," said Wenger.
He concedes that Valencia have helped make La Liga stronger than the premier league, but says he prefers the English top-flight for entertainment and satisfaction.
"Technique has made (Spanish clubs) the dominant force in the Champions League," said Wenger, whose side lost on away goals to Valencia in the 2001 Champions League quarter-finals.
"Spanish clubs know how to play, they don't hide and they are never scared. But I prefer the premier league -- for the speed, commitment and forward play."
Captain Patrick Vieira is doubtful for Arsenal through injury, but goalkeeper David Seaman returns after a groin strain.