Leaving the European Union would be an act of economic "self harm", Nick Clegg has said.
It comes after a Liberal Democrat Cabinet minister indicated David Cameron's commitment to an in/out referendum would make a second coalition with the Tories "incredibly difficult" to agree.
The Lib Dem leader, launching his party's election campaign, said it was "stating the flaming obvious" that the UK needed to be at the heart of Europe.
He said large parts of the Tory party were "straining at the leash" to leave the EU, potentially putting the economic recovery at risk.
Speaking in Abingdon, part of a key marginal seat the Lib Dems are hoping to win back from the Conservatives, Mr Clegg also hit out at Labour, saying he was "dismayed" by Ed Miliband's failure to spell out details of when he would balance the nation's books.
Energy Secretary Ed Davey, tipped as a potential successor to Mr Clegg if he is forced out after the election, said the issue of Europe could be a stumbling block in any fresh coalition negotiations with the Tories.
He told the Observer the Tory policy on Europe amounted to "economic and environmental irresponsibility of the highest order" and any deal that involved supporting an in/out referendum would be "incredibly difficult" to envisage.
Speaking in the Oxford West and Abingdon seat, won by the Tories with a majority of just 176 in 2010, Mr Clegg said: "Of course we have different views to the Conservatives. I'm the leader of the Liberal Democrats, David Cameron is the leader of the Conservatives.
"They have a completely different attitude towards Europe. They have this hokey cokey where one day they want to leave, the next day they don't want to leave.
"We are unambiguous as a party that says 'yes, Europe needs to be reformed, yes there needs to be a referendum when a decision needs to be made about the transfer of powers to the European Union, we guaranteed that in law'.
"We are not like the Conservative Party, half of whom are straining at the leash to leave. We think that would be a terrible thing for the British economy.
"But in the same way I strongly disagree with Ed Miliband and Ed Balls's failure to spell out when they would actually balance the books. I am dismayed the Labour Party want to borrow £70 billion more than we think is needed."
He added: "Of course it would be an economic act of self harm to leave the European Union.
"Large parts of the Conservative Party and Ukip appear to want Britain to cut itself off from what is the world's largest marketplace with hundreds of millions of customers for British goods and services.
"For me it's stating the flaming obvious that we need to secure Britain's presence at the heart of the European Union in the same way it's flaming obvious that we should secure the British economic recovery by finishing the job of balancing the books but doing it in a fair and sensible way - not taking an ideological approach to cuts, which is the Conservative approach, nor taking such an irresponsible approach to borrowing, which is Labour's approach."
Mr Clegg indicated that the rise of parties such as Ukip and the SNP could make any coalition negotiations more complicated than in 2010.
Asked if he would rule out a fresh deal with the Tories, he said: "I don't think it's for me to rule out how this country is governed. It's for the people voting in the ballot box to rule in what they want.
"Last time they ruled in only one combination - the Conservatives and Liberal Democrats together were the only combination of the sufficient numbers of MPs to govern in a stable and sensible way.
"Next time the cards might be dealt to us in a different way by the voters. But it's totally presumptuous for me, or indeed anybody else, to assume that we now know how people are going to vote."
Party strategists believe that winning over the female vote will be crucial to the Lib Dems' chances of success across a range of key battleground constituencies.
Despite their dire opinion poll ratings, they insist that their own internal polling shows women voters coming over to them in a series of must-win seats.
Their analysis of 18 marginals shows that the number of women who say they are undecided as to which way they will vote on May 7 has fallen from 30.6% in 2014 to 22.6% - a drop of 8%.
Over the same period they say that support for the Lib Dems in those seats has risen from 15.2% to 24.9% - an increase of 9.7%.
They say younger women particularly are attracted by their policy on shared parental leave, while headline plans to prioritise mental health, raise tax thresholds and cut the deficit are popular among women generally.
Party officials said that Mr Clegg's battlebus has been fitted with a satellite radio transmitter so that he can do interviews and radio phone-ins while he is on the road.
The bright yellow vehicle is decorated with abstract images by two graphic design students and the slogans "stronger economy, fairer society" and "opportunity for everyone", in contrast with 2010's campaign bus which featured prominent photographs of Mr Clegg and Vince Cable.