The UK's EU referendum represents a "once-in-a-lifetime decision", Chancellor George Osborne has said.
He told Newsnight it was "unrealistic" to assume the poll - likely to be held later this year - would be repeated.
Mr Osborne, who described himself as a Eurosceptic, said he was "optimistic" about reaching a deal on EU reforms.
Jean-Claude Juncker, head of the European Commission, has suggested a deal is likely in February - which would allow for a referendum in June.
Speaking in Brussels, Mr Juncker said the negotiations were not easy but they had to deliver an agreement satisfactory to all sides.
"I'm quite sure that we'll have a deal - not a compromise, a solution, not a mere compromise - a permanent solution in February."
David Cameron has promised an in/out referendum on EU membership by the end of 2017 but the exact timing of the poll hinges on when agreement is reached with the EU's 27 other members over the UK's proposed package of reforms, including curbs on EU migrants' welfare entitlements.
The EU referendum
Timeline: What will happen when?
Explained: What does Britain want from Europe?
Analysis: Cameron tries to avert slanging match
Ministers are hopeful of getting a deal at next month's European Council meeting. If this does not happen, that would effectively rule out the referendum taking place until the autumn at the earliest.
The BBC's political correspondent Ben Wright said the European Commission was a key player in the process and while progress could easily be derailed over the next six weeks, Mr Juncker was "clearly confident" of a consensus next month.
'Falling into place'
Mr Osborne told the BBC that he saw "the essential pieces of the deal falling into place" and insisted the Treasury was "100% focused" on the talks, rather than planning for a UK exit from the EU.
While some of the alternative arrangements to remaining within the EU "do not look very attractive", it was vital the UK was not "discriminated" against outside the eurozone and would be demanding a permanent guarantee that it would not have to contribute to future eurozone bailout payments.
He added: "I've been concerned about some of the things that have happened in the European Union, that's why I want to make those changes.
"It's a perfectly respectable position to say 'let's seek those changes, let's achieve those changes, let's have that new settlement, and then we can have the best of both worlds'.
"We can be in the European Union, but not run by the European Union."
Mr Osborne, a key member of the UK's negotiating team, quashed talk of the possibility of a second referendum if the outcome of the poll was close.
"I think anyone who votes out on the assumption that a year or two later you can have another vote to vote back in... is being unrealistic about the nature of the choice," Mr Osborne said.
"And I think it's really important that the British people focus on the fact this is the once-in-a-lifetime decision."
Mr Cameron has said his ministers will be able to campaign for either side in the referendum, but must back the government until negotiations are complete.
In an intervention seen as the first sign of a minister preparing to campaign to leave the EU in the UK's referendum. Leader of the Commons Chris Grayling said staying in the EU under the current terms would be disastrous,.
Mr Grayling said the UK was at "a crucial crossroads" and "cannot be left in a position where we have no ability to defend our national interest" within the EU".
He backed Mr Cameron to secure the reforms he is demanding, a stance supported by Mayor of London Boris Johnson.
Mr Johnson has said he remains "very confident" the prime minister will "get a good deal" for the UK in his renegotiation with EU leaders.
Asked about Mr Grayling's article in the Telegraph, Mr Johnson said: "He's totally right to say unless you get reform, Europe will continue to be a zone of low growth and stagnation."
You can watch Newsnight's full interview with George Osborne on iPlayer .