European Commission president Jose Manuel Barroso said states breaking away from existing EU countries would struggle to gain EU membership, further complicating Scottish nationalists' already uncertain plans for independence.
Mr Barroso said in an interview today it would be nearly impossible for the European Union to grant membership to such states - days after the British government said an independent Scotland would not be able to keep sterling as its currency.
Scotland is due to hold a referendum on independence in September. Polls show around 29 per cent of voters in favour and 42 per cent against, with 29 per cent undecided.
Mr Barroso, interviewed on BBC television, declined to comment directly on whether an independent Scotland would be welcome to join the EU.
But he said all EU states would need to back the membership of any new country that emerged from a current member state.
“It would be extremely difficult to get approval of all the other member states ... I believe it’s going to be extremely difficult, if not impossible,” he said.
The Scottish National Party (SNP), which is fronting the independence campaign, is banking on retaining both EU membership and the pound.
John Swinney, an SNP deputy in Scotland’s parliament, told the BBC Barroso’s comments were “preposterous” and that no EU state had indicated it would veto Scottish membership.
But secession is a sensitive subject for several other countries that have regions seeking to form their own states.
Spain, which Mr Barroso said in the interview had been "opposing even the recognition of (former Serbian province) Kosovo", is for instance wary that a vote for Scottish independence might encourage separatists in its Catalonia region.
Scotland's First Minister, Alex Salmond, also went on the offensive today against critics of the independence campaign.
Writing in the Sunday Times newspaper, he accused the British government of bullying over the currency issue and said he had asked British prime minister David Cameron to rein in his campaign to keep Scotland's 307-year union with the rest of Britain intact.
On Thursday, Mr Cameron's finance minister, George Osborne, warned Scotland it would have to give up the pound if it voted to end the union, declaring the currency could not be divided up.
The leader of the campaign to keep Scotland in the UK, former British finance minister Alistair Darling, said the independence campaign was beginning to unravel.
"Alex Salmond is a man without a plan on currency and Europe. The wheels are falling off the independence wagon," Mr Darling said.
Mr Barroso has previously said that any newly independent state would have to re-apply to join the EU. His comments are at odds with Scotland’s blueprint for independence, published last year, which says that it hoped to agree a “smooth transition” to membership of the EU as an independent state.
The Scottish government paper said they believed transition could be agreed without interrupting its EU membership in time for a potential independence declaration in March 2016.
Mr Barroso and other officials on the current Commission are due to step down when their term ends at the end of October but there is no evidence to suggest a new Commission would take a different view of Scotland’s membership rights.
Agencies