The Scottish Liberal Democrats leader argued that independence would create a “border affect” with the rest of the UK that would damage existing trade relationships with other nations.
And he suggested a Yes vote would lead to the loss of some of the £85 billion worth of trade north of the Border currently supported by the UK government.
Mr Rennie also announced that the party’s former UK leader, Sir Menzies Campbell, would head a new commission setting out the party’s plans for more powers for Holyrood.
Sir Menzies will set out a “timetable for implementing lasting, permanent constitutional change after the referendum” in the event of a No vote, Mr Rennie said.
Speaking at the David Hume Institute last night, he claimed that independence would lead to a “whole new variety of red tape” for businesses that would deter companies from investing in Scotland.
He highlighted the example of the border between the US and Canada which, he argued, reduces trade involving the nations by 44 per cent.
Mr Rennie said: “Trade with Europe is worth £356bn to the UK economy. Three million jobs in Britain depend on it.
“Similarly, trade between Scotland and the rest of the UK is worth £85bn every year. These trade relationships are being put fundamentally at risk by the euro-sceptics and the nationalists.”
Mr Rennie also said “imposing borders that hit trade” would have a damaging affect on jobs and investment.
He said: “As part of Britain, Scotland has trade across that border worth £85bn. The recently published Scotland analysis paper from the UK government shows how that trade supports tens of thousands of jobs in Scotland.
“That report shows there is a ‘border effect’ which damages trade across any border, however allied the economies and benevolent the relationships. It happens without a physical border.
“If you have just spent years building your business and getting to grips with the red tape, the last thing you need is a whole new variety of red tape.”
However, Linda Fabiani, SNP MSP, insisted a Yes vote would be good for the economy.
She said: “The people of Scotland can achieve the powers we need to build a fairer society and more prosperous economy, and a new relationship of equality with our friends and neighbours south of the Border, by voting Yes in September.”