Support for remaining in the EU has fallen among business leaders, dropping from 60% to 54.1%, according to a survey from the British Chambers of Commerce.
In its final poll before the referendum, the BCC also said the percentage of those wishing to leave has risen from 30% to 37% since the last survey was conducted in February.
The data on voting intentions among 2,200 businesspeople highlighted divisions based on size and export interests.
Firms trading with other EU markets expressed the strongest support for remain, while the strongest levels of support for leave was seen among those that do not.
Those representing big businesses are "significantly more likely" to vote remain, the BCC said.
Dr Adam Marshall, BCC acting director general, said: "As the EU referendum campaign enters the final straight, the race for the business vote has clearly tightened.
"While only a minority of businesspeople report that the referendum campaign has had a material impact on their firms to date, much larger numbers say they expect significant impacts in the aftermath of the vote."
The Brexit campaign has revealed deep divisions in the world of business, with both sides wheeling out big hitters to make their case.
Most recently, former HSBC boss Michael Geoghegan and CMC Markets founder Peter Cruddas were among 110 signatories to a letter warning that Brussels poses a threat to Britain's financial services industry.
It followed a similar letter, signed by the likes of WPP's Sir Martin Sorrell and BT chairman Sir Mike Rake, backing Britain to remain a member of the EU.
Dr Marshall added: "Whichever outcome prevails, Westminster must shift its attention back to the economy on June 24 without delay.
"Growth is softening, and Westminster's referendum 'tunnel vision' over the past year has meant that far too many key economic issues have been given short shrift or delayed altogether."