Harriet Green said both votes are "unsettling" for companies seeking to create jobs and attract investment.
Ms Green said: "There are two political uncertainties that are most unsettling for business. The first is the Scottish referendum and the second is the European referendum. Both create massive uncertainty.
"For me, what we should be focused on is creating jobs. If you take the European market, the biggest trading market that we are part of and we are contemplating exiting that market? How could that be good for jobs?
"Many companies look at Britain and are questioning whether they will make some of the very sizable investments that they need to and want to until they know whether Britain is staying in the EU or not."
Ms Green also dismissed British Airways boss Willie Walsh's suggestion that Scottish independence could be a "positive development", due to the Scottish Government's pledge to cut and eventually scrap air passenger duty.
She said air passenger duty was a "relatively small part of that debate".
"It's not unimportant. If that issue was taken in isolation, I guess it would be a no- brainer," Ms Green said.
Voters in Scotland will go to the polls on September 18 to decide whether the country should leave the UK, while the Conservatives have pledged to hold an in/out referendum on a new settlement with the EU if the party wins the next general election.
Liberal Democrat MEP George Lyon said: "This is a significant intervention from the CEO of Thomas Cook.
"As a heavyweight organisation which trades across the UK and Europe, the SNP need to take these remarks seriously. Scotland is better off in Britain and in Europe."