In a speech in Florence, the head of the EU's executive arm warned of tough negotiations to come but also struck a more conciliatory tone after the first round of contacts on a Brexit deal degenerated into a cross-Channel war of words.
"Our British friends have decided to leave, which is a tragedy," the former Luxembourg Prime Minister told the European University Institute's State of the Union conference.
"I don't want anyone to underestimate the real importance of this sovereign decision of the British people. It is not a small thing but we must and will negotiate in all 'fairness' with our British friends.
"But I would like to recall here that there should be no doubt that it is not the EU abandoning the UK, it is the oppposite. They are abandoning the EU and this is a difference of status that must and will be felt over the next few years."
In a nod to longstanding British complaints about Brussels bureaucracy and perceived meddling in national affairs, Juncker acknowledged that the EU "has some weaknesses which can partly explain the outcome of the referendum in the UK."
"In the past, the EU has done a little too much, even the Commission: too many rules, too much interference in the daily lives of our citizens."
Juncker said his Commission had slashed the number of legislative proposals put to EU governments from 130 a year to 23 and was concentrating on the environment and programmes to boost trade, growth and jobs.
The only comment that could have been interpreted as a barb at Britain came at the start of his speech, when he briefly spoke in English to say it would be delivered in French.
"Slowly but surely English is losing importance in Europe," he said with a rueful smile, before explaining: "The French will have elections on Sunday and I would like them to understand what I am saying about Europe and about nations."
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Juncker's speech came at the end of a week which has seen Prime Minister Theresa May accuse Brussels of meddling in the British election campaign and another government minister describe the Commission as trying to "bully" London.
The spat was sparked by the leaking of details of a Downing Street dinner May hosted for Juncker which exposed frustration in Brussels about the British government's "delusions" over the divorce proceedings.
