The Louvre is facing accusations that it overcleaned a masterpiece by Leonardo da Vinci, leaving it with a brightness that the Renaissance master never intended.

Two of France's top art experts have voiced their protest over the cleaning of The Virgin and Child with Saint Anne – a jewel of western art – by resigning from the Paris museum's advisory committee responsible for its "restoration", the Guardian has learned.

Such was their concern for the 500-year-old painting that Ségolène Bergeon Langle and Jean-Pierre Cuzin – eminent former specialists in conservation and painting respectively at the Louvre – could no longer associate themselves with its treatment.

Bergeon Langle is regarded as France's national authority on the art and the science of restoring paintings. She was director of conservation for all of France's national museums.

She said: "I can confirm that I have resigned from the international consultative committee, but my reasons I am reserving for a meeting with the president-director of the Louvre, Henri Loyrette."

Cuzin, the Louvre's former head of paintings, declined to comment beyond confirming his resignation. But a senior museum source said the experts believed the restoration had gone too far, and that steps had gone ahead without adequate tests. The restoration has divided the committee between those who believe the painting is now too bright and those who regard the cleaning as moderate. There were also disputes over whether an area dismissed as removable repaint was in fact a glaze applied by Leonardo.

Two such resignations are a major embarrassment for the Louvre as well as for fellow colleagues of the international committee, whose 20 members include two specialists from the National Gallery in London, Larry Keith and Luke Syson.

The Louvre source said that Keith and Syson were particularly keen on this restoration: "The English were very pushing, saying they know Leonardo is extremely delicate but 'we can move without any danger to the work'. There was a row a year ago about solvents because they said they were safe and Bergeon Langle said they're not safe. It took a long time before the committee really had explanations on the chemicals used on the picture. Details were asked for [by the critics on the committee], but didn't come for months …

"There are people who are very much for bright hues and strong cleaning. Those people are in charge."

The Louvre source, too, has concerns that it has been overcleaned, but awaits the reaction once the painting is viewed again by experts on 3 January. They will then decide on the retouching.

Jacques Franck, consulting expert to the Armand Hammer centre for Leonardo studies at the University of California, Los Angeles, is another member of the committee. He described the resignations as a loss, saying Bergeon Langle was "totally irreplaceable as a technical adviser to the committee".

Seventeen years ago, the Louvre abandoned an earlier attempt to clean the painting amid fears over how the solvents were affecting the sfumato, Leonardo's trademark painterly effect for blurring contours. Since then, the British influence on restoration has helped to sway the Louvre.

Michael Daley, director of ArtWatch UK, the restoration watchdog that has repeatedly criticised the National Gallery for its "overzealous" cleaning of paintings, said of the resignations: "Implicitly, this is a vote of no confidence in the National Gallery cleaning policy because the most pro-active members of the [Louvre] committee have been the advisers from the National Gallery."

The Louvre declined to comment on the two resignations, but defended its cleaning process. Vincent Pomarède, the Louvre's head of paintings, said: "Rarely has a restoration been as well prepared, discussed and effected, and never will it have benefited from such effective techniques. The first assessment revealed the excellent state of conservation … comforting us in the choices made."

The National Gallery declined to comment.