Boris Johnson described the protection of school playing fields as vital for the health of young people today.
The mayor of London said the policy within the capital was to preserve existing playing fields, except when replacements are to be provided elsewhere.
He said: "We don't believe in selling off playing fields and I think if you look at what we do in London, we have very clear rules that you can't sell off green space, you can't sell off playing fields.
"What you can do if you do need to expand a school - and there's huge pressure on school places - you can do that provided you increase the availability of playing fields, so you've got to have new provision of playing fields elsewhere."
Mr Johnson, who recently called for schoolchildren to do two hours of sport every day, added that London's schools do not have sufficient outside space.
"Playing fields are absolutely vital for the health and wellbeing of young Londoners.
"Many schools don't have enough, many parts of inner London simply don't have enough playing space, and it's vital that we conserve what we have."
The mayor made the comments at the Sea Life London Aquarium, where he was promoting a range of summer discounts on attractions, hotels, restaurants and theatres in the capital.
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