The fine ebonised oak "ladderback" chairs cost only 17 shillings and six pence each when the famous tea rooms' owner, Miss Kate Cranston, ordered them more than a century ago.
Mackintosh designed the distinctive chairs in 1903 for the opening of the Willow Tea Rooms, in Sauchiehall Street, the following year.
The chairs were estimated to fetch at £10,000 to £15,000 as part of Lyon & Turnbull's sale of Scottish Design.
But yesterday they sold for £37,460 including buyers premium and the VAT on buyers premium at the Edinburgh auction house.
John Mackie, director and head of decorative arts at Lyon & Turnbull, said: "The chairs were used by Mackintosh to divide the space in the tea rooms and form an intimate environment within a much larger space. The chairs' design is informed by vernacular 'ladder back' chairs from the past. However, Mackintosh has taken this traditional chair form and made it his own."
The chairs in the auction came from a private seller in London.
The seller of the chairs, who has not been named, said: "I feel that I have been extraordinarily lucky as I was given the chairs by a friend when I was 21 years old, in the early 1960s.
"Since then I have used them every day, but after some careful thought and discussions with my wife, we have decided to sell the chairs."