The Bon Accord in Glasgow won the title for the first time after impressing a series of drinkers who sampled its wares anonymously before reporting their findings back to the publication.
The pub sells about 380 different whiskies, which range in price from a few pounds for a nip to a £165 dram of 40-year-old Glenfiddich single malt.
It has won 27 different awards over the years for its range of beers and whiskies, but this is the first time it has gained the award bestowed by the Good Pub Guide.
Owner Paul McDonagh said: "What we have is quality and quantity, so there are whiskies for those who just want a nip or the real connoisseurs.
"There are two big growth areas in the industry at the moment - malt whiskies and cask beers, and we have both in abundance."
However, while the pub in North Street, near Charing Cross, is celebrating, the overall picture for UK hostelries is not so rosy.
According to research by the publication, up to 4000 pubs will close in the next year. The guide says pubs that are "stuck in the 1980s" offering indifferent drink and food will fall by the wayside and advises landlords to move with the times.
The guide says standards are improving in pubs, amid increasing vocational training among staff, but it wants bars to name their chefs on menus as part of moves to close the "absurd status gap" between TV chefs and those working in pubs.
The UK Pub Of The Year was the Olive Branch in Clipsham, Rutland. Other awards included the Best Landlord, which went to Tim Gray of the Yew Tree, in Lower Wield, Hampshire; Best Brewery was Fuller, Smith & Turner; and Best New Pub was Bulls Head, Mottram St Andrew, Cheshire;