Finishing with a time of 55.49 seconds, Peaty beat the previous record of 55.61 set by South African Cameron van der Burgh in 2009.
The 25-year-old also set a British record of 25.48 in the 50m Breaststroke during the event, as the London Roar franchise finished second in the competition and qualified for next week’s Grand Finale.
Peaty also secured another first-place finish in the Men's 4x100m Medley Relay, hitting the wall in 3:19.50.
Elsewhere, fellow Loughborough swimmers once again made their mark on the competition.
Marie Wattel helped Roar’s cause by finishing second in the Women’s 100m Butterfly with a time of 56.09 and also secured second-place finishes in both the Women's 4x100m Freestyle (3:26.64) and the Women's 4x100m Medley Relay (3:46.59).
New York Breakers’ Abbie Wood continued her recent fine form by claiming third place in the Women's 200m Individual Medley with a time of 2:05.27. She also finished in the top three of the Women's 400m Individual Medley, clocking 4:25.65.
Representing London Roar, Andreas Vazaios finished third in the Men's 200m Individual Medley with a time of 1:52.60. The Greece international also secured third place in the Men's 200m Butterfly, finishing in 1:52.13.
Rounding up a successful weekend for Loughborough, Max Litchfield (Energy Standard) took second place in the Men’s 400m Individual Medley (4.03.76), Molly Renshaw (New York Breakers) secured third place in the Women’s 200m Breaststroke (2:17.80), Luke Greenbank (London Roar) claimed fourth in the Men’s 200m Backstroke (1:49.31), and Felix Aubock (New York Breakers) finished in fifth in Men's 400m Freestyle (3:44.02).
The Grand Finale of the International Swimming League will take place on the 21st and 22nd November 2020.