The Japanese, who shares the number seven TS050 hybrid car with France's Stephane Sarrazin and Britain's Mike Conway, lapped with a best time of three minutes 18.793 seconds.
The number eight Toyota of Britain's Anthony Davidson, Japan's Kazuki Nakajima and Switzerland's Sebastien Buemi was second and 0.638 slower.
Toyota have never won Le Mans, with Mazda the only Japanese manufacturer to have triumphed back in 1991.
Champions Porsche, winners for the past two years, were third fastest with the number two car of Germany's Timo Bernhard and New Zealanders Brendon Hartley and Earl Bamber.
The session was shortened by 20 minutes to allow for barrier repairs at the Porsche Curves after a crash in an earlier practice.
(Reporting by Alan Baldwin in London, editing by Nick Mulvenney)
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