The Spaniard, who has won five times at the circuit, set a track record on the resurfaced circuit the day before in practice and on Saturday was the only rider under 1 minute 47 seconds.
Marquez, who rides for Ducati satellite team Gresini, recorded a best lap of 1min 46.766sec a crushing 0.84sec ahead of his young compatriot Pedro Acosta of GasGaS.
World championship leader Francesco Bagnaia was a further 0.002sec back in third on his factory Ducati.
Jorge Martin crashed his Ducati-Pramac at the start of the session but raced back to the pits where he changed his bike but not his tattered leathers and returned to the track to finish fourth fastest.
Bagnaia leads Martin by five points in the riders' standings ahead of this 12th round of the season.
