Pramac rider Martin, who is 23 points ahead of reigning champion Francesco Bagnaia, clocked one minute and 31.707 seconds at Misano Adriatico, 0.037sec behind fellow title pretender Marc Marquez.
The 26-year-old has not won a Grand Prix since the French back in May but he looked in good form to repeat his triumph at Misano from last season.
Martin has also finished second in both the sprint and main race in four of the last five GPs.
Bagnaia, riding injured after crashing out of last weekend's Aragon MotoGP, finished 0.462sec back in eighth after a difficult session in which his bike spent most of the time in the Ducati garage with a mechanical issue.
The second practice session, from which the top 10 riders go straight into Saturday's second round of qualifying, gets underway at 1500 local time (1300 GMT).
Qualifying shapes the grid for both the sprint race on Saturday and the main event GP on Sunday afternoon, with a maximum of 37 points available.
