Shod with fresh tyres, Miller’s best time came on his 19th lap in the dying moments of the first hitout.
He led Ducati satellite rider Alex Marquez for Gresini, who was just 0.036s in arrears on a 1:28.317s.
Marquez had a scare with Miller’s Pramac teammate Miguel Oliveira exiting the first turn, Doohan Corner, though the incident was not investigated.
Pedro Acosta showed some speed early, going third quickest for the factory KTM team with a 1:28.416s to end up 0.099s away from Miller.
Gresini’s Fermin Aldeguer was fourth ahead of factory Yamaha rider Fabio Quartararo.
Francesco Bagnaia was seventh behind Aprilia’s Marco Bezzecchi. The factory Ducati rider looked as if he was struggling with stability as he gesticulated to his team.
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The session was punctuated by Joan Mir’s Honda expiring late on, spilling oil through Doohan Corner.
Franco Morbidelli was the only rider to fall, dropping his VR46 Ducati at the Southern Loop.
MotoGP practice continues on Friday afternoon at 3pm AEDT, which will determine who goes through to the second leg of qualifying on Saturday.
Results: MotoGP Australian Motorcycle Grand Prix, Free Practice 1
| Pos | Num | Rider | Team | Bike | Lap | Diff | Gap |
| 1 | 43 | Jack Miller | Prima Pramac Yamaha MotoGP | Yamaha | 1:28.281 | ||
| 2 | 73 | Alex Marquez | BK8 Gresini Racing MotoGP | Ducati | 1:28.317 | 0.036 | 0.036 |
| 3 | 37 | Pedro Acosta | Red Bull KTM Factory Racing | KTM | 1:28.416 | 0.099 | 0.135 |
| 4 | 54 | Fermin Aldeguer | BK8 Gresini Racing MotoGP | Ducati | 1:28.466 | 0.05 | 0.185 |
| 5 | 20 | Fabio Quartararo | Monster Energy Yamaha MotoGP Team | Yamaha | 1:28.520 | 0.054 | 0.239 |
| 6 | 72 | Marco Bezzecchi | Aprilia Racing | Aprilia | 1:28.532 | 0.012 | 0.251 |
| 7 | 63 | Francesco Bagnaia | Ducati Lenovo Team | Ducati | 1:28.556 | 0.024 | 0.275 |
| 8 | 5 | Johann Zarco | CASTROL Honda LCR | Honda | 1:28.618 | 0.062 | 0.337 |
| 9 | 49 | Fabio Di Giannantonio | Pertamina Enduro VR46 Racing Team | Ducati | 1:28.631 | 0.013 | 0.35 |
| 10 | 33 | Brad Binder | Red Bull KTM Factory Racing | KTM | 1:28.654 | 0.023 | 0.373 |
| 11 | 25 | Raul Fernandez | Trackhouse MotoGP Team | Aprilia | 1:28.760 | 0.106 | 0.479 |
| 12 | 36 | Joan Mir | Honda HRC Castrol | Honda | 1:28.877 | 0.117 | 0.596 |
| 13 | 10 | Luca Marini | Honda HRC Castrol | Honda | 1:28.976 | 0.099 | 0.695 |
| 14 | 21 | Franco Morbidelli | Pertamina Enduro VR46 Racing Team | Ducati | 1:28.986 | 0.01 | 0.705 |
| 15 | 23 | Enea Bastianini | Red Bull KTM Tech3 | KTM | 1:29.019 | 0.033 | 0.738 |
| 16 | 44 | Pol Espargaro | Red Bull KTM Tech3 | KTM | 1:29.038 | 0.019 | 0.757 |
| 17 | 42 | Alex Rins | Monster Energy Yamaha MotoGP Team | Yamaha | 1:29.239 | 0.201 | 0.958 |
| 18 | 88 | Miguel Oliveira | Prima Pramac Yamaha MotoGP | Yamaha | 1:29.329 | 0.09 | 1.048 |
| 19 | 32 | Lorenzo Savadori | Aprilia Racing | Aprilia | 1:29.791 | 0.462 | 1.51 |
| 20 | 35 | Somkiat Chantra | IDEMITSU Honda LCR | Honda | 1:30.309 | 0.518 | 2.028 |
| 21 | 51 | Michele Pirro | Ducati Lenovo Team | Ducati | 1:30.645 | 0.336 | 2.364 |
| 22 | 79 | Ai Ogura | Trackhouse MotoGP Team | Aprilia | 1:30.755 | 0.11 | 2.474 |
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