Valentino Rossi began his home grand prix weekend with the fastest time in Friday practice at Mugello.
The Italian battled with his Yamaha teammate Jorge Lorenzo throughout the hour, but was always able to respond to the Spaniard's challenges, ending up 0.4sec ahead with his time of 1m49.751sec.
Loris Capirossi, who traditionally shines at Mugello, jumped to third late on, only to crash his Suzuki moments later when he lost the front end at the first corner, San Donato. He was unhurt.
Casey Stoner crashed early on, running into the gravel and staying upright at first, before allowing the bike to fall to avoid hitting the barriers. Thereafter he only completed a series of short runs, popping in and out of the Ducati pit for adjustments, and ended up seventh.
To complete a good day for Yamaha, Tech 3 riders Colin Edwards and Ben Spies claimed fourth and sixth, split by Honda's Dani Pedrosa.
Andrea Dovizioso was eighth on the second Honda, followed by Ducati's Nicky Hayden and a strong performance from Hector Barbera on the Aspar Ducati.
Pos Rider Team Time Gap
1. Valentino Rossi Yamaha 1m49.751s
2. Jorge Lorenzo Yamaha 1m50.120s + 0.369s
3. Loris Capirossi Suzuki 1m50.359s + 0.608s
4. Colin Edwards Tech 3 Yamaha 1m50.772s + 1.021s
5. Dani Pedrosa Honda 1m50.773s + 1.022s
6. Ben Spies Tech 3 Yamaha 1m51.004s + 1.253s
7. Casey Stoner Ducati 1m51.128s + 1.377s
8. Andrea Dovizioso Honda 1m51.255s + 1.504s
9. Nicky Hayden Ducati 1m51.387s + 1.636s
10. Hector Barbera Aspar Ducati 1m51.582s + 1.831s
11. Randy de Puniet LCR Honda 1m51.605s + 1.854s
12. Marco Simoncelli Gresini Honda 1m51.717s + 1.966s
13. Marco Melandri Gresini Honda 1m51.782s + 2.031s
14. Aleix Espargaro Pramac Ducati 1m52.202s + 2.451s
15. Hiroshi Aoyama Interwetten Honda 1m52.399s + 2.648s
16. Mika Kallio Pramac Ducati 1m52.418s + 2.667s
17. Alvaro Bautista Suzuki 1m55.257s + 5.506s