Rose, who trails Harrington by $948 for the European Tour money title, nailed the ace with a 6-iron at the third hole. The Englishman also had three birdies and four bogeys at the Valderrama course.
Order of Merit leader Ernie Els is $313,892 in front of Harrington, last year's winner, but the South African is playing in Singapore on the Asian Tour and Rose and Harrington are likely to pass him with their winnings at the Volvo Masters.
'At the top of its flight I said 'go' and when it landed I said 'in' and it disappeared,' Rose said of his sixth hole-in-one.
Harrington recovered from being 4 over after the front nine to hole five of his six birdies and shoot a 71. He would have finished tied with Rose if not for a penalty stroke he received at No. 7 when his ball moved after he had grounded his putter.
'I'm not going to change my (putting) routine,' the Irishman said. 'These things, they happen.'
Graeme McDowell of Northern Ireland played the swirling winds best to head the field with a 3-under 68, leading 2005 champion Paul McGinley of Ireland by one stroke. The wind gusted to 25 mph and is expected to pick up for Friday's second round.
Peter O'Malley of Australia (70) was the only other player in the 55-man field to finish under par.
Rose, who was on an intravenous drip before his round because of a virus, was at 3 under by the fifth before bogeys at the seventh and ninth dropped him to 1 under.
At the 547-yard 11th, the Englishman pulled his second shot into the left rough, but recovered by sticking his wedge from 86 yards to within 15 inches for a birdie.
After three-putting for bogey at No. 14, Rose sank his birdie at the par-3 15th, but missed a par-saving putt at the last.
Harrington stuck his approaches at the 10th and 11th for easy birdies, and then curled in birdie putts at No. 16 and 17 from about seven feet to climb back from being six shots behind Rose after five holes.
'I think he helped me, no question about it. The fact that he was 3 under par through eight, nine holes ... certainly showed me that it was possible to do it,' the British Open champion said. 'He hit some good shots and no question, that was very positive affirmation to me that it could be done.'
Swedish players Niclas Fasth (75) and Henrik Stenson (76) are also in contention for the Order of Merit, while defending Volvo Masters champion Jeev Milkha Singh of India shot a 5-over 76.