The pair joked that they had a wager on which royal tumbles from their bike the most during the eight-day off-road adventure.
"It's going to be very challenging and we're expecting to fall off many a time," said Harry, 24, before more than 80 riders began the ride at the holiday resort of Port Edward on South Africa's southeast coast.
A crowd cheered the riders as they hit the winding dust road bound for the southern coastal city of Port Elizabeth.
William, 26, who next year will begin training to be an RAF search and rescue helicopter pilot, said he and his brother had decided to take part in the rally "because it's a mixture of adventure and charity."
Added Harry, "There may be an opinion that we are just riding bikes, but we are not. There is the whole background scenario of the charity side is hugely important to us."
Funds raised through the princes' participation in the Enduro Africa '08 event will be divided among UNICEF, the Nelson Mandela Children's Fund and Sentebale, a charity established by Prince Harry to help disadvantaged children in Lesotho.
They've helped raise more than $500,000 so far, according to CBS News correspondent Sheila MacVicar.
It also can't hurt their public images, she notes. The last time Harry, an officer in the Household Cavalry regiment, was seen riding a bike was in the desert of Afghanistan, where he had been deployed with his regiment.
Most often, MacVicar observes, they're pictured coming out of one expensive London nightclub or another, sometimes in the company of their girlfriends -- or sometimes in the company of a celebrity or two: William was snapped two days ago partying with Paris Hilton.
Harry said he was looking forward to spending some rare time together with his brother.
Both brothers ride high performance motorbikes when in the UK, but William said they had not trained for the event.
"We both ride bikes at the moment, but on-road biking is completely different. All the off-road stuff is up hills, down slopes, across rivers and is all rocky and hilly," William said.
Questioned about which royal was the better rider, William said, diplomatically, "We're both quite good, actually," while his brother quipped, "We'll have to wait and see in a couple of days time."