The Prince arrived in Cape Town on Monday with his Walking with the Wounded teammates for a brief stop-over before beginning the 200-mile challenge.
Harry will trek alongside four injured service personnel representing Britain in a race across Antarctica.
They are competing against a team from the United States and one from the Commonwealth to be the first to finish the 335 km (210 mile) race. Alexander Skarsgard and The Wire actor Dominic West are also taking part as team ambassadors.
After landing in South Africa on a flight from London, adventurous Harry, 29, travelled with his team mates to a local hotel to relax for the evening.
On Tuesday, they will fly to an airbase near Russia's Novolazarevskaya Station, which is located at the edge of the Antarctic continent.
Once they have become acclimatised to the altitude and the freezing temperatures, the teams will begin the race. They aim to set off on 29 November, and reach the South Pole around 17 December.
Prince Harry is a patron of Walking with the Wounded and has shown great determination in championing the charity, which raises funds to retrain injured troops and help them find careers outside the military.
Last week, he launched the South Pole Allied Challenge in London's Trafalgar Square.
"All the men and women behind me have achieved so much just to get here, let alone actually walk to the South Pole, " he said. "It's just a wonderful display of courage on their part.
"These guys aim to achieve something quite remarkable, and in doing so will prove to everybody else that even though you've lost a leg or lost an arm… that you can pretty much achieve anything if you put your mind to it."
He added, "This trip is another way of reminding everybody that as Afghanistan draws down and these guys aren't in the news anymore… the support must continue."
The Walking with the Wounded teams will be trekking around 20km a day, pulling 70kg sleds. They are expected to endure temperatures as low as -35 degrees Celsius (-31 degrees Fahrenheit) and wind speeds of around 50mph (80km).
Harry also took part in the 2011 trek to the North Pole in 2011, but had to come home early so he could act as best man at his brother Prince William's wedding.