MacPherson was discovered while competing in the Kurnell triathlon in southern Sydney when he was 16. He soon landed the role of Joel Samuels in the soap opera Neighbours, starting in 1998. Soon after, MacPherson won a Logie Award in 1999 for Most Popular New Talent. After leaving the soap in 2002, MacPherson travelled to the United Kingdom to star in a British production of the musical Godspell in which he alternated the lead roles of Jesus and Judas with English entertainer Jonathan Wilkes. During this time MacPherson resided in Notting Hill with close friend Robbie Williams. After completing the show, MacPherson was offered the role of PC Cameron Tait in the British drama The Bill, which he played from 2003 until late 2004. Upon leaving The Bill, he took a role playing Jesus opposite Edward Woodward as God in The Mysteries. This was a drama based on a medieval mystery play cycle in which every scene moved to a different part of Canterbury Cathedral.

MacPherson starred in the pantomime Jack and the Beanstalk in 1999 at The Victoria Theatre in Halifax. He also starred in two pantomimes at the Marlowe Theatre in Canterbury alongside Paul Hendy and Leila Birch, Cinderella in 2000 and Aladdin in 2001.

In 2005, MacPherson returned to Australia to present the Australian version of The X Factor. MacPherson received very positive feedback, despite the series attracting low ratings. After the season had finished, reports stated that before one of the live shows MacPherson was accidentally knocked unconscious by a crew member. After regaining consciousness several minutes later, and being rushed to hospital, MacPherson went on to do the show that night despite being heavily concussed. He says he "remembers nothing at all" from that night's broadcast. This was undetected by the public. The following year, MacPherson hosted a three-part series called Killer Sharks on Australia's Network Ten in February 2006. In late 2006, he appeared in a British/Australian production entitled Tripping Over. The show appeared on Channel 5 in the United Kingdom and Network Ten in Australia. Tripping Over featured MacPherson's ex-Neighbours co-star Brooke Satchwell. Tripping Over received numerous positive reviews and the show was a hit with Australian critics; however due to funding issues it was not picked up for a second season.

In 2006, MacPherson co-hosted the Sydney New Year's Eve 2006-07 telecast alongside Big Brother host Gretel Killeen. The telecast caused much controversy after former Big Brother 2004 housemates Ryan Fitzgerald and Bree Amer appeared to be drunk throughout their segments of the evening. Notably, MacPherson received much praise for being a "complete professional" and left with his reputation unscathed.

MacPherson competes in World Triathlon Corporation (WTC) and Ironman 70.3. He competed in Ironman Australia at Port Macquarie where he missed out on qualifying for the Ironman World Championship in Hawaii by only one spot. His swim time of 49:30 in Ironman New Zealand 2008 was the fastest in his age group. His fastest Ironman remains 9-hour and 42 minutes in Port Macquarie in 2007.

In April 2009, MacPherson completed the China Ironman in Haikou. He completed the race in 11 hours and one minute, after a 3.8 km swim, 180 km bike ride and 42 km run in 45C heat. He came first in his 25–29 age group, qualifying him for the Hawaiian Ironman in October. MacPherson competed in the Hawaiian Ironman in 2009, finishing in 10 hours and 32 minutes. Macpherson has twice qualified for the Ironman 70.3 world championships, in 2013 and 2014.

MacPherson grew up in Sydney's beachside suburb of Cronulla, and has listed swimming, cycling, rugby, surfing, golf and watching the Cronulla Sharks, a rugby league football team, as his favourite pastimes. MacPherson went on to live in Melbourne for many years before relocating to Sunshine Beach, Queensland before returning to Sydney for work in 2010. He has been based in the USA since 2013.

He began dating his Wild Boys co-star Zoe Ventoura in 2011. They became engaged in December 2014, and married on the Sunshine Coast, Queensland in November 2015.

In 2016, MacPherson was a fill-in co-host on The Project with Carrie Bickmore.