Irish horse The Cliffsofmoher has been euthanased on the Flemington track after breaking its shoulder during the Melbourne Cup.
The race was won by Cross Counter, ahead of Marmelo, A Prince of Arran and Finche.
The Cliffsofmoher suffered its injury early in the race and quickly fell behind the field, failing to finish.
As the remaining 23 horses passed the post, The Cliffsofmoher received medical attention before vets determined it was "unable to be saved".
"This was an unfortunate incident that happens infrequently, with Victoria having one of the best safety records in world racing," a Racing Victoria statement read.
"Our sympathies are extended to Coolmore and the Williams family, the owners of The Cliffsofmoher, jockey Ryan Moore, trainer Aidan O'Brien and his staff, who cared for the horse and are greatly saddened by their loss."
Mr O'Brien said he was relieved Moore was unharmed.
"Unfortunately these things can happen to a horse galloping around the field at home," he said.
"It's very sad. It could have been worse, Ryan [Moore] could have taken a fall off him, someone could have been seriously injured."
The news sparked an instant and angry reaction from animal welfare organisations and protesters alike.
People for the Ethical Treatment of Animals (PETA) released a statement demanding an investigation into the death of The Cliffsofmoher.
"He's the latest in a long line of dead horses. Cliffsofmoher was spotted looking very agitated moments before the start and has now been killed after fracturing his right shoulder in the Melbourne Cup," the statement read.
"Before they've even finished maturing, these 500-kilogram animals are forced to race at breakneck speeds while being whipped and pushed past their limits, supported on ankles as small as those of humans.
"Of course, horses die at lower-profile racing events all the time: during the last racing year 119 were pronounced dead on Australian tracks between August 2017 and July 2018 - that's one animal every three days.
"They die of cardiac arrest, haemorrhaging, ruptured aortas, and broken necks, legs, or pelvises, and that's without mentioning the thousands of horses bred for the industry who don't make the grade and are abandoned, neglected, or sent to slaughter.
"Considering Australians hate cruelty to animals, commemorating a day on which horses routinely die in the Melbourne Cup is fundamentally un-Australian. While public holidays give Aussies a break, horses are breaking legs."
The Coalition for the Protection of Racehorses followed suit in a Facebook post.
"As only a five-year-old, [The Cliffsofmoher] had won $1.7 million for his owners. Today he broke his shoulder in the Melbourne Cup and was killed on the track," the post read.
"No horse deserves to die for gambling profits, but it happens every three days, and we are so sorry."
The Cliffsofmoher becomes the 6th horse to die as a result of the #melbournecup since 2013. Highlights the very real risks to horses from racing.— RSPCA Australia (@RSPCA) November 6, 2018
The CliffsofMoher's shoulder broke early on in the race.— Animals Australia (@AnimalsAus) November 6, 2018
He was killed on the track.
RIP to another beautiful horse. He was 5 years old. #MelbourneCup #LoveHorsesNotracing #NuptotheCup #HorseRacingKills pic.twitter.com/ZmOLIPMSRR
The Cliffsofmoher is the latest horse to have died on Melbourne Cup day in recent years, joining Verema (euthanased on track in 2013), Admire Rakti (suffered a heart attack in his stall in 2014), Araldo (euthanased after breaking his leg post-race in 2014), Red Cadeaux (euthanased two weeks after falling in 2015) and Regal Monarch (euthanased after a fall in Race Four of 2017).
Earlier on Tuesday, the hashtag #NupToTheCup was trending in Australia as protests were held across the country.
One, led by the Coalition for the Protection of Race Horses and called the Melbourne Cup Protest and Picnic, was promoted as "a great way for those who don't want to support animal cruelty to still enjoy a celebration".
"While those in the bird cage at Flemington Racecourse will be sipping on champagne and nibbling on canapes, Nup to the Cup attendees will do the same and will at the same time be drawing attention to the plight of racehorses who don't have a say," a post on the Animals Australia website said.