Driver Layne Bremner's right lead horse, Duke, was injured in the sixth heat of the night.
"A veterinarian was on the scene immediately and determined the horse suffered a broken right hind leg – specifically a fractured cannon bone," wrote the Stampede in a release Saturday night.
Duke was a 10-year-old thoroughbred who had a previous career as a racehorse before Bremner bought the animal three years ago.
The injury appears to be due to a collision with another wagon. Bremner received a $2,500 penalty as well as a five-second penalty for interference.
The chuckwagon races are the deadliest Stampede event for horses, with more than 50 deaths over the last 20 years.
Animal rights groups have long called for the end of the rodeo and chuckwagon races being part of the Stampede.
In 2011, the organization changed its rules in an effort to make the sport safer after four horses died the previous year.
The number of outriders for each chuckwagon was cut from four to two to make the track less congested.
Pre-race health checks by veterinarians and rest days for the horses were also made mandatory.
But in 2012, three horses died in a crash after the lead suffered an aneurysm pulling the others down with it.
The Stampede says it is reviewing this weekend's incident to see if it was preventable.