A number of protesters unloaded large hay bales ahead of the riders and sat on the road in front of official vehicles. Gendarmes came in to remove the protesters and shortly after a number of riders were left clutching at their eyes in visible distress.
Teams and French media said police used tear gas to disperse the protesters but it ended up blowing back into the main bunch of riders, which led to the race being halted with 187km to go.
Several riders were later seen washing out their eyes before it resumed some 10 minutes later.
Current leader Geraint Thomas, four-time champion Chris Froome, both of Team Sky, and world champion Peter Sagan (BORA-hansgrohe) were among those who had to seek medical treatment after tear gas was used to break up a protest.
The farmers were said to be protesting recent administrative changes to disadvantaged and difficult-to-farm areas.
Gendarmes remove hay bales that were placed on the stage's route. (Getty) Source: Getty
The race continued to its final destination of Bagnères-de-Luchon, with an eventful stage won by Julian Alaphilippe (Quick-Step Floors).
After the finish, riders were perplexed at the circumstances of the protest and the resultant gassing of the riders.
"What was it, mace? That's a new experience," said tenth-placed Dan Martin (UAE Team Emirates). "I'm always open to new experiences but pepper spray during the race... you could feel it in the lungs there for a bit - it wasn't exactly pleasant.
Gendarmes detain a protester on Stage 16 of the 2018 Tour de France (Getty) Source: Getty
"But they dealt with it pretty well and the medical team gave the guys who needed it some attention. Bizarre though."
Race leader Geraint Thomas was one of those gathered by the medical car after the incident but suffered no lasting effects.
Geraint Thomas was one of the riders affected by the pepper spray used to quell protests on Stage 16 of the 2018 Tour de France (Getty) Source: Getty
"Obviously it was unfortunate that whatever they used was lingering when we came through," said Thomas, "but fortunately it didn't affect me too much. I had a bit of a tingle in the eyes and the back of the throat, so I used a bit of water to have a quick wash out but I was ok in the end."
The Tour de France continues to Stage 17 in the Pyrenees, with a short,spectacular stage set to be run with a trial grid system start.
Coverage of Stage 17 will start from 10.35 pm (AEST) on SBS HD, also avaliable on the SKODA Tour Tracker App and via live web streaming on SBS Cycling Central.
