Race director Jean-Francois Pescheux said the small metal tacks were spread out over the course of about one kilometre during the climb up the Mur de Peguere about 40 kilometres (25 miles) from the finish, with defending champion Cadel Evans among those to suffer a puncture.
However, the Sky Team - whose leader, Bradley Wiggins, is in the yellow jersey - encouraged the pack to not speed ahead or take advantage of the mishaps of the affected riders, and Evans was eventually able to catch up to the peloton after getting a new wheel.
"Sky immediately stopped the pack so that everyone could finish in the best conditions," Pescheux said. "Sky were very sporting, they slowed things down and everything returned to order."
Astana rider Robert Kiserlovski had to drop out of the race after breaking his collarbone in an accident related to the tacks. Kiserlovski was helping teammate Janez Brajkovic change a punctured tire when American rider Levi Leipheimer crashed into him, Astana press officer Valentina Quattro said.
Leipheimer was able to continue and finished in the main pack with Evans and Wiggins. Kiserlovski had been nearly 21 minutes behind Wiggins overall going into the stage, and was not a contender for the title.
Pescheux said the search for the culprit would be "difficult" because thousands of fans were on the edge of the road up to the Mur de Peguere climb as the riders came by.
Though race organizers and local officials roll out street sweepers ahead of the peloton to clear the roads of debris, the tack incident underscores the difficulty of securing the Tour route - this year 3,500 kilometres (2,175 miles) long - where tens of thousands of fans line the course every day.
From time to time, stray dogs or photograph-snapping fans get hit by speeding riders on the route. On Friday, Wiggins was hit on the arm and received minor burns by a flare being waved by a spectator.
Stage winner Luis Leon Sanchez and several other breakaway riders were many minutes ahead of the pack and appeared to be spared the sabotage during Sunday's 191-kilometre (119-mile) stage between Limoux and Foix.
Evans had to stop right under the banner marking the summit of the Mur de Peguere to take his punctured rear wheel off. As he waited for a team car to give him a replacement, he flapped his hand in frustration, then clapped - apparently trying to keep his spirits up.
By the time Evans got back on his bike, he'd lost about 2 minutes to the pack, and Sky riders were seen pressing their earpieces - as if learning of the Australian rider's misfortune, and slowed up.
Evans, thanks to an escort from BMC riders who waited for him, eventually caught the main pack.
"We're going to see with police gendarmes at the finish line whether we can find the person who threw all that," Pescheux told France-2 television, as the pack was finishing.
Race etiquette generally dictates that when top riders face mishaps beyond their control - Evans is in fourth place overall, 3:19 back of Wiggins - their biggest rivals for the yellow jersey don't try to exploit them.
BMC team manager Jim Ochowicz said Evans actually got three flats because of the tacks. French TV pictures showed shiny, metal tacks imbedded in a tire of a motorcycle in a convoy that accompanies riders.
"There were people that even crashed as a result," said Ochowicz. "I wasn't angry - there was no one to be angry at - it was more important to get Cadel back in the race."
"It's a criminal act, I think you're taking people's lives in your hands ... and creating a very dangerous situation," he added.
Evans took it a bit more in stride.
"That's the way things go in life, karma hopefully comes around," he said. "Couldn't see them on the road ... this has happened to me before - two times in Spain" where he lost races like the Vuelta as a result, he said.
"That's why I don't race in Spain very often," Evans added. "Sorry for the good Spanish people and my Spanish friends and people in Spain who support me but there's a few people that just take things too far."
Associated Press writher Jamey Keaten in Toulouse, France, contributed to this report.