Day, however, made three bogeys over his last four holes and had to settle for a 70.
And the 37-year-old American joins 26-year-old Hughes at the top of the leaderboard after the 2016 RSM Classic victor fired a bogey-free round which included five birdies.
"I wouldn't have expected a round like that", said Hughes, who has missed four of his last six cuts. "I was thinking actually seven (or) eight-under after (my 11th hole) and I did give myself opportunities coming in, just, unfortunately, had a couple of mistakes", Day told reporters.
South Africa's Louis Oosthuizen and American Kyle Stanley are the leaders at nine-under, but numerous higher-rated players are off the pace, including world No. 1 Dustin Johnson, who is even-par.
Masters champion Sergio Garcia marked his return to action in spectacular fashion with a hole-in-one in the first round of the Players Championship.
Stanley has one PGA Tour win, the 2012 Waste Management Phoenix Open.
After playing just 13 tournaments previous year due to a foot injury, Poulter had 10 events this season to earn 218 FedEx Cup points or United States dollars 347,634 to remain fully exempt.
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Adam Scott admits he is "way back" on the leaderboard at the Players Championship in Florida but he is not ruling himself out of winning golf's unofficial fifth major for a second occasion.
Defending champion Jason Day plays with past champions Rickie Fowler and Henrik Stenson, while world No. 1 Dustin Johnson tees it up with Rory McIlroy and Justin Thomas in Thursday's opening round of The Players.
Stanley got his trouble out of the way early with three bogeys in four holes, and then was bogey-free the rest of the way. This is a course you have to just execute and hit great shots, and rather than go out and validate what I already know, I just worked on my game.
McGirt's flawless front nine, which yielded three birdies, inspired the American as he eagled the 11th and 16th holes in between bogeys at the 10th and the 18th. - You know what you expected out of this Players Championship. "Thanks to that and some great putts ... we were able to shoot a very good 5 under".
"I hit a good shot, but it can go there, it can go 3 yards past".
"It was very solid, but yeah, to go bogey-free around this place, very hard to do, obviously, and I started the day, I wouldn't have expected a round like that, but I knew my game was trending", Hughes added.
"It was very windy - not very consistent with the wind either", Garcia told Sky Sports after a round containing six birdies, three bogeys and an eagle on the 16th. Things got interesting down the stretch, though, as Holmes' final four tee shots weren't just off the fairway - they were off the map. I'd rather just get a nice practice session. "It's simple. When we get to the Masters next year, they're going to take it away, so I think I'm going to hold on to it as long as I can for this year". He managed to play a low punch shot with a draw, and it ran up to about 5 feet for a birdie.