The 29th-seeded American lost 6-4, 4-6, 6-2 to Spanish player Paula Badosa.
Stephens reached the quarterfinals at Roland Garros last year and the final in 2018.
She had reached at least the third round every year since 2012 except for 2017 when she was injured. Stephens won the U.S. Open later that year.
Stephens led Badosa 27-20 in winners but made 36 unforced errors compared to 18 for her unseeded opponent.
Badosa next faces 2017 champion Jelena Ostapenko.
Roberto Carballes Baena reached the third round at a Grand Slam tournament for the first time by beating ninth-seeded Denis Shapovalov 7-5, 6-7 (5), 6-3, 3-6, 8-6 in a French Open match that lasted five hours.
Shapovalov committed 106 unforced errors in the match compared to 42 for Carballes Baena and twice served for the match. The Canadian also had more winners than Carballes Baena with 65 to the unseeded Spaniard’s 31.
Carballes Baena sank to his knees after Shapovalov hit a forehand long and let out a loud shout of delight.
Carballes Baena will next play 18th-seeded Grigor Dimitrov.
Novak Djokovic’s biggest concern seemed to be dealing with a rare bit of sunshine during his second-round victory at Roland Garros.
The No. 1-ranked Djokovic got his team to procure him a hat so he could wear that while on the side of Court Philippe Chatrier that was brightest during his 6-1, 6-2, 6-2 victory over Ricardas Berankis.
Djokovic has dropped a total of only 10 games so far on his way to the third round in Paris. He won the title at Roland Garros in 2016 to complete a career Grand Slam.