Feliciano Lopez equalled Roger Federer's record of 81 main-draw appearances at grand slam events in the Open era but he was beaten in the first round at Wimbledon.
The 40-year-old made his grand slam debut at the 2001 French Open and had appeared in every major since the 2002 event in Paris until he failed to progress through qualifying at Roland Garros this year.
Spaniard Lopez went down in straight sets to Botic van de Zandschulp at SW19 on Tuesday.
Ranked 214 in the world, Lopez is a three-time quarter-finalist at the All England Club, while he also reached the quarter-finals of the US Open in 2015.
Since then, the veteran has never progressed further than the third round at a grand slam.
Fabrice Santoro is third on the list behind Federer and Lopez with 70 main-draw appearances at majors, with Mikhail Youzhny and Fernando Verdasco on 69.
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