Defending US Open champion Murray, the world number two, started strongly but faded thereafter and crashed to a 6-3, 6-4 defeat against Tomas Berdych of the Czech Republic.
World number one Djokovic had the chance to make history this week by becoming the first player to win all nine Masters titles, but his hopes of completing the set in Cincinnati were dashed after home favourite John Isner came through 7-6 (7/5), 3-6, 7-5.
It is a bitter blow for Djokovic’s US Open preparations, although arguably his biggest rival at Flushing Meadows, Murray, fared no better as he came off second best against Berdych.
World number six Berdych had triumphed in the pair’s last encounter at the Madrid Masters in May but Murray defeated the Czech en route to his Flushing Meadows win last year.
And it seemed the Wimbledon champion was at the top of his game yesterday after Berdych barely escaped his opening service game before the Scot opened his account with four aces.
The two-time grand slam champion continued to pressurise his opponent’s serve, but Berdych held firm and then broke in the eighth game after Murray netted a forehand.
Berdych served to love to wrap up the first set and then staved off three break points in his next service game. It proved to be a crucial hold as Berdych then broke 2008 and 2011 Cincinnati winner Murray in the next game at the second attempt.
Despite fleeting moments of quality, Murray failed to force his way back into the contest and a composed Berdych held his nerve to set up a clash with either Rafael Nadal or defending champion Roger Federer.
Djokovic needed to save a break point in his opening service game against the big-hitting Isner, who suffered a surprise first-round loss to Vasek Pospisil in last week’s Rogers Cup.
The match went with serve for the remainder of the set and it was only a Djokovic double fault in the tiebreak that handed Isner the initiative, with the 28-year-old taking full advantage.
Isner once again had the opportunity to break his opponent in the opening game of the next set, but reigning Australian Open champion Djokovic showed his mettle.
Indeed, the Serbian would save seven break points before converting one of his three chances to square the match.
Despite Djokovic taking the momentum into the decider, Isner refused to wilt. The American made his march in the 12th game, but while Djokovic saved the first match point with an ace, he double faulted at the crucial time once more and Isner sealed a memorable win.
Juan Martin del Potro rallied after losing the second set to roll past qualifier Dmitry Tursunov 6-4, 3-6, 6-1 to reach the semi-finals. The seventh-seeded Del Potro, who won the 2009 US Open, reached the semi-finals at the event for the second consecutive season.
The 44th-ranked Tursunov was trying to become the first qualifier to reach the tournament’s semi-finals since David Wheaton in 1994. He won the second set after Del Potro double-faulted to set up break point. Del Potro then volleyed a short Tursunov lob into the net, giving the qualifier a 5-3 lead.
Del Potro responded by breaking Tursunov in the second game of the third set on his way to a 3-0 lead. Another break gave del Potro a 5-1 lead, and he clinched the win with an ace at love.
Meanwhile, Maria Sharapova has confirmed she has severed ties with coach Jimmy Connors after one match together. The Russian world No 3 said in a statement emailed to Tennis.com that the pairing had not been “the right fit for this time in my career”.
Sharapova had hoped her partnership with the fiery eight-time grand slam champion would produce the same type of success Murray has enjoyed since recruiting Ivan Lendl, another former playing great, as his coach. But instead, the brief experiment ended after one match, a second-round loss to Sloane Stephens in Cincinnati on Tuesday.
Looking on from the stands on Tuesday, Connors, 60, could only hang his head as he watched his student unravel in a three-set defeat. The split leaves Sharapova without a coach heading into the US Open which starts on 26 August.