Alexandr Dolgopolov refused to continue his match with Matt Ebden over a controversial call involving his shoe.
The World No. 37 was too good for the Aussie on Wednesday, battling out a 7-6, 6-3, 6-4 win.
However Dolgopolov was left fuming early in the second set when he came charging in on a drop shot and his shoe came off.
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The Ukrainian managed to return the ball, but Ebden hit a clean winner as Dolgopolov gave up on the point.
The chair umpire originally called a let and ordered the point be replayed, but after Ebden protested that the point should be his, the official changed her mind.
"I was too quick to call a let," she told Dolgopolov, sparking an angry reaction from the Ukrainian.
"I'm not going to play until you call the supervisor," Dolgopolov said.
"This is a joke."
Dolgopolov was then involved in a heated discussion with the match supervisor, who finally convinced him to continue playing.
Post-match Ebden, who had never encountered a loose shoe before, said the rule was clear.
"I don't know why she called a let as it's obstruction of play - and if obstruct play you lose a point, it's bad luck."
While Ebden won that game, Dolgopolov regained his focus to steam through the second set and take full control.
The Australian No.2 actually served for the first set, up 6-5, but lost 11 points in a row to hand it to his opponent.
"I didn't do all that much wrong so it was frustrating," Ebden said.
"I felt I was still playing well enough to come back in the second or third set ... there's fine lines."
Ebden, who upset 16th seed John Isner in round one, hoped he had done enough to earn a Davis Cup recall for next month's tie against Germany.
"I haven't been asked," the 30-year-old said.
"Could I be a chance, sure, but that's for the captain and coach to decide.
"I'd love to go - I'd love five or six of us to go and whoever is playing or feeling the best than choose the team from there."
With John Millman's loss earlier on Wednesday, Nick Kyrgios is now the last Aussie man standing at Melbourne Park.