The two Americans were virtually even off the blocks, but Crocker already had a sizable lead when their heads emerged from the water.
He was about a half-body length ahead at the turn, and didn't have to worry about Phelps making one his patented charges in the final 50 meters.
With arms pumping furiously, Crocker stretched out his advantage all the way to the wall. He touched in 50.40 seconds - easily eclipsing the mark of 50.76 that he set in beating Phelps at last year's U.S. Olympic trials.
But Phelps won the race that really mattered, overtaking Crocker at the Athens Olympics.
That didn't sit well with Crocker, who was determined to regain the upper hand in his trademark event, even though it meant beating the world's best swimmer.
"It was definitely my goal to break the record," he said. "I didn't know it would be by that much. When you're racing against Phelps, you always have to assume it's going to take a world record to win. My time definitely surprised me."
Crocker didn't have to worry about Phelps, who took the silver but wasn't even close to the winner at 51.65. Ukraine's Andriy Serdinov claimed the bronze.
"I haven't trained much fly this year," Phelps said. "But I don't want to use that as an excuse. That was a horrible swim for me." It was another big night for the Africans, an emerging force at these championships.
South Africa's Roland Schoeman captured his second gold by winning the 50 free in 21.69. That was just five-hundredths of a second off Alexander Popov's world record, but did beat Popov's meet record from 2003.
Croatia's Duje Draganja took the silver and Poland's Bartosz Kizierowski the bronze.
Zimbabwe's Kirsty Coventry completed a sweep of the 100-200 backstroke, winning the longer event in 2:08.52. She also has a silver from the 200 individual medley, making her one of the biggest female stars of the meet.
Margaret Hoelzer of the United States claimed silver, Japan's Reiko Nakamura the bronze.
Australia's Danni Miatke won the 50 butterfly, a non-Olympic event. The silver and bronze went to a pair of Swedish swimmers, Anna-Karin Kammerling and Therese Alshammar.
While Phelps has won four golds medals and, now, a silver at the world championships, he's not happy with his performance. He failed to qualify for the final of the 400 freestyle and was a disappointing seventh in the 100 free.
Two of his four golds have come on relay teams, and Grant Hackett is likely to win the most individual medals on the men's side. The Australian star already has two golds and a silver, and he's an overwhelming favorite in Sunday's 1,500 free.
"Everything that has happened this week is a wakeup call," Phelps said. "It's all going to be used for motivation. I don't want to be in a position where I'm disappointed and upset with a race. I want to be happy with the way I'm swimming. This year, hopefully, there will be big improvements."
Crocker wants to become the first swimmer to break the 50-second barrier in the 100 fly. He is only looking forward - not back to Athens.
"The Olympics didn't end quite the way I wanted them to," Crocker said. "I didn't happen to be the best on that day."