The five-times Olympic gold medallist's Asian World Cup series ended in more disappointment, although he shaved nearly a second off his time from his efforts in Beijing earlier this week.
Australian Thorpe clocked 53.59 seconds, way adrift of fastest qualifier Takeshi Matsuda of Japan (51.39).
Thorpe, returning after a five-year absence, remained confident he was on schedule in his bid to qualify for next year's London Olympics.
"This week has been challenging," he said. "I wish I could have done it with no one watching -- but that's unfortunately not the case."
Thorpe, who also failed to qualify for the men's 100 metre freestyle, added: "I am very comfortable with where I'm at."
The Australian also pulled out of the 100 metre individual medley in Tokyo after his setback in the men's 100 metre freestyle, an event he once dominated.