Thomas beat Virginia’s Emma Weyant by 1.75 seconds with a time of four minutes and 33.24 seconds on Thursday, falling short of Katie Ledecky’s NCAA record time of 4:24.06.
Erica Sullivan, a silver medallist in the 1,500 metre event at last year’s Tokyo Olympics, finished third.
“It means the world to be here,” Thomas told ESPN.
Last month, USA Swimming unveiled a new policy to allow transgender athletes to swim in elite events by setting out criteria that aims to mitigate any “unfair advantages”.
Thomas competed on Pennsylvania’s men’s team for three years before transitioning and moving to the women’s team, setting multiple programme records. Her eligibility has come under considerable scrutiny, including from several Pennsylvania team mates.
“I try to ignore it as much as I can,” Thomas added. “I try to focus on my swimming, what I need to do to get ready for my races. And just try to block out everything else.”
Thomas will also compete in the 200-yard and 100-yard NCAA championships this week.