The vote was 52 to 48 along party lines.
Senators convened at the unusual, early hour -- 6:30 a.m. ET -- because of Democratic stalling tactics that reflect their unhappiness with Trump's cabinet picks and the overall confirmation process.
DeVos has come under fire from Democrats and two key Republicans for her positions on public and charter schools, as well as statements made during her confirmation hearing last month.
Despite losing support from Republican Sens. Susan Collins of Maine and Lisa Murkowski of Alaska, she is expected to be confirmed Monday with the help of Vice President Mike Pence, who will be in the chamber to break an expected 50-50 tie.
Collins and Murkowski, who both have said they intend to vote against DeVos' final conformation, voted to end the debate and advance her confirmation, as expected.
In an impassioned show of opposition, Senate Minority Leader Chuck Schumer implored other senators to "look into their conscience" over the weekend and reconsider their support for DeVos, saying she is "one of the worst nominees that has ever been" brought before the Senate.
"Sometimes loyalty to a new president demands a bit too much," he said. "With this nominee it does."
Following the vote, Sen. Patty Murray, a Democrat from Washington State and the ranking member on the education committee that held a hearing on DeVos' nomination, took to the Senate floor to deliver a 25-minute speech blasting the nominee, a Michigan billionaire, not only on her positions but over questions about her finances and conflicts of interest.
Republican Sen. Lamar Alexander, the chairman of the education committee, also appeared on the floor to defend DeVos, citing her work in the conservative education reform movement over the years. "I'm pleased to support her," he said, adding that he looks forward to the final vote on her nomination early next week.
In an interview with CNN after the vote, Sen. John Cornyn of Texas -- the No. 2 Republican in the Senate -- said the Democrats' concerns weren't "particularly fair" and argued that their ties to teachers unions are influencing them politically.
"If people think our public education system is perfect, then I guess they don't think we need to have any changes or any choices for students and their families," he said. "I certainly think we do."