The former president's legal battle to have a $110,000 (£90,300) contempt of court fine that he was required to pay to the office of New York Attorney General Letitia James dismissed.
His failure to turn over documents during an investigation into his business practices led to the penalty, which was upheld by an appeals court.
For disobeying a court order, Mr. Trump was fined $10,000 per day.
If Mr. Trump inflated the value of his assets is being investigated by Ms. James' office.
Mr. Trump was required to provide business records as part of the civil investigation, which Ms. James launched in 2019. He also had to provide a thorough explanation of how he searched for the records and failed to locate them. He was found to be in contempt of court last year after failing to comply.
The former president eventually complied with the court order and paid the $110,000 fine, but he then filed an appeal in the hopes that the punishment would be overturned.
The fine "was not excessive or otherwise improper, under the particular circumstances," according to a ruling made on Tuesday by a New York state appeals court.
Ms. James, a Democrat, said in a statement on Tuesday that "the courts have once again ruled that Donald Trump is not above the law.".
He fought against our lawful investigation into his business dealings for years, but today's ruling makes it clear that abusing the legal system has repercussions. No intimidation or deterrents will deter us from seeking justice. ".
The investigation has been dubbed a "witch hunt" by Republican Mr. Trump. Alina Habba, Mr. Trump's attorney, did not immediately respond to a request for comment.
According to the lawsuit filed against Mr. Trump, his real estate business, and three of his children, they overstated the value of assets like real estate in order to obtain better financial terms and tax breaks.
The attorney general is requesting a $250 million fine as well as a lifetime ban on Mr. Trump acting as a corporate officer in the state.
The case brought by New York authorities is only one of the former president's numerous ongoing legal matters, which also include a Manhattan criminal investigation into the Trump Organization's practices, civil lawsuits over the 6 January 2021 Capitol riot, a criminal investigation into allegations he improperly retained classified documents, a defamation case brought by a journalist who claims Mr. Trump sexually assaulted her in the 1990s, and an investigation into allegations he engaged in obstruction of justice.