The President appeared to be responding to the Air Force's decision to order a review of all global layover stays over the weekend.
Trump tweeted on Monday that he knew "nothing" about one trip to the resort, but it was unclear which one he was discussing.
The crew, which consisted of active duty and national guard members from Alaska, was charged $136 per room, which was less expensive than a Marriott property's rate of $161.
Still, the Air Force acknowledged that "service members lodging at higher-end accommodations, even if within government rates, might be allowable but not advisable".
"In order to ensure compliance with the Emoluments Clause of the Constitution, the Department of Defense should not allow any of its officials to stay at Trump properties using taxpayer funds", they wrote.
CNN said it raises "ethics questions about government use" of the president's private dealings, while NBC News pointed out that the Turnberry resort is the "subject of an ethics investigation" by the House Oversight and Government Reform Committee.
A decision by the Pentagon to increase its use of the airport to refuel Air Force flights in 2015 had local airport officials sending flight crews to hotels in the area for overnight stays, including Turnberry, according to the paper.
In the years leading up to the 2016 election, it explains, Mr Trump spent hundreds of millions of dollars to purchase and renovate the Turnberry golf course, which continues to suffer financial losses and has not turned a profit for the President or his companies.
"Therefore, we are reviewing all associated guidance".
Trump said he has successfully pressured several countries to pay more of their North Atlantic Treaty Organisation commitments, as the USA routinely pays more than its own minimum requirement per year. The property had lost $4.5 million in 2017, but revenue went up $3 million in 2018. He said the airport is attractive because it conducts refueling 24 hours a day, typically has better weather conditions than other nearby airports and has less parking congestion than other European airports for crews stopping for fuel between the Middle East and the United States.
The Air Force defended the stop in Glasgow on the way to Kuwait on Saturday and offered additional details about the trip, which took place from March 13 to March 19. The vice-president stayed at the Trump International Golf Club in Doonbeg, on the west coast.
Speaking to reporters at the White House this evening, Mr Trump referred to Mike Pence's family links to Doonbeg and the fact that the US Vice President had previously stayed at the hotel years ago, before it was owned by the Trump Organisation.
"A long time ago", said Trump.
In the letter, the two Democrats also took issue with Trump's contention in a tweet Monday that he has "nothing to do with" the Glasgow Prestwick Airport. "Every time you find a person landing in an airplane within 500 miles of something I own - Mike Pence, as an example - his family lives in, he actually told me he stayed there many years ago".
