Police warned the crowd multiple times to clear the intersection of High and State streets in downtown Columbus to make way for traffic, according to ABC News affiliate WSYX.
Video posted on Twitter around midnight showed police officers in gas masks spraying the crowd that gathered near the Ohio Statehouse brandishing anti-Trump signs and chanting.
"Several of our members were part of the group who were pepper-sprayed, but many other groups showed up in solidarity," Kyle Landis, one of the event's organizers, told ABC News on Tuesday morning.
The Columbus Police Department did not immediately respond to ABC News' requests for comment on the matter.
His controversial executive order on immigration temporarily bars entry into the U.S. for people from seven predominantly Muslim countries and has been meet with protests in several cities.
The demonstrators began gathering on Monday evening at City Hall as Columbus Mayor Andy Ginther announced that the city will continue to accept refugees regardless of their immigration status.
"Our immigrant population is part of the fabric of what makes Columbus so vibrant," he said in a statement. "They are our neighbors. We must not turn our backs on them — now or ever."
Ginther said he will issue an order of his own this week that will "support the resettlement of refugees to Columbus and prohibit the detention of anyone unless a warrant exists or a criminal violation was observed," according to the city's website.