The guided-missile destroyer USS Dewey conducted a patrol within 20 kilometres of Mischeef Reef, part of the Spratly Islands over which several countries, including China, have competing claims.
The exercise is the first since October.
Pentagon spokesman Jeff Davis said, "We operate in the Asia-Pacific region on a daily basis, including in the South China Sea. We operate in accordance with international law."
The "freedom of navigation" operation is a signal intended by the US to assert its intention to keep critical sea lanes open, The Hill newspaper reported.
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"In conducting the freedom of navigation patrol, President Trump is likely to anger China at a time when the US is seeking increased cooperation with the country to help rein in North Korea," it said.
China claims almost all of the South China Sea. But Taiwan, the Philippines, Brunei, Malaysia, and Vietnam have rival claims over the region.
In February USS Carl Vinson Strike Group arrived in South China Sea but did not conduct Freedom of Navigation Operations (FONOPS) against Chinese maritime claims around its artificial-island bases in the Spratly and Paracel islands.
Early this month, Davis told foreign journalist that the FONOPS is a routine activity carried out by the US around the world.
"We did last fiscal year, freedom of navigation assertions against 22 different countries all over the world. Many of those countries are friends and allies," he said.
"We do these. We will continue to do them," the Pentagon spokesman said.
In an annual FNOPS report released by the Pentagon in February, the Department of Defence said that in 2016 it carried out freedom of navigation operations against 22 countries, including India. Other major countries were Brazil, Indonesia, Iran, Italy, Japan, Malaysia, Oman, Pakistan, Philippines, South Korea, Thailand, and Vietnam.