The surprise disclosure triggered immediate calls for US action by members of Congress who advocate deeper American involvement in Syria's bitter civil war.
The White House said the US intelligence community assessed with varying degrees of confidence that the chemical agent sarin was used by forces allied with President Bashar al-Assad. But it noted that "the chain of custody is not clear."
While Obama has declared that Syrian use of chemical weapons would be a game-changer, his administration made clear it would move carefully - mindful of the lessons of the start of the Iraq war 10 years ago.
Then, the George W. Bush administration used faulty intelligence to justify invading Iraq in pursuit of nuclear, chemical and biological weapons that turned out not to exist.
"Given the stakes involved and what we have learned from our own recent experiences, intelligence assessments alone are not sufficient - only credible and corroborated facts that provide us with some degree of certainty will guide our decision-making," Miguel Rodriguez, White House director of the office of legislative affairs, said in a letter to lawmakers.
One senior US defense official told reporters that, "We have seen very bad movies before," where intelligence was perceived to have driven policy decisions that later, in the cold light of day, were proven wrong.
The term "varying degrees of confidence" used to describe the assessment usually suggests debate within the intelligence community about the conclusion, the defense official noted.
The White House said the evaluation that Syria probably used chemical weapons was based in part on physiological samples, but a White House official declined to say what kind of evidence it had, such as soil samples or blood or hair from victims.
The scale of the sarin use appeared limited, with one US intelligence official noting that nobody was "seeing any mass casualties" from any Syrian chemical weapons use.
The United States has so far resisted being dragged militarily into Syria's conflict, providing only non-lethal aid to rebels trying to overthrow Assad, given concerns that weapons end up the hands of al Qaeda-linked opposition fighters.
But acknowledgement of the US intelligence assessment appeared to move the United States closer to some sort of action, military or otherwise.
A White House official told reporters "all options are on the table in terms of our response" and said the United States would consult with allies.
SURPRISE ANNOUNCEMENT
The Obama administration's sudden disclosure caught many off-guard, coming just two days after Defense Secretary Chuck Hagel and other US officials appeared to play down an Israeli assessment of chemical weapons use.
France and Britain have also concluded evidence suggests chemical arms have been used in Syria's conflict.
"The intelligence community has been assessing information for some time on this issue and the decision to reach this conclusion was made within the past 24 hours," Hagel said.
The White House said it wanted to provide a "prompt response" to an April 24 query from lawmakers about whether Syria had used chemical weapons. The lawmakers' letter to Obama cited the assessments by Israel, France and Britain.
Greg Theilmann, a former weapons proliferation expert in the State Department's intelligence bureau, said, "I'm just a little surprised the US rolls out an assessment in this way."
In the last 24 hours, "did hard evidence suddenly appear?" said Theilmann, now at the private Arms Control Association.
Republican Senator John McCain of Arizona, one of the leading advocates of deeper US involvement in the Syrian conflict, said the US intelligence assessment demanded a US response.
"The president of the United States said that if Bashar Assad used chemical weapons, it would be a game changer, that it would cross a red line," he said.
"I think it's pretty obvious that red line has been crossed."
Senator Dianne Feinstein, head of the Senate Intelligence Committee, voiced concerns that the public US acknowledgement of its intelligence assessment could embolden Assad, prompting him to calculate "he has nothing more to lose."
"Syria has the ability to kill tens of thousands with its chemical weapons. The world must come together to prevent this by unified action," she said.
Syria's deputy foreign minister, Faisal Mekdad, told Reuters that US aid to the rebels may backfire and lead to attacks on American soil like those of Sept. 11, 2001.
"Once the fire of terrorism spreads in Syria it will go everywhere in the world," he said in an interview in Damascus.
