Supporters knew they wouldn't achieve the two-thirds vote needed to approve a constitutional amendment, but they had predicted a majority of votes. Instead, they fell one short, 49-48.
That was one vote more than they got last time the Senate voted on the matter, in 2004.
"We were hoping to get over 50 percent, but that didn't happen today," said Sen. David Vitter, R-La., one of the amendment's supporters. "Eventually, Congress is going to have to catch up to the wisdom of the American people or the American people will change Congress for the better."
"We're not going to stop until marriage between a man and a woman is
protected," said Sen. Sam Brownback, R-Kan.
President Bush said he was disappointed by the vote, but it often takes time to amend the Constitution.
"Our nation's founders set a high bar for amending our Constitution and history has shown us that it can take several tries before an amendment builds the two-thirds support it needs in both houses of Congress," the president said in a statement.
While there is no chance for this constitutional amendment to move forward, that won't stop House Republicans from holding their own debate and vote on the measure next month, CBS News correspondent Bob Fuss reports.
Supporters lost three key "yes" votes. Two Republicans changed their votes from yes in 2004 to no this time: Sens. Judd Gregg of New Hampshire and Arlen Specter of Pennsylvania. The third was Sen. Chuck Hagel, R-Neb., who did not vote this time because he was traveling with Bush.
All told, seven Republicans voted to kill the amendment. The four others were Sens. Lincoln Chafee of Rhode Island, Susan Collins of Maine, John McCain of Arizona, Olympia Snowe of Maine and John Sununu of New Hampshire.
Gregg said that in 2004, he believed the Massachusetts Supreme Court decision legalizing same-sex marriage in that state would undermine the prerogatives of other states, like his, to prohibit such unions.
"Fortunately, such legal pandemonium has not ensued. The past two years have shown that federalism, not more federal laws, is a viable and preferable approach," Gregg said in a statement.
"Most Americans are not yet convinced that their elected representatives or the judiciary are likely to expand decisively the definition of marriage to include same-sex couples," said Sen. John McCain, R-Ariz., a possible presidential candidate in 2008. He told the Senate on Tuesday he does not support the amendment.