"Airstrikes are not acceptable," Karzai told CNN on Friday.
"We believe strongly that air strikes are not an effective way of fighting terrorism, that air strikes rather cause civilian casualties," he said.
The New York Times and CNN on Friday quoted unnamed Pentagon sources saying a preliminary inquiry investigation indicated that US raids resulted in some of the civilian casualties in the western Farah province.
Afghan police have maintained that more than 100 people - about 70 of them civilians - were killed in raids and ground fighting in two villages on Monday.
Colonel Greg Julian, a spokesman for US forces in Afghanistan, refused to comment on the reported outcome of the investigation, but said the death toll given by Afghan police was "grossly exaggerated".
"There were civilian casualties no doubt," he said on Friday after US military and Afghan teams returned from a joint investigation in Farah.
"But the conclusion from the investigation has not been reached, and it's inappropriate to indicate one way or the other how they were caused."
Karzai dismissed the possibility that Taliban had caused the civilian casualties and said the toll could be even higher than 100.
"I got definitive word from the government this morning that there were more than 100 casualties - nearly 125 to 130 civilians lost. Deaths - children, women and men - and it was done by the bombings," Karzai said.
'Civilians as cover'
Julian said earlier that the Taliban had used civilian positions as cover for attacks on US and coalition troops.
"[Civilian casualties] are always a possibility when conducting counterinsurgency activity," he said.
"We are here to protect the civilian population and we take this very seriously. The last thing we want to have happen is innocent civilians to be injured or killed."
During a visit to Kabul on Thursday, Robert Gates, the US secretary of defence, said he heard reports that grenades were thrown into homes by the Taliban during clashes, which created casualties that could be blamed on US forces.
Hundreds of Afghans gathered to denounce the killings and demanded that US troops leave Afghanistan.
The US government has come under increasing criticism during the past year for civilian deaths during operations against Taliban and al-Qaeda fighters in Afghanistan.
Barack Obama, the US president, and other Washington officials have repeatedly expressed regret for civilian casualties and vowed to take measures that would avoid future attacks.