"Altogether, signs do not give me much hope," he was quoted as saying by Iran's semiofficial Fars News Agency.
U.N. Secretary-General Ban Ki-moon rescinded an invitation to Iran to attend the Geneva II conference in Montreaux, Switzerland, after Washington and members of the Syrian opposition expressed frustration over Iran's allegiance to Syrian President Bashar Assad.
Rouhani said Wednesday it was the parties to the conference that were causing instability in Syria.
Talks on the Syrian crisis got off to a testy start Wednesday with U.S. and Russian representatives sparring over Assad's future. The United States and Russia, two permanent members of the U.N. Security Council, have been at odds over the best way out of the crisis.
Russia is considered a Syrian ally.
In his opening remarks to the conference, U.N. Secretary-General Ban Ki-moon said he was optimistic about the prospects given the wide range of representatives in attendance.
"For the first time, the Syrian government and the Syria opposition, countries of the region, and the wider international community are convening to seek a political solution to the death, destruction and displacement that is the dire reality of life in Syria today," he said in a statement.
Rouhani, for his part, said the Iranian government would be "pleased" with the outcome provided it brings relief to the region and the Syrian people.