The 100-minute debate was marked by several sharp exchanges and finger-pointing, but the two shook hands cordially when it ended, maintaining a controlled tone for most of the session.
As they traded accusations, Yanukovych tried to be more conciliatory.
"Whoever wins, I think, we should hold a forum of national accord," Yanukovych said, urging Yushchenko to pledge not to contest the official results of the Dec. 26 rerun vote.
"If you win, I will recognize (your victory), if I win — you will," Yanukovych said. "And then, you and I, are working to form a normal government of national concord."
Yanukovych several times proposed forming a unity government but Yushchenko ignored the proposals.
After the debate, Yushchenko's close adviser Oleh Rybachuk dismissed Yanukovych's unity government idea: "Yanukovych was trying to strike a deal ... he was begging."
The debate was the first since the Supreme Court annulled the Nov. 21 presidential runoff following allegations of widespread fraud.
The electoral campaign has been marked by tension following massive street protests and revelations that Yushchenko was poisoned by dioxin in September. During the debate, however, neither candidate mentioned the poisoning that left Yushchenko's face pockmarked and disfigured.
Yushchenko was more aggressive during the encounter, pointing his finger at his rival and clenching his fists. At times, he slashed his hand through the air and expressed incredulity toward his opponent.
Yanukovych appeared more defensive, standing up straight and occasionally shifting his weight.
"You're a religious person, right? Thou shalt not steal ... And then you stole three million votes," Yushchenko said, wearing a tie and handkerchief in his campaign color of orange.
Yanukovych, with a tie in his trademark blue, spoke in Russian instead of Ukrainian in his introductory remarks.
"Your accusations toward me and toward my voters don't give us the chance to look into the future optimistically," he said, wagging his finger at Yushchenko.
Yanukovych suggested a Yushchenko victory would endanger the country's unity.
"If you win the vote you will only be the president of part of Ukraine," he told his rival. "I am not struggling for power — I am struggling against bloodshed."
The bitter campaign has split the country between the eastern, Russian-speaking industrial heartland that backs Yanukovych, and other regions backing the Western-leaning reformist Yushchenko.
"We have to discuss how to unite Ukraine and not divide it," said Yanukovych, who enjoys strong support from Russia.
Rules for the debate allowed the two to question each other directly after their opening statements. Yushchenko used his first question to quiz his opponent about what he described as Yanukovych's mistakes in economic policy.
Yanukovych defended his record, recalling a recent one-time increase in pensions, and promised he would increase benefits to retirees again. He later tried to focus on campaign financing, hinting that his rival received funding from abroad.
Yushchenko, showing his hands, said: "These hands have never taken anything."
Yushchenko's questions focused on economic matters, pensions, the budget and salaries, while Yanukovych emphasized voting and changes in election law.
Both candidates exceeded the time limits allowed for questions and answers, prompting warnings from the moderator.
Yushchenko also criticized Yanukovych for calling his supporters "goats" and "orange rats" during the electoral campaign.
Yanukovych shot back: "If I said an offensive emotional word — I ask that I be forgiven."
The two candidates prepared for their debate in markedly different styles, Ukraine's daily Segodnya reported. Yushchenko read books about economics and history, while Yanukovych visited Kiev's Orthodox Monastery of Caves, where he prayed, the paper said.