Roberto Mancini's side overcame a one-goal deficit after the first leg with Dutch side Utrecht to record a 2-1 aggregate win.
Roberto Mancini's side overcame a one-goal deficit after the first leg with Dutch side Utrecht to record a 2-1 aggregate win, and top the domestic league, three points clear of Lokomotiv Moscow.
"It was a very necessary win against very difficult opponents," Mancini said.
"The European League play-off round is a difficult hurdle because it's just the beginning of the season when some of our players have yet to hit their top form."
"We're creating a new team after a number of skilled footballers left Zenit. Now we should focus completely on the domestic league as winning the Russian title is our main goal this season."
Rostov boss Leonid Kuchuk knows his men are in for a tough encounter with Zenit.
"Any team in the Premier League is dangerous especially at home," he said.
"But I demand my players to fight hard against any rivals regardless of when and where the match takes place."
Second-placed Lokomotiv, who beat reigning champions Spartak 4-3 last week, will face Ural Yekaterinburg, who remain unbeaten this season after drawing six of seven matches.
Lokomotiv boss Yury Semin is demanding there be no drop-off after some impressive wins to start the campaign.
"We looked good in the matches with Spartak and CSKA," he told Lokomotiv's website.
"But we experienced problems with concentration in the games with lowly Arsenal Tula and promoted Tosno. We're working on it but have failed to overcome this problem completely."
Title-holders Spartak Moscow hope to bounce back from consecutive defeats when they visit promoted SKA Khabarovsk on Sunday.
However SKA beat Spartak 1-0 in their only previous meeting in the last 32 round of the Russian Cup last year.
Coach Massimo Carrera insists there is no reason to sound the alarm just yet though.
"There's no panic in my team. The defeats are part of football," Carrera told the Spartak website.
"We need to rise from our knees and to continue our work. Our immediate task is to prepare for the next match the best possible way."
Fixtures (all times GMT)
Anzhi Makhachkala v Ufa (1700)
Arsenal Tula v Amkar Perm (1200), Rubin Kazan v Tosno (1430), Lokomotiv Moscow v Ural Yekaterinburg (1700)
SKA Khabarovsk v Spartak Moscow (0800), CSKA Moscow v Akhmat Grozny (1430), Zenit St Petersburg v Rostov (1700), Krasnodar v Dynamo Moscow (1700)