The 41-year-old former Ferrari driver, who has started more races than anyone in Formula One history and won 11 of them, reluctantly left the sport in 2011 when he was dropped by Williams.
Talk of a possible comeback was fuelled on Thursday when Kaltenborn appeared to leave the door open to him in comments to reporters at the Korean Grand Prix but on Saturday she closed it firmly.
"I am ruling out that he would drive for us. Maybe it is better if one is that clear about it," she told reporters after both her current drivers qualified in the top 10 for Sunday's race.
Kaltenborn explained that Barrichello had never been in consideration but she had tempered her earlier comments out of respect for him and courtesy.
"I have a lot of respect for Rubens, and I don't think it is right that I just so abruptly say 'No, we are not going to talk'," she said. "I fully understand that he wants to drive, and that he is trying his best.
"I talk a lot to him when he's here, we discuss young drivers and so on, but we have never discussed a seat. That's just reality," she added.
"It's a bit unfair also towards him because it doesn't look good for him, so just to be very clear on it: We have not discussed a seat for next year, and also no drive in Brazil (this season's final race)."
Barrichello has raced in IndyCar and Brazilian stock cars since he left Formula One and has also been attending some races as a television pundit.
On Saturday he told his more than two million followers on Twitter that he wanted to return if an opening emerged.
"Lots of things have being said this week about a possible F1 return. The truth is: "if there is an opportunity, I'm in," said the Brazilian.
(Reporting by Alan Baldwin, editing by Amlan Chakraborty)
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