Pfeiffer was born in Santa Ana, California, the second of four children of Richard Pfeiffer, an air-conditioning contractor, and Donna (nee Taverna), a housewife. She has one elder brother, Rick (born 1955), and two younger sisters, Dedee Pfeiffer (born 1964), a television and film actress, and Lori Pfeiffer (born 1965). Her parents were both originally from North Dakota. Her paternal grandfather was of German ancestry and her paternal grandmother was of English, Welsh, French, Irish, and Dutch descent, while her maternal grandfather was of Swiss-German descent and her maternal grandmother of Swedish ancestry.

Pfeiffer chose to begin the process of dissolving her film production company, Via Rosa Productions, in 1999, and move into semi-retirement in order to spend more quality time with her children and family, meaning that she would continue to star in films sporadically into the 2000s and beyond. Pfeiffer handed her producing partner Guinzburg one final film to produce under the Via Rosa Productions header. The film was called Original Sin (2001). It was originally intended to star Pfeiffer, who later changed her mind as she was looking to work less for a while. The film was produced by her company, but instead starred Angelina Jolie and Antonio Banderas.  In the Hitchcockian thriller What Lies Beneath (2000), Pfeiffer and Harrison Ford starred as a well-to-do couple who experience a strange haunting that uncovers secrets about their past. While critical response towards the film was mixed, it opened atop at the box office in July 2000, and went on to gross US$291 million worldwide. She then accepted the role of Rita Harrison, a highly strung lawyer helping a father with a developmental disability, in the drama I Am Sam (2001), opposite Sean Penn. Despite grossing $97.8 million worldwide, the movie received unfavorable reviews; Seattle Post-Intelligencer wrote: "Pfeiffer, apparently stymied by the bland cliches that prop up her screechy role, delivers her flattest, phoniest performance ever". Meanwhile, SF Gate observed: "In one scene, she breaks down in tears as she unburdens herself to him about her miserable life. It's hard not to cringe, watching this emotionally ready actress fling herself headlong into false material".  Pfeiffer took on the role of a murderous artist named Ingrid Magnussen in the drama White Oleander (2002), alongside Alison Lohman (in her film debut), Renee Zellweger and Robin Wright. The film was an arthouse success and Pfeiffer garnered a substantial amount of critical praise; Stephen Holden of The New York Times wrote that "Ms. Pfeiffer, giving the most complex screen performance of her career, makes her Olympian seductress at once irresistible and diabolical". Kenneth Turan of the Los Angeles Times described her as "incandescent," bringing "power and unshakable will to her role as mother-master manipulator" in a "riveting, impeccable performance". She earned Best Supporting Actress Awards from the San Diego Film Critics Society and the Kansas City Film Critics Circle, as well as a Screen Actors Guild Award nomination.  Pfeiffer lent her voice for the character of goddess of chaos Eris in Sinbad: Legend of the Seven Seas (2003), an animated film featuring Brad Pitt as the voice of Sinbad the Sailor. She had struggles with finding the character's villainies. Initially the character was "too sexual," then she lacked fun. After the third rewrite, Pfeiffer called producer Jeffrey Katzenberg and told him "You know, you really can fire me", but he assured her that this was just part of the process. Following the release of the film, she took a four-year hiatus from acting, during which she remained largely out of the public eye to devote time to her husband and children. At the time, she turned down the role of the White Witch in the fantasy film The Chronicles of Narnia: The Lion, The Witch and The Wardrobe (2005), which went to Tilda Swinton.

Did she ever come back after her hiatus?
In the Hitchcockian thriller What Lies Beneath (2000), Pfeiffer and Harrison Ford starred as a well-to-do couple