 | Ben Stiller | | Alex (voice) |
| Chris Rock | | Marty / Additional Zebras (voice) |
| David Schwimmer | | Melman (voice) |
| Jada Pinkett Smith | | Gloria (voice) |
| Sacha Baron Cohen | | Julien (voice) |
| Cedric the Entertainer | | Maurice (voice) |
| Andy Richter | | Mort (voice) |
| Bernie Mac | | Zuba (voice) |
| Alec Baldwin | | Makunga (voice) |
| Sherri Shepherd | | Mom (voice) |
| Will.i.am | | Moto Moto (voice) (as Will.I.Am) |
| Elisa Gabrielli | | Nana (voice) |
| Tom McGrath | | Skipper / Lemur (voice) |
| Chris Miller | | Kowalski (voice) |
| Christopher Knights | | Private (voice) |
| Conrad Vernon | | Mason (voice) |
| Quinn Dempsey Stiller | | Baby Alex (voice) |
| Declan Swift | | Baby Alex (voice) |
| Fred Tatasciore | | Teetsi / Poacher #1 / Elephant (voice) |
| Eric Darnell | | Joe the Witch Doctor / Poacher #2 (voice) |
| Willow Smith | | Baby Gloria (voice) |
| Thomas Stanley | | Baby Marty (voice) |
| Zachary Gordon | | Baby Melman (voice) |
| Meredith Vieira | | Newscaster (voice) |
| Lesley Stahl | | Newscaster (voice) |
| Al Roker | | Newscaster (voice) |
| David Soren | | Lemur (voice) |
| Phil LaMarr | | Guide (voice) |
| Stephen Kearin | | Stephen the Giraffe / Tourist with Video Camera / Rhino (voice) |
| Dan O'Connor | | Tourist with University Shirt / Cape Buffalo (voice) |
| Edie Mirman | | Telephone Recording (voice) |
| Fergie | | Hippo Girlfriend (voice) |
| Harland Williams | | Additional Giraffe (voice) |
| Danny Jacobs | | Tourist with New York T-Shirt (voice) |
| Bridget Hoffman | | Female Tourist (voice) |
| Terrence Hardy | | Cub (voice) |
| Conner Rayburn | | Little Giraffe (voice) |
| Holly Dorff | | Fish (voice) |
| David P. Smith | | Bobby the Dik Dik (voice) |
| Lynnanne Zager | | Lioness (voice) |
