I would do a top five list, but that would be impossible.
Here are my favorite villains and villainesses:
Resident Evil: The DC version of Alfred Ashford (though he was okay in Code: Veronica/Code: Veronica X, his psyche was fleshed out in greater detail and he was far more threatening in DC), the DC version of Alexia Ashford (cannot stand her in Code: Veronica/Code: Veronica X), Jack Krauser (after DC, I like him), Nicholai Ginovaef, and James Marcus.
Pinocchio: The Coachman
Disney's Peter Pan: Captain James Hook (MUCH better than the cold, serious Hook in other adaptations
) and Smee
The Little Mermaid: Ursula
Sleeping Beauty: Malefacent (her role is the film's only saving grace
)
Silent Hill: Pyramid Head, Maria, and Claudia Wolfe. Actually, I might as well list the town itself XD
Parasite Eve: Eve
Eternal Darkness: Sanity's Requiem: Pious Augustus
Devil May Cry: Vergil Sparda
Tales of Series: Mithos Yggdrasill, Dist the Rose/Reaper, and Yeager.
Heavy Rain: Scott Shelby
Yu Yu Hakusho: Elder Toguro and Sakyo. While Younger Toguro is one of my favorite characters from the series and is categorized as a villain, he is more of an anti-hero than a villain.
Yu-Gi-Oh! Duel Monsters: Maximilian Pegasus (when he was a villain, that is), Dartz, Yami Bakura/Thief King Bakura, Rafael (same as with Pegasus), and Valon (again, same as with Pegs).
Star Wars: Darth Vader and Count Dooku/Lord Tyrannous.
Power Rangers: Lord Zedd after he married Rita (he was a boring archetypical villain before he got laid XD), Rita Repulsa, Rito Revolto, Goldar, Ecliptor, and Villamax.
Batman: The Joker, Harley Quinn, Two Face, and Mr. Freeze.
X-Men: Magneto. (Depending on the storyline. Sometimes, he is a villain. Others, an anti-hero)
Buffy the Vampire Slayer: Spike in seasons 2 - 4 (when I think about how his character was destroyed in season five, I feel like vomiting >_>) and Drusilla.
Carrie: Margaret White
Psycho: Norman's "Mother" personality and Norma Bates.
Fullmetal Alchemist original anime series: Lust (she is a villainess for the majority of the series, anyway)
One Piece: Buggy the Clown (though from what I understand, he is now more of an anti-hero than a villain) and Captain Kuro.
Harry Potter: Lucius Malfoy pre-Deathly Hallows (hate the bad-*** decay he went through) and Barty Crouch, Jr.
The Interview with a Vampire film: Does Claudia count? If so, then she is one of my favorites.
The Queen of the Damned film: Akasha
The first Blade film: Deacon Frost
The Fright Night remake: Jerry Dandrige
The Nightmare Before Christmas: Oogie Boogie
Invader Zim: Zim
SpongeBob SquarePants: Plankton
True Blood: Marnie Stonebrook. I like Antonia, but she is an anti-heroine, not a villainess.
Pokemon: Jessie, James, and Meowth (though I view them as wannabe villains as opposed to real villains XD)
Long list, eh? There are probably more, but these are the ones that come to mind.