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Challenge Mode is an alternative mode unlocked after collecting The Key in the Citadel of Spiders. It features 30 stages with pre-set Pikmin totals in a series of challenge caves, to collect various treasures within a time limit.

Completion of a stage awards a white flower, and completion of a stage with no Pikmin deaths (as well as all Pikmin being plucked) will award the player a pink flower. Only five stages are initially unlocked, and after the completion of one level another will open.

Merely getting to the end of stage is not the only objective for completion. Beating the stage with the most Pokos, in the fastest time, with the most Pikmin, will increase a player's final score. The equation to calculate end score is:

Once all stages have been completed with a pink flower, a special cutscene will be unlocked to view at any time.