Takagoyama Nature Zoo in Chiba said DNA testing showed the monkeys had been crossbred with the rhesus macaque.
The non-indigenous rhesus macaque is banned under Japanese law.
A local official said they had to be killed to protect the native environment.
The zoo's operator held a memorial service for the snow monkeys' souls at a nearby Buddhist temple.
Japanese macaques, commonly known as snow monkeys, are native to Japan and are one of the country's major tourist attractions.
Japan prohibits the possession and transport of invasive species, including crossbreeds.
An official from the Office for Alien Species Management, part of the country's environment ministry, told local media that the culling was unavoidable because there were fears they might escape and reproduce in the wild.
Junkichi Mima, a spokesman for conservation group WWF Japan told AFP news agency that invasive species cause problems "because they get mixed in with indigenous animals and threaten the natural environment and ecosystem".
A zoo in northern Japan has culled 57 of its snow monkeys by lethal injection after discovering they carried the genes of an "invasive alien species".