Maharashtra, one of India's most populous states, has introduced a ban on beef so strict that even possession could land a person in jail for five years, officials have said.
The country's Hindu majority considers cows sacred, and several states have already banned their slaughter.
But the latest measures in the western state of Maharashtra - home to India's commercial centre Mumbai - go even further, making the sale or possession of beef an offence punishable by a five-year jail term or a $160 fine.
The Indian Express newspaper said the measures became law after President Pranab Mukherjee gave his assent to a legal amendment - which was passed by the state parliament two decades ago, but was never sent to a president for approval.
The measures include a ban on the slaughter of bulls and bullocks, hitherto legal with a vet's certificate, although it will still be legal to slaughter buffalo.
'Dream come true'
Maharashtra's Chief Minister Devendra Fadnavis tweeted his thanks to the president, saying "our dream of ban on cow slaughter becomes a reality now".
The ban did not go unnoticed and led to a debate on the social media with #BeefBan top-trending on India's twitter.
Right-wing Hindu groups in India have long demanded a complete ban on the slaughter of all cattle, citing religious scriptures.
The main players in the beef industry are Muslims, the country's largest religious minority, who make up some 13 percent of India's 1.25 billion population.
Maharashtra state is ruled by Prime Minister Narendra Modi's Hindu nationalist Bharatiya Janata Party in alliance with the far-right Shiv Sena party.