The rhino, named Tam, was about 30 years old and lived at a wildlife reserve in Sabah state on Borneo island since his capture in 2008, Christina Liew, state minister for tourism, culture and environment, said in a statement.
The Sumatran rhino, the smallest species of rhinoceros, was declared extinct in the wild in Malaysia in 2015.
Iman, a female captured in 2014, is now the only surviving member of the subspecies left in the country.
Another female rhino, Puntung, died in captivity in 2017.
Wildlife experts estimate that only about 30 to 80 Sumatran rhinos remain in the world, mostly on the Indonesian island of Sumatra and on the Indonesian side of Borneo.
Their isolation, caused by habitat loss and poaching, means they rarely breed and may go extinct in a matter of decades, according to conservation group International Rhino Foundation.
Since 2011, Malaysia has tried to breed the species in captivity through in vitro fertilisation, but without success.
Liew said Tam’s genetic material has been preserved for future attempts to reproduce Sumatran rhinos.