The festival, organised in the town of Pobla de Farnals in the Valencia region on Saturday, attracts huge crowds and has seen a number of incidents involving spectators and participants being badly injured in recent years.
The victim, whose identity has not been revealed, was gored in his side by a bull called Cocinero during the festival on Saturday and sustained severe injuries.
He was rushed to the hospital but died soon after, officials said.
This was the second injury reported during the festival this week alone. Earlier another man was also gored in the leg by the same bull.
However, he survived the incident and was in a stable condition in hospital, officials said.
The festival, where bulls are released into the streets and runners dash ahead of them, remains a controversial topic in Spain with activists saying it amounts to animal cruelty aside from the clear dangers to people involved.
Festivals involving running of the bulls are nonetheless still widespread in Spain, with a survey by rights groups AnimaNaturalis and CAS International finding that more than 1,820 events take place across all Spanish municipalities every year.
The most famous is held in the northern city of Pamplona in July.
Despite the risks, the traditional runs remain popular with both locals and tourists and attract huge crowds. However, accidents are also frequent.
Last year two men, aged 50 and 46, died after being violently struck by bulls in Valencia while a French tourist in his 60s sustained serious injuries.
In 2015 two US citizens and a Briton were gored and eight others injured during a bull run during the San Fermin festival.