MEP Francesca Donato said she could not support the Italian government's "discriminatory" measure to make COVID-19 health passes mandatory for workers.
An Italian MEP has left the far-right League party over its decision to back so-called "green" COVID health passes.
Francesca Donato, who was elected to the European Parliament in 2019, announced her departure on Tuesday.
The Eurosceptic MEP has long been opposed to obligatory vaccinations and has regularly criticised the COVID-19 restrictions enforced by Italy's government.
Donato said she had resigned over the decision by party leader Matteo Salvini to support new measures that make coronavirus health passes mandatory for all workers.
In a statement on her website, she stated that the new restrictions were "liberticidal and discriminatory".
"After a very long and in-depth reflection, I have come to the painful decision to leave the party in which I was elected," Donato said.
"The values in which I firmly believe - those of equality, individual freedom and human dignity - were increasingly trampled on by the measures taken by the national government, of which the League is a member," she added.
"Despite the reassurances and internal battles of our leader, liberticidal and discriminatory decrees have been passed which - in my opinion - are incompatible with the fundamental principles of our legal system."
But Donato said she would remain in the European Parliament as a member of the Identity & Democracy group.
The MEP added that she did not want to create "further embarrassment or problems" for the League by remaining at odds with Salvini's decisions.
"I thank Matteo Salvini for the battles he continues to fight in his delicate and difficult role, as well as for the space granted to me, without ever censoring my personal opinions," Donato said.
"My esteem and affection for him remain unchanged," she added.
Her resignation from the League could add pressure on Salvini, who has already faced criticism from Italian Economic Development Minister Giancarlo Giorgetti over his decision to back "green passes".
When asked about Donato's resignation, Salvini told reporters that he does not have any concerns about party unity.
"Those who leave, I thank, greet and wish them well," he said on Tuesday.