Peru's foreign affairs ministry said Friday that it had given the ambassador, Diego Molero, five days to leave Peru.
'The Peruvian Government reaffirms its strong willingness to continue contributing to the restoration of democracy in Venezuela,' the ministry added in a statement.
The announcement marks the first time that a country has moved to expel a Venezuelan ambassador since Venezuela's July 30 vote to form the assembly amid allegations of fraud.
Peru has taken the toughest stance In Latin America toward Maduro as he seeks to consolidate power and crack down on anti-government protests and unrest in which over 120 have died since early April.
Earlier this week, Peru summoned top diplomats from the region to discuss Venezuela in the Peruvian capital Lima, where 12 nations condemned the 'rupture of democratic order' in Venezuela and said they would not recognize any action taken by the constituent assembly.
Peru's Foreign Minister told Reuters on Wednesday that expelling the ambassador was under evaluation daily but that Peru first wanted to gauge the response from Venezuela and the world to the so-called Lima Declaration.
Venezuela sent a note of protest regarding the Lima Declaration that Peru dismissed because it contained 'unacceptable terms' that the foreign ministry did not specify.
Venezuela's embassy in Lima did not immediately respond to requests for comment.
Maduro has said the constituent assembly is the country's only chance at securing peace and prosperity amid the unrest.
Peru, which withdrew its ambassador to Venezuela in March after the Venezuelan Supreme Court tried to take over the functions of Congress, maintains consular support to protect Peruvians in Venezuela.
U.S. President Donald Trump on Friday threatened a military intervention in Venezuela.