The four MPs and a senator, elected in April, arrived at the two chambers early Monday in military police vehicles. They were barred from speaking to reporters.
The five, who have been in custody for more than a year, will return to their cells at the Soto del Real prison near Madrid after carrying out the formality, Presstv Reported.
The Supreme Court, which has been trying the men for their role in an October 2017 secession attempt in Catalonia, ruled last week that they could also attend the opening session of parliament and take the oath of office on Tuesday.
But the court ruled against definitively releasing the five, who include the former vice president of the wealthy northeastern region, Oriol Junqueras.
And parliament is to decide whether to suspend their functions as lawmakers until the top court reaches a verdict on the rebellion charges against them.
Junqueras, 50, was elected an MP as a member of the Republican Left of Catalonia (ERC). He faces 25 years in prison in Catalonia.
The others include civic leader Jordi Sanchez and Jordi Turull, Catalonia's government spokesman at the time of the failed attempt to break from Spain.
Both were candidates for Together for Catalonia, the party of Catalonia's former president Carles Puigdemont, who fled Spain and avoided arrest.
Josep Rull, also formerly part of Catalonia's regional government, was another elected to the lower house, while Raul Romeva, in charge of Catalonia's foreign affairs at the time of the secession bid, was elected to the upper house Senate.
They were all part of a push to hold an independence referendum in October 2017 in defiance of a court ban.
That sparked Spain's deepest political crisis in decades, with the referendum followed by a short-lived declaration of independence.
Spain's then conservative prime minister moved in, taking direct control of the region, sacking the Catalan executive and calling snap polls.
Spain's April elections saw Prime Minister Pedro Sanchez's socialists win but without the necessary majority to govern solo in a fragmented political landscape marked by the far right's entry into parliament.
As such, Sanchez could need the backing of Catalan separatist lawmakers like the five jailed politicians.
During the parliament's opening session, the five Catalan lawmakers will have to promise to honor the Spanish constitution which calls for the "unbreakable unity" of Spain.