The defendants are on trial over an independence referendum which was held in October 2017 despite a court ban, as well as a short-lived declaration of independence.
The events sparked Spain's worst political crisis since the death of dictator Francisco Franco.
Separatist officials demonstrated near the Madrid courthouse as the trial got underway. They dismiss the trial as a politically-motivated "farce".
Protests are also scheduled at 7pm local time in the Catalan capital, Barcelona.
But many Spaniards support the decision to try the separatists.
Chief separatist absent
Carles Puigdemont, Catalonia's former president who fled to Belgium days after the declaration of independence on 27 October 2017, is not among those on trial as Spain does not try suspects in absentia for major offences.
The key protagonist in court is therefore Puigdemont's former deputy, Oriol Junqueras, who chose to remain in Spain. He faces up to 25 years in jail.
He and eight other defendants are charged with rebellion.
Under Spanish law, rebellion is defined as "rising up in a violent and public manner", but supporters of independence deny the accusation of violence.
They instead denounce police violence during the referendum.
Some of the defendants have been in pre-trial detention for more than a year.
The trial is expected to last three months.