The demonstrations began after Spanish authorities, which outlawed the vote scheduled for October 1, detained on Wednesday 14 regional officials and seized referendum materials, including some 10m ballot papers, in an unprecedented raid of regional government offices.
After a series of rulings from the Constitutional Court decreed the vote illegal, Prime Minister Mariano Rajoy is imposing financial curbs on the regional administration and has promised tougher sanctions still if officials refuse to back down.
Wednesday's episode is the latest development in the Spanish government's ongoing efforts to stop Catalonia's upcoming vote.
Puigdemont described the raids as "a coordinated police assault", which he said showed that Madrid "has de facto suspended self-government and applied a de facto state of emergency" in Catalonia.
Why is Spain cracking down?
Spain did not stop an earlier vote taking place in November 2014, but this time the Catalan leadership plans a declaration of independence within 48 hours of a Yes vote.
A number of protests against the policy of Spanish authorities with regard to the preparations for the independence referendum in Catalonia continue across Span's northeastern region on Thursday.
On Wednesday, Catalan President Carles Puigdemont urged residents in the region to turn out for the October 1 vote.
Several ministries in Barcelona were raided on Wednesday, including the economy, foreign affairs, telecoms, social affairs and presidency buildings.
Police were searching for computer equipment and any documentation linked to the planned vote.More news: Tesla 'running tests' on in-house AI chips
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The operation comes a day after police seized a trove of documents related to the independence referendum from the offices of private delivery company Unipost in the Catalan city of Terrasa.
Madrid has even taken direct control of Catalonia's finances, arguing that it wants to prevent public money from being spent on the vote.
How are Catalan leaders responding?
The independence movement, led by the regional president, Carles Puigdemont, argues that Catalonia has a moral, cultural, economic and political right to self-determination. A top treasury official is being held on sedition charges, the BBC reports.
Demonstrators clash with Spanish National Police officers as they leave the road outside the Catalan Pro-Independence Lefty party CUP headquarters on September 20, 2017, in Barcelona, Spain. "We will not allow it", he said.
While the court assessed the claim, it called for a suspension of the referendum, but the Catalan government said it would press on regardless.
"Just 35% said Catalonia should be independent in a July survey by the Catalan government's polling agency", says Bloomberg.
An influential pro-independence citizens' organisation, the Catalan National Assembly (ANC), urged Catalans to hit the streets in protest.
The nation's constitution states the country is bound in "indissoluble unity" and therefore regions can not unilaterally declare independence.
Catalonia has long sought independence from Spain, but the country's central government has repeatedly declined to grant the region independence, saying it would violate the country's constitution.