The government plans to remove the dedicated north-south cycleway on College Street in favour of a bus lane. It will be replaced by two new cycleways down Castlereagh Street and Liverpool Street.

Bicycle NSW chief executive Ray Rice criticised the government for failing to take into account cyclists' safety.

"What they haven't really considered is the 2000 riders per day who will continue to ride this area as it's the best route from the eastern suburbs to the Macquarie Street area."

"The government has proposed that riders should be able to use the Kent Street cycleway. Well, that's a bit like closing the Western Distributor to cars and telling the cars to use the Cahill Expressway. It's just not practical."

Mr Rice said cyclists held the protest "to show the government there are many, many riders who are concerned about safety issues".

The plan was previously criticised by Liberal MP Shayne Mallard who said cycling in the city would become more dangerous as a result.

"They are life-saving pieces of infrastructure for cyclists," Mr Mallard said.

"We don't want cyclists riding on footpaths with pedestrians, pedestrians get very concerned about that."

CBD transport coordinator Marg Prendergast said the College Street cycleway would remain open until the alternative cycleways were built along Castlereagh, Liverpool and Kent streets.

"College Street cycleway will be removed once the new connected cycleway network, linking the Sydney Harbour Bridge and Central, is operational," Ms Prendergast said.

Barbara Davis, from northern Sydney, said she was protesting to stop cycling infrastructure from being removed, arguing it should be expanded instead.

"I'm here in solidarity because I'm really anxious and angry about the fact that we don't have any cycling infrastructure around the suburbs where I am and my kids are," Ms Davis said.

"For me it's really outrageous to remove a perfectly good cycleway when we should be building more."

In April, Fairfax Media revealed advice commissioned by Transport for NSW urging the government not to allow traffic to use the cycleways.

The protest follows another in February which attracted 300 cyclists.