The small group, from the People for the Ethical Treatment of Animals (PETA), was in Fishergate calling on the travel agent to stop selling tickets to SeaWorld in Florida as part of a wider campaign.
Police were made aware of the protest, though a spokesman said officers were not called out.
PETA director Elisa Allen said demonstrations were also planned at Thomas Cook's branches in Blackpool and Lancaster.
She said: “PETA is calling on Thomas Cook to join STA Travel and Responsible Travel in rejecting SeaWorld’s orca prisons, where intelligent marine mammals have no choice but to swim in endless circles around tiny concrete tanks.”
SeaWorld said in a statement its 'animal welfare standards are amongst the highest in the world'.
It added: "Despite the views of animal activists, who want to see an end to all zoological display, the role of accredited zoos and aquariums is more important now than ever before.
"Through educating guests about animals and the plights they face in the wild, facilitating vital scientific research to help wild populations and funding conservation projects in the field, SeaWorld Parks and other leading accredited facilities are working to protect species for generations to come. "
Thomas Cook, which said its Preston branch was open as usual today, said in a statement: “We are the first organisation to remove animal excursions from sale as the direct result of auditing against independent welfare standards.
“So far this year, 12 attractions are on notice to improve their animal welfare arrangements or face Thomas Cook removing them from sale.
“By the end of next year all of our animal related excursions will have been audited and we will not be selling those which do not meet the required standards.
“We take our policy to raise animal welfare standards very seriously so that our customers can be confident in all of the trips that they take when on holiday with us.”
PETA, which has campaigned throughout this week, said today’s ‘day of action’ would see coordinated demonstrations outside Thomas Cook branches across the UK.
On its website, it said protestors would be ‘holding signs, handing out leaflets, and engaging with the public’.