An Instagram post on the flotilla's official page showed activists chanting "Free Palestine" as their boats departed into the open sea.
The post read "We've set sail. Our mission to break the Gaza siege continues."
The Global Sumud Flotilla, sumud being the Arabic term for "resilience", originally scheduled to depart on Sunday, carried Greta Thunberg, Game of Thrones star Liam Cunningham, American actress Susan Sarandon, South African Mandla Mandela, German human rights activist Yasemin Acar, Spanish actor Eduard Fernandez, and organisers Thiago Avila, Saif Abukeshek, and Muhammad Nadir Al-Nuri.
The new Gaza-bound flotilla convoy, which is expected to reach the shores of Gaza in mid-September, is the third attempt by global activists to break the Israeli blockade on the strip amid growing global condemnation of Tel Aviv's 23-month-old genocidal war and starvation campaign agains the strip.
Israel had blocked and intercepted two similar flotillas in June and July, arresting and deporting all those who were on board.
The new effort comes two weeks after the United Nations declared a famine in Gaza, warning that 500,000 people face "catastrophic" conditions.
More than 340 Palestinians, including tens of children, have lost their lives to starvation and accute malnutrition amid a six-month Israeli blockade on all aid to 2.4 million Palestinians under siege and bombardment in the strip.
To date, the Israeli war on Gaza has killed at least 63,459 Palestinians and wounded more than 160,000 others, mostly women and children.
In a reel published on the flotilla's Instagram page, Mandla Mandela, grandson of South African Leader Nelson Mandela, said, "Ours is simply to end the blockade and break the siege on Gaza. We are brining humanitarian aid that is much needed to the Palestinians."
"Who is the one that is one that is perpetuating the violence? Who is the one that is a threat, not only to Palestinians but to you and I, to the entire global community? There is only one threat, apartheid Israel and Netanyahu and his regime,' added Mandela.
In another reel, environmental campaigner and activist Greta Thunberg said, "The story here is about Palestine. The story here is how people are being deliberately deprived of the very basic needs to survive."
Thunberg, who was deported by Israel in June after trying to reach Gaza waters earlier this year, stressed that "Israel are very clear about their genocidal intent. They want to erase the Palestinian Nation. They want to take over the Gaza Strip."
"Personally, I am terrified of how people can go on with their everyday life, accepting this genocide, watching a livestreamed genocide on their phones and then pretend like nothing is happening."
Unlike previous attempts to break the Israeli blockade and siege on Gaza, forcing Palestinians deeper into famine, the Global Sumud Flotilla is the "biggest mission ever," including"dozens of boats and massive demonstrations in more than 44 countries."
The initial launch, including some 20 ships, will be joined by dozens more as the convoy moves towards Gazaan waters. The grand total is estimated at some 70 boats, containing civilians and public figures to break the Israeli blockade on Gaza.
The Global Sumud Flotilla will be the fourth attempt to break the maritime blockade so far this year. The Conscience first tried in May, but was attacked by drones after setting sail from Malta. After the Madleen, the Israeli military stopped another aid ship, the Handala, in late July, detained 21 international activists and reporters and seized its cargo, including baby formula, food and medicine, according to the Freedom Flotilla Coalition.
Around 20 vessels originally left the Spanish city on Sunday, aiming to "open a humanitarian corridor and end the ongoing genocide of the Palestinian people", said the Global Sumud Flotilla.
But "due to unsafe weather conditions", the flotilla returned to port "to allow the storm to pass," the organisation said in an earlier statement, without specifying when exactly the boats returned to Barcelona.
"We made this decision to prioritize the safety and well-being of all participants and to safeguard the success of our mission," it added, citing gusts that exceeded 55 kilometres (34 miles) per hour.
Organisers said they were especially concerned about the risk the weather posed to the smaller boats.
Spain's national weather agency AEMET had issued warnings of rainfall and strong storms for the northeastern region of Catalonia, which includes Barcelona.
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