Macron did not refer to the US president by name but his address to the UN General Assembly outlined positions that were polar opposites to Trump's world view.
"Some have chosen the law of the most powerful, but it cannot protect any people," said Macron, who reaffirmed his strong backing for multilateralism embodied by the United Nations and its global peace efforts.
Earlier at the UN podium, Trump vowed to "never surrender America's sovereignty to an unelected, unaccountable global bureaucracy" and to reject "the ideology of globalism."
Since Trump took office in 2017, the United States has ditched the Iran nuclear deal and the Paris climate deal -- two international accords that France has championed.
The US administration has cut funding to the United Nations, aid agencies and to UN peacekeeping whose missions in Mali and the Central African Republic are working with French troops to bolster stability.
Macron pointed to growing economic inequality as one of the drivers of the crisis in the world order, with some 783 million people living under the threshold of poverty and 250 million children out of school.
Touching on the Israeli-Palestinian conflict, the French president said there was no credible alternative to the two-state solution that Trump has imperiled with his recognition of Jerusalem as Israel's capital.
Macron also touched on climate change and migration as global issues that "no one can tackle" on their own.
There was even a warning that trade agreements could be contingent on membership in the Paris climate pact, in a clear bid to pressure the United States to rethink its intention to quit the accord.
Defending the Iran nuclear deal, Macron urged dialogue with Tehran, again clashing with the US president who a few hours earlier called on world governments to isolate Iran.
"What will bring a real solution to the situation in Iran and what has already stabilized it? The law of the strongest? Pressure from only one side? No!" Macron said in his address.
"We know that Iran was on a nuclear military path but what stopped it? The 2015 Vienna accord."
Trump in May withdrew from the seven-nation agreement negotiated under his predecessor Barack Obama, calling it a "disaster" and instead ramping up pressure on Iran including through renewed sanctions.
Supported by Israel and Saudi Arabia, Trump has sought to roll back Iranian influence around the Middle East, including in war-ravaged Syria.
"We should not aggravate regional tensions but rather through dialogue and multilateralism pursue a broader agenda that allows us to address all the concerns caused by Iranian policies -- nuclear, ballistic, regional," said Macron.
Speaking at a press conference, Macron said Iran should be able to sell oil to bring down prices, challenging Trump's plan to tighten an economic vise around Iran.
"It would be good for the price of oil for Iran to be able to sell it. It's good for peace and it's good for the shape of the international price of oil," said Macron.
France and other European powers are setting up a way to allow businesses to keep doing business in Iran in hopes of avoiding sanctions by the United States.
The French president was due to hold talks later in the day with his Iranian counterpart Hassan Rouhani, who has refused to meet Trump.
Despite their vastly differing world views, Macron and Trump -- both political outsiders who defied the odds in winning power -- have forged an unlikely relationship.
After meeting with Macron on Monday, Trump said the two men have had "some very good experiences" but acknowledged, "on occasion not so good, but 99 percent very good."
And after Trump's latest espousal of his "America First" foreign policy, Macron again painted himself as an upholder of a more internationalist approach to diplomacy.
"Do not accept the erosion of multilateralism," Macron told the gathering on Tuesday. "Don't accept our history unraveling. I'm not getting used to it and I'm not turning my head," he said.