Mr Spicer, when asked to clarify, then continued to shock at the White House press briefing yesterday, by correcting himself to say they were only used in "Holocaust centres".
Six million Jews were killed by the Nazis, with many being slaughtered in gas chambers at concentration camps.
He was speaking in reference to the April 4 attack in Syria that killed at least 87 people.
During the controversial press briefing Mr Spicer said: “You had someone as despicable as Hitler who didn't even sink to using chemical weapons.”
Sean Spicer at a White House press briefing claimed Hitler did not sink to using chemical weapons
Mr Spicer later issued an apology saying his remarks were inappropriate
When asked to clarify, he added: "I think when it comes to sarin gas he (Hitler) was not using the gas on his own people the same way that Assad is doing.
"There was clearly not…he (Hitler) brought them into the Holocaust centre and I understand that. What I'm saying is Assad used them where he went into towns and dropped them down into the middle of towns…the use of it, I appreciate the clarification, that was not the intent."
His comments caused fury, particularly as they were made during the Jewish holiday of Passover.
Steven Goldstein, executive director of the Anne Frank Centre for Mutual Respect, said: "Sean Spicer now lacks the integrity to serve as White House press secretary, and President Trump must fire him at once."
But later on Tuesday, Mr Spicer issued an apology on CNN and said he should not have made that comparison.
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It was inappropriate and insensitive
The White House spokesman said: "I apologise, it was a mistake to do that. It was a mistake. I shouldn't have done it and I won't do it again.
"It was inappropriate and insensitive. There's really is no explaining it at this point.
“It's just to say that especially this week [the Jewish holiday of Passover], it was not something that was appropriate.”
Washington has blamed the chemical weapons attack on the Syrian president Bashar al-Assad’ government and President Trump launched a missile attack on one of the nations air bases, reportedly destroying 20 percent of Syria’s operational aircraft.
The Trump administration has blamed Syrian president Assad for the chemical gas attack
Shocking photographs depict the horrors of the Holocaust
Fri, January 27, 2017
Holocaust Memorial Day 2017 marks the liberation of Auschwitz and remembers the Jewish people killed in World War II, as well as the victims and survivors of other genocides.
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The infamous German inscription reads 'Work Makes Free' at the main gate of the Auschwitz I extermination camp.
Steven Goldstein, executive director of the Anne Frank Centre for Mutual Respect, has called for President Donald Trump to sack the White House spokesman over the remarks.
He said: ”Sean Spicer now lacks the integrity to serve as White House press secretary, and President Trump must fire him at once.”
His sentiment was also backed by US House Democratic leader Nancy Pelosi.
She said: ”Sean Spicer must be fired, and the President must immediately disavow his spokesman's statements.”
The White House did not immediately respond when asked to comment on Pelosi's statement
The Anne Frank Centre has called for Sean Spicer to be fired as White House press secretary
The US Holocaust Museum, while not mentioning Mr Spicer's comments directly, sent out a tweet shortly after the briefing which showed graphic footage of dead bodies discovered by US forces when liberating the Nazi’s Buchenwald concentration camp after the war.
The video was retweeted more than a thousand times, with many Twitter users referencing Mr Spicer's comments.
Shortly after the controversial White House briefing, Spicer had emailed a further statement, which said: "In no way was I trying to lessen the horrendous nature of the Holocaust.
“I was trying to draw a distinction of the tactic of using airplanes to drop chemical weapons on population centres. Any attack on innocent people is reprehensible and inexcusable.”
Horrific images depict sickening chemical gas attack in Syria
Tue, April 4, 2017
At least 58 people, including nine children, were killed in an air raid that released "toxic gas" on the rebel-held Syrian town of Khan Sheikhoun
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A child receiving treatment at a field hospital after an alleged chemical attack in Idlib, northen Syria
This is not the first time the White House has been criticised for statements over the Holocaust.
The Trump administration’s statement marking International Holocaust Remembrance Day in January was received with disapproval after it omitted mentioning of the millions of Jewish victims.
At the time, Mr Spicer defended the statement by saying it had been written in part by a Jewish staff member whose family members had survived the Holocaust.
Last month Mr Spicer also had to issued an apology to GCHQ after he accused the British intelligence agency of wiretapping Mr Trump at the alleged request of former President Barack Obama.