The traditional Muslim invocation to worship was performed in front of 1,000 people at the launch of a multi-cultural Faith Exhibition at Gloucester Cathedral.
Imam Hassan of the local Masjid-e-Noor mosque was invited by church leaders to carry it out in the cathedral's 11th century Chapter House.
Many worshippers welcomed the "inclusive" act but some traditionalists questioned why a "different God" should be worshipped in such an iconic Christian house.
The Muslim invocation to worship was performed in front of 1,000 people at Gloucester Cathedral
A clip of the call to prayer was initially shared on the cathedral's Facebook page but then taken down as "inappropriate" as tensions rose.
We are never to worship other Gods in a house built for our saviour
One post on the removed thread by Isabel Farmer said: "It's wonderful to be multicultural, but faith is set apart.
"We are never to worship other Gods in a house built for our saviour.
"My ancestors built this cathedral and to allow a practising Muslim pray to another God is insanely naive. What did you think it would do? Encourage them to convert?
A row has broken out after an Islamic call to prayer was held inside the historic cathedral
"This is why England is on the downslide. Culture and Race have nothing to do with the first commandment as God made all races and he cares about souls only.
"Stand firm Christians. Bring people to the faith by telling them the truth.
"God is the same yesterday, today, forever, so stop with the multicultural appeal and open a history book and a Bible for yourself."
The row spread as far as the USA and Twitter user 'Sir Anglo' tweeted: "Very disappointed that we are desecrating our gems. Gloucester Cathedral is magnificent."
Another Twitter user with the name '& drew' added: "Blasphemy at Gloucester Cathedral as Muslims abuse another 'interfaith' attempt'
Church leaders yesterday defended the call to prayer and said it was designed to promote religious tolerance and understanding.
The Very Revered Stephen Lake, Dean of Gloucester, said: "The community event on Saturday was to launch an art exhibition, 'Faith'.
"The exhibition portrays a number of individuals' stories and gives a glimpse of who they are as people of faith.
Artist Russell Haines with his exhibition in the cloisters at Gloucester Cathedral
The 'Faith' exhibition by artist Russell Haine runs until February 26
"The exhibition promotes religious tolerance and understanding.
"Over 1,000 people from different Gloucester communities attended to share and learn about each other's spiritual practices and traditions."
He added: "It is important to stress that neither the art exhibition nor the gathering on Saturday afternoon took place in a sacred space but were held in the Cathedral Cloisters and Chapter House.
"Furthermore, the launch was not in the context of worship.
"Whilst the Cathedral's primary purpose is as a place of Christian witness and worship, it is also a place for all people – everyone is welcome, whatever their faith or background.
"We are proud to be holding the exhibition and would encourage everyone to visit to learn more about people of different faiths."
According to the 2011 Census in Gloucester 63.5 per cent of the population were Christian and 1.0 per cent were Muslim.
The 'Faith' exhibition by artist Russell Haine runs until February 26 and features 37 portraits of individuals from different belief systems.
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Organiser Reverend Ruth Fitter said: "We live in a world that is becoming more and more polarised by people who claim to have the truth.
"No one has any proof of God – that's what faith is about.
"I happen to believe in Jesus Christ as the Son of God who came to dwell with me and save me from my sins.
"That doesn't mean I expect others to change their faith or believe wholeheartedly.
"It does mean, however, that I hope they will offer me the same respect as I seek to offer them.
"My parish is made up of such an eclectic diverse population and is, therefore, a beautiful image of God's creation in my view."
The incident is the second Muslim prayer row involving a British cathedral in as many weeks.
Last week a verse from the Koran denying the divinity of Christ was read during a multi-faith event at Glasgow Cathedral.
The Queen's chaplain spoke out against the event and said cathedral should apologise to Christians "suffering dreadful persecution at the hands of Muslims".