While Spanish is spoken by some 400 million people around the world and is one of the official UN languages, it is used sparingly on UN internet sites and in documents issued by its public information services, the letter charged.
In a letter to UN Secretary-General Kofi Annan, envoys from 20 nations said there was a "growing imbalance" in favour of English in UN affairs and called the situation "disturbing and alarming to those we represent".
While Spanish is spoken by some 400 million people around the world and is one of the official UN languages, it is used sparingly on UN internet sites and in documents issued by its public information services, the letter charged.
The other five official languages at the UN are Arabic, Chinese, English, French and Russian.
UN officials say it is costly to provide every document in all six languages.
Equal
The ambassadors said there was "a growing imbalance" between the six languages and "a tendency to favour the use of only one language" within the UN system.
They said there was a discernible trend toward English being used increasingly in the drafting of major publications, negotiations on resolutions and even in decision by governing bodies.
They appealed to Mr Annan to "ensure that the necessary measures are taken, without delay whatever, to mitigate the alarming increase in the imbalance".
"This is not only a matter of principle but also an effective way for the United Nations to present itself as a global institution that is truly representative of the entire world," the ambassadors' letter said.
The BBC's UN correspondent Greg Barrow says privately that the organisation could do better, but they say the cost of providing every document and every speech in all six official languages would be prohibitive.
They say that alongside official languages, the UN has adopted working languages for specific institutions. At the UN headquarters in New York, the working languages are English and French; at UN institutions in the Middle East they would be English and Arabic.
The 20 Spanish-speaking ambassadors want to reverse this trend but unless they can find the money to fund the extra translating staff, that seems an unlikely prospect, says our correspondent.