The United Kingdom has informed European Council President Donald Tusk's office of May's intention, the spokesman said.
Brexit Secretary David Davis said in a statement that the UK is "on the threshold of the most important negotiation" for a generation.
Monday's announcement means that May will meet her self-imposed deadline of the end of March to get the withdrawal process underway.
She was cleared to take the step when the European Union (Notification of Withdrawal) Act gained royal assent last week, after a Supreme Court ruling forced her to seek the approval of both Houses of Parliament.
May will address MPs in a statement to the House of Commons following her regular weekly session of prime minister's questions on March 29.
Tusk has previously said he expects to release an initial response to the Article 50 notification within 48 hours, and an extraordinary summit of the remaining 27 EU member states is due to be called within four to six weeks.
That summit will draw up a mandate for the European Commission's chief negotiator, Michel Barnier, probably allowing talks to begin in earnest in May.
The European Commission says it's ready to begin Brexit negotiations.
"Everything is ready on this side," Margaritis Schinas, the chief spokesman for the EU executive, told a regular news briefing.
He noted that negotiations would begin once other EU states had met to confirm the Commission's negotiating mandate.
It will be the first time that the provisions of Article 50 - which sets out the process for any EU member state "to withdraw from the Union in accordance with its own constitutional requirements" - have been activated.
Notification comes 279 days after the referendum of June 23 last year delivered a 52 per cent to 48 per cent majority in favour of withdrawal.
May, who was visiting Swansea on Monday, is expected to conduct visits in all four nations of the UK before notification takes place.
May's spokesman said the UK wanted to start withdrawal negotiations "promptly", but accepts that "it is right that the 27 have a chance to agree their position" before talks start.
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