"In the current context of tight public budgets, it is particularly important that large multinationals pay their fair share of taxes," EU Competition Commissioner Joaquin Almunia said.
Apple and Starbucks -- as well as a number of other multi-national companies including Amazon and Google -- have come under intense pressure from politicians and campaigners over their tax affairs.
The tax policies ruling the financial arm of Italian automaker Fiat, based in Luxembourg, is also included in the probe.
Almunia said the investigation would focus on transfer pricing payments, an accounting technique where units of a multinational pay 'royalties' to another unit of their business.
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The European Union strictly has no jurisdiction over national tax policies, a cherished prerogative of member states, and must limit its investigation to rules governing free competition.
"Under the EU's state aid rules, national authorities cannot take measures allowing certain companies to pay less tax than they should if the tax rules of the Member State were applied in a fair and non-discriminatory way," Almunia said.
California iPad maker Apple has shifted billions in international earnings through Ireland using such loopholes, but the government said it had not breached EU rules.
"Ireland is confident that there is no state aid rule breach in this case and we will defend all aspects vigorously," a government spokesman said.