Negotiations over the miniature took around a year and a half after London-based agents Omnia (previously Robert Holden) instigated the sale.
“We are so proud and honoured to be involved in the partnership that has brought about such an exceptional acquisition that is so important for Welsh, British and indeed European heritage,” said National Trust director for Wales Justin Albert.
The National Trust was able to acquire the object eventually with the help of £1.5m from the National Heritage Memorial Fund and £300,000 from Art Fund.
National Trust raised the remaining £300,000 required through donations, which included one a ‘generous legacy bequest’ as well as a contribution from the fund of the late Simon Sainsbury.
The miniature was sold by a descendant of the sitter, Edward Herbert, 1st Baron Herbert of Cherbury (1583-1648).
The National Trust plan to return the piece to its previous home of Powis Castle following conservation work.
Royal miniaturist Isaac Oliver completed the portrait c.1602-17 in watercolour on vellum. It is 23 x 19cm and is in a contemporaneous 17th century tortoiseshell and ebony frame.
“We are very happy to have been able to support this extraordinary miniature, which constructs a fascinating vision of courtly life in Renaissance England,” said Art Fund’s director of programmes Sarah Philp. “It is one of the undisputed gems of the National Trust and belongs at Powis Castle, for everyone to see and enjoy.”