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William Mackinnon was born on 13 March 1823 in Campbeltown, Argyleshire. He was educated in Campbeltown and trained in the grocery trade there. Early in his life he went to Glasgow, where he was employed in a silk warehouse and afterwards in the office of a merchant engaged in the Eastern Trade.
MacKinnon began his business career in 1847, when he joined an old school fellow Robert MacKenzie who was engaged in the coasting trade in the Bay of Bengal. Together they founded the firm of Mackinnon, MacKenzie & Company. On 29 September 1856, the Calcutta and Burmah Steamship Navigation Company was founded, mainly through Mackinnon's exertions. In 1862 the rapidly expanding company was renamed the British India Steamship Navigation Company. What had begun as a single steamer plying between Calcutta and Rangoon, became one of the greatest shipping companies in the world. Under Mackinnon's guidance it developed and created a vast trade around the coast of India and Burma, the Persian Gulf and East Coast of Africa, besides establishing subsidiary lines of connection with Great Britain, the Dutch East Indies and Australia. In 1873, the company established a mail service between Aden and Zanzibar. Mackinnon gained the confidence of Sultan Seyyid Barghash and in 1878 opened negotiations with him for the lease of a territory extending 1,150 miles along the coastline from Tungi to Warsheik, and extending inland as far as the eastern province of the Congo Free State. The British Government however, declined to sanction the concession, which if ratified would have secured for England the whole of what became German East Africa. In 1886 the British Foreign Minister availed himself of Mackinnon's influence to secure the coast line from Wanga to Kipini, a charter was granted and the Imperial British East Africa Company was formally incorporated on 18 April 1888 with Mackinnon as Chairman. The territory was finally taken over by the British Government on 1 July 1895, and became British East Africa. Mackinnon was also instrumental in promoting and funding Sir Henry Morton Stanley's expedition for the relief of Emin Pasha in 1886.
In 1858 Mackinnon became a Director of the City of Glasgow Bank, resigning that position in 1870, eight years before the Bank's complete collapse. Mackinnon did not escape from the consequences of the failure of the Bank, one of the most serious crises in modern Scottish financial history. The liquidators sued him for close to a quarter of a million pounds on a claim connected with advice Mackinnon was said to have given on American railway securities. Following protracted litigation, Mackinnon was completely exonerated by the court from the charges against him when it was demonstrated that the course of action taken by the remaining directors was contrary to his express advice.
Mackinnon was one of the chief supporters of the Free Church of Scotland. However, towards the end of his life the passage of the Declaratory Act, of which he disapproved, led to a difference of opinion between him and the leaders of the Church and he materially assisted the seceding members in the Scottish Highlands. In 1891 he founded the East African Scottish Mission.
In 1882 he was nominated C.I.E. He was created a Baronet on 15 July 1889. He married Janet Colquhoun (d 1894) on 12 May 1856. They had no children. William Mackinnon died on 22 June 1893, in the Burlington Hotel, London. He was buried at Clachan, Argyleshire.