His introduction to business was as clerk in the employ of Samuel Hicks & Sons. One of the most important episodes of this period of his life was his experience during the cholera of 1832, when he was the only one of the employees who remained at his post during that fatal time. Upon one occasion he, in company with a carman in the employ of the firm, left the building at the same time. Within four hours his companion was dead and buried; Mr. Colgate himself was seized with the disease, but fortunately survived the attack.

In 1833 he went into business on his own account, in company with his Uncle Charles and George P. Pollen, under the firm-name of Colgate, Pollen & Colgate, manufacturers and dealers in dye-stuffs and paints, their place of business being at 177 Water Street. In 1845 he built the Atlantic White Lead Works in Brooklyn, having previously purchased the store property at 287 Pearl Street, New York, which, at that time, was surrounded by the private residences of many wealthy and prominent citizens. To the works in Brooklyn he added the manufacture of linseed oil, and the firm, which is now known by the name of Robert Colgate & Co., are among the most extensive dealers in paint and oil in the country, and bear a high reputation in the commercial world. They have stood unshaken through all the financial reverses which have visited the city, and never failed to meet all obligations with promptness. It has always been the policy of the firm to secure the services of trusty and faithful employees and to retain them as long as they are willing to remain. As an illustration, it may be mentioned that the bookkeeper, James B. Carr, has been in the employ of the firm for fifty years, and the cashier for thirty years.