Over a period of several years a number of studies of co₂ fixation in roots of various sources have related mineral assimilation to the synthesis of organic acids, notably malic acid. Further, investigations have been made of the proportion of CO₂ from the soil fixed by roots compared with the total CO₂ fixation by the whole plant. The aim of the investigations reported here was to extend information of CO₂ fixation to roots grown in sterile tissue culture. The root tissue chosen was comfrey , Symehytum uplandicum, Nyman, (S. peregrinum of various authors), this being easily and reproducibly cultured in bulk. Further, it was thought that by varying the time of exposure to a source of C¹⁴O₂ and subsequently measuring the total and specific activities of the various radioactive compounds, data indicating the pathway of incorporation might be obtained. It was realized that such data would be of little significance to problems concerning the effects of sterile culture procedure on the basic metabolism of the tissue unless a comparison between the incorporation rates of cultured roots and whole plant roots was made. For this reason the fixation of O₂ was measured under similar conditions with roots taken from whole plants. Incidental to this aspect was the measurement of the total amount s of a limited number of substances in both cultures roots and whole plant roots.