The 24-desalkyl and 24-alkylsterols synthesized by leaves of flowering sorghum plants were identified by a combination of glc, hplc, ms, and nmr. Leaves from plants in the same developmental stage were treated with [2-14C]-acetate, and the radioactivity was associated with individual components of the 4,4-desmethyl sterol fraction determined by a combination of hplc-radioactivity counting and glc/ms. The specific activities of sorghum leaf sterols are in the following decreasing order: cholesterol (1036 dpm/μg)>24α-ethylcholesterol (416 dpm/μg)>24ξ-methylcholesterol (389 dpm/μg)ξ24α-ethylcholesta-5,22-dienol (39 dpm/μg). While all four of these sterols were detected by glc as components of steryl glycosides and steryl esters, only the 24-alkysterols were present as [14C]-labeled steryl glycosides. The differential regulation of sterol biosynthesis and derivatization is discussed in terms of differences in their functions.