The inability of previous workers to recover completely the radioactivity from ingested [4-14C] cholesterol has led to the hypothesis that the colonic flora of some individuals degrade the sterol nucleus to volatile hydrocarbons, particularly CH4. In the present investigation, the production of radioactive volatiles was measured following incubation of [4-14C] cholesterol with 8 human fecal homogenates or after instillation of the labeled sterol into the cecum of 3 rats housed in a closed rebreathing system. Three of the 8 homogenates and each of the 3 rats produced copious CH4. However, analysis by combustion demonstrated no radioactivity above background in the volatile headspace of the homogenates or the gas space of the closed system housing the rats, indicating that <0.001% of the number 4 carbon of [4-14C] cholesterol could have been converted to volatile hydrocarbons. This study, therefore, provides no support for the concept that volatile products account for the incomplete recovery of ingested sterols observed in certain subjects. However, this hypothesis can not be excluded entirely until similar results are obtained with subjects who can be shown to degrade cholesterol.