1. Methods are described for the extraction and assay of acetyl-CoA and of total acid-soluble and total acid-insoluble CoA derivatives in rat epididymal adipose tissue. 2. The concentration ranges of the CoA derivatives in fat pads incubated in vitro under various conditions were: total acid-soluble CoA, 0·20–0·59mm; total acid-insoluble CoA, 0·08–0·23mm; acetyl-CoA, 0·03–0·14mm. 3. An investigation was made of some postulated mechanisms of control of fatty acid and triglyceride synthesis in rat epididymal fat pads incubated in vitro. The concentrations of intermediates of possible regulatory significance were measured at various rates of fatty acid and triglyceride synthesis produced by the addition to the incubation medium (Krebs bicarbonate buffer containing glucose) of insulin, adrenaline, albumin, palmitate or acetate. 4. The whole-tissue concentrations of glucose 6-phosphate, l-glycerol 3-phosphate, citrate, acetyl-CoA, total acid-soluble CoA and total acid-insoluble CoA were assayed after 30 or 60min. incubation. The rates of fatty acid and triglyceride synthesis, calculated from the incorporation of [U−14C]glucose into fatty acids and glyceride glycerol respectively, and the rates of glucose uptake, lactate plus pyruvate output and glycerol output were measured over a 60min. incubation. 5. The rate of triglyceride synthesis could not be correlated with the concentrations of either l-glycerol 3-phosphate or long-chain fatty acyl-CoA (measured as total acid-insoluble CoA). Factor(s) other than the whole-tissue concentrations of these recognized precursors appear to be involved in the determination of the rate of triglyceride synthesis. 6. No relationship was found between the rate of fatty acid synthesis and the whole-tissue concentrations of the intermediates, citrate or acetyl-CoA, or with the two proposed effectors of acetyl-CoA carboxylase, citrate (as activator) or long-chain fatty acyl-CoA (as inhibitor). The control of fatty acid synthesis appears to reside in additional or alternative factors.

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