Aided by the analysis of induced, single gene mutants in barley, independent elongation systems were inferred for the synthesis of β-diketones (98% hentriacontan-14,16-dione) and hydrocarbons (primarily hentriacontane). This proposal has been substantiated by comparing the effects of preincubations of inhibitors on the ability of whole spikes to incorporate [2-14C]acetate into the various epicuticular wax lipids. Dithiothreitol and mercaptoethanol inhibited the incorporation of label into hydrocarbons, but not into β-diketones. Cyanide blocked the synthesis of β-diketones, while stimulating hydrocarbon formation more than twofold. β-diketone synthesis was far more sensitive to arsenite than was synthesis of hydrocarbons. Degradation of the asymmetric β-diketone molecules by base hydrolysis and determination of the amount of label in the resulting fragments revealed a specific inhibition by arsenite of label incorporation into the C-31 end, i.e., the end from which previous studies have shown elongation to proceed. Tissue slices prepared from spikes minus awns were able to incorporate into β-diketones 1abel from [1-14C]palmitate and shorter evenchained fatty acids, but not from [1-14C]stearate. However, all fatty acids tested served equally well as hydrocarbon precursors. The elongation systems leading to the β-diketones and hydrocarbons are thought to diverge when the chain has 16 carbons. Thus, when a C16 chain is elongated by the addition of a C2 unit to form a C18 β-keto acyl chain, the β-keto group is not reduced as in normal fatty acid synthesis, but both carbonyl groups are protected and retained during further elongation. After reaching a C32 chain length, decarboxylation and release of the protected carbonyl groups yield the β-diketone.