<jats:p>Incorporation of <jats:sup>13</jats:sup>C‐labelled acetate into the hopanoids of the purple non‐sulfur bacteria <jats:italic>Rhodopseudomonas palustris</jats:italic> and <jats:italic>Rhodopseudomonas acidophila</jats:italic> and the facultative methylotroph <jats:italic>Methylobacterium organophilum</jats:italic> showed that the bacteriohopane skeleton is built from an unique carbon/carbon linkage between the triterpenic hopane moiety and the C‐5 carbon of a <jats:sc>d</jats:sc>‐ribose derivative arising from the non‐oxidative pentose phosphate pathway.<jats:p>Furthermore a probable compartmentation of the acetate metabolism could be observed in these bacteria. Whereas exogenous acetate was directly incorporated into the glucose derivatives and poly‐(3‐hydroxybutyrate), the isoprenic units were apparently solely synthesized from two acetate units arising from the glyoxylate cycle and a third one issued either from the glyoxylate cycle or from the Entner‐Doudoroff pathway of glucose catabolism. Although an unknown biosynthetic pathway different from that usually proposed for isoprenoid biosynthesis can not be excluded, the former hypothesis explained all labelling patterns observed on the triterpenic skeleton.