Partial beta‐oxidation of gamma‐hydroxybutyrate (GHB) in perfused rat livers

GHB is both a brain metabolite and a drug of abuse (date rape drug). We conducted a metabolomic study of GHB metabolism liver, using GC‐MS, GC‐combustion‐IRMS, and LC‐MS‐MS. Rat livers were perfused with recirculating albuminated buffer containing 4 mM glucose and 2 mM of either unlabeled GHB, [U‐ 13 C 4 ]GHB, or [ 2 H 6 ]GHB. In the perfusate, we identified by GC‐MS 2,4‐dihydroxybutyrate, 3,4‐dihydroxybutyrate, and 3‐hydroxypropionate. The mass isotopomer distribution of these compounds in livers perfused with GHB, [U‐ 13 C 4 ]GHB, or [ 2 H 6 ]GHB confirms that they are derived from the partial β‐oxidation of GHB. 3‐Hydroxypropionate may derive from the omega‐oxidation of 3,4‐dihydroxybutyrate (or its CoA ester). GC‐combustion‐IRMS detected 5 unknown metabolites of [U‐ 13 C 4 ]GHB (to be identified). In livers perfused with [U‐ 13 C 4 ]GHB, acetyl‐CoA was unlabeled (even in the absence of glucose), confirming the incomplete βoxidation of GHB. Citric acid cycle intermediates were labeled from [U‐ 13 C 4 ]GHB consistent with entry into the cycle via succinate. LC‐MS‐MS of liver acyl‐CoAs detected an unknown compound (m/z 934 with CoA signature daughter ion at m/z 428). This compound, which becomes M4 with [U‐ 13 C 4 ]GHB, and M6 with [ 2 H 6 ]GHB, contains the 4 C and the 6 H of GHB. In livers perfused with unlabeled GHB and either [U‐ 13 C 6 ]glucose or [1,2‐ 13 C 3 ]acetate, the CoA ester was not labeled. Based on the above and on its rapid migration on the reverse phase LC column, the unknown CoA ester was tentatively identified as 4‐phospho‐GHB‐CoA. Supported by NIH grant ES013925.