High-performance liquid chromatography–tandem mass spectrometry of enterostatins in biological samples

The three forms of enterostatin (Ala–Pro–Gly–Pro–Arg, Val–Pro–Gly–Pro–Arg, and Val–Pro–Asp–Pro–Arg), pentapeptides known to inhibit fat-intake, were resolved on a C18 reversed-phase column using a ternary mobile phase consisting of methanol, acetonitrile, and water. Coupled with MS/MS detection, the method has been applied to identify enterostatin sequences in human cerebrospinal fluid and rat brain tissue. Ala–Pro–Gly–Pro–Arg (APGPR) was found to be the predominant enterostatin sequence in both cases. The levels of APGPR were 98.3(plus-minus)16.3 ng/ml in human cerebrospinal fluid and 30.1(plus-minus)12.6 ng/g wet tissue in rat brain, respectively.