Rapid diagnosis of Zellweger syndrome and infantile Refsum's disease by fast atom bombardment—mass spectrometry of urine bile salts

A method is described for the rapid determination of urinary bile salt profiles by fast atom bombardment-mass spectrometry (FAB-MS). Urine was passed through a reverse-phase octadecylsilane bonded silica cartridge and the bile salts eluted with methanol. Negative ion FAB spectra could be obtained from the equivalent of 10 μl of urine loaded onto the target probe with glycerol as matrix. In samples from normal infants and children bile salt peaks were rarely detectable above the background whereas peaks produced by steroid sulphates and glucuronides and bile alcohol glucuronides could usually be identified. In samples from infants and children with cholestasis the major peaks were produced by the taurine and glycine conjugates of di-, tri- and tetrahydroxycholanoic acids (and their monosulphates). In samples from patients with Zellweger syndrome and infantile Refsum's disease, a unique ion at m/ z 572 indicated the presence of taurine-conjugated tetrahydroxycholestanoic acid(s). The amide linkage to taurine was cleaved by alkaline hydrolysis but not by cholylglycine hydrolase. Capillary gas chromatography-mass spectrometry (GC-MS) of the bile acids liberated by alkaline hydrolysis indicated the presence of at least two nuclear-tetrahydroxylated cholestanoic acids, probably the 6α- and 1β-hydroxylated derivatives of 3α, 7α, 12α-trihydroxy-5β-cholestan-26-oic acid.