Faeces and urine of Studies 1, 3 and 4 were analysed as described previously (except neutral sterols and bile acids) [11 (link),19 (link),20 (link)]. In Study 2, mineral concentrations in faeces were determined following pressure digestion using ICP-OES (Optima 3000, Perkin Elmer, Waltham, MA, USA).
Faecal neutral sterols (cholesterol and its metabolites coprostanol, coprostanone, cholestanol, cholestanone, and cholestenone) and bile acids (iso-lithocholic acid: iLCA, lithocholic acid: LCA, iso-deoxycholic acid: iDCA, deoxycholic acid: DCA, cholic acid: CA, chenodeoxycholic acid: CDCA, and 12keto deoxycholic acid: 12keto DCA) were analysed as described previously [22 (link)]. Briefly, neutral sterols were extracted with cyclohexane following a mild alkaline hydrolysis. Analysis was performed by GC-FID (GC17A-AF Vers. 3, Shimadzu, Kyoto, Japan). Bile acids were extracted with diethyl ether following a strong alkaline hydrolysis. Thereafter, extracts were methylated, silylated, and analysed by GC-MS (GC17-QP5000, Shimadzu, Kyoto, Japan).
Faecal neutral sterols (cholesterol and its metabolites coprostanol, coprostanone, cholestanol, cholestanone, and cholestenone) and bile acids (iso-lithocholic acid: iLCA, lithocholic acid: LCA, iso-deoxycholic acid: iDCA, deoxycholic acid: DCA, cholic acid: CA, chenodeoxycholic acid: CDCA, and 12keto deoxycholic acid: 12keto DCA) were analysed as described previously [22 (link)]. Briefly, neutral sterols were extracted with cyclohexane following a mild alkaline hydrolysis. Analysis was performed by GC-FID (GC17A-AF Vers. 3, Shimadzu, Kyoto, Japan). Bile acids were extracted with diethyl ether following a strong alkaline hydrolysis. Thereafter, extracts were methylated, silylated, and analysed by GC-MS (GC17-QP5000, Shimadzu, Kyoto, Japan).
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