The quantitative analysis of bile acids (CA, DCA, GCA) was performed on a ZORBAX Eclipse XDB-C18 column (150 mm×2.1 mm, 3.5 μm, Agilent, CA, USA) and detected by a triple quadruple TSQ Quantum mass spectrometer with ESI source (Thermo Fisher, Palo Alto, CA, USA). Blood sample preparation and analytical methods were shown in Supporting Information Methods 1.3 .
For ketogenic amino acids (Phe, Lys, Trp) and BHB quantitative analysis, samples were separated on a BEH HILIC column (100 mm×2.1 mm, 1.7 μm, Waters, Ireland) and detected by a triple quadruple TSQ Quantum mass spectrometer with ESI source (Thermo Fisher, Palo Alto, CA, USA). Blood sample preparation and analytical methods were shown inSupporting Information Methods 1.4 .
All the quantification methods were developed and validated in accordance with FDA guidance for Bioanalytical Method Validation (2013) (http://www.fda.gov/downloads/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/UCM368107.pdf ) in terms of accuracy, precision, linearity, matrix effects, extraction recovery and stability.
For ketogenic amino acids (Phe, Lys, Trp) and BHB quantitative analysis, samples were separated on a BEH HILIC column (100 mm×2.1 mm, 1.7 μm, Waters, Ireland) and detected by a triple quadruple TSQ Quantum mass spectrometer with ESI source (Thermo Fisher, Palo Alto, CA, USA). Blood sample preparation and analytical methods were shown in
All the quantification methods were developed and validated in accordance with FDA guidance for Bioanalytical Method Validation (2013) (