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The amino acids (AAs) and organic acids (OAs) are closely interrelated in metabolic pathways.In this study, simultaneous metabolic profiling analysis of AAs and OAs in urine as ethoxycarbonylated, and tertbutyldimethylsilylated derivatives was performed by gas chromatography (GC).After metabolic profiling analysis, the enantioseparation of urinary chiral AAs and OAs as target metabolites to determine their absolute configurations was performed by using two-phase extractive ethoxycarbonylation (EOC) in alkaline aqueous solution with subsequent conversion into volatile diastereomeric EOC-O-trifluoroacetylated (S)-(+)-3-methyl-2-butyl esters after acidification and extraction.The resulting diastereomeric derivatives were analyzed by achiral dual-columns with different polarities.The clinical application of the present methods to urine specimens from breast cancer patients before-and after-mastectomy will be presented as compared to normal control.