Validation of a Microwave-Assisted Derivatization Gas Chromatography-Mass Spectrometry Method for the Quantification of 2-Hydroxybutyrate in Human Serum as an Early Marker of Diabetes Mellitus
María Rodríguez-García,
Guillermo Fernández-Varo,
Susana Hidalgo,
Gabriela Rodríguez,
Irene Martínez,
Muling Zeng,
Eudald Casals,
Manuel Morales-Ruiz,
Gregori Casals
Affiliations
María Rodríguez-García
Service of Biochemistry and Molecular Genetics, Hospital Clinic Universitari, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), 08036 Barcelona, Spain
Guillermo Fernández-Varo
Service of Biochemistry and Molecular Genetics, Hospital Clinic Universitari, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), 08036 Barcelona, Spain
Susana Hidalgo
Service of Biochemistry and Molecular Genetics, Hospital Clinic Universitari, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), 08036 Barcelona, Spain
Gabriela Rodríguez
Service of Biochemistry and Molecular Genetics, Hospital Clinic Universitari, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), 08036 Barcelona, Spain
Irene Martínez
Service of Biochemistry and Molecular Genetics, Hospital Clinic Universitari, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), 08036 Barcelona, Spain
Muling Zeng
School of Biotechnology and Health Sciences, Wuyi University, Jiangmen 529020, China
Eudald Casals
School of Biotechnology and Health Sciences, Wuyi University, Jiangmen 529020, China
Manuel Morales-Ruiz
Service of Biochemistry and Molecular Genetics, Hospital Clinic Universitari, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), 08036 Barcelona, Spain
Gregori Casals
Service of Biochemistry and Molecular Genetics, Hospital Clinic Universitari, Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), 08036 Barcelona, Spain
Circulating levels of 2-hydroxybutyrate (2HB) are highly related to glycemic status in different metabolomic studies. According to recent evidence, 2HB is an early biomarker of the future development of dysglycemia and type 2 diabetes mellitus and may be causally related to the progression of normal subjects to impaired fasting glucose or insulin resistance. In the present study, we developed and validated a simple, specific and sensitive gas chromatography-mass spectrometry (GC-MS) method specifically intended to quantify serum levels of 2HB. Liquid–liquid extraction with ethyl acetate was followed by 2 min of microwave-assisted derivatization. The method presented acceptable accuracy, precision and recovery, and the limit of quantification was 5 µM. Levels of 2HB were found to be stable in serum after three freeze-thaw cycles, and at ambient temperature and at a temperature of 4 °C for up to 24 h. Extracts derivatized under microwave irradiation were stable for up to 96 h. No differences were found in 2HB concentrations measured in serum or plasma EDTA samples. In summary, the method is useful for a rapid, precise and accurate quantification of 2HB in serum samples assessed for the evaluation of dysglycemia and diabetes mellitus.
