title: A sensitive and practical RP-HPLC-FLD for determination of the low neuroactive amino acid levels in body fluids and its application in depression.
abstract: Ion-exchange high performance liquid chromatography (HPLC) generally fails as a method to determine low levels of free amino acids (AAs) in body fluids. Here we present a modified reversed-phase HPLC (RP-HPLC) protocol for the determination of AAs in body fluids and its application in mood disorder patients. We improved a previous research protocol by modifying i) sample preparation, including deproteination, ii) derivitization, including derivating agent and condition, and iii) sample separation, which is mainly determined by the pH value, the components and the additives of the mobile phases. The combination of these modifications, together with fluorescence detection (FLD), allows sensitive and practical determination of free AA levels in body fluids of depressive patients. This protocol was validated by determining the postmortem cerebrospinal fluid (CSF) glutamic acid (Glu) and γ-aminobutyric acid (GABA) levels of 8 major depressive disorder (MDD) patients, 9 bipolar disorder (BD) patients, and 19 well-matched controls, while also testing the plasma and CSF AA levels of living MDD patients. CSF Glu and GABA levels were both significantly decreased in MDD but not in BD patients. The data indicate that this RP-HPLC-FLD protocol is applicable for detection of low levels of neuroactive AAs in body fluids, as well as for routine clinical applications.
authors.affiliation: Department of Neurobiology, Key Laboratory of Medical Neurobiology of Ministry of Health of China, Zhejiang Province Key Laboratory of Neurobiology, Zhejiang Province Key Laboratory of Mental Disorder's Management, Zhejiang University School of Medicine, Hangzhou 310058, China.
authors.forename: Juan-Li
authors.initials: JL
authors.lastname: Wu
authors.affiliation: Department of Neurobiology, Key Laboratory of Medical Neurobiology of Ministry of Health of China, Zhejiang Province Key Laboratory of Neurobiology, Zhejiang Province Key Laboratory of Mental Disorder's Management, Zhejiang University School of Medicine, Hangzhou 310058, China.
authors.forename: Si-Yang
authors.initials: SY
authors.lastname: Yu
authors.affiliation: Research Center of Siyuan Natural Pharmacy and Biotoxicology, College of Life Sciences Zhejiang University, Hangzhou 310058, China.
authors.forename: Shi-Hua
authors.initials: SH
authors.lastname: Wu
authors.affiliation: Department of Neurobiology, Key Laboratory of Medical Neurobiology of Ministry of Health of China, Zhejiang Province Key Laboratory of Neurobiology, Zhejiang Province Key Laboratory of Mental Disorder's Management, Zhejiang University School of Medicine, Hangzhou 310058, China. Electronic address: baoaimin@zju.edu.cn.
authors.forename: Ai-Min
authors.initials: AM
authors.lastname: Bao
mesh_terms: D000368:Aged; D000369:Aged, 80 and over; D000596:Amino Acids / Q000097:blood* / Q000134:cerebrospinal fluid*; D001714:Bipolar Disorder / Q000134:cerebrospinal fluid; D001826:Body Fluids / Q000737:chemistry*; D016022:Case-Control Studies; D002851:Chromatography, High Pressure Liquid; D003863:Depression / Q000097:blood* / Q000134:cerebrospinal fluid*; D003866:Depressive Disorder / Q000097:blood / Q000134:cerebrospinal fluid; D005260:Female; D018698:Glutamic Acid / Q000134:cerebrospinal fluid; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008297:Male; D008875:Middle Aged; D019964:Mood Disorders / Q000134:cerebrospinal fluid; D005680:gamma-Aminobutyric Acid / Q000134:cerebrospinal fluid
chemical_list: D000596:Amino Acids; D018698:Glutamic Acid; D005680:gamma-Aminobutyric Acid
keywords: Amino acids; Cerebrospinal fluid; HPLC; Major depressive disorder; Plasma