A novel liquid-liquid extraction and stable isotope dilution NCI-GC-MS method for quantitation of agmatine in postmortem brain cortex.
The group of biologically important amines includes putrescine, spermidine and spermine, as well as agmatine, which is a guanidino-amine. There is considerable evidence supporting a role of these amines in the etiology and pathology of mental disorders. We have previously developed a quantitative GC-MS method for simultaneous measurement of three major polyamines to support our studies linking polyamines to mental disorders. However, a unique GC-MS method is required for agmatine. To efficiently extract agmatine from postmortem brain tissues, we developed an isopropanol based liquid-liquid extraction protocol using potassium carbonate as a salting-out agent which showed a much greater recovery than n-butanol used in earlier methods. The GC-MS analysis employed hexafluoroacetylacetone as derivatization reagent and was carried out using negative chemical ionization with total ion and selected ion monitoring. (15)N(4)-agmatine was synthesized from (15)N(4)-L-arginine and used as internal standard in a conventional stable isotope dilution assay. This method accurately measures the level of agmatine from very small quantities (10-20 mg) of postmortem brain tissue, with a quantitation limit down to 1 ng/g of wet tissue. The limit of detection is 0.01 ng/g of wet tissue.
Chen
Gary Gang
GG
McGill Group for Suicide Studies, Douglas Mental Health University Institute, McGill University, Montreal, Quebec, Canada.
Turecki
Gustavo
G
Mamer
Orval A
OA
D019840:2-Propanol / Q000737:chemistry; D000376:Agmatine / Q000032:analysis*; D002138:Calibration; D002540:Cerebral Cortex / Q000737:chemistry* / Q000473:pathology; D005591:Chemical Fractionation / Q000379:methods*; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006845:Hydrocarbons, Fluorinated / Q000737:chemistry; D007553:Isotope Labeling / Q000379:methods*; D016014:Linear Models; D009587:Nitrogen Isotopes / Q000737:chemistry / Q000378:metabolism; D010422:Pentanones / Q000737:chemistry; D012680:Sensitivity and Specificity
D006845:Hydrocarbons, Fluorinated; D009587:Nitrogen Isotopes; D010422:Pentanones; C518705:hexafluoroacetylacetonate; D000376:Agmatine; D019840:2-Propanol