title: Direct detection of tetrahydrobiopterin (BH4) and dopamine in rat brain using liquid chromatography coupled electrospray tandem mass spectrometry.
abstract: A simple and rapid liquid chromatography tandem mass spectrometry (LC-MS/MS) method was developed for the quantification of tetrahydrobiopterin (BH4) and dopamine in rat brain using epsilon-acetamidocaproic acid (AACA) as an internal standard. Proteins in the samples were precipitated with acetonitrile and then the supernatants were separated by a Sepax Polar-Imidazole (2.1 × 100 mm, i.d., 3 μm) column using a mixture of 10mM ammonium formate in acetonitrile/water (75:25, v/v) as the mobile phase at a flow rate of 300 μl/min. Quantification was performed on a triple quadrupole mass spectrometer employing electrospray ionization with the operating conditions as multiple reaction monitoring (MRM) and positive ion mode from m/z 242.1 → 166.0 for BH4, m/z 154.1 → 90.0 for dopamine and m/z 174.1 → 114.0 for AACA (IS). The total chromatographic run time was for 5.5 min. The method was validated for the analysis of samples: the limit of detection was 10 ng/g. The calibration curve was linear between 10-2000 ng/g for BH4 (r(2)=0.995) and 10-5000 ng/g for dopamine (r(2)=0.997) in the rat brain. Thus, good correlated LC-ESI/MS/MS results were obtained and found to be a powerful tool for the quantitative analysis of BH4 and dopamine in the rat brain.
authors.affiliation: Department of Pharmacology, College of Medicine, Dankook University, San 29, Anseo-dong, Dongnam-gu, Cheonan, Chungnam 330-714, Republic of Korea.
authors.forename: Hak Rim
authors.initials: HR
authors.lastname: Kim
authors.forename: Tae-Hyun
authors.initials: TH
authors.lastname: Kim
authors.forename: Sung-Hyun
authors.initials: SH
authors.lastname: Hong
authors.forename: Hyung-Gun
authors.initials: HG
authors.lastname: Kim
mesh_terms: D000818:Animals; D001708:Biopterins / Q000031:analogs & derivatives* / Q000032:analysis; D001923:Brain Chemistry; D002853:Chromatography, Liquid / Q000379:methods*; D004298:Dopamine / Q000032:analysis*; D051381:Rats; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
chemical_list: D001708:Biopterins; C003402:sapropterin; D004298:Dopamine