MONICA CULEA*, MARIA CHIRIAC**, E. CULEA***, CAMELIA LEHENE*, ANDREEA IORDACHE*
*”Babeş-Bolyai” University of Cluj-Napoca, 1, M. Kogălniceanu st., 400084 Cluj-Napoca, Romania; email@example.com
**National Institute for Research and Development of Isotopic and Molecular Technologies, 65–103, Donath st. Cluj-Napoca, Romania
***Technical University of Cluj-Napoca, 15, C. Daicoviciu st., Cluj-Napoca
Abstract. This paper has developed an isotopic dilution gas chromatography-mass spectrometric (ID-GC-MS) rapid method for diagnosing inborn error of metabolism of some neonatal diseases. Small volumes of dry plasma or blood spots were used for neonatal blood screening for diagnosis of phenylketonuria (PKU) and maple syrup urine disease (MSUD) metabolic diseases. The blood samples were derivatized as trifluoroacetyl butyl esters and analyzed by gas chromatography coupled with mass spectrometry in the selected ionization monitoring (SIM) mode. Regression curves for standard amino acids are used for quantitative determination of valine, leucine, proline, phenylalanine and tyrosine, by using 15N-glycine as internal standard. GC/MS analyses were performed on a Trace DSQ ThermoFinnigan quadrupole mass spectrometer coupled with a Trace GC gas chromatograph. Samples were separated on a Rtx-5MS capillary column, 30 m x 0.25 mm, 0.25 µm film thickness, using a temperature program from 50 oC (1 min), then 20 oC/min to 310 oC, in the selected ion monitoring (SIM) mode. The following important ions from the mass spectra of Phe, Pro, Val, Leu and Tyr were used: m/z 91, 148, 204 for Phe, m/z 166 for Pro, m/z 168 for Val, m/z 182 Leu, m/z 203, 260, 316 for Tyr and m/z 155 for the internal standard. The following conditions were followed: transfer line temperature: 250 oC, injector temperature: 200 oC; ion source temperature 250 oC; Splitter: 10:1. Electron energy was 70 eV and emission current 100 µA.
Key words: isotopic dilution, blood, amino acids, diagnosis.
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