Phenylketonuria (PKU) and Maple Syrup Urine Disease (MSUD) are two inborn metabolic diseases which are carried by autosomal recessive genes in man. These genetic errors result in accumulation of phenylalanine (in PKU) or valine, leucine and isoluecin (in MSUD). At high concentrations, amongst other problems, these amino acids cause mental retardation. However if detected early after birth, using special diets and other forms of therapy, mental abnormalities can be prevented. As a result in many countries screening of infants for MSUD and PKU, by measuring plasma amino acids has become a routine neonatal test. Capillary Electrophoresis (CE) assays have a number of advantages over the traditional chromatography techniques (such as GC or HPLC). These include low cost, high speed of analysis and high resolution. These characteristics, make CE an ideal method for the screening of inborn errors of metabolism. We developed a CE assay based on pre-column derivatisation of amino acids with phenylisothiocyanate. This conjugate has strong absorbance at 254 nm. CE was carried out using a Spectraphoresis 1000 instrument, fitted with 40 cm of a 25 microm capillary, at 17 degrees C. A running voltage of 18KV was used to separate the amino acid mixture in an electrophoretic buffer containing 45 mM imidazole, 6 mM borate and 208 mM SDS, fixed at pH 9 with 2-N-morpholino ethane sulfonic acid. The assay was calibrated using various concentrations of amino acid standards. LOD, LOQ, recovery, inter-day and intra-day variations of the assay were determined. Also, levels of the 4 amino acids in normal and abnormal plasma were determined and compared with HPLC.

This study was carried out to evaluate the effects of noscapine, a benzylisoquinoline alkaloid from opium poppy, on oligodendrocyte during ischemia/reperfusion-induced excitotoxic injury. Changes in intracellular calcium levels due to chemical ischemia and nitric oxide (NO) production during ischemia/reperfusion were evaluated as the hallmarks of ischemia-derived excitotoxic event. OLN-93 cell line (a permanent immature rat oligodendrocyte) was used as a model of oligodendrocyte. 30-or 60-minute-oxygen-glucose deprivation/24 hours reperfusion were used to induce excitotoxicity. MTT (3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) assay was used to evaluate cell viability. Ratiometric fluorescence microscopy using Ca 2+-sensitive indicator Fura-2/AM was utilized to assess intracellular calcium levels. NO production was evaluated by Griess method. Noscapine (4 μM) significantly attenuated intracellular Ca 2+ elevation (P < 0.001). Also, noscapine significantly decreased NO production during a 30-minute oxygen-glucose deprivation/reperfusion (P < 0.01). The inhibitory effect of noscapine (4 μM) on intracellular Ca 2+ was greater than ionotropic glutamate receptors antagonists. Noscapine is protective against ischemia/reperfusion-induced excitotoxic injury in OLN-93 oligodendrocyte. This protective effect seems to be related to attenuation of intracellular Ca 2+ overload and NO production.