In vivo simultaneous monitoring of gamma-aminobutyric acid, glutamate, and L-aspartate using brain microdialysis and capillary electrophoresis with laser-induced fluorescence detection: Analytical developments and in vitro/in vivo validations.
gamma-Aminobutyric acid (GABA), glutamate (Glu), and L-aspartate (L-Asp) are three major amino acid neurotransmitters in the central nervous system. In this work, a method for the separation of these three neurotransmitters in brain microdialysis samples using a commercially available capillary electrophoresis (CE) system has been developed. Molecules were tagged on their primary amine function with the fluorogene agent naphthalene-2,3-dicarboxaldehyde (NDA), and, after separation by micellar electrokinetic chromatography, were detected by laser-induced fluorescence using a 442 nm helium-cadmium laser. The separation conditions for the analysis of derivatized neurotransmitters in standard solutions and microdialysates have been optimized, and this method has been validated on both pharmacological and analytical basis. The separation of GABA, Glu, and L-Asp takes less than 10 min by using a 75 mmol/L borate buffer, pH 9.2, containing 70 mmol/L SDS and 10 mmol/L hydroxypropyl-beta-cyclodextrin and + 25 kV voltage. The detection limits were 3, 15 nmol/L and, 5 nmol/L for GABA, Glu, and L-Asp, respectively. Moreover, submicroliter samples can be analyzed. This method allows a simple, rapid and accurate measurement of the three amino acid neurotransmitters for the in vivo brain monitoring using microdialysis sampling.
D000073738:2-Hydroxypropyl-beta-cyclodextrin; D000818:Animals; D001224:Aspartic Acid / Q000032:analysis*; D001923:Brain Chemistry; D002021:Buffers; D020374:Chromatography, Micellar Electrokinetic Capillary / Q000379:methods; D003505:Cyclodextrins / Q000737:chemistry; D015314:Dialysis Solutions / Q000737:chemistry; D019075:Electrophoresis, Capillary / Q000379:methods*; D005453:Fluorescence; D018698:Glutamic Acid / Q000032:analysis*; D006863:Hydrogen-Ion Concentration; D007834:Lasers; D017551:Microdialysis / Q000379:methods*; D009281:Naphthalenes / Q000737:chemistry; D018377:Neurotransmitter Agents / Q000032:analysis*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry; D013997:Time Factors; D047392:beta-Cyclodextrins; D005680:gamma-Aminobutyric Acid / Q000032:analysis*; D047408:gamma-Cyclodextrins
D002021:Buffers; D003505:Cyclodextrins; D015314:Dialysis Solutions; D009281:Naphthalenes; D018377:Neurotransmitter Agents; D047392:beta-Cyclodextrins; D047408:gamma-Cyclodextrins; C479659:hydroxypropyl-gamma-cyclodextrin; D000073738:2-Hydroxypropyl-beta-cyclodextrin; D001224:Aspartic Acid; D012967:Sodium Dodecyl Sulfate; D018698:Glutamic Acid; D005680:gamma-Aminobutyric Acid; C027717:2,3-naphthalenedicarboxaldehyde