Purification of GABA on small columns of Dowex 50W; Combination with a method for separation of biogenic amines.
Elution of GABA has been included in the method for separation of biogenic amines on small Dowex 50W columns described by Atack & Magnusson (1978). After extraction with perchloric acid and neutralization to pH 3.0, the sample is passed through the column. 5-HIAA might thereafter be eluted as described by Lindqvist (1971). After washes with water and 8 ml of a 0.025 M sodium citrate buffer pH 4.5, GABA is eluted in 3 ml of a 0.05 M sodium citrate buffer pH 5.3-5.4. The elution of tryptophane and the above mentioned amines can thereafter be performed. The recovery of GABA from the column is 100%. Silica thin-layer chromatography of the GABA eluate gives only one ninhydrine positive spot with a Rf value identical to GABA standard. When the GABA eluate from a whole brain and spinal cord is run on an automatic amino acid analyser, only one peak is detectable, eluted in the same position as GABA standard. Fluorescence analysis of the GABA eluate gives similar values if a( ninhydrine, b) acetylacetone plus formaldehyde or c) fluorescamine is used to form the flurophore. The GABA values are identical whether the samples reacted with ninhydrine are diluted with water or copper tartrate. The GABA concentrations found in some parts of the rat brain are of the same order of magnitude as reported by other authors.
D006916:5-Hydroxytryptophan / Q000302:isolation & purification; D000818:Animals; D001679:Biogenic Amines / Q000302:isolation & purification*; D001923:Brain Chemistry; D002852:Chromatography, Ion Exchange; D008297:Male; D009638:Norepinephrine / Q000302:isolation & purification; D051381:Rats; D014020:Tissue Extracts / Q000032:analysis; D014364:Tryptophan / Q000302:isolation & purification; D015091:beta-Alanine / Q000302:isolation & purification; D005680:gamma-Aminobutyric Acid / Q000032:analysis / Q000302:isolation & purification*
D001679:Biogenic Amines; D014020:Tissue Extracts; D015091:beta-Alanine; D005680:gamma-Aminobutyric Acid; D014364:Tryptophan; D006916:5-Hydroxytryptophan; D009638:Norepinephrine