Abstract: The catalytic activity of metalloporphyrins in the chemiluminescent reaction of luminol and H2O2 is inhibited by the presence of an amino-acid and this effect has been investigated as a possible sensitive post-column detection procedure for amino-acids. The relative response of >20 amino-acids were measured and significant responses were obtained only for L-cysteine, L-tyrosine, L-tryptophan and L-cystine, with detection limits from 68 nM to 22 µM. Four amino acids were determined on the basis of the finding that the catalytic activity of mimetic peroxidase (metalloporphyrin) in the chemiluminescence reaction between luminol and hydrogen peroxide is inhibited in the presence of an amino acid. The degree of chemiluminescence inhibition is a measure of the amino acid concentration. The electrostatic interaction between amino acid and metalloporphyrin was confirmed by comparing the degree of inhibition of cationic and anionic metalloporphyrin-catalyzed chemiluminescence reactins. More than 20 amino acids were tested, and only L-cysteine, L-tyrosine, L-tryptophan and L-cystine significantly quenched the chemiluminescence intensity. The detection limits were 6.8 x 10^-8, 1.3 x 10^-7, 8.5 x 10^-6 and 2.2 x 10^-5M, respectively. The detection approach is demonstrated with a silica-based liquid chromatography separation of amino acids using phosphate buffer (pH 7.3) as mobile phase. Compared with other chemiluminescence analyzes, this method is faster, can be run at room temperature and, in favorable cases, has a lower detection limit.

Abstract: An automatic stopped-flow method is proposed for the routine determination of microgram amounts of Image-cysteine, based on its oxidation by 2,6-dichlorophenolindophenol (DPIP) in a weakly basic medium. The oxidation reaction is monitored by measuring the rate of the absorbance decrease at 615 nm, the wavelength of the maximum absorption for DPIP. Under optimal conditions, the calibration graph is linear over the range 2-32 µg/ml and the detection limit is 300 ng/ml. Most of the amino-acids tested have no effect on the rate of the oxidation reaction and are tolerated at high concentration levels. The sample throughput achieved, 80 samples per hr, allows the proposed method to be used for the routine determination of L-cysteine.

Abstract: Amino-acids were found to accelerate the Cu(II)-catalyzed oxidation of di-2-pyridyl ketone hydrazone (I) to form a fluorescent compound in acidic medium. With use of this enhancement effect, L-histidine, L-cysteine, L-glutamic acid, glycine, DL-serine and L-arginine were determined by flow injection analysis (diagram given). Sample was injected into a carrier stream of water which was merged with stream of 0.5 µM I - 0.1 M NaOH, passed through a reaction coil operated at 25°C, and mixed with a stream of 0.4 M HCl before fluorimetric detection at 435 nm (excitation at 349 nm). The detection limit of L-histidine was 2 pmol. The method was also used to determine proteins which were found to decrease the catalysis; the detection limit for bovine serum albumin was 20 ng.