抄録
It was found that amino sugars and amino acids reacted with pyridoxal and zinc (II) ion in pyridirle-methanol solution to yield highly fluorescent zinc (II) chelates of N-pyridoxylidene ami+no sugars or amino acids. The recommended procedures are as follows:
Method A: 0.5ml each of20% pyridine-methanol solution and of 0.05% pyridoxal solution in methanol were added to 0.5ml of the samplesolution (containing about 5μg of amino sugar). After having allowed it to stand for 30 minutes at room temperature, 0.5ml of 2% Zn (NO3) 2 solution was added. And the nafter further 15 minutes of standing at room temperature, 3.0ml of methanol was added. The fluorescence intensity was measured by a fluorophotometer (Ex. 390 mμ, Em. 470 mμ).
Method B: A portion of mucopolysaccharide was dissolved in 2N HCl, sealed in an ampoule under N2 gas and hydrolyzed at 100°C for 5-17 hours. The tube was opened and usually the aliquot amount of the content was evaporated to dryness in a desiccator in vacuo (P2O5 and KOH). The residue is finally dissolved in a suitable amount of redistilled water. The resultant solution was analyzed by Method A.
Method C: 0.5ml of the sample solution (containing 5-25×10-9 moles) was mixed with 5ml of pyridoxal-Zn (II) reagent (10mg Pyridoxal HCI and 100 mg zinc acetate were dissolved in 2% pyridine methanol solution), and then allowed to stand for one hour at 37°C. The fluorescence intensity of the reaction mixture was measured at Ex. 390mμ and Em. 470 mμ.
This fluorescence reaction is sensitive and selective for amino sugars and amino acids.
The zinc (II) chelate of N-pyridoxylidene amino sugar or amino acid contains the 1: 1 molar ratio of ligand to metal ion.
