Spectrophotometric characteristics and assay of pure pyronin Y
SummaryThe spectrophotometric characteristics of analytically pure pyronin Y have been investigated. Addition of metal ions (Fe3+, Zn2+, and Mg2+) and of dextrin were shown not to influence the absorption characteristics. The composition of the solvent strongly influenced the value of the extinction coefficient. Aqueous ethanolic solutions with a content of about 50% ethanol gave higher ε-values than those found for more concentrated ethanol solutions. The difference can be explained by the existence of a solvent-solute complex in the less concentrated ethanol solutions. A new spectrophotometric assay is proposed using the ε-value 11.7×104 lmol−1 cm−1 found in aqueous ethanol (52%) as standard.
Purity of commercial non-certified European samples of Pyronin Y
SummaryThe purity of six European non-certified samples of Pyronin Y was compared with that of two American samples certified by the Biological Stain Commission. The methods used were spectrophotometry and a Methyl Green-Pyronin staining test (both as applied by the Biological Stain Commission), thin layer chromatography, mass spectrometry, determination of pH, and content of some electrolytes. It was found that none of the European batches of Pyronin Y passed the complete test as prescribed by the Biological Stain Commission. Their dye content was uniformly low (between 5 and 19%). Furthermore, thin layer chromatography and mass spectrometry revealed that two of the dye samples contained no Pyronin Y or only traces.It is concluded that assessment of an unknown sample of a dye labelled Pyronin Y should be initiated with thin layer chromatography. The pH and content of electrolytes in an aqueous solution of the dye should also be determined in order to obtain reproducible staining results. Finally, the value of the work performed by the Biological Stain Commission is underlined, although more sophisticated methods are necessary for testing the purity of dyestuffs.
Dextrin and salt as impurities of histological dyes
SummaryThe dextrin and salt contents of 18 common histological dyes (usually obtained from more than one supplier) were estimated by solvent extraction of dye from the commercial sample with organic solvents such as ethanol, iso-propanol and methanol. The procedure is also suitable for preparing large quantities of dextrin and salt-free dyes. The dye contents of the commercial dye samples varied widely:many commercial dyes were pure but nearly half the samples contained less than 75% dye. The solvent insoluble residues from 10 commercial pyronin and rhodamine dyes were studied in more detail. Tests for dextrin and for NH4+, Cl− and SO42−ions were applied and the dextrin:salt ratio determined by combustion. Only 3 of the residues contained appreciable amounts of salt. Certain samples contained organic materials other than dextrin.