Pyronines

For uniformity with the stmctures given in the Colourindex the ammonium radical (9) is used for the amino-substituted xanthenes and the keto form for the hydroxy derivatives. The xanthene dyes may be classified into two main groups diphenylmethane derivatives, called pyronines, and triphenylmethane derivatives (eg, (4)), which are mainly phthaleins made from phthaUc anhydride condensations. A third much smaller group of rosamines (9-phenylxanthenes) is prepared from substituted ben2aldehydes. The phthaleins may be further subdivided into the following fluoresceins (hydroxy-substituted) rhodamines (amino-substituted), eg, (6) and mixed hydroxy/amino-substituted. 

Pyronines. Pyronines are diphenylmethane derivatives synthesized by the condensation of y -dialkylarninophenols with formaldehyde, followed by oxidation of tiie xantiiene detivative (12) to the coiiesponding xanthydiol (13) which in the presence of acid forms the dye (14). If R is methyl, the dye produced is... 

Succineins. Succineins are carboxyethyl-substituted pyronines made by substituting succinic anhydride for formaldehyde in the basic synthesis,... 

Fig. 5. Stmctural interrelationships among diaryl and triaryl carbonium dyes. Pyronine = xanthene (61), R = CH3 is Bindschedler s Green.

Pyronin Y [3,6-bis(dimethylamino)xanthylium chloride] [92-32-0] M 302.8, m 250-260, Cl 45005, Xmax 522nm, pKesi Commercial material contained a large quantity of zinc. Purified by dissolving Ig in 50mL of hot water containing 5g NaEDTA. Cooled to 0, filtered, evapd to dryness and the residue extracted with EtOH. The soln was evaporated to 5-lOmL, filtered, and the dye pptd by addition of excess dry diethyl ether. It was centrifuged and the crystals were washed with dry ether. The procedure was repeated, then the product was dissolved in CHCI3, filtered and evapd. The dye was stored in a vacuum. 

Pyronin B [di-(3,6-bis(diethylamino)xanthylium chloride) diFeCIs complex] [2150-48-3] M 358.9 (Fe free), m 176-178° (diFe complex). Cl 45010, X.max 555nm, pK 7.7. Crystd from EtOH. Forms Fe stain. 

This technique was employed to study the binding dynamics of Pyronine Y (31) and B (32) with /)-CD/ s The theoretical background for this particular system has been discussed with the description of the technique above. Separate analysis of the individual correlation curves obtained was difficult since the diffusion time for the complex could not be determined directly because, even at the highest concentration of CD employed, about 20% of the guest molecules were still free in solution. The curves were therefore analyzed using global analysis to obtain the dissociation rate constant for the 1 1 complex (Table 12). The association rate constant was then calculated from the definition of the equilibrium constant. 

Table 12 Association and dissociation rate constants for pyronine/CD complex at 21 °C65...

Many fluorophores of practical interest are heterocyclic coumarins, rhodamines, pyronines, fluoresceins, oxazines, etc. A few examples will now be presented. 

In pyronines, replacement of the carbon atom of the central ring (opposite to the oxygen atom) by a nitrogen atom produces oxazine dyes, which are used as laser dyes emitting in the range 600-750 nm. 

Pyronin G Pyronin Y N-[6-(Dimethylamino)-3H-xanthene-3-ylidene- dene]-N-methyl-methanaminium chloride... 

The microsomal fraction was first obtained by Claude in 1943. In addition to lipid in the fraction, he noted the presence of RNA-rich granules, consistent with reports from Brachet that cytoplasm stained for RNA by the methyl-green/pyronin procedure. Glucose-6-phos-phatase was a prominent enzyme when the fraction was prepared from liver. Since density gradient sedimentation showed G-6-P-ase was absent from mitochondria and lysosomes, it was used as a marker for liver microsomes. 

Pyronine Y (PY) with beta cyclodextrin by using the temperature-jump technique. It was found that the kinetic data could be adequately explained by a mechanism involving the stepwise association of two beta cyclodextrin molecules (C) to the dye. Thus,... 

On addition of Na -saponite to the rhodamine-ethanol solution and the pyronine-ethanol solution, the color of the solutions gradually faded within a few hours, even at room temperature. All the composites were intensely colored, namely bright red for Rhodamine 590 and cardinal for Pyronine Y. 

From the X-ray diffraction data and the calculated sizes of the dye molecules, the conformation of the dye molecules in the interlayer was briefly estimated. Fig. 1 shows the conformation of the dyes, in which the xanthene nucleus of pyronine or rhodamine was positioned parallel, and the phenyl group of rhodamine perpendicular, to the silicate layers of the clay. 

Fig. 1. Schematic illustration on the conformations of rhodamine 590 and pyronine Y in the interlamellars of clay. The values indicated in the figure were estimated from X-ray diffraction patterns.

From the value of the CEC, the interlamellar surface area per cation equivalent could be calculated to be ca 91.7 A. With the assumption that the cross section of the dye molecule was a square, the approximate areas were estimated to be ca 90 A for pyronine and ca 180 A for rhodamine. In this... 

As a function of the dye content intercalated, the relative intensity of fluorescence, the maximum wavelength in the fluorescence spectrum of lax d-spacing are illustrated in Fig. 2(a) for the pyronine Y-saponite composite (referred to as PY-SA), and Fig. 2(b) for the rhodamine 590-saponite composite (referred to as R590-SA). 

Fig. 2. Intensity, in fluorescence and li-spacing vs the contents of pyronine Y (a) and rhodamine S90...

Pyronin B, fluorometry, 640 Pyruvate, lactate determination, 625, 626 Pyruvic acid, hydrogen peroxide determination, 655... 

Pyronin Y (Pyronin G, C.l. 45005) moves some faster than bromophenol blue and thus better indicates the electrophresis front. A further benefit is the strong binding to nitrocellulose which allows the identification of the electrophoresis front after immunochemicl reactions in Western blotting. 

Dissolve samples in buffer G. Electrode buffer is Soln H. Applied voltage should be 8 -10 V per centimeter of gel length. Tracking dye has to be an anionic one intensively colored at pH 2.4 (e.g., cresol red or Pyronin Y). 

Basic fuchsin (rosaniline, basic violet 14) or Pyronin Y are suitable as tracking dye. 

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