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Nile Blue Dyes

Oxazine

Oxazine is a heterocyclic compound containing one oxygen and one nitrogen atom in a double unsaturated six-membered ring. The presence of isomers depends on the relative position of the heteroatom and the relative position of the double bond. By extension, these derivatives are also called oxazines. Examples include ifosfamide and morpholine (tetrahydro-1,4-oxazine). Commercially available dihydro-1,3-oxazine is a reagent in the Meyers synthesis of aldehydes. Fluorescent dyes such as Nile Red and Nile Blue are based on aromatic benzoxazines. Cinnabar and cinnabaric acid are two naturally occurring dioxazines derived from the biodegradation of tryptophan. Nile red is an oxazine derivative fluorescent dye.

Nile blue Dyes
Figure 1. The 8 existing isomers of oxazine.

Dioxazine

Dioxazine is a pentacyclic compound consisting of two oxazine subunits. A commercially important example is Pigment Violet 23.

Nile blue Dyes
Figure 2. Synthetic route to dioxazine dyes.

Benzoxazine

The benzoxazine formed by the reaction of phenol, formaldehyde, and primary amine polymerizes to form a polybenzoxazine network when heated to about 200 ° C (~ 400 ° F). The obtained high molecular weight thermosetting polymer-based composites can be used in applications requiring higher mechanical properties, flame resistance and fire resistance than epoxy resins and phenolic resins.

Nile Blue

Nile Blue (or Nile Blue A) is a stain used in biology and histology. It can be used with live or fixed cells and gives the nucleus a blue color. It can also be used in combination with fluorescence microscopy to stain the presence of polyhydroxybutyrate particles in prokaryotic or eukaryotic cells. Boiling the Nile Blue solution with sulfuric acid produces Nile Red (Nile Blue Oxazolone). Nile blue is a fluorescent dye. The fluorescence shows especially in nonpolar solvents with a high quantum yield.

Nile blue Dyes
Figure 3. Chemical structure of Nile Blue.

Nile Blue in DNA Electrophoresis

Nile blue is also apparently used in a variety of commercial DNA staining formulations for DNA electrophoresis, as it does not require the ultraviolet transillumination illumination visible in agarose gels like agarose gels, so it can be used to observe DNA. It can be isolated or used as a dye to help gel extract DNA fragments without being damaged by UV radiation.

Nile blue Dyes
Figure 4. Nile blue in water. Left to right: pH 0, pH 4, pH 7, pH 10, pH 14.

Nile Blue in Oncology

Nile blue derivatives are potential photosensitizers in the photodynamic therapy of malignant tumors. These dyes accumulate in tumor cells, especially in lipid membranes, and / or are sequestered and concentrated in subcellular organelles. Using Nile Blue derivative N-ethyl-Nile Blue (EtNBA), it is possible to distinguish between normal and precancerous tissue in animal experiments. In fluorescence imaging by fluorescence spectroscopy. EtNBA has no phototoxic effect.

Synthesis

Nile Blue and related naphthoxazinium dyes can be prepared by acid-catalyzed condensation of either 5-(dialkylamino)-2-nitrosophenols with 1-naphthylamine, 3-(dialkylamino)phenols with N-alkylated 4-nitroso-1-naphtylamines, or N,N-dialkyl-1,4-phenylenediamines with 4-(dialkylamino)-1,2-naphthoquinones. Alternatively, the product of an acid-catalyzed condensation of 4-nitroso-N,N-dialkylaniline with 2-naphthol (a salt of 9-(dialkylamino)benzo[a]phenoxazin-7-ium) can be oxidized in the presence of an amine, installing a second amino substituent in 5-position of the benzo[a]phenoxazinium system. The following scheme illustrates the first of these four approaches, leading to Nile Blue perchlorate:

Nile blue Dyes
Figure 5. Synthesis of Nile Blue.

Reference:

  1. Lin, CW.; et al. Lysosomal localization and mechanism of uptake of Nile blue photosensitizers in tumor cells. Cancer Research. 1991, 51 (10): 2710–9.

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