Fuchsin, preparation

Stains.—One of the most useful bacteriologic stains is ZiehVs Carbol Fuchsin, prepared as follows ... 

The first triaryknethane dyes were synthesized on a strictiy empirical basis in the late 1850s an example is fuchsine, which was prepared from the reaction of vinyl chloride with aniline. Thek stmctural relationship to triphenylmethane was estabHshed by Otto and Fmil Fischer (5) with the identification of pararosaniline [569-61-9] as 4,4, 4 -triaminotriphenyknethane and the stmctural elucidation of fuchsine. Several different stmctures have been assigned to the triaryknethane dyes (6—8), but none accounts precisely for the observed spectral characteristics. The triaryknethane dyes are therefore generally considered to be resonance hybrids. However, for convenience, usually only one hybrid is indicated, as shown for crystal violet [548-62-9] Cl Basic Violet 3 (1), for which = 589 nm. 

In a variation of this method, isolation of the ben2hydrol derivative is not required. The methane base undergoes oxidative condensation in the presence of acid with the same or a different arylamine direcdy to the dye. New fuchsine [3248-91 -7] Cl Basic Violet 2 (16), is prepared by condensation of two moles of o-toluidine with formaldehyde in nitrobenzene in the presence of iron salts to give the corresponding substituted diphenylmethane base. This base is also not isolated, but undergoes an oxidative condensation with another mole of o-toluidine to produce the dye. 

Measurements show some variation depending upon the staining solution used and the method of application. In dried and fixed smears, the cell wall and slime layer do not stain with weakly staining dyes such as methylene blue but do stain with the intensely staining pararosaniline, new fuchsin, crystal violet, and methyl violet. The great majority of bacteria have been measured in fixed and stained preparations. In some instances dried, negatively stained smears have been used. Therefore, the method employed should be specified when measurements of bacteria are reported otherwise the results will be of doubtful v alue. 

The technical preparation of crystal violet and of its methyl-free parent substance, parafuchsine, almost the oldest of the triphenylmethane dyes, is not so easily explained. As is well known, in this process aniline and p-toluidine are united by oxidation in an acid melt. (In the preparation of fuchsine itself, which contains a methyl group attached to one of the benzene rings, o-toluidine is an additional ingredient.) Although all the phases of this important synthesis have not yet been experimentally established, we may nevertheless explain it on the basis of a dehydrogenation similar to that involved in the formation of malachite green. Moreover, the union of several molecules of base proceeds exactly according to the principle on which indamines are formed (p. 321) (Bucherer). 

Tin(lV) chloride is a mordant for dying fabrics a stabihzer for perfume in soap used in weighting silk in ceramic coatings in manufacturing blue print papers and to produce fuchsin. Also, tin(IV) chloride is used in preparing many organotin compounds. 

The last compound 4 4 -tetramethyl-di-amino-benzophenone is also known as Michler s ketone, and is an important intermediate in the preparation of dyestuffs of the fuchsine series, e.g., crystal violet. For the preparation of this ketone by treatment of dimethylaniline with carbon dioxide in presence of aluminium chloride, see E.P., 353464. 

Photochemical preparation of stable dyes by irradiation of aminotriarylacetonitriles Photosensitive leucocyanide compositions Photoactivation of leucocyanides Solid photosensitive compositions Purification of triarylrnethane leucocyanides Fuchsine cyanide... 

Procedure Transfer 10.0 mL each of the Standard Preparation and the Test Preparation to separate 25-mL volumetric flasks. Add 5.0 mL of Potassium Permanganate and Phosphoric Acid Solution to each, and mix. After 15 min, add 2.0 mL of Oxalic Acid and Sulfuric Acid Solution to each, stir with a glass rod until the solutions are colorless, add 5.0 mL of fuchsin-sulfurous acid TS (see Solutions and Indicators), dilute with water to volume, and mix. After 2 h, using a suitable spectrophotometer, concomitantly determine the absorbances of both solutions in 1-cm cells at the wavelength of maximum absorbance at about 575 nm, using water as the blank. The absorbance of the solution from the Test Preparation is not greater than that from the Standard Preparation. PH Determine as directed under pH Determination, Appendix IIB. 

The fuchsin reagent is prepared by dissolving 0-015 g fuchsin in 100 ml water. 

Fig. 1. Bovine preparations stained with Weigert s resorcin-fuchsin (Partridge and King, 1961).

Bromine may be detected by the bluish-violet colour which it imparts to a paper impregnated with Schilfs reagent. This paper may be prepared by soaking a narrow strip of filter paper in an aqueous solution containing 0 025% fuchsine decolorised with sulphur dioxide, and allowing to dry. 

Formaldehyde and Phosgene Methods.—Two other processes for the commercial preparation of fuchsin are the formaldehyde and the phosgene, COCI2, processes. For details in regard to them special books on dyes should be consulted, e.g. Cain Thorpe. 

Add four drops of a freshly prepared solution of NTB/BCIP in stain buffer, and incubate at 37°C. Examine for development of blue stain after 90 min. If color IS detected, stop the reaction by nnsing slides in T-E, pH 7.4. If blue color IS not detected, resume incubation at 37°C, and reexamine every 10 mm. Counterstain with Basic Fuchsin for 5 mm. 

Schiff s reagent is prepared by dissolving 0.1 g Basic Fuchsin (p-rosaniline hydrochloride) in too mL of water and then adding 4 mL of a saturated aqueous solution of sodium bisulfite. After I h add 2 mL of concentrated hydrochloric acid. 

The preparation of fuchsine illustrates the formation of a typical triphenylmethane dye but does not show the relations of the colored substance to the compounds which are formed in the intermediate steps. These may be represented by equations. When p-toluidine is oxidized it forms p-aminobenzaldehyde, which condenses with aniline to form triaminotriphenyhnethane. 

In the preparation of commercial fuchsine aniline is heated with a mixture of ortho and para toluidine. p-Toluidine is oxidized to the aminoaldehyde which condenses with one molecule of both aniline and o-toluidine to form homorosaniline hydrochloride- At the same time there is also formed pararosaniline hydrochloride owing to the reaction of two molecules of aniline with the aldehyde. The differences in the structure of leuco base, color base, and dye are shown by comparison of their respective formulas (1), (2), and (3). 

C) Preparation of Rosaniline Hydrochloride (Fuchsine). Place an eight-inch test tube attached to a wire on a horn balance and... 

Experiment Fuchsine-sulphurous acid is prepared by dissolving fuchsine in water a sufficient quantity of the latter is taken to prevent the solution from being too intense in colour. Sulphur dioxide is conducted into this until a complete decolouration takes place. To a few cubic centimetres of this solution add several drops of aldehyde. On shaking, a violet-red colour will be produced. 

Preparation of Fuchsine-Paper A crystal of fuchsine, the size of a lentil, is pulverised, dissolved by heating in 100 c.c. of water, and the solution filtered. 

Michler s ketone, tetramethyldiamidobenzophenone is of technical importance it is obtained from dimethyl aniline and phosgene, and is used in the preparation of dyes of the fuchsine series (see Crystal Violet) ... 

V. Airila states that mixtures of fuchsine and methyl violet have the same color as phenolphthalein in its transformation region, and may therefore be used as comparison solutions. The latter can be prepared from a 0.0125% aqueous fuchsine solution and a saturated solution of methyl violet. 

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