Millon’s test

The tryptophane is precipitated by adding a solution of 200 g. of mercuric sulfate (Note 5) in a mixture of i860 cc. of water and 140 cc. of concentrated (95 per cent) sulfuric acid. After standing for twenty-four to forty-eight hours, the clear liquid is siphoned out and the yellow precipitate is filtered off and washed (Note 6) with a solution of 100 cc. of concentrated sulfuric acid in 1900 cc. of distilled water containing 20 g. mercuric sulfate, until the filtrate is colorless and Millon s test is atypical (Note 7) about 1500 cc. is necessary. The precipitate is washed with three successive 500-cc. portions of distilled water to remove most of the sulfuric acid. 

Proteins respond to the following color tests (a) biuret, pink to purple with an excess of alkali and a small amount of copper sulfate (b) ninhydrin. a blue color when boiled with ninhydrin (triketohydrindene hydrate), which is intensified by the presence of pyridine (c) Millon s test for tyrosine, a brick-F color or precipitate when boiled with mercuric nitrate in an excess of nitric acid (d) Hopkins-Cole test for tryptophan, a violet zone with a salt of glyoxylic acid and stratified over sulfuric acid and (e) xanthoproteic test, a brilliant orange zone when a solution in concentrated nitric acid is stratified under ammonia. Almost all so-called alka-loidal reagents will precipitate proteins in slightly acid solution. 

The Millon s test (27) is negative when all the tyrosine in a protein is iodinated (278-280). This serves as a convenient method of detectii completion of the reaction. 

Salts of Millon s Base.—Suspend a little of the base in water and shake with a dilute solution of potassium iodide. The solution turns alkaline. The brown precipitate is the same as that produced by Nessler s solution in tests for ammonia. 

This coloration may, however, be due to maltol derived from torrefied malt. If, then, the reaction with ferric chloride is obtained, a portion of the aqueous solution of the residue from the ethereal extract should be tested with Millon s reagent2 a red coloration indicates salicylic acid, whilst no coloration shows that the reaction with ferric chloride is due to maltol. 

Strong Acid Fraction Aromaticity Ferric Chloride Folin-Ciocalteu Reagent Liebermann s Test Millon s Reagent Nessler s Reagent... 

Colour Tests. Ferric Chloride—brown Folin-Ciocalteu Reagent—blue Liebermann s Test—black Millon s Reagent— pink— red. 

Colour Tests. Ammoniacal Silver Nitrate—black p-Dimethyl-aminobenzaldehyde—red/- Ferric Chloride—green-brown Folin-Ciocalteu Reagent—blue Methanolic Potassium Hydroxide—red Millon s Reagent—red-orange Nessler s Reagent—black Palladium Chloride—orange - brown Potassium Dichromate—brown. 

Colour Tests. Liebermann s Test—orange Marquis Test— yellow Millon s Reagent—red Sulphuric Acid—yellow. 

Colour Tests. Folin-Ciocalteu Reagent—blue Mandelin s Test—green- blue rim Millon s Reagent—red. 

Colour Tests. Aromaticity (Method 2 —yellow/brown Folin-Ciocalteu Reagent—blue Liebermann s Test—violet-brown Millon s Reagent—red. 

Colour Tests. Ammoniacal Silver Nitrate—yellowbrown/ black Ferric Chloride—green Folin-Ciocalteu Reagent— blue Methanolic Potassium Hydroxide—pinkred-brown Millon s Reagent (cold)—brown-red Nessler s Reagent—black Palladium Chloride—orange (- brown) Potassium Dichromate (Method 1)—green brown. 

III. Millon s Reaction.—The reagent for this test is a solution of mercury in nitric acid containing nitrous acid. When the reagent is added to a solution or suspension of the protein in water, there is obtained either in the cold, or after boiling, a pink coloration of the fluid, or a pink to brownish red coloration of the precipitated protein. The reaction is given by all derivatives of benzene in which one of the hydrogen atoms has been replaced by a hydroxyl group. It thus serves as a delicate test for the presence of tyrosine in a protein. 

If PgCy is slowly added to a hot mixture of NH3 and the fusible white precipitate , Pg(NH3)2]Cl2, is formed, an example of (1) above. However, if NH3 is added to PgCy the infusible white precipitate , HgNH2Cl, is obtained, an example of (2) above. The addition of NH3 to Pgy or, more readily, the reaction between NH3 and Nessler s reagent , pgU] and OH, precipitates the reddish-brown iodide of Millon s base [an example of (3) above], i.e., Hg2Nl. This is a test for NH3 and is sometimes inconveniently over-sensitive. Rather similar bromides are Pg(NH3)2]Br2, HgNH2Br and Hg2NBr. 

In addition to the precipitation reagents, a large variety of color-tests is in use for the detection of proteins. (A) In Millon s Reaction, the material is treated with nitric acid, in which a small... 

In these reactions the tested organic substance and an inorganic salt give a more or less stable strong color due to a complex salt. In Part 2 of this monograph a series of such reactions is described, such as, for example, the reaction of hydroxamic acids with ferric salts (p. 276), the reaction of phenols with ferric chloride (p. 188), the reaction of molybdenum with o-dihydro-xybenzenes (p. 191), of diacetyl dioxime with nickel salts (p. 227), of phenols with Millon s reagent (p. 196), alcohols with ceric ammonium nitrate (p. 170), alcohols with vanadium-hydroxy quinoline complex (p. 171), and the reaction of cis-enols of )5-dicarbonylic compounds and a-dicarbonyl compounds with ferric chloride, (p. 294.)... 

Millon et al. performed unconfined compressive tests on 10 wt% PVA-C samples through 1, 3, and 6 FTCs. The samples were tested using strain-rates of 1, 10, and 100 %/s at 0-45 % strain at 37 °C to mimic the physiological conditions of cartilage. Figure 3 shows the strain-rate dependency of 10 % PVA at 1 and 6 FTCs. The authors concluded that PVA-C exhibits the same exponential characteristic in stress-strain behavior as cartilage. However, it has weak strain-rate dependency. Only the six-cycle samples showed a statistically significant difference between the strain rates tested. The elastic modulus measured was 1.18 MPa at 45 % strain and at 100 %/s strain rate [45]. 

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