Nitroprusside reaction

Nitroprusside reaction. Add 1 ml. of a freshly prepared solution of sodium nitroprusside to the aldehyde or its solution. Add dil. NaOH solution in excess a red coloration is produced. 

Testing for Sulphur.—The qualitative test for sulphur is carried out in the same way as that for nitrogen. Ignite the substance in a small tube with sodium, dissolve the product in water, and add to one half of the cooled solution a few drops of sodium nitroprusside solution freshly prepared by shaking a few particles of the solid salt with cold water. A violet colour indicates the presence of sulphur. Since the nitroprusside reaction is extremely sensitive and does not allow any estimate of the amount of sulphur to be made, filter the second half of the liquid, add lead acetate solution to the filtrate, and acidify with acetic acid. According as the amount of sulphur is small or large, a dark turbidity or a more or less heavy precipitate will form. 

Cool to 20 C, add to each liter 40 gm iodoacetic acid in 150 ml water adjusted to pH 7 to give pH 9 in mixture stand until nitroprusside test is negative. Add sufficient thioglycolate to give positive nitroprusside reaction. Dialyze against tap water. Then either... 

Cystine, or cysteine hydrochloride (which has been neutralized with KOH), and KCN in water, treated with H202 until the nitroprusside reaction is negative, whereby the temp, rises to 30-40°, more H202 added, allowed to stand for 3 hrs., and isolated as the Cu-salt —Cu-a-amino-y5-thiocyano-propionate. Y 90 and 79% respectively.—When prepared from cystine (II), H202, slowly added, serves to reoxidize the cysteine (I) which has been formed. (A. Schoberl and R. Hamm, Chem. B. 81, 210 (1948).)... 

The nitroprusside reaction is noteworthy for being highly specific for —SH groups (if carried out in faintly alkaline solutions) but notoriously difficult for defining detail of distribution. The color developed fades quickly also fairly thick sections are required to give adequately robust... 

These disadvantages of the otherwise excellent nitroprusside reaction are not present in the case of the Prussian blue method which has been applied to the same problems by Chevremont and Frederic (25). However, the Prussian blue methodjs not specific for —SH groups, but... 

Spectrophotometric Complex formation with metals (e.g. ferric ions) Colorimetric reaction e.g. nitroprusside reaction)... 

Fig. 1. Nitroprusside reaction of glutathione in water and 7 M urea. The concentrations of glutathione and urea are on the basis of the volume before the addition of the color reagents. This applies to all concentrations given in this paper.

Fig. 3. Effect of urea concentration on the nitroprusside reaction of glutathione.

Effect of Urea and Guanidinium Salts on the Nitroprusside Reaction of Glutathione and Thioqlycolic Acid... 

Rapkine s quantitative data are limited to the description of the fluctuation of soluble SH in the course of division. His proposal that the rise in soluble SH during the predivision stage is caused or at least accompanied by an increase in protein SH due to denaturation is not based on quantitative evidence and is not a logical consequence of the soluble SH picture. Nevertheless, it is the latter hypothesis that has attracted the most attention. The hypothesis led a number of workers to anticipate an increase, in connection vnth cell division, of total SH demonstrable by cytochemical procedures. Rapkine himself reported that he observed an increase in the intensity of the nitroprusside reaction during Phase II of the division of the sea-urchin egg and in certain Protozoa. 

The nitroprusside reaction has been used to study the localization of sulfhydryl groups in the adrenal gland (149, 152). The adrenal cortex shows a more intense nitroprusside staining reaction than does the medulla (149). In young guinea pigs the inner zone of the cortex (Z-reticularis) gives a more intense nitroprusside reaction than does the rest of the cortex (152). These histochemical methods do not differentiate between protein-bound sulfhydryl, GSH, and other diffusible sulfhydryl compounds. 

The nitroprusside reaction will not work (i.e., it will not produce colored products) with primary and secondauy amines if the pH is acidic. Why ... 

It should be stressed that in order to use the delayed nitroprusside reaction for the detection of acetyl CoA it was necessary first to remove the large amounts of the free mercaptans, mostly glutathione, from the yeast Kochsaft which otherwise would have strongly interfered with the color reaction. We achieved this by titration of the Kochsaft with iodine, thereby oxidizing thiols to disulfides which do not any longer react with nitroprusside at alkaline pH. The identity of the sulfhydryl component of our active preparations with coenzyme A was finally proved in various ways which should not be discussed here in detail. 

The nitroprusside reaction was positive but developed very slowly. This delayed reaction is characteristic of acylated mercaptans, the positive reaction depending on prior hydrolysis of the S-acyl link. 

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