5-Mercapto-2-nitrobenzoic acid

The application of CPO, HRP and CiP is limited to sterically unencumbered substrates and all these peroxidases produce the same absolute configuration of the chiral hydroperoxide. To overcome this limitation, the semisynthetic enzyme selenosubtilisin, a mimic for glutathione peroxidase, with the peptide framework of the serine protease subtilisin was developed by Bell and Hilvert. This semisynthetic peroxidase catalyzes the reduction of hydrogen peroxide and hydroperoxides in the presence of 5-mercapto-2-nitrobenzoic acid. It was utilized by Adam and coworkers and Schreier and coworkers for the kinetic resolution of racemic hydroperoxides (equation 17) . The results obtained were very promising. 

Thiols can be detected on insoluble supports by treatment with the symmetric disulfides of 4-mercaptonitrobenzene or 5-mercapto-2-nitrobenzoic acid ( Ellman s reagent [69,70]). These reagents are reduced by thiols to the corresponding thiophe-nolates, which are intensely colored. 

FIGURE 58.1. Acetylthiocholine is hydrolyzed by AChE to acetic acid and thiochohne, which forms yellow 5-mercapto-2-nitrobenzoic acid after reaction with Ellman reagent. 

Electrochonical Sensors, Biosensors and Their Biomedical Applications 5 -Mercapto -2 -nitrobenzoic acid (yellow)... 

A very sensitive and commonly used method for ChE determination was described by Ellman et al. (1961). This method uses thiocholine esters as substrates (acetyl- and butyrylthiocholine or others). After enzymatic hydrolysis, the relevant acid and thiocholine are released and thiocholine by its SH-group is detected using 5,5 dithiobis-2 nitrobenzoic acid (DTNB) forming 5-mercapto-2-nitrobenzoate (3-thio-6-nitrobenzoate, TNB) with an absorbance maximum of approximately 412nm (Figure 65.1). 

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