Dabsyl chloride

Dansyl chloride (5-dimethylaminonaphthlene-l-sulphonyl chloride) and the related dabsyl chloride (4-dimethyl-aminoazobenzene-4 -sulphonyl chloride) have also been used. Sensitivity of the latter method is poor and both suffer problems from interference with reagent excess. 

Derivatization with dabsyl chloride (94) was applied for the separation of primary amino acids in physiological samples, prior to determination of their specific radioactivity. The derivatization is easy to perform and the derivatives are stable252. 

The remaining dabsyl chloride is not removed, since it reacts with Tris of Soln. C to a dye acting as tracking dye like bromophenol blue. 

Various procedures are used to analyze protein primary structure. Several protocols are available to label and identify the amino-terminal amino acid residue (Fig. 3-25a). Sanger developed the reagent l-fluoro-2,4-dinitrobenzene (FDNB) for this purpose other reagents used to label the amino-terminal residue, dansyl chloride and dabsyl chloride, yield derivatives that are more easily detectable than the dinitrophenyl derivatives. After the amino-terminal residue is labeled with one of these reagents, the polypeptide is hydrolyzed to its constituent amino acids and the labeled amino acid is identified. Because the hydrolysis stage destroys the polypeptide, this procedure cannot be used to sequence a polypeptide beyond its amino-terminal residue. However, it can help determine the number of chemically distinct polypeptides in a protein, provided each has a different amino-terminal residue. For example, two residues—Phe and Gly—would be labeled if insulin (Fig. 3-24) were subjected to this procedure. 

Dabsyl Chloride (4-dimethyl-aminoazobenzene-4-sulfonyl chloride) Aabs = 420 nm. Derivatives are very stable (days) and can be formed from both primary and secondary amino acids. Detection is by absorption only. Detection limits are in the high picomole range. Reaction time is typically around 10 minutes at 70°C. Completeness of reaction can be adversely affected by the presence of high levels of various salts. Because reaction efficiency is highly matrix dependent and variable for different amino acids, standard amino acid solution should be derivatized under similar conditions/matrix for accurate calibration. Commercial systems available uti-... 

EHJM Jansen, RH van den Berg, R Both-Miedema, L Doom. Advantages and limitations of precolumn derivatization of amino acids with dabsyl chloride. J Chromatogr 553 123-133, 1991. 

A,iV-dimethylaminoazobenzene-4 -sulfonyl — usually as the chloride, DABSYL chloride... 

Figure 4.20. Determination of the Amino-Terminal Residue of a Peptide. Dabsyl chloride labels the peptide, which is then hydrolyzed with the use of hydrochloric acid. The dabsyl-amino acid (dabsy 1-alanine in this example) is identified by its chromatographic characteristics.

Dabsyl chloride A-N,N-dimethylaminoazobenzene-4 -sulfonyl chloride) Visible 436 nm Low fmol 18-44 min precolumn Multiple products, critical concentration Stable product, good separation, high sensitivity... 

Dimethylamino)azobenzene-4-sulfonyl chloride (dabsyl chloride or DABS-Cl) [dabsyl derivatives]... 

Dabsyl chloride is an alternative to DABITC as a derivatization reagent to be used for manual sequencing. Dabsyl chloride reacts with primary and secondary amino acids forming red-orange derivatives that are stable for months. The method offers excellent sensitivity, ease, and speed of preparation and high-resolution capabilities. However, it suffers from interferences with ammonia present in biological samples. Furthermore, it results in a relatively reduced column lifetime due to the utilization of excess of Dabsyl chloride [9]. 

Another labelling agent suitable for determination by HPLC of histamine, tyramine, phenylethylamine, tryptamine, cadaverine, putrescine, spermidine and spermine in wines is 4-(4-dimethyl-aminophenylazo)benzenesulfonyl (dabsyl) chloride (see Figure 4.14) (Bockhardt et al., 1996 Romero et al., 2000). 

A peptide composed of 12 amino acids does not react with dabsyl chloride or with phenyl iso thiocyanate. Cleavage with cyanogen bromide yields a peptide "with a carboxyl-terminal homoserine lactone residue, which is readily hydrolyzed, in turn yielding a peptide whose sequence is determined by the Edman procedure to be... 

Why does the untreated peptide fail to react with dabsyl chloride or phenyl isothiocyanate ... 

The sequences of the peptides produced by the two cleavage methods are circular permutations of each other. Thus, the peptide is circular, so it has no free a-amino group that can react with dabsyl chloride or with phenyl isothiocyanate (see Figure 4.5). 

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