Spectrophotometric reagents

Reagents which give the colour reactions, upon which spectrophotometric methods are based, are called spectrophotometric reagents. Most spectrophotometric methods are based on organic reagents [1-4], 

This Section deals with the more important groups of spectrophotometric reagents. Some important reagents are presented, with formulas, in chapters devoted to individual elements. 

Pyridylazo- and thiazolylazo reagents have become of great importance in the methods of determination of metals [10,11]. 

The selectivity of the methods using PAN is enhanced by suitable selection of pH and masking agents. Iron, cobalt, and nickel, for example, react with PAN at pH = 4. At such a low pH the reagent does not form complexes with Mn, Zn and Cd. Cyanide enables manganese to be determined in the presence of Ni, Zn, Cd, Co, and Cu, which form stable cyanide complexes. Zinc and cadmium can be demasked from their cyanide complexes with formaldehyde. 

Unlike PAN, 4-(2-pyridylazo)resorcinol (PAR) (formula 4.2) is water-soluble and forms water-soluble complexes with metal ions [15,16]. In the range from 90% H2SO4 to pH 2 the following protonated forms are present HsR, H4R and H3R. The neutral PAR molecule exists between pH 2.1 and 4.2 (A,max 385 nm). The anion HR (A,max 413 nm) occurs over the pH range 4.2-7. In alkaline solutions (pH 11-13) both hydroxyl groups are dissociated. The R form has Xmax 490 nm. 

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A number of other spectrophotometric reagents and combinations thereof have been used and compared with NBP 4-p3ridinecarboxaldehyde 4-nitrophenylhydra-zone 4-pyridinecarboxaldehyde 2-benzothiazolylhydrazone 4-acetylpyridine 4-nitrophenylhydrazone 4-acetylpyridine 2-benzothiazolylhydrazone [62]. S -Dodecylisothiouronium bromide has been used with primary and secondary alkyl halides [63]. 4-Nitrothiophenol has been claimed to be better than NBP, but only when used in combination with HPLC and not as a stand-alone reagent [64]. 

Materials. Olefins were Phillips pure grade, appearing to be better than the reported 99% purity by gas chromatographic analysis on an AgN03 column. Methylene blue was U.S.P. basic blue No. 9, and methanol was Baker spectrophotometric reagent. All other solvents were either chromatographic or spectroscopic grade. 

The same three spectrophotometric reagents were compared for their abilities to differentiate mono- and polynuclear hydroxy-aluminium complexes in solutions typical of those used in phytotoxicity studies (Parker et al., 1988b). Methods based upon each of the three reagents yielded estimates of the mononuclear aluminium fraction of adequate precision for most purposes. Studies using ferron demonstrated its utility for characterising the non-mononuclear aluminium fraction using kinetic analyses. The ferron spectrophotometric procedure was preferred for its simplicity, level of precision and moderate rate of reaction with aluminium. 

As the pH is increased, spectrophotometric reagents ionize and their electronic structure becomes deformed, which often leads to a bathochromic shift of the absorption maximum. Ionization causes polarization of the chromophoric system. The formation of a chelate complex disturbs the electronic state of the organic molecule to produce, as a rule, a bathochromic shift. 

The value of e is most readily determined under conditions where the spectrophotometric reagent has zero absorbance at Xmax of the complex, and only one stable coloured complex is formed in the system. In cases where the complex is formed stepwise [as in the system Fe(IlI) - SCN ] the value of e depends on the excess of the reagent. 

Most of the colour systems, used as a basis of spectrophotometric methods are formed as a result of complexing reactions. The most important spectrophotometric reagents are presented in Chapter 4. 

The reference (comparison) solution in one-colour spectrophotometric measurements is usually the solvent (if the reagent is colourless), and in two-colour methods a reagent solution is used. As well as the spectrophotometric reagent the reference should contain other reagents and have pH identical to the analyte solution. It happens that at A-mm the absorbance of the colour reagent is zero and pure solvent may be applied as the reference. 

Thiazole azo compounds have similar properties to PAN and PAR as spectrophotometric reagents. Their reactions with metals are more selective, principally as a consequence of the lower stability of their complexes. The following reagents are examples l-(2-thiazolylazo)-2-naphthol (TAN) [22] (formula 4.6) and 4-thiazolylazo)resorcinol (TAR) [23, 24] (formula 4.7). 

Dithizone (H2DZ, diphenylthiocarbazone, 3-mercapto-l,5-diphenylformazane) is one of the foremost organic spectrophotometric reagents [83-85]. It provides the basis of sensitive methods for the determination of Pb, Zn, Cd, Ag, Pd, Hg, Cu, Bi, and other metals. It has often been used in the extractive separation of traces of metals before their determination. 

Reagents related to dithizone are 1,5-diphenylcarbazide (formula 17.1), a reagent for Cr(VI) and Os, and diphenylcarbazone which is used, inter alia, for determining mercury. To the same family of reagents belong 1,5-diphenylthiosemicarbazide (formula 42.1) and 2,4-diphenylthiosemicarbazide, which are valuable spectrophotometric reagents for Ru, Re, Se, and Te. The application of thiosemicarbazones and semicarbazones in spectrophotometry has been reviewed [94,95]. 

Other dithiocarbamates which have been applied as spectrophotometric reagents are dibenzyl dithiocarbamate, pyrazoline dithiocarbamate, glycine dithiocarbamate, and pyrrolidine dithiocarbamate [106]. 

P-Diketones are a group of commonly applied spectrophotometric reagents, including acetylacetone (formula 1.6), thienoyltrifluoroacetone (formula 1.7), and dibenzoylmethane (formula 54.1) used as a reagent for uranium. The reagents of this group are discussed in a monograph [123]. Fluorinated P-diketones are applied in the determination of lanthanides [124]. 

Thiocyanate is one of the most important spectrophotometric reagents. The availability of the reagent and the simplieity of thioeyanate methods are responsible for its great popularity in analytical laboratories [131,132]. Thiocyanate is principally used for determination of Fe(m), Mo, W, Nb, Re, Co, U, and Ti. 

Other organic spectrophotometric reagents used for the determination of Al. were Alizarin S (formula 57.1) [66,67], ferron [68], salicyloylhydrazones [69], and haematoxyline in the presence of CTA [70,71]. The use of Bromopyrogallol Red in the presence of diphenylguanidine [72] and some cationic surfactants [73,74] has been proposed. 

As with EDTA (Complexone III), DCTA ( Complexone IV) has also been used as a spectrophotometric reagent for chromium [13,14,17,31,32]. The Cr(III)-DCTA complexes have been extracted with chloroform solutions of Aliquat 336 [13,14]. 

A number of other oximes have also been recommended as spectrophotometric reagents for cobalt, namely 2,2 -dipyridylketoxime (e = 2.1 10 at 388 nm) [58], 2-pyridyl-2-thienyl-[l-ketoxime [59], and 2,2-diquinolyl ketoxime (e = 5.3-10 ) [60]. 

Many hydrazone derivatives have been proposed as spectrophotometric reagents for Co, namely 2,2 -dipyridyl-2-pyridylhydrazone (e = 4.2-10 ), 2-furaldehyde-2-pyridylhydrazone [79], benzil-2-pyridylketone-2-pyridylhydrazone [80], and 2,2 -dipyridyl-2-benzothiazolyl-hydrazone [81]. Co was determined in the presence of Hg [82], and in the presence of Ni [83] by the derivative spectrophotometry using benzil-2-pyridylketo-2-quinolylhydrazone. 

One of the more often used spectrophotometric reagents for gold is p-dimethylaminobenzylidenerhodanine (rhodanine, formula 46.3) [33,60]. The reagent reacts with gold ions in weakly acidic media e.g., 0.1 M HCl) to form a pink-violet complex, which is either stabilized in the aqueous phase, or extracted into a mixture of chloroform and benzene (3+1) or isoamyl acetate. In aqueous-pyridine medium, e = 3.8-IO at 515 nm [60]. Rhodanine and its derivatives were applied for determination of Au and other noble metals in the presence of surfactants [61-63]. 

Apart from PAR, many other azo reagents have been applied as spectrophotometric reagents for indium. l-(2-Pyridylazo)-2-naphthol (PAN) has been applied widely as analytical reagent for indium [24,36-38]. The indium chelate with PAN is extractable into chloroform from solutions of pH 6 (e = 1.9-10 ). Indium has also been determined with PAN by derivative spectrophotometry [39]. Other azo reagents proposed for indium include 2-(2-pyridylazo)-l-hydroxynaphthalene-4-sulphonic acid [40], TAR (formula 4.7) [41], Lumogallion (e = 5.4-10 at 510 nm) (formula 21.1) [42], Sulpharsazen [43], Thoron I [44], and Eriochrome Black T (extraction into n-butanol in the presence of diphenylguanidine, = 3.6-10 ) [45], Picramine M [46], 2-(2-thiazolylazo)-p-cresol (TAC) [47], and 2,4,6-tris(2-hydroxy-4-sulphonaphthylazo)l,3,5-triazine (e = 8.4-10 [48]. 

The following spectrophotometric reagents were applied for determining indium 2-(2-thiazolylazo)-p-cresol (TAC), in catalysts [47] Malachite Green, in gallium metal and in ZnGeAs2 [53] and Pyrogallol Red, in zinc alloys [7]. 

The most impoitant spectrophotometric reagents for determining nickel are dioximes, which give specific and fairly sensitive methods. An example of a very sensitive method is one using the azo reagent 5-Br-PADAP. 

A review of spectrophotometric reagents for determination of nickel has been presented... 

Several other azo compounds have been employed as spectrophotometric reagents for niobium. The reactions are carried out in the presence of complexants (tartrate, oxalate, H2O2). The coloured species produced are generally ternary niobium complexes. PAR (e = 3.6-10 ) [45-53] and Sulphochlorophenol S (e = 3.3-10 ) [54-61] are often employed. 

Other organic spectrophotometric reagents for Nb include 8-hydroxyquinoline-5-sulphonic acid [75], 5,7-dichloro-8-hydroxyquinoline (e =1.3-10 at 400 nm) [76], 5-chloro-7-iodo-8-hydroxyquinoline [77], Lumogallion [78], and thioglycolic acid [79], Nb has also been determined after extraction of its complex with 3-hydroxyflavone [80] and N-a-phenylstyrylacrylhydroxamic acids [81]. 

Many organic spectrophotometric reagents for Pd incorporate sulphur as a ligand atom. Apart from dithizone and thio-Michler s ketone, which have been discussed above, examples include the thiourea derivatives [33-36] p-dimethylaminobenzylidene-rhodanine (rhodanine) (e = 4.9 10 ) [37] and its derivatives [38], thiodibenzoylmethane [39,40], and thiosemicarbazone derivatives [41—47]. 

Certain nitroso compounds are sensitive spectrophotometric reagents for Pd. Of particular value is p-nitrosodimethylaniline (formula 38.1), which reacts with palladium in the cold at pH 2-2.5 to form a red complex (e = 8.6-10 at 535 nm). The corresponding platinum complex is not formed unless the solution is heated. Further reagents of this group are /r-nitrosodiphenylamine [13], nitrosodibenzylaniline [51], and 2-nitroso-5-diethyl-aminophenol [52]. 

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