Test Reagents Test for Result Reaction/PrincipleBiuret Test CuSO4 sol'n, diluted
NaOHIntact proteins and protein hydrlysates (at least tripeptide, 2 peptide bonds
Pink to violet blue color
ComplexationForm. Of coordination complex Cu+2 and 4 N atoms (2 from each 2 polypeptide chain)
Ninhyrin Test 1,2,3-indanetrione monohydrate or triketohydintene hydrate, ethanol
Alpha- amino group (usually a general test for amino acids)
Blue to blue-violet color (proline:hydroxyproline gives a yellow color)
Oxidative decarboxylation & deamination followed by condensation
Xanthoproteic Test
Conc. HNO3, conc. NaOH
For W,F,Y (aromatic except for H)
Yellow sol'n/ppt. with conc. HNO3,
orange with excess NaOH
Nitration via SEAr
Millon's Test Millon's rgt.(Hg2+ & Hg2
2+nitrates& nitriles/Hg(NO3)2 in conc. HNO3, w/ trace of HNO2/HgSO4,H2SO4,NaNO2)
Phenolic group of Y Old rose/flesh to red ppt.
Complexation(mercuration & nitration or nitrosation/complexation of nitrohydroxyphenyl derivatives with Hg2+)
Hopkins-Cole Test Glyoxylic acid (Mg powder, oxalic acid, HOAc), conc. H2SO4
Indole group in W Pink to violet interface
Reduction of oxalic acid to glyoxolic acid and acid-catalyzed condensation of two tryptphans w/glyoxolic acid
Sakaguchi Test Alpha-naphthol, NaOBr, NaOH, urea(to stabilize color & destroy excess OBr ions)
Un-or monosubstituted guanidines in R
Red to red-orange color
Complexation (base-catalyzed condensation of alpaha-naphthol w/ the guanidine group of R)
Fohl's Test Pb(CH3COO)2 , NaOH
Sulfur-containing (C, C-C)
Brown to black ppt. Degradation & substitution rxn to form PbS
Nitroprusside Test Na2Fe(CN)3 in dil. NH3
For cys Red coloration Complexation rxn
Test for Amide NaOH 10,20,30 amides and nitriles ; peptide bonds
Red litmus to blue Basic hydrolysis
Pauly's Test Diazo rgt. (1% sulfosalicylic acid with 5% NaNO2), 10% Na2CO3
Detects tyr and his Red coloration Formation of azo dyes
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