Proteins, Peptides and Amino Acids: Lab Experiments

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Test
Biuret Test

Reagents
CuSO4 sol'n, diluted
NaOH

Test for
Intact proteins and
protein hydrlysates (at
least tripeptide, 2
peptide bonds

Result
Pink to violet blue
color

Ninhyrin Test

1,2,3-indanetrione
monohydrate or
triketohydintene
hydrate, ethanol

Alpha- amino group
(usually a general test
for amino acids)

Xanthoproteic
Test

Conc. HNO3, conc.
NaOH

For W,F,Y (aromatic
except for H)

Blue to blue-violet
Oxidative decarboxylation
color
& deamination followed by
(proline:hydroxypr condensation
oline gives a yellow
color)
Yellow sol'n/ppt.
Nitration via SEAr
with conc. HNO3,
orange with excess
NaOH

Millon's rgt.(Hg2+ &
Hg22+nitrates&
nitriles/Hg(NO3)2 in
conc. HNO3, w/ trace
of
HNO2/HgSO4,H2SO4
,NaNO2)
Hopkins-Cole Test Glyoxylic acid (Mg
powder, oxalic acid,
HOAc), conc. H2SO4
Millon's Test

Sakaguchi Test

Fohl's Test

Nitroprusside Test
Test for Amide
Pauly's Test

Alpha-naphthol,
NaOBr, NaOH,
urea(to stabilize
color & destroy
excess OBr ions)

Pb(CH3COO)2
NaOH

Na2Fe(CN)3 in dil.
NH3
NaOH
Diazo rgt. (1%
sulfosalicylic acid
with 5% NaNO2),
10% Na2CO3

Reaction/Principle
Complexation
Form. Of coordination
complex Cu+2 and 4 N
atoms (2 from each 2
polypeptide chain)

Phenolic group of Y

Old rose/flesh to
red ppt.

Complexation(mercuration
& nitration or
nitrosation/complexation of
nitrohydroxyphenyl
derivatives with Hg2+)

Indole group in W

Pink to violet
interface

Reduction of oxalic acid to
glyoxolic acid and acidcatalyzed condensation of
two tryptphans w/glyoxolic
acid

Un-or monosubstituted
guanidines in R

,

Red to red-orange Complexation (basecolor
catalyzed condensation of
alpaha-naphthol w/ the
guanidine group of R)

Sulfur-containing
C-C)

Brown to black ppt.

Degradation & substitution
rxn to form PbS

For cys

Red coloration

Complexation rxn

10,20,30 amides and
nitriles ; peptide bonds
Detects tyr and his

Red litmus to blue

Basic hydrolysis

(C,

Red coloration

Formation of azo dyes