ame: NOBLE, KYRA EIRIS M. Date performed: Jan 16, 2018
BS CHEMISTRY III Group No. 01
Chemistry 125
Biochemistry
Experiment No. 7
COLOR REACTION OF
... [Show More] PROTEINS
Introduction
Proteins are highly complex substance that is present in all living organisms. Amino acid
is the building block of proteins and they are linked in a series of peptide bonds to form the
primary structure of proteins. Proteins react with different reagents and forms colored products.
These test are important in qualitative detection and quantitative estimation of proteins and of
their constituent amino acids in body fluids and other biological materials.
Objectives:
1. To identify specific amino acids based on their chemical group present.
2. To identify which sample is complete or incomplete protein.
Material & Reagents:
10% NaOH Conc. H2SO4 Conc. NaOH 2% gelatin
1% CuSO4 Hopkin’s- Cole reagent 1% lead acetate 2 marbles
0.1% ninhydrin sol’n Sulfanilic acid Bromine water
Conc. HNO3 0.5% NaNO2 n-amyl alcohol
Millon’s reagent 10% Na2CO3 Blue & red litmus paper
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Data & Results
Sample
Test
Albumin Gelatin Inference
Biuret Cloudy white to
light violet
solution
Green gelatin to
blue green colored
solution
Tripeptide protein is present in
albumin and absent in gelatin.
Ninhydrin Cloudy white to
dark blue violet
Green solution (no
changes)
Small peptides and proteins are
present in albumin but not in
gelatin.
Xanthoproteic
a. With HNO3
Yellow
precipitate
Clear yellow
solution
Tyrosine and phenylalanine,
and tryptophan are present in
albumin and gelatin.
b. With
NH4OH
Yellow orange
precipitae
No changes
Millon’s ------- ------- ----------------------------
Hopkin’s- Cole Cloudy white
ring formed
Yellow-green
colored ring formed
Tryptophan is not present.
Lead acetate Cloudy white to
blackish brown
color
Green color to
yellow color
solution
Cysteine is present in albumin.
Pauly Orange solution Light green
solution
Tyrosine and histidine are
present in albumin.
Bromine water ---------- --------- -------------------
Sakaguchi ---------- ---------- ------------------
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Questions:
1. Write the positive result, chemical group responsible for the positive and the importance
of each test.
Test Positive result Chemical group Importance
Biuret Violet solution Peptide linkage Detects presence of
tripeptides and up
Ninhydrin Lavender solution Free amino and free
COOH
Detects presence of
a.a peptides and
amino acids
Xanthoproteic Lemon yellow Benzene ring Detects presence of
tyrosine,
phenylalanine, and
tryptophan
Millon’s White ppt to brick
red
Phenolic amino acids Detects presence of
tyrosine
Hopkin’s-Cole Violet ring Indole group Detects presence of
tryptophan
Lead acetate Red orange to lighter
orange
Sulfide containing
amino acids
Detects presence of
histidine and tyrosine
Pauly Orange solution Imidazole group Detects presence of
tyrosine and histidine
Bromine water Pink solution Indole group Detects presence of
tryptophan
Sakaguchi Intense red color Guanidine Detects presence of
arginine
2. What color will be produced by proline with ninhydrine test? Why?
Proline will give yellow color solution because the secondary amino group (imino group) of
proline residues is held in unbending conformation that reduces the structural flexibility of
polypeptide regions containing proline. Proline does not give ninhydrin reaction as this
reagent requires free alpha amino group but proline have imino group.
3. Which test can be used to show the extent of hydrolysis of proteins? Why?
Biuret test can show the extent of hydrolysis of proteins due to the coordination complex
with the peptide bond of the protein from the CuSO4.
4. Classify proteins according to nutritional value. To which classification does albumin and
gelatin belong?
Proteins nutritional values can be classified as complete and incomplete. For complete, a
protein that consists of all essential amino acids while incomplete is a protein that is
insufficient of one or more essential amino acid. Albumin is classified as a complete protein
while gelatin is classified as incomplete protein.
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