Biochem lab (midterm)
Proteins
- Proteins consist of amino acids which are characterized by the- CH(NH2) COOH
substructure. It is also a classification of peptides consisting of 50 or more amino acids.
- Amino acids are the combination of the amino group (nitrogen + 2 hydrogen = -NH2) and
the acid entity is the carboxylic group (-COOH). It is the basic building blocks of
enzymes, hormones, proteins and body tissues.
- Peptide bond -C(=O)NH- is the link between amino acids when the carboxyl group of
one molecule reacts with the amino group of another molecule. Hence, releasing water
molecules (H2O).
- Peptide is a compound consisting of 2 or more amino acids.
- Oligopeptides have 10 or fewer amino acids.
- Polypeptides are chains of 10 or more amino acids.
● Biuret Test is used for the quantitative photometric determination of total protein
concentration. The intensity of the color produced in this test is proportional to the
number of peptide bonds participating in the reaction.
➢ Positive result: Peptides and proteins (long chain polypeptides) react with Cu2+ in
alkalinity to create a positive result, blue violet colored complex.
➢ Amino acids and copper (II) ions form a blue complex.
● Xanthoproteic Test is when the nitro derivatives (nitrated aromatic rings of amino acids
like tyrosine and tryptophan with the use of 65% nitric acid) show an intensely yellow
color. It is because nearly all proteins contain aromatics, taken as a protein-test either.
● Millon’s Test (Millon’s reagent) an analytical reagent used to detect soluble proteins. It
consists of mercury dissolved in nitric acid. Millon’s test is not specific for proteins, it
actually detects phenolic groups which are present in amino acid tyrosine.
➢ Positive result: a reddish-brown coloration or precipitate indicates the presence of
tyrosine residue which occurs in nearly all proteins.
● Sulfur Test is a process where sulfur-containing amino acids are boiled with an alkali, the
sulfhydryl or disulfide groups are converted to an inorganic sulfide, Na2S. This reacts
with lead acetate to form a black precipitate of PbS. Cysteine and cystine in the free
state and in proteins give a positive result to the sulfur test.
● Hopkins-Cole Test is where the indole ring of tryptophan condenses with glyoxylic acid in
the presence of sulfuric acid to form a violet-colored complex.
● Ninhydrin Test is a test for amino acids and proteins with a free -NH2 group. When such
an amino group reacts with ninhydrin, a purple-blue complex is formed.
Proteins
- Proteins consist of amino acids which are characterized by the- CH(NH2) COOH
substructure. It is also a classification of peptides consisting of 50 or more amino acids.
- Amino acids are the combination of the amino group (nitrogen + 2 hydrogen = -NH2) and
the acid entity is the carboxylic group (-COOH). It is the basic building blocks of
enzymes, hormones, proteins and body tissues.
- Peptide bond -C(=O)NH- is the link between amino acids when the carboxyl group of
one molecule reacts with the amino group of another molecule. Hence, releasing water
molecules (H2O).
- Peptide is a compound consisting of 2 or more amino acids.
- Oligopeptides have 10 or fewer amino acids.
- Polypeptides are chains of 10 or more amino acids.
● Biuret Test is used for the quantitative photometric determination of total protein
concentration. The intensity of the color produced in this test is proportional to the
number of peptide bonds participating in the reaction.
➢ Positive result: Peptides and proteins (long chain polypeptides) react with Cu2+ in
alkalinity to create a positive result, blue violet colored complex.
➢ Amino acids and copper (II) ions form a blue complex.
● Xanthoproteic Test is when the nitro derivatives (nitrated aromatic rings of amino acids
like tyrosine and tryptophan with the use of 65% nitric acid) show an intensely yellow
color. It is because nearly all proteins contain aromatics, taken as a protein-test either.
● Millon’s Test (Millon’s reagent) an analytical reagent used to detect soluble proteins. It
consists of mercury dissolved in nitric acid. Millon’s test is not specific for proteins, it
actually detects phenolic groups which are present in amino acid tyrosine.
➢ Positive result: a reddish-brown coloration or precipitate indicates the presence of
tyrosine residue which occurs in nearly all proteins.
● Sulfur Test is a process where sulfur-containing amino acids are boiled with an alkali, the
sulfhydryl or disulfide groups are converted to an inorganic sulfide, Na2S. This reacts
with lead acetate to form a black precipitate of PbS. Cysteine and cystine in the free
state and in proteins give a positive result to the sulfur test.
● Hopkins-Cole Test is where the indole ring of tryptophan condenses with glyoxylic acid in
the presence of sulfuric acid to form a violet-colored complex.
● Ninhydrin Test is a test for amino acids and proteins with a free -NH2 group. When such
an amino group reacts with ninhydrin, a purple-blue complex is formed.
