The document describes a practical experiment using the ninhydrin screening test to detect amino acids. Ninhydrin reacts with amino acids in a pH range of 4-8, producing a purple colored product. The student conducted the test on solutions of arginine, tryptophan, tyrosine, cysteine, glycine, and glutamic acid. All solutions turned purple upon mixing with ninhydrin and heating for two minutes, except the distilled water control. The color change occurs because ninhydrin breaks down amino acids into aldehydes, ammonia, and carbon dioxide, then condenses with the breakdown products to form an intensely colored pigment.

1. KENYATTA UNIVERSITY
SCHOOL OF MEDICINE.
DEPARTMENT OF MEDICAL
BIOCHEMESTRY
NAME: LANDO ELVIS OTIENO
REG NO: P29S/16344/2015.
COURSE: MBCHB
LECTURER: Dr OKUN
HANDING DATE: 8 /10/2015
SUBJECT: PRACTICAL REPPORT ON NINHYDRIN
SCREENING TEST FOR AMINO ACIDS.
HANDING DATE: /10/2015

2. 2.NINHYDRIN SCREENING TEST FOR AMINO ACIDS.
Introduction:
-Ninhydrin (2,2-Dihydroxyindane-1,3-dione) is a chemical used to
detect ammonia or primary and secondary amines.
-amino acids are building blocks of all proteins, and are linked in series
by peptide bond(CONH) to form the primary structure of a proteins.
Amino acids possess an amine group, carboxylic acid group and a
varying side chain that differs between different amino acids.
-There are 20 different naturally occurring amino acids, which vary from
one another with respect to their side chains. Their melting point are
extremely high(usually exceeding 200°c), and at their PI, they exist as
zwitterions, rather than unionized molecules.
-Amino acids responds to all typical chemical reactions associated with
compounds that contain carboxylic acid and amino groups, usually
under conditions where zwitterions form is present in only small
quantities. All amino acids(except glycine)exhibit optical activity due to
the presence of an asymmetric § -carbon atom. Amino acids with an L-
configuration are present in all naturally occurring proteins, whereas
those with D-forms are found in antibiotics and in bacterial cell walls.
-PRINCIPLE:
- In the pH range of 4 – 8, all amino acids react with ninhydrine(
triketohydrine), a powerful oxidizing agent to give a purple colored

3. product (diketohydrin) termed Rhuemann’s purple.All primary amines
and ammonia reacts similarly but without liberation of carbon dioxide.
The amino acid proline and hydroxyproline also react with ninhydrine.
But give a yellow colored complex instead of a purple one. Beside
amino acids, other complex structures such as peptides,pepetones, and
proteins also reacts positively when subjected to ninhydrin reaction.
-REAGENTS AND MATERIALS:
-Solution prepared; 1% Arginine, tryptophan, tyrosine,cystein,glycine,
glutamic acid and distilled water.
-Reagent; Ninhydrine(in2% W/V) in acetone.
-METHODOLOGY:
-set up a number of test tubes and into each put 2.0ml of each solution
of amino acids
-Into each tube add 2drops of ninhydrin reagent.
-mix the solutions and boil in water bath for 2 minutes.
-Record your observations in each test tube.

4. -OBSERVATIONS AND RESULTS
Reagent color Color when
mixed with
ninhydrine
before
heating
Color after
heating the
solution for
2minutes.
Arginine colorless colorless purple
tryptophan colorless colorless purple
Glycine colorless colorless Deep purple
Glutamic acid colorless colorless purple
Distilled water colorless colorless Colorless
-All the reagents except water turned purple when mixed with
ninhydrine and heated for two minutes.
-After the heated solutions of the above reagents and ninhydrine were
cooled down, the color(purple) they formed did not change.
-Discussion/Analysis:
Ninhydrin degrades amino acids (1% Arginine, tryptophan, tyrosine,
cystein, glycine, glutamic acid) into aldehydes, ammonia,
andCO2 through a series of reactions; the net result is ninhydrin in a
partially reduced form hydrindantin.

5. Ninhydrin then condenses with ammonia and hydrindantin to produce
an intensely blue or purple pigment, sometimes called Ruhemann's
purple.
The color varies slightly from acid to acid, probably because unreacted
acids complex with the pigment.
To differentiate between Carbohydrates (-ve) and Amino Acids &
Proteins (+ve). Ninhydrin Reacts with α-amino acids (–NH2) in proteins
giving a purple colored complex, except Proline and hydroxy proline
gives yellow color(no –NH2).
-Conclusion
-Since all that is required for color development is ammonia and
partially
reduced ninhydrin, the ammonium salts of weak and strong acids, as
well as certain amines, can give a false positive result to the ninhydrin
test.
The striking color change is due to the large change in electron
confinement on formation of the anion.
-Distilled water was used as a control in this experiment that is why
there was no change in color.

6. -references
1.https://en.wikipedia.org/wiki/Ninhydrin
2.http://chemwiki.ucdavis.edu/Biological_Chemistry/Proteins/Proteins
_and_Amino_Acids
3. Kenyatta university-laboratory-practical-manual- for-MBCHB-Year 1-
semester1.
4.http://antoine.frostburg.edu/chem/senese/101/organic/faq/amino-
acid-test.shtml