CHEMISTRY PROJECT

FILE
2014-2015

CHIRAG KUMAR
Roll no:

CERTIFICATE
This is to certify that Chirag Kumar of class XII-A of
DAV PUBLIC SHOOL, Vasant kunj carried out this
project under the guidance of Mrs.Swastika. The
work on this project is his original work.

SIGNATURE:
(Mrs.Swastika)

ACKNOWLEDGEMENT
I would like to thank our chemistry teacher
Mrs.Swastika & lab assistant Mr.Kamlesh for their
constant support and encouragement throughout
this project.This project couldnt have been
possible without their support.

BLOOD STAIN DETECTION

BY VARIOUS CHEMICAL
METHODS.

INDEX
Materials required
Introduction
Procedure & Results
Bibliography

MATERIALS
REQUIRED
Apparatus:
1) Microscope
2) Cover slip
3) Test tubes
4) Beakers
5) Glass rod
6) Water

Chemicals:
1) Blood
2) Alkaline solution of methylamine
3) Phenolphthalein
4) Zinc powder
5) Sodium hydroxide

Introduction
Blood is a specialized bodily fluid in animals that delivers necessary
substances such as nutrients and oxygen to the cells and transport metabolic
waste products away from same cells.
Blood consists of cellular material (99% red blood cells, with white blood cells
And platelets making up the remainder), water, amino acids, proteins,
Carbohydrates, lipids, hormones, vitamins, electrolytes, dissolved gases, and
Cellular wastes.
The most abundant cells in vertebrate blood are red blood cells. These contain
Haemoglobin, an iron-containing protein, which facilitates transportation
Of oxygen by reversibly binding to this respiratory and greatly its solubility in blood.
In contrast,

is almost entirely transported extracellularly dissolved in plasma as

biacarbonate ion.

The presence or absence of blood stains often provides important information for
those investigating criminal cases and forensic scientists. The detection ofblood is
usually based on one of classes of methods.

CRYSTAL TESTS
Haem forms crystals when reacted with certain reagents. The most common such
reagent is pyridine, which forms pink crystals.

CATALYTIC TESTS
These tests rely on the fact that haem can catalyse the breakdown of hydrogen
peroxide. As the

breaks down, another substance in the reaction mixture is

oxidised, producing a colour change.

INSTRUMENTAL METHODS
Chromatography can be used to identify the presence of haemoglobin.
All of the methods are in some way dependent on the presence of haemoglobin, and
will therefore give positive results for both human and animal blood. The discussions
is confined to chemical methods and therefore and does not consider biological
methods such as antigen- antibody reactions. The biological methods are generally
slower than chemical methods but more specific.
These tests are used practically for several different purposes. These include both
the confirmation of the nature of visible stains, the detection of non-visible and the
enhancement of hard to see stains.

PROCEDURE
CRYSTAL TEST
The crystal tests, are all based on the formation of haemoglobin derivative crystals
such as haematin, haemin and haemochromogen. The test is carried out on a
microscopic slide, with an alkaline solution of methylamine as reagent being added
to the stain under the cover slip, and crystals formation observed microscopically. If
blood is present, pink crystals of a complex between methylamine and haem form as
the slide is warmed.

As well as methylamine, a large number of other nitrogenous bases, including

nicotine, pyridine, histidine, and glycine can be used in variations of this test.

The best known of the crystal test is that developed by TAKA YAMA about 80
years ago using an alkaline solution of pyridine as reagent.

(The complex formed in Taka Yama test)

The sensitivity is about 0.001 mL of blood or 0.1 mg of haemoglobin. Blood stains

up to 20 years old have given positive results in crystal tests.

CATALYTIC TESTS
These methods depend on the fact that the haem group of haemoglobin possesses
a peroxidase-like activity which catalyses the breakdown of hydrogen peroxide. The
oxidising species formed in this reaction can then react with a variety of substrates to
produce a visible colour change.
Among substrates in common use are benzidine and various substituted benzidines,
ortho-tolidine, leucomalachite green, leucocrystal violet and phenolphthalein.

The Kastle-Meyer test

It is a presumptive blood test, first described in 1903, in which the chemical
indicator phenolphthalein is used to detect the possible presence of haemoglobin. It
relies on the peroxidase-like activity of haemoglobin in blood to catalyse the
oxidation of phenolphthalein (the colourless reduced form of phenolphthalein) into
phenolphthalein, which is visible as a bright pink colour. The KastleMeyer test is a
form of catalytic blood test.
In the kastle-Meyer test the reduced phenolphthalein is kept in alkaline solution in
the presence of zinc. This solution is colourless.
(To prepare reduced phenolphthalein: In a test tube, dissolve 0.1 g of
phenolphthalein in 10.0 mL of 25% sodium hydroxide solution. Add 0.1g mossy zinc
to the tube. The solution should be bright pink. Add a boiling chip and gently boil the
solution until it changes colour to become colourless or pale yellow.)

Oxidation with haemoglobin and peroxide causes an instant colour change to the
well- known bright pink.

(Oxidation of reduced phenolphthalein by haemoglobin and peroxide)

(Test results)
The catalytic test are extremely accurate. The general principle is that if the test is
negative, blood is absent, but that if the test is positive, blood is probably present.
For this reason the test are often described as presumptive tests.

INSTRUMENTAL METHOD
High performance liquid chromatography (HPLC) can be used to confirm the identity
of blood using the absorbance of haemoglobin for detection. This method can also
be used to identify the species of origin from variations in the globin chain, to
distinguish foetal haemoglobin from adult haemoglobin, and to give an estimate of
the age of bloodstain.

RESULT

The crystal and catalytic test confirm

the presence of haemoglobin in the
stain, and hence confirm the presence
of blood.

BIBLIOGRAPHY
http://nzic.org.nz/chemprocesses/biotech/12A.pdf

http://en.wikipedia.org

http://chemistry.about.com

http://www.bluestar-forensic.com

THE END