DR.

APJ ABDUL KALAM TECHNICAL

UNIVERSITY

Pharmacy
Pharmacognosy & Phytochemistry-II
Lecture – 4

Radioactive Tracers in Biogenetic

Studies
 Radiotracer Techniques
• Radiotracer technique involves the use of labelled compounds to
trace out various precursors and intermediates involved at
different stages of the biogenetic pathways of plants.
• A radioisotope is incorporated into the presumed precursor of
the plant constituent. This labelled compound is introduced into
plant and can be used as marker in biogenetic studies.
 RADIOACTIVE ISOTOPES
• Radioactive properties of a radioisotope remain stable during
the biogenetic studies.
• Isotopes are defined as two or more forms of same elements
which contain equal number of protons but differ in number
of neutrons and thus have different atomic masses but same
• chemicalcompounds
Labelled properties.can be prepared using
either radioactive or stable isotopes.
 RADIOACTIVE ISOTOPES
• In biogenetic investigations, radioactive Carbon and Hydrogen
12
are used, e.g Carbon - C , 13C , 14C Hydrogen - 1
H , 2H , 3H
(Hydrogen, Deuterium and Tritium).
• For specific metabolic studies, Sulphur, Phosphorus, alkali and
alkaline earth metals are used.
• For specific study on proteins, amino
acids and alkaloids labelled nitrogen
gives more specific information.
 RADIOACTIVE ISOTOPES
• Stable isotopes can be detected using Mass spectrometery (14N,
15
N), NMR spectrometry (13C, 2 H).
• Stable isotopes do not involve radiations whereas radioactive
isotopes decay after emitting radiation.
 Steps involved in Tracer
Technique
Selection of suitable radioisotope tracer.
• A tracer can either be a radioactive isotope or a stable isotope
depending on the system under study. Stable isotopes are used
in several chemical and biochemical systems.
• The initial concentration of the isotope should be able to
withstand the dilutions during the whole process.
• Physical and chemical properties of
the tracer should be compatible with
the system under study.
 Steps Involved in Tracer
• Half life of the tracerTechnique
should be long enough till the completion
of investigation, e.g Tritium has a half life of 12 years.
Labelling of the Precursor or interrmediate
1. Growing Chlorella in an atmosphere of 14CO2.
When the green algae, Chlorella is grown in an atmosphere
rich in CO2,all the carbon compounds of this organism become
labelled.
Physical and chemical properties of the tracer
should be compatible with the system under study.
 Steps Involved in Tracer
Technique
2. Tritium labelling can be done by
i) catalytic hydrogenation in aqueous media using platinum
as catalyst
ii) by exposing organic compounds to Tritium gas,
iii) hydrogenation of unsaturated compounds of unsaturated
compounds can also be done with Tritium gas.
3. Nuclear reactorsofcan
Bombardment be usedover
neutrons to prepare
the radioactive isotopes.
atom causes its fission, as a result it
gets radioactive labelled.
 Steps involved in Tracer
Technique
4. By Organic synthesis
CH 3MgBr + 14CO2 → CH3 14COOMgBr + H2O
↓
CH314COOH + Mg(OH)Br
(radio-labelled acetic acid)
 Steps involved in Tracer
Technique
Introduction of Radioactive Labelled Compound into the
plant
A radioactive tracer can be introduced into the plant body by
several ways as under-
• Direct injection : This method can be used for plants bearing
capsular fruits with hollow stems. This method is not suitable
for rigid tissues.
 Steps involved in Tracer
• Floating method : Technique
Chopped pieces of leaves are allowed to
float in a radioactive labelled reagent.
• Root and Stem feeding : This is the most common method used.
Selection of plant part is determined according to the site of
biosynthesis of the concerned metabolite, e.g Nicotine is
mainly synthesised in the roots, so is the main site of
application of radioactive substance.
 Steps involved in Tracer
Technique
For this, the radioatracer reagent may be injected into the soil
at a specific depth and lateral width around the root of the
plant.
Radiotracer substance may even be employed to plants grown
in hydroponic solution(mineral nutrient solution).
• Spraying method : Radioactive reagent which are easily
absorbed from plant surfaces like leaves can be used as
spraying agents.
 Steps involved in Tracer
• Technique
Wick Method : Long cotton strands are passed through the
stem of the plant and the terminal end of the strands are
dipped in radioisotope labelled reagent. The radioactive
material travels throughout the plant by way of stem.
 Steps involved in Tracer
Technique
Isolation and detection of radioactive labelled Compound
Extraction/Isolation of a particular metabolite depends on the
chemical nature and source of the crude drug:-
e.g for alkaloids, glycosides, flavonoids – slightly polar solvents
for phenols _ polar solvents
for unorganised drugs _ maceration
for organised drugs with soft tissues _ infusion , maceration
for organised drugs with hard tissues _ hot percolation,
decoction
THANK YOU