FAR 458 : Nuclear Pharmacy

Radiolabelling
Compound
Mohd Ikhwan Hashim RPh
Nuclear Pharmacy Unit, AMDI, USM
mohdikhwan@usm.my
Presentation Outline
Methods of Radiolabeling

Factors Affecting Radiolabeling

Radiolabeled Compound Tc-99m

 What Is Radiolabeling??????
Tagging substance with
radiotracer / radioisotope
METHODS OF RADIOLABELING
Radiolabel compounds
 Substance where atoms or group of atoms in
the molecule are substituted with similar or
dissimilar but radioactive atoms or groups of
atoms.
 A radioactive cation is chelated with a carrier
molecule.
 Labeling can be carried out by using
radioisotopes of the same element or of a
different element.
LABELING TECHNIQUES
Isotope
Exchange
Reactions
Introduction
Excitation
of a Foreign
Labeling
Label
RADIOLABELED
COMPOUND
Labeling with
Recoil
Bifunctional
Labeling
Chelates
Biosynthesis
or Chemical
Synthesis
LABELING TECHNIQUES
Isotope Exchange Reactions

Here, one or more atoms in a molecule are replaced by isotopes of the same
element having different mass numbers.
Since the radiolabled and the parent molecules are the same except for the
isotope effect, they are expected to have similar chemical and biological
effects. Example:
T3 labelled with 125I
T4 labeled with 125I
The labeling reactions are reversible

125I
 LABELING TECHNIQUES
Introduction of a Foreign Label
 A radionuclide is incorporated into a molecule with known
biologicals effects principally via the formation of covalent or
covalent coordinate bonds.
 The radionuclide used to tag the molecule is foreign to the
molecule and not by isotope exchange.
 Majority of radiopharmaceutical
 Examples:
99mTc labeled albumin
99mTc DTPA
51Cr labeled RBCs
Iodinated proteins, enzymes
LABELING TECHNIQUES
Labeling with Bifunctional Chelates
Proteins form complexes with bifunctional chelates like EDTA, DTPA and
desferoxamine
The complex is then labeled by chelation with an appropriate radionuclide
Because of the presence of the chelate, the biological properties of the
molecule may be altered and need to the assessed before use
Examples:
111In labeled DTPA-albumin
67Ga labeled desferoxamine-albumin
99mTc labeled DTPA-antibody
LABELING TECHNIQUES
Biosynthesis or Chemical Synthesis
 Biosynthesis: A living organism is grown in an environment or
culture media which contains a radionuclide. The radionuclide is
incorporated into metabolites produced by the biological
process of the organism and the metabolites are then separated
and purified. Example: Vitamin B12 labeled by 60Co or 57Co by
adding the respective radionuclide into the culture medium of
Streptomyces griseus.

 Chemical Synthesis: Complex molecules are synthesized from

simple radiolabel molecules. Often multi-step and the final
product will possess physico-chemical properties different from
the non-labeled compounds.
 LABELING TECHNIQUES
Recoil Labeling
In a nuclear reaction, when particles are emitted from the
nucleus, recoil atoms or ions are produced that can form a bond
with molecules in the target material.
This method is of limited interest as it not used on a large scale.
Often, the high energy of the recoil ions results in poor yields and
low specific activity.
Example: formation of tritiated compound in the reactor by 6Li(n,
α)3H reaction. the target compound is mixed with a lithium salt
and irradiated in the reactor. Tritium produced will then label the
target compound.
 LABELING TECHNIQUES
Excitation Labeling
Involved the use of radioactive daughter ions which are
reactive and produces from a nuclear decay process.
During the beta-decay or electron capture process, high
energy charged ions are produced which are capable of
labeling target compounds.
Example: 77Kr decays to 77Br. If the target compound is
exposed to 77Kr, then energetic 77Brions will label the
compound to form a brominated product.
 What are the factors to be taken into
consideration when labeling a compound
Efficiency of the A high yield is always desirable, although not always achieved.
labeling process A lower yield is sometimes acceptable if the product is pure
and not damaged by the labeling method, the expense is low
and there are better ways to perform the labeling

Chemical stability This refers to the stability of the bond between the
of the product radionuclide and the labeled molecule.
Compounds labeled by covalent bonds are more stable under
physiological conditions.

Denaturation or The structure and or biological properties of the labeled

alteration compound may be altered during labeling
For example, proteins may be altered by heating, extremes of
pH and excessive iodination.
Red blood cells are denatured by heating.
 What are the factors to be taken into
consideration when labeling a compound
Isotope effect The labeling of compounds with radioisotopes may give rise to
radiolabels compounds having different physical (and perhaps
biological) properties due the difference in isotope weights.
For example., the property of tritiated water is different from
normal water under physiological conditions.
The isotope effect is not serious except when the isotopes are
heavier.

Carrier free or no Radiopharmaceutical tend to be adsorbed onto glassware if

carrier added they are in a carrier free or no carrier added state (NCA).
state The molar concentration of carrier free compounds are in the
nanomolar range and is thus difficult to handle.
Storage Many radiolabeled compounds, especially biological are
conditions susceptible to heat.
Proteins and labeled dyes are degraded by heat.
Light may also break down labeled compounds and carrier-free
tracers should be stored in silicone-coated vials to reduce losses.
 What are the factors to be taken into
consideration when labeling a compound
Radiolysis Labeled compounds may be decomposed by the radiations emitted by the
radionuclides present in them
The higher the specific activity, the greater the radiolysis effect.
When a chemical bond breaks down by radiation from itself, this is called
autoradiolysis.
Radiation may also break down the solvent, producing free radicals that
breakdown the bonds of the labeled compounds. This is indirect radiolysis.
To prevent indirect radiolysis, the pH of the solvent should be neutral as
more reactions of this nature occur at extremes of pH.
The longer the half-life of the radionuclide and the more energetic the
radiations, the more breakdown is observed.
Thus, radiolysis introduces radiochemical impurities into the preparation.
These factors are taken into consideration when setting the expiration date
of a radiopharmaceutical.

Specific activity Defined as the activity per gram of the labeled material
High specific activity in most cases is desirable
However sometimes high specific activity may cause radiolysis
 What are the factors to be taken into
consideration when labeling a compound
Purification Radioactive contaminant arise when producing radionuclides
and analysis Nuclear fission produces many impurities due to the numerous
modes of fission occurring
Target impurities also present a problem
Impurities also arise from incomplete labeling and from radiolysis.
The removal of contaminants can be done using various chemical
separation methods like solvent extraction, ion exchange etc.

Shelf life The shelf life of a labeled compound is the time period which the
preparation can be safely used.
Usually a period of six months is suggested or three t ½ .
99mTc preparation half shelf lives of 0.5 to 18 hours.

The most commonly used radionuclide is 99mTc and 131I, the former
accounting for about 80% and the latter about 15% of all nuclear
medicines.
FACTORS AFFECTING LABELING
Efficiency of the labeling process
Chemical stability of the product
Denaturation or alteration
Isotope effect
Factors which Carrier free or no carrier added state
affect labeling
Storage conditions
Specific activity
Radiolysis
Purification and analysis
Shelf life
 RADIOLABELING COMPOUND
TECHNETIUM-99m
 Its favorable physical and radiation characteristics
 The 6hr t½ and the little amount of electron emission permit the administration of
milicurie amounts of 99mTc without significant radiation dose.
 The 140keV photons monochromatic photons give good images and resolution
 Technetium is readily available in a sterile, pyrogen free and carrier free state from
99Mo- 99mTc generators.

 The chemical form of 99mTc available form the molybdenum generator is the
pertechnate (99mTcO4-) ion with a +7 oxidation state. This species is relatively non-
reactive. Before labeling is possible, a reduction in the oxidation state must occur.
Commonly this is done using stannous chloride under acidic condition, thus:
 3Sn2+  3Sn4- + 6e-
 2 99mTcO4- + 16H+ + 6e-  99mTc4- +8H2O
RADIOLABELING COMPOUND
TECHNETIUM-99m
 Technetium – group VIIB transition element which has seven electrons
beyond the noble gas electronic configuration.
 Members of the group: Manganese (Mn), Technetium (Tc) and Rhenium(Re)
 Elements easily lose the seven electron to yield the +7 oxidation state of
permetallate anions (MnO4-,TcO4-, ReO4-)
 Technetium exhibits multiple oxidation states from -1 to +7 and variable
coordination members and geometries.
 In aqueous solution, the most stable oxidation states: +7(TcO4-) and
+4(TcO2)
 The other oxidation states are stabilized by different ligands.
 Disproportionation reaction:
 3Tc(VI)  Tc(IV) + 2 Tc(VII)
 3Tc(V)  2Tc(IV) + Tc(VII)
RADIOLABELING COMPOUND
TECHNETIUM-99m
CHEMICAL CHARACTERISTIC
 Its variable oxidation states and
coordination numbers have led to the
preparation of wide variety of 99mTc
radiopharmaceuticals.
 Its ease of formation of complexes and
chelates makes its suitable for the ‘in-
hospital’ preparation of many 99mTc
radiopharmaceuticals.
 Its separation from its radionuclide
(99mMo) is accomplished in an enclosed
and sterile generator system.
 Its generator eluate (99mTcO4-) does not
need further processing or purification
and can either be directly administered
or used for the preparation of other
99mTc radiopharmaceuticals.
PHYSICAL CHARACTERISTIC
It monoenergetic gamma ray of 140keV is
easily collimated and yields a sufficient
number of photons.
Its half-life of 6.0h is long enough to
obtain the desired diagnostic information
in many studies and is short enough to
cause minimal radiation exposure to the
patient and personnel.
Its lack of beta emissions lowers the
radiation dose to both patient and
personnel.
 RADIOLABELING COMPOUND
TECHNETIUM-99m
 Technetium-99m Pertechnetate
(99mTcO4-)
 Technetium-99m d,l-
hexamethylpropyleneamine oxime
(99m-d,l-HMPAO)
 Technetium-99m l,l-ethylcysteine
dimer (99mTc-l,l-ECD)
 Technetium-99m macroaggregated
albumin (99mTc-MAA)
 Technetium-99m human albumin
microspheres (99mTc-HAMs)
 Technetium-99m red blood cells
(99mTc-RBCs)
 Technetium-99m heat-denatured
red blood cells (99mTc-HD-RBCs)
 Technetium-99m hexakis-2-
methoxy-2-isobutyl isonitrile (99mTc-
Sestamibi)
 Technetium-99m phosphorous
compound
 Technetium-99m colloids and
colloid solid material
 Technetium-99m iminodiacetate
derivatives (99mTc-IDAs)
 Technetium-99m
diethylenetriaminepentaacetic acid
(99mTc-DTPA)
 Technetium-99m
dimercaptosuccinic acid (99mTc-
DMSA)
 Technetium-99m human serum
albumin (99mTc-HAS)
RADIOLABELING COMPOUND
TECHNETIUM-99m - MECHANISM OF LABELING
 The reduced 99mTc species are chemically reactive and combine
with a wide variety of chelating agents, thus:
 Reduced 99mTc + chelating agent  99mTc-chelate
 The chelating agent donates lone pair of electrons to form
coordinate covalent bonds with 99mTc. Normally the chelating
agent is added in excess to prevent the hydrolysis of free
reduced 99mTc and stannous ion in aqueous media.
 Labeling with 99mTc has been made more convenient with the
introduction of commercial kits. 99mTc labeling is accomplished
by adding 99mTcO4- to the kits. The kits are prepared from a
master solution consisting of the compound to be labeled mixed
with acidic stannous compound in appropriate proportions. The
pH of the solution is adjusted to 5 – 7 with NaOH and aliquots
are dispensed into individual vials for use or lyophilized.
Technetium-99m Pertechnetate
99m
( TcO4 )-

 99mTcO4- is most often eluted from 99mMo-99mTc column

type generator with 0.9% sodium chloride
solution(saline).
 The volume of saline used for elution depends upon
the activity size of the generator and/or the
concentration of 99mTcO4-required for subsequent use.
 The first eluate after a few days ‘standing’ of the
generator should be discarded because it may contain
excess 99mTcO4-
 QC: Radionuclidic test, Radiochemical test, Chemical
test.
Labelling with Tc99m Process
◦ Tagging
◦ Pharmaceutical agent/drugs being prepared in colour coded vial shields
◦ Withdraw tc-99m from the elution vial with the right activity
◦ Top up tc-99m in the syringe with NACL to desired volume
◦ Syringe in asepticly the tc-99m into the pharmaceutical vial. Maintain negative pressure
inside the vial and carefully withdraw the empty syringe
◦ Shake gently the colour coded vial for 45 to 60 second
◦ Leave it still for at least 10 minutes before performing quality control.
 mohdikhwan@usm.my

Mohd Ikhwan Hashim mohdikhwanhashim

PRESENTED BY
M O H D I K H WA N B I N H A S H I M R P H