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BIOLOGICAL
(MOLECULAR AND CELLULAR) EFFECTS OF
IONIZING RADIATION
THE EFFECTS OF RADIATION ON THE
CELL AT THE MOLECULAR LEVEL
When radiation interacts with target atoms, energy
is deposited, resulting in ionization or excitation.
The absorption of energy from ionizing radiation
produces damage to molecules by direct and
indirect actions.
For direct action, damage occurs as a result of
ionization of atoms on key molecules in the biologic
system. This causes inactivation or functional
alteration of the molecule.
Indirect action involves the production of reactive
free radiacals whose toxic damage on the key
molecule results in a biologic effect.
DIRECT EFFECTS
Ionization of atoms in molecules is a result
of absorption of energy by photoelectric
and Compton interactions. Ionization
occurs at all radiation qualities but is the
predominant cause of damage in
reactions involving high LET radiations.
Absorption of energy sufficient to remove
an electron can result in bond breaks.
 Ionizing radiation+RH
R - + H+

INDIRECT ACTION
These are effects mediated by free radicals.
 A free radical is an electrically neutral atom
with an unshared electron in the orbital
position. The radical is electrophilic and
highly reactive. Since the predominant
molecule in biological systems is water, it is
usually the intermediary of the radical
formation and propagation.

INDIRECT ACTION- RADIOLYSIS OF
WATER
H-O-H  H OH+ + e (ionization)
H OH+ + e H0+OH0
(free radicals)
Free radicals readily recombine to electronic and
orbital neutrality. However, when many exist, as in
high radiation fluence, orbital neutrality can be
achieved by:
1.Hydrogen radical dimerization (H2)
2.The formation of toxic hydrogen peroxide (H2O2).
3.The radical can also be transferred to an organic
molecule in the cell.
INDIRECT ACTION
H0 + OH0 HOH (recombination)
 H0 + H0  H2 (dimer)
 OH0 + OH0  H2O2 (peroxide dimer)
 OH0 + RH  R0 + HOH (Radical transfer)

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The presence of dissolved oxygen can modify the
reaction by enabling the creation of other free
radical species with greater stability and lifetimes
H0+O2  HO20 (hydroperoxy free radical)
 R0+O2 RO20 (organic peroxy free radical)

INDIRECT ACTION - THE LIFETIMES OF FREE
RADICALS

The lifetimes of simple free radicals (H0 or OH0)
are very short, on the order of 10-10 sec. While
generally highly reactive they do not exist long
enough to migrate from the site of formation to
the cell nucleus. However, the oxygen derived
species such as hydroperoxy free radical does
not readily recombine into neutral forms. These
more stable forms have a lifetime long enough
to migrate to the nucleus where serious
damage can occur.
INDIRECT ACTION- FREE RADICALS
The transfer of the free radical to a biologic
molecule can be sufficiently damaging to
cause bond breakage or inactivation of key
functions
 The organic peroxy free radical can transfer
the radical form molecule to molecule
causing damage at each encounter. Thus a
cumulative effect can occur, greater than a
single ionization or broken bond.

BIOCHEMICAL REACTIONS WITH IONIZING
RADIATION
BIOCHEMICAL REACTIONS WITH IONIZING
RADIATION
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DNA is the most important material making up
the chromosomes and serves as the master
blueprint for the cell. It determines what types
of RNA are produced which, in turn, determine
the types of protein that are produced.
The DNA molecule takes the form of a twisted
ladder or double helix. The sides of the ladder
are strands of alternating sugar and phosphate
groups. Branching off from each sugar group is
one of four nitrogenous bases: cytosine,
thymine, adenine and guanine.
DNA
BIOCHEMICAL REACTIONS WITH IONIZING
RADIATION-DNA DAMAGE
There is considerable evidence
suggesting that DNA is the primary target
for cell damage from ionizing radiation.
 Toxic effects at low to moderate doses
(cell killing, mutagenesis, and malignant
transformation) appear to result from
damage to cellular DNA. Thus, ionizing
radiation is a classical genotoxic agent.

BIOCHEMICAL REACTIONS WITH IONIZING
RADIATION-DNA DAMAGE
The lethal and mutagenic effects of moderate
doses of radiation result primarily from damage
to cellular DNA.
 Although radiation can induce a variety of DNA
lesions including specific base damage, it has
long been assumed that unrejoined DNA
double strand breaks are of primary
importance in its cytotoxic effects in
mammalian cells.

BIOCHEMICAL REACTIONS WITH IONIZING
RADIATION-DNA DAMAGE
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Active enzymatic repair processes exist for the repair
of both DNA base damage and strand breaks. In many
cases breaks in the double-strand DNA can be
repaired by the enzymes, DNA polymerase, and DNA
ligase.
The repair of double strand breaks is a complex
process involving recombinational events, depending
upon the nature of the initial break.
BIOCHEMICAL REACTIONS WITH IONIZING
RADIATION-DNA DAMAGE

Residual unrejoined double strand breaks
are lethal to the cell, whereas incorrectly
rejoined breaks may produce important
mutagenic lesions. In many cases, this
DNA misrepair apparently leads to DNA
deletions and rearrangements. Such
large-scale changes in DNA structure are
characteristic of most radiation induced
mutations.
RADIATION INDUCED CHROMOSOME
DAMAGE
Chromosomes
are composed of
deoxyribonucleic acid (DNA), a
macromolecule containing genetic
information. This large, tightly coiled,
double stranded molecule is sensitive to
radiation damage. Radiation effects range
from complete breaks of the nucleotide
chains of DNA, to point mutations which
are essentially radiation-induced chemical
changes in the nucleotides which may not
affect the integrity of the basic structure.
RADIATION INDUCED CHROMOSOME
DAMAGE
 After
irradiation, chromosomes may appear to be
"sticky" with formation of temporary or permanent
interchromosomal bridges preventing normal
chromosome separation during mitosis and
transcription of genetic information. In addition,
radiation can cause structural aberrations with pieces
of the chromosomes break and form aberrant
shapes. Unequal division of nuclear chromatin
material between daughter cells may result in
production of nonviable or abnormal nuclei.
RADIATION INDUCED MEMBRANE
DAMAGE

Biological membranes serve as highly specific
mediators between the cell (or its organelles) and the
environment. Alterations in the proteins that form part of
a membrane’s structure can cause changes in its
permeability to various molecules, i.e., electrolytes. In
the case of nerve cells, this would affect their ability to
conduct electrical impulses. In the case of lysosomes,
the unregulated release of its catabolic enzymes into
the cell could be disastrous. Ionizing radiation has been
suggested as playing a role in plasma membrane
damage, which may be an important factor in cell death
(interphase death)
DNA- mutation:Unusual permanent change in
the primary structure of DNA ( gene mutation).
 Chromosomal mutation: change in the amount
of DNA in chromosomes (aberrations) leads to
abnormalities in cell division.
 Genotoxic:damging the genetic information

STRATEGIES FOR GAMMA STERILIZATION
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During use of this type of radiation, high-energy photons
bombard the product, causing electron displacement within.
These reactions, in turn, generate free radicals, which aid in
breaking chemical bonds.
Disrupting microbial DNA renders any organisms nonviable
or unable to reproduce .
However, these high-energy reactions also have the potential
to disrupt bonds within the pharmaceutical formulation, to
weaken the strength of packaging materials, and to cause
changes in color or odor in some materials
APPLICATION OF RADIATION STERILIZATION
A. pharmaceutical preparation
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Injectable preparations supplied in the dry state to be
reconstituted just prior to administration ;
Injectable aqueous solutions of some electrolytes;
Dusting powders of antibiotics and steroids
Ophthalmic preparations of ointments, paper strips
impregnated with a diagnostic agent like fluorescein
sodium ;
Enzymes and other pharmaceutical materials which need
not be sterile but should not contain some non-pathogenic
microorganisms above certain limits;
APPLICATION OF RADIATION STERILIZATION
B. Cosmetic materials
 Such as; Talc, fatty acid esters, proteins, etc. which may be a
source of microbial contamination ;
C. Health care items Such as sanitary pads.
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Drugs in dry state are generally more stable to radiation
than in aqueous media .
Some drugs in ophthalmic ointment basses are also stable
to sterilization doses of radiation.
APPLICATION OF RADIATION STERILIZATION
The radiolysis products of water are well characterized today
1. The main radiolytic products of water, are Hydroxyl radicals
(OH.), Hydrogen atoms (H.), and hydrated electrons (éaq.).
2. The destructive effect of radiolytic products of water, either
collectively or individually on cortisone acetate during
gamma irradiation and the protective effect of different
types of free radicals scavengers as well as surfactants,
cationic, anionic or non-ionic, in irradiated aqueous
solutions of cortisone acetate was studied.
APPLICATION OF RADIATION STERILIZATION
3. Comparing the effect of the three radiolytic products of
water, OH., H. and éaq. on cortisone acetate, it can be
observed that only 3.95% of the drug was degraded by the
hydrated electron at a dose of 2.29 kGy, while
approximately 72% of the drug was degraded by OH. or H.
at the same dose of radiation indicating that the hydrated
electron has very little destructive effect on the drug
compared to OH. or H. .
APPLICATION OF RADIATION STERILIZATION
4. However, the effect of OH. radicals can be seen to be
slightly greater than that of H. atom
5. The slight greater effect of OH. than that of H. may
be result of an inadequate removal of the OH. from
the system because of the low solubility of hydrogen
gas in water
6. It has been reported that the addition of free radical
scavengers to aqueous solutions of organic material
may markedly affect the radiolytic yield of that system
through their reaction with the primary radiolytic
products resulting from radiation.
APPLICATION OF RADIATION STERILIZATION
7. For example, methanol acts as a scavenger for OH.
radical, H. and éaq. resulting from radiolysis of aqueous
solutions, while 2-propanol acts as a scavenger only for
OH. and H.
8. Both methanol and 2-propanol have a protective effect
on cortisone acetate when irradiated in aqueous solution.
9. On increasing the percentage of both alcohols in
aqueous solutions of cortisone acetate, it is apparent
from that 2-propanol is more effective as a stabilizer for
the drug than methanol.
APPLICATION OF RADIATION STERILIZATION
10. This result is expected because 2-propanol
is considered to be more reactive to OH.
and H. than methanol
11. This result would indicate the important role
of OH. radicals and H. atom in the degradation
of cortisone acetate in the irradiated aqueous
solution
APPLICATION OF RADIATION STERILIZATION
Thus, the calculation of a sterilizing dose will depend on a
number of factors ;
1. The final degree of contamination that can be
tolerated in a given application,
2. The initial count of viable microorganisms,
3. The radiosensitivity of the contaminating
organisms under the conditions of the sterilizing
process, and
4. The minimization of radiation damage .
APPLICATION OF RADIATION STERILIZATION
Ionizing radiation was proposed in the British pharmacopoeia
as a suitable sterilization method for sterilizing certain
surgical materials and equipments with a dose requirement
of 25 kGy, but no specific guidance is given on how to
estimate doses of less than 25 kGy.
 While in International Atomic Energy Agency (IAEA), it was
recommended that a 25 kGy radiation dose may be used as
practical dose for radiation sterilization of medical supplies.
 The choice of sterilizing radiation dose is no longer fixed at
25 kGy but rather is based on the initial microbial load
coupled with the desired sterility assurance level .
APPLICATION OF RADIATION STERILIZATION
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A dose of 25 kGy was established as a dose of
irradiation for the sterilization of medical devices.
This was based on extensive studies and more
important, on mutual international agreement. The
use of 25 kGy, as a standard dose, has been
criticized as being too low to satisfy the
requirements of sterility.
APPLICATION OF RADIATION STERILIZATION

The USP XXIV states that “Although 25 kGy of absorbed
radiation was historically selected, it is desirable and
acceptable in some cases to employ lower doses for
devices, drug substances, and finished dosage forms. In
other cases, however, higher doses are essential…”. While
international generally accepted regulation dose not exist
yet for the radiation treatment of pharmaceuticals, it can
be said, that the radiation dose is depend on the radiation
resistance of the microorganisms, the sterility assurance
level desired and the radiation sensitivity of the products.