 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.

 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 +

 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.

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 (H 2 ) 2.The formation of toxic hydrogen peroxide (H 2 O 2 ). 3.The radical can also be transferred to an organic molecule in the cell. H-O-H  H OH + + e (ionization) H OH + + e  H 0 +OH 0 (free radicals)

 H 0 + OH 0  HOH (recombination)  H 0 + H 0  H 2 (dimer)  OH 0 + OH 0  H 2 O 2 (peroxide dimer)  OH 0 + RH  R 0 + HOH (Radical transfer)  The presence of dissolved oxygen can modify the reaction by enabling the creation of other free radical species with greater stability and lifetimes  H 0 +O 2  HO 2 0 (hydroperoxy free radical)  R 0 +O 2  RO 2 0 (organic peroxy free radical)

 The lifetimes of simple free radicals (H 0 or OH 0 ) are very short, on the order of 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.

 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.

 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. I S-AT-S I P I S-CG-S I P I S-GC-S I P I S-TA-S I

 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.

 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.

 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.

 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.

 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.

 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.

 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

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

A. pharmaceutical preparation 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;

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. 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.

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.

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..

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.

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.

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

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.

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.

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.

 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.