Evaluation of the efficiency of sterilization method and sterility indicators

A detailed overview of the Evaluation of the efficiency of sterilization method and sterility indicators

1. UNIT II 2.2 Evaluation of the efficiency of sterilization method and sterility indicators Presented by: Mohammad Abuzar( M. Pharm ) Assistant Professor School of Pharmacy AIKTC, New Panvel .

2. CONTENTS 2

3. 3 INTRODUCTION Sterility criteria Bioburden is normally defined as the number of bacteria living on a surface that has not been sterilized. The term is most often used in the context of bioburden testing, also known as microbial limit testing, which is performed on pharmaceutical products and medical products for quality control purposes. Time and temperature relationship for steam sterilization to ensure that a large number of the most resistant pathogens would be killed.

4. Sensitivity of microorganisms Microorganisms shows resistance to heat, radiation and chemicals. The vegetative forms of bacteria and fungi are most sensitive. The thermophilic bacteria, smaller viruses and mould spores are killed at temperature between 70 to 90°C, while bacteria spores may be destroyed at 90 to 120°Ctemperatures Death rate or Survivor curve It is determined by assessing the reduction in the number of viable microorganisms resulting from contact with a given destructive force. This can be represented graphically with a 'survivor curve' drawn from plot of the log of the fraction of survivor against the exposure time or dose. 4

5. 5

6. 6 D- Value or Decimal reduction time Time in minutes at any defined temperature to destroy 90% viable microorganisms is called D- value

7. 7 Z- value or Thermal reduction time The slope of the TDT curve is defined as "Z" which is equal to the number of degrees on the temperature scale when the curve traverse one log cycle. Z is the change in temperature necessary to cause a ten fold change in D-value. The value of Z for C. botulinum is 10°C Every 10°C change in temperature there is a ten fold change In its death rate. B. subtilis has Z value of 6.5°

8. 8 f- value Used in the food industry, also called "Unit of Lethality" has been devised called as F- Value This can be defined as the equivalent in minutes of 121°C of all heat consider with respect to its capacity to destroy spores or vegetative cell of a particular organisms The F value for a process is the number of minutes required to kill a known population of microorganisms in a given food under specified conditions. This value is also used to calculate the probable number of bacteria remaining after the process in food Q10 Value or Temperature Coefficient It is defined as the increase in rate of reaction or killing rate of a sterilization process brought after increasing the temp. by 10°C.

9. Inactivation Factor It is the degree to which the viable population of organisms is reduced by applying a sterilization process. It is obtained by dividing the initial viable count by final viable count. Inactivation factor (IF) = 10 t/D Where, t - exposure time in minutes, D = Decimal reduction time for the same temperature and conditions. 9

10. Sterilization indicators Sterilization procedures should be monitored through a combination of physical, chemical, and biological techniques designed to evaluate the sterilizing conditions and the procedure's effectiveness Physical indicators Chemical indicators Biological indicators Physical indicators Observing the gauges or displays on the sterilizer for Assessing the cycle time Temperature Pressure of sterilization equipment Relative humidity

11. 11 Heat sterilization A temperature record chart is made - part of the batch documentation Compared against a master temperature record (MTR) Information on heat distribution and penetration within a sterilizer can be gained by the use of thermocouples Gaseous sterilization Measurement of elevated temperatures using temperature probes Routine leak tests to ensure gas-tight seals Pressure and humidity measurements are recorded. Gas concentration is measured, often by reference to weight of gas used Radiation sterilization A plastic (often Perspex) dosimeter is used It gradually darkens in proportion to the radiation absorbed

12. 12 Integrity testing sterilizing filters Membrane filters have been used successfully for many years to remove yeast, bacteria and particulate from fluid streams To test the integrity of filters, the following tests are done: a)Destructive test b)Non - destructive integrity test   a)Destructive test- Destructive challenge testing is the best way to determine a sterilizing filter's ability to retain bacteria. During the bacterial retention test, 0.22 µm filter discs and devices are challenged with a solution of culture medium containing bacteria ( Brevundimonas diminuta ATCC 19146) at a minimum challenge of 107 per cm 2 . The effluent is then passed through a second 0.45 µm filter disc that is placed on an agar plate and incubated. The filter cannot be used for filtration purposes again

13. b)Non - destructive integrity test , may be done on filters before and after filtration process This is done to ensure that the filter meets specification, is properly installed and intact during filtration, and to confirm the rating of the filter Integrity testing before filtration process monitors filter integrity, preventing use of a non-integral filter. Integrity testing after a batch has been filtered can detect if the integrity of the filter has been compromised during the process There are 3 major tests used to determine the integrity of a membrane filter: T he Bubble Point Test The Forward Flow Test, T he Pressure Hold Test All three tests are based on the same physics, the flow of a gas through a liquid-wetted membrane under applied gas pressures They differ in which part of the flow/pressure spectrum they examine This is an indirect method for the measurement of pore size of filter 13

14. 14 WWW.PHARMANOTES.ORG 1)Bubble point pressure test Bubble point test is based on the fact that liquid is held in the pores of the filter by surface tension and capillary forces The minimum pressure ( as seen on the pressure dial) required to force liquid out of the pores is a measure of the pore diameter The bubble point is expressed as : Where k = shape correction factor Ύ = surface tension cos ɵ = liquid solid contact angle d = pore diameter Consists of immersing the filter candles in water or filling the funnel with the liquid in the case of a membrane filter or sintered glass filter Air or gas is passed from the bottom of the filter The pressure of Air/ gas is gradually increased until the first bubble is seen at the filter/liquid interface

15. Filtration sterilization Bubble point pressure test

16. 16 Chemical indicators Chemical indicators are designed to respond to a characteristic change to one or more of the physical conditions within the sterilizing chamber The principle is based on the ability of heat, steam, gases and ionization radiation to alter the chemical and/or physical characteristics of chemical substances Change should take place only when satisfactory conditions for sterilization prevail Used for monitoring a sterilization process Chemical indicators generally undergo melting or colour changes Included in strategically placed containers or packages Monitors the conditions prevailing at the coolest or most inaccessible parts of a sterilizer

17. 17 Bowie Dick test Organic chemical in a printing ink base impregnated into a carrier material. A combination of moisture and heat produces a darkening of the ink Brown’s tube Sealed tubes partly filled with a solution which changes colour at elevated temperatures; rate of colour change is proportional to temperature

18. 18 Thermalog S Consists of a blue dye in a waxy pellet. At autoclaving temperatures, and in the continued presence of steam, the pellet melts and travels along a paper wick forming a blue band the length of which is dependent upon both exposure time and temperature Thermalog G Same as Thermalog S. Used for gaseous sterilization Formaldehyde sterilization indicators

19. 19 Chemical indicators for radiation sterilization

20. Biological indicators Consist of standardized bacterial spore preparations Usually in the form either of suspensions in water or culture medium or of spores dried on paper, aluminium or plastic carriers. As with chemical indicators, they are usually placed in dummy packs located at strategic sites in the sterilizer After the sterilization process, the aqueous suspensions or spores on carriers are aseptically transferred to an appropriate nutrient medium, which is then incubated and periodically examined for signs of growth. The bacterial species to be used in a BI must be selected carefully, as it must be non-pathogenic and should possess above-average resistance to the particular sterilization process. 20

21. Filtration sterilization Measuring the ability of a filter to produce a sterile filtrate from a culture of a suitable organism Serratia marcescens has been used for filters of 0.45mm pore size, Brevundimonas diminuta (formerly Pseudomonas diminuta ) having a minimum dimension of 0.3mm is applied to filters of 0.22mm pore size. 21

22. 22 Summary Validation means demonstrating that a process will consistently produce the results that it is intended to Sterilization indicators - Physical indicators, Chemical indicators, Biological indicators Physical indicators validate the temperature, pressure, radiation dose or filter pore size Chemical indicator - Based on the ability of heat, steam, sterilant gases and ionizing radiation to alter the chemical and/or physical characteristics of a variety of chemical substances Biological indicators - Consist of standardized bacterial spore preparations

23. 23 W.B. Hugo and A.D. Russel: Pharmaceutical Microbiology, Blackwell Scientific publications, Oxford London. Prescott and Dunn., Industrial Microbiology, 4th edition, CBS Publishers & Distributors, Delhi. Pelczar , Chan Kreig , Microbiology, Tata McGraw Hill edn . Malcolm Harris, Balliere Tindall and Cox: Pharmaceutical Microbiology. Rose: Industrial Microbiology. Probisher , Hinsdill et al: Fundamentals of Microbiology, 9th ed. Japan Cooper and Gunn’s: Tutorial Pharmacy, CBS Publisher and Distribution. Peppler : Microbial Technology. I.P., B.P., U.S.P.- latest editions. Ananthnarayan : Text Book of Microbiology, Orient-Longman, Chennai Edward: Fundamentals of Microbiology. 12. N.K.Jain : Pharmaceutical Microbiology, Vallabh Prakashan , Delhi REFERENCES

24. 24