validation of autoclave

industrial operation and validation of double door autoclave

1. INDUSTRIAL OPERATION AND VALIDATION OF A DOUBLE DOOR AUTOCLAVE PRESENTED BY B.KARTHIK M . Sc MICROBIOLOGY

2. INTRODUCTION OF AN AUTOCLAVE:- An autoclave is a pressure chamber used to carryout industrial processes requiring elevated temperature and pressure different from ambient air pressure. It is used for the sterilization of media, glass were, gloves, gowns etc., It is a moist type of sterilization unit. Normal temperature and pressure required for autoclave is 121⁰C and 15lbs pressure.

3. Aim:- To operate and validate the industrial double door autoclave.(Pharmalab) Objective:- Operate and Validate the industrial double door auto clave. Dehydrated media, glass were and accessories sterilization for analysis .

4. HOW DOES AN AUTOCLAVE WORKS Steam enters the chamber jacket, passes through an operating valve and enters the rear of the chamber behind a baffle plate. It flows forward and down through the chamber and the load, exiting at the front bottom. A pressure regulator maintains jacket and chamber pressure at a minimum of 15 lbs , the pressure required for steam to reach 121ºC (250ºF). Overpressure protection is provided by a safety valve.

5. What is validation ? Validation shows that a process (or item of equipment) does what it is claimed to do. Why validate? Need to show that sterile equipment is being taken into sterile core - do not want to contaminate production (which could lead to adulterated product) For micro testing (both bacteriological and Viable Environmental Monitoring), using sterile equipment helps reduce false positives. Need to show that decontamination cycles are effective

6. How To Validate The usual – URS. IQ, OQ, PQ. This discussion will focus on PQ. IQ: does equipment meet the URS requirements? Is everything that was on the box, in the box? Is the unit installed properly. Are support programs in place for ongoing operation of A/C? OQ: does the A/C operate properly? Does the unit hold temp and pressure correctly? PQ: validation of autoclave cycles and loading patterns – need to show sterilization.

7. What is D value? Refers to decimal reduction time - The time required at a certain temperature to kill 90% of the organisms being studied. Thus after an organism is reduced by 1 D, only 10% of the original organisms remain. Dependant on microbe and initial numbers. E.g. D value of 1.5 = 1CFU . What is Z value? Refers to the temperature change required to produce a 1 log reduction in D value. What is F ? The number of minutes to kill a specified number of microbes with a Z value of 10 o C at a temp of 121.5 o C. Often confused with the time the chamber is held at elevated temperature and pressure and in practice is the same thing.

8. Overkill Use many more microbes than would find on items typically autoclaved. Negates the need to test sample for bio load before running the cycle. Use a sterilisation time exceeding what is necessary to kill a large number of microbes. Negates the need to determine D value of microbe.

9. Pro cedure to operate an autoclave :- Switch on auto clave Fill the water tank with water line Switch O n the heater switch Open the door – 1 Load the chamber with media or glass were or with accessories Arrange the sensors as we required Close the door- 2 login with user id and password

10. Select the type of recipe Start the cycle Set up the strip chart Observe the temperatures on the display After completion of cycle open the door – 2 Give print to the printer Calculate the F0- value, D-value and Z- value with the use of computer software

11. Recipes for validation of a double door autoclave There are mainly 5 recipes present in the validation of a double door auto clave. They are :- VLT (Vacuum leak test) Bowie – dick test cycle Steam In Process (SIP) Gravity test cycle HPHV test cycle

12. Vacuum leak test:- Vacuum leak test is performed to check any leakage in autoclaves. This test is also called chamber integrity test. In VLT, first vacuum is applied inside the chamber and then wait for the vacuum to reach at set vacuum point then hold the vacuum for required time and when hold time completed vacuum breakage occurs. After that check the results whether autoclave leakage is within the limits or not.

13. PARAMETERS:- Chamber level test vacuum :- -0.007bar Stabilization time :- 5min Vacuum hold time :- 10min Acceptable leakage:- 0.0013bar/min Print interval :- 60sec Frequently test :- 15 days ± 2 days Acceptable criteria :- 0.0013 bar/Min No. of cycles run for validation :- 3

14. Bowie Dick test :- This is the cycle done for the removal of contaminated air pockets in an autoclave. In this the chemical indicator Bowie Dick chemical indicator papers will be used. The Bowie Dick indicator paper will turn from yellow to black(or) brown in colour.

15. Bowie – Dick test papers

16. Parameters :- Pre vacuum :- -0.700Bar Pre vacuum hold time :- 01 min Pre pressure :- 0.300Bar Pre pressure hold time :- 01 min Heat up 1 :-107⁰C Heat up 1 hold :- 01min Heat up 2 :- 112⁰ Heat up 2 hold :- 01min Heat up 3 :- 115⁰ Heat up 3 hold :- 01 min Heat up 4:-117⁰ Heat up 4 hold :- 1min Sterile hold temp :- 121.3⁰ Control band :- 000.3 Sterile hold time :- 11 min Sterile hold reset :- 120.6⁰ Sterile hold stop count :- 121⁰ Post vacuum :- -0.700Bar Post vacuum hold time :- 02 min Door open temp :- 90 Print interval :- 60 sec Control temp :- min for all

17. SIP (Steam In Process) cycle:- It is the cycle done for vent filter sterilization of an autoclave. It is the test done once per a month Acceptance criteria :- For SIP of filter temperature should be more than 121⁰C during sterilization.

18. Parameters :- Heat up 1 :-107⁰C Heat up 1 hold :- 01min Heat up 2 :- 112⁰C Heat up 2 hold :- 01min Heat up 3 :- 115⁰C Heat up 3 hold :- 01 min Heat up 4:-117⁰C Heat up 4 hold :- 1min Sterile hold temp :- 122⁰C Sterile hold time :- 15 min Sterile hold reset :- 120.6⁰C Sterile hold stop count :- 121⁰C Sterile temp over shoot :- 130 ⁰C Post vacuum :- -0.700Bar Post vacuum hold time :- 02 min Door open temp :- 90 Print interval :- 60 sec

19. Gravity cycle :- This is the cycle used for the sterilization of the media and water. This test is done as per our requirement. Temperature should be in between 121⁰– 124⁰ during sterilization hold time. The steam pressure should be 1.04-1.24 bar during the sterilization hold time. In this the chemical indicator paper and biological indicator will be used. The biological indicator is Geobacillus sterothermophillus . This is a spore producing bacteria which can resist up to 120⁰C temperature.

20. Parameters :- Heat up 1 :-105⁰C Heat up 1 hold :- 02min Heat up 2 :- 108⁰ Heat up 2 hold :- 02min Heat up 3 :- 112⁰ Heat up 3 hold :- 02 min Heat up 4:-117⁰ Heat up 4 hold :- 03min Sterile hold temp :- 121.3⁰ Control band :- 000.3 Sterile hold time :- 20 min Sterile hold reset :- 120.5⁰ Sterile hold stop count :- 121⁰ Sterile temp over shoot :- 124⁰ Jacket control temp :- 121⁰ Door open temp :- 96⁰ Print interval :- 60 sec Control temp :- min for all

21. Biological indicator for gravity cycle Geobacillus sterothermophillus Before test After test

22. HPHV cycle :- High pressure and high vacuum test cycle is used for the sterilization of a glass were, accessories etc., Temperature should be in between 121⁰– 124⁰ during sterilization hold time. The steam pressure should be 1.04-1.24 bar during the sterilization hold time.

23. Parameters :- Pre vacuum :- -0.700Bar Pre vacuum hold time :- 02 min Pre pressure :- 0.300Bar Pre pressure hold time :- 02 min Heat up 1 :-107⁰C Heat up 1 hold :- 02min Heat up 2 :- 112⁰C Heat up 2 hold :- 02min Heat up 3 :- 115⁰C Heat up 3 hold :- 02 min Heat up 4:-117⁰C Heat up 4 hold :- 1min Sterile hold temp :- 121.3⁰C Control band :- 000.3 Sterile hold time :- 30 min Sterile hold reset :- 120.6⁰C Sterile hold stop count :- 121⁰C Sterile temperature over shoot :- 124⁰C Post vacuum :- -0.700Bar Post vacuum hold time :- 02 min Door open temp :- 90 Print interval :- 60 sec

24. Observations and Results: Vacuum leak test: Run - I Run - II Run –III Date 25/05/2016 25/05/2016 25/05/2016 Cycle start time 14;53;50 15:16:37 15:59:19 Vacuum hold start time 15:01:16 15:24:04 16:06:49 Vacuum hold end time 15:11:16 15:34:04 16:16:49 Cycle completion time 15:12:35 15:35:23 16:18;09 Vacuum hold start pressure -0.723 bar -0.724 bar -0.725 bar Vacuum hold end pressure -0.717 -0.717 -0.717 Actual leakage 0.006bar 0.007bar 0.008bar

25. Bowie Dick test Run –I Run – II Run – III Date 25/05/2016 25/05/2016 25/05/2016 Cycle start time 16:38:01 18:04:20 19:11:44 Sterile hold start time 17:18:58 18:29:25 19:36:13 Sterile hold end time 17:29:58 18:40:25 19:47:1 3 Cycle completion time 17:47:32 19:00:08 20:05:42 No leaks observed Bowie Dick kit turns blue colour to black colour

26. Steam in process (SIP) test: Run – I Run – II Run - III Date 27/05/2016 27/05/2016 27/05/2016 Cycle start time 13:36:00 14:29:42 16:14:40 Sterile hold start time 13:45:00 14:39:05 16:24:40 Sterile hold end time 14:00:00 14:54:05 16:39:40 Cycle completion time 14:03:45 14:57:35 16:42:20 Min. Temp during hold 121.1 121.1 121.1 Max. Temp during hold 124.4 124.5 124.5 Avg. Temp during hold 123.25 123.0 123.8 Min F 22.79 22.64 27.52 Max F 26.25 23.41 28.66

27. Conclusion After operating and validating the autoclave all the parameters which are given by USP is satisfied and the results are also satisfied for all the recipes and all the cycles what we done for validating the autoclave. So we concluded that this autoclave is working very efficiently and sterilizes the media and accessories properly without any contamination. So now it is used for regular usage of sterilization.

28. Thank you