Autoclave validation | Sterilization Validation

OBJECTIVE

To ensure that the desired temperature and pressure required to sterilize the   

load is achieved at different places in the load placed in an autoclave.

SCOPE    

This protocol is intended to prove with a high degree of confidence that the sterilization process is consistent, reproducible, under control and able to supply a final product which fulfils all the sterilisation specification attributes. This will be demonstrated by monitoring the critical sterilisation parameters, according to what established in this protocol.

A concurrent validation approach has been chosen to perform three consecutive runs for each consisting empty load, full load and third full load with biological indicators according to the validation protocol. All the equipment, procedures and personnel involved in the validation process will be the same as those used for normal production batches. Procedures and documentation will comply with the current GMP and Global Quality Standards.

The validation of the autoclave shall be divided into three parts as follows:

             Part A   :    Thermal  Studies

             Part B   :    Biological  Studies

             Part C   :    Correlation of Thermal and Biological studies to determine the

                                Fo delivered by the process.

Part A - Thermal Studies (Heat penetration & heat distribution)

The studies shall be performed to validate the "Performance" of the sterilizer for   delivering the desired lethality at all the points in the sterilizer equally and 

uniformly.

The studies, therefore shall include the following aspects: -

Use of Thermocouples (T type) min. 12 nos.  for thermal mapping of the    

steam sterilizer using a data logger. It will include

• Empty Run
• Maximum Load

Since these are trial runs for 3220D type granule with small batch size Minimum Load validation runs are not performed. However, these will be performed on regular batch size.                            

• Monitoring the pressure inside the autoclave using a calibrated sensor mounted on the sterilizer.
• Check for correlation between 'RTD’ s already in place and additional thermocouples together with the gauges mounted for correct operating parameters     
• Studying the effect of various loads and loading patterns on the thermal distribution and heat penetration.

Determination of the following points for critical evaluation: -

                              a.    'Slow to heat' points

                              b.    'Cold' point

                              c.    'Most-difficult-to-sterilize’ point.

(Note: All relevant calibration documents will be made available.)

Part B - Biological Studies

This aspect involves the challenging of the sterilizer with an appropriate biological load to evaluate the efficacy of the sterilization process.

Under the present condition where low temperature sterilization is being carried out viz. at 112.5 ºC for 50 mins. the most appropriate organism would be “Bacillus subtilis" ATCC: 35021 or ‘5230' with a D - value of 0.3 mins. Based on the data obtained from the thermal studies the Biological Indicators (BI) would be placed appropriately to evaluate the lethality delivered by the process. After the exposure of the BI’s it will take 48 hrs to obtain the results. (As per diagram attached)

(Certificate of Analysis will be made available mentioning D - value and the empirically derived survival/kill time and Fo values)

Part C - Correlation of Thermal and Biological Studies to determine the  

Process Fo

This will be the final step wherein the Process Fo value would be derived from the thermal data.

Note: If sufficient lethality is not being achieved at all the points in the sterilizer uniformly, relevant recommendations will be given to achieve the   same.

Procedure of Validation: - For each autoclave following runs shall be taken.

• Empty Run - to setup the thermocouples at various locations and run the cycle.
• Maximum Load - with the thermocouples kept inside the bottles (by cutting   the bottle at the top and sealing the bottle with the help of cellophane tape) at various locations and run the cycle. In the third cycle of full load, BI’s shall be placed adjacent to thermocouple (inside the bottles) to verify lethality at the locations.                                    

(generally the maximum thermal load will be employed as a "WORST

CASE MODEL.                                    

In this validation since batch size is 5000 L for 1000 ml, 1200 L for 500 ml and 600 L for 100 ml, number of bottles produced may not be sufficient for maximum l load, * so for making full load bottles from another batch shall be taken. This is being done especially for this validation studies. In normal validation studies or production cycles such condition shall not prevail.
Instruments:

Digital Temperature Indicator & controller:   Calibrated 

Pressure Gauge: Calibrated

Calibrated Data logger

Calibrated RTD’S –  Min. Twelve numbers.

PROCESS DESCRIPTION

BRIEF DESCRIPTION OF OPERATION

• Calibrated temperature sensors shall be connected to autoclave and then to data logger before starting autoclave validation. 
• Data logger shall be started and temperature shall be recorded at interval of 1 minute during each sterilization cycle.
• Pressure and temperature on the dial shall be noted manually. (indicated    temperature and pressure)
• Biological validation shall be done on third full load of sterilization of each autoclave. Ampoules of biological indicators shall be kept inside the bottles by cutting the bottle at the top and then by sealing it with the cellophane tape in which the thermocouples are kept in previous cycles. One ampoule of biological Indicator shall be kept outside the autoclave as a positive control during each sterilization cycle.
• For negative control one ampoule of negative control shall be incubated along with these ampoules.
• After completion of sterilization cycle, ampoules of spores shall be collected and incubated at 35-39 °C for 72 hours. And results shall be recorded in the   separate protocol.

ACCEPTANCE CRITERIA:

• The holding time as determined from the measured temperature shall not be less than 50 minutes for Minimum Load and Maximum Load.
• During the holding time -
• The measured temperature shall be within the average sterilization temperature of 112.5 to 114.5 °C.
• The measured temperature in each load item does not fluctuate by more than 2 °C.   The indicated chamber pressure shall be within 0.05 bar of set pressure.
• The F0 value for each load should be greater than or equal to 8 and the Sterility Assurance Level (SAL) should be 10`   ⁶. for Minimum Load and Maximum Load.
• Ampoules of spores exposed to sterilization shall not show any evidence of growth in all the sterilization cycles during specified incubation period i.e. colour of the ampoule shall remain as it is. (red colour)
• Positive control should show growth in terms of turbidity and change in colour from red to yellow within 24 hours of incubation.
• Negative control should not show any evidence of growth i.e. red colour of the ampoule shall remain red only.
• If the growth resulted from organism other than the test organism, the test is inconclusive and should be repeated.

AT THE END OF THE CYCLE:

The temperature sensors shall have remained in position.

The items containing sensors should be intact.

Printouts of data logger shall be checked for appropriate temperature.

EVALUATION AND CERTIFICATION:

Data obtained from data logger and Sterility reports of biological indicators shall be evaluated by Quality Assurance Department.

After confirming the satisfactory result, Quality Assurance head shall certify the validation.

CORRECTIVE ACTION IN CASE OF VALIDATION FAILURE:

The Quality Assurance Head shall decide on corrective action to be initiated.

The corrective action shall be based on investigation of various factors affecting the study like:

Points related to Autoclave,

Steam leakage, Valve adjustment, Air pockets, Condition of heating element

Calibration of instruments (in consultation with Head of Maintenance department.) 

An investigation report shall be prepared jointly by Quality Assurance and Maintenance specifying the corrective actions to be taken. The validation process shall be repeated after taking the corrective actions. The autoclave   shall not be used till the autoclave is revalidated.

Frequency of Steriliser Validation: -  Once in a Year.