(1) Sterility testing (2.5 points)
(a) Within which systems and under which cleanroom class must the sterility test be performed?
(b) The culture medium is of central importance in sterility testing. What test besides checking the sterility of the culture medium must be carried out for each batch of culture medium?
(c) What is the acceptance criterion for sterility testing?
(d) What is the maximum number of containers of a parenteral that must be examined for sterility testing (see table)?
a) under aseptic conditions ISO 5 (Class A) in isolator
= sterility testing isolator class A with surrounding area class D
b) Positive growth promotion test (with test microorganisms for each batch)
= prove that special test organisms cannot grow in the nutrient medium
-> Test principle: - very small number (100 CFU) of colonies is taken & placed in the nutrient medium (TBS or TM)
- different microorganisms (aerobic/ anaerobic/ fungi/ spores) get incubated to detect if they grow
c) acceptance criterion for sterility testing
= No visible growth after 14 days the test sample
= a product is sterile if no microbes are detected in the sample after incubation
d) maximum number of containers of a parenteral
= 20 containers
(2) (a) What is described by the SAL value?
(b) For a sterilization process, the probability of sterility after the process is P = 0.99999. How many non-sterile containers would have to be expected for a batch size of one million containers?
(2 points)
a) SAL = Sterility Assurance Level
= probability, that germs may be alive
= is expressed as the probability of a non-sterile item after sterilisation
= describes the probability of a unit being non-sterile after undergoing sterilization process
b) P = 0.99999. How many non-sterile containers would have to be expected for a batch size of one million containers?
SAL of 10^−6
= means a probability of not more than 1 viable germ in 1 ×10^6 sterilised items of final product
= of 1 million product containers 1 container is contaminated after sterilization
Calculation:
SAL = 1-0,99999 = 1*10^-5
10^6/ 10^5 = 10
=> 10 non-sterile containers woulb be expected for a batch size of 1 mio. containers (with P = 0,99999)
(6) Steam sterilization processes (4 points)
(a) How is the air removal carried out in the autoclave shown?
Note: the items to be sterilzed are placed in the grid basket.
(b) Name two disadvantages of the autoclave shown in (a) compared to an autoclave with an external steam generator.
(c) Draw the pressure curve of a steam sterilization process using fractionated vacuum for air elimination and a single vacuum phase for drying.
(d) Which steam sterilization process is suitable for the sterilization of bottled water for injections besides the hot water spray process?
(e) The figure shows an autoclave tape. What is it used for?
(5) (a) A requirement for pure steam is the “correlation with dry saturated steam”. Why does pure steam have to be dry and saturated?
(b) Why is there also a limit value for bacterial endotoxins in pure steam?
(1.5 points)
(3) You have the following data on the heat killing of a microorganism at a temperature of 126.5 °C: a microbial population is reduced by 5 log levels in a time of 240 s. z = 5.5 °C.
(a) What D-value at 121°C results from the data?
(b) After what total time at 121 °C is a SAL of 10^-9 achieved if 100 microorganisms are present at the beginning?
(2.5 points)
a) D-value = decimal reduction value [mins]
= time needed at a certain temperature to kill 90% of germs
= time required to reduce the number of viable organisms to 10 % of the original number
= time that is needed to reduce cfu number by factor 10 or by 1 log 10
Calculation: 240s = 4min
D-value = mins / log levels = 4mins / 5 = 0,8 mins
b) Total time = F-value = time (holding time) at a temperature T in minutes to reduce the population (N0) with the specific D value by the desired number of powers -> often log10 levels (n)
n = from 100 germs to 10^-9 = 11 log levels
F (total time) = 11 * 0,8 mins = 8,8 mins
(4) (a) What does the z-value tell us about the heat resistance of a microorganism?
(b) What is the relationship between the F value and the SAL value?
(c) For a steam sterilization process at 134 °C, an F0 value of 50 min is achieved. How is the achieved F0 value of 50 min to be interpreted?
a) Z-value = Temperature change in °C required to alter D-value by factor of 10
high z-value = means large increase in °C is required to achieve 1 log reduction of D-value = means that germs are more heat resistant to changes of temeprature
low z-value = low increase in °C is required to achieve 1 log reduction in the D-value = germs are less heat resistant
e.g. spores = z-value 10°C & vegetative cells = 5,5 °C
b) Relationship between the F value and the SAL value
F-value = thermal death time = holding time [mins] at a temeprature T to reduce population N0 with specific D-value by desired number of poweres (log levels) to final value N
SAL = sterility assurance level = probability that germs may be alive
Relationship: with sufficient F-value [holding tims in mins] a sterilisation process can ensure that the SAL-value achieves the required standard
high F-value = high holding time = low probability of items remaining non-sterile, low probability that germs stay alive = means a better (lower) SAL-value
low F-Value = low holding time = high probability that sterilisation is not sufficient & germs stay aliba = high SAL-value
F0-value [mins] = 1 min means the lethality of a method that works for 1 minute at 121.1 °C (250 °F) and a z-value of 10 °C (18 °F) or an equivalent temperature-time combination
= makes different steam sterilization comparable -> important for pharmacy
50min (F0-value) at 134°C = process achieves the same microbial lethality as the sterilisation at 121°C for 50 mins
(10) Sterile filtration (2 points + 0.5 bonus points for part b)
(a) What type of filtration (in terms of separation limits) does sterile filtration belong to? Tick the correct box.
(b) Bonus question: Specify one common membrane material for sterile filters for the filtration of aqueous products.
(c) What is the unit of the bubble point?
(d) What is the acceptance criterion for the bacterial challenge test that is performed as one of the validation activities for a sterile filtration process?
(e) Sterile filters must be analyzed with regard to extractables, among other things. What are extractables?
(9) Describe the steps that are needed to biodecontaminate a cleanroom with hydrogen peroxide vapour (VHP process)? (2 points)
(18) Why must the monitoring results always be taken into account when releasing an aseptically filled product batch, even though a sterility test is carried out as part of the final product quality control? (1 point)
(11) Validation of sterilization processes (2.5 points)
(a) How do you call the release of terminally sterilized products without batch-related sterility testing, based solely on the measurement of physical process parameters?
(b) What must be considered in the PQ of a steam sterilization process with regard to loading - in addition to worst-case considerations?
(c) Which third key specification of a bioindicator besides the z-value and D-value must be known for the validation of a steam sterilization process?
(d) What should not be included in the URS of a steam sterilizer? Mark with a cross
(16) Preparation of equipment and primary packaging material (2.5 points)
(a) Which medium is used for the final rinse in the washing process of primary packaging materials?
(b) What is the key advantage of using single-use systems in aseptic manufacturing from a patient protection perspective?
(c) What is shown in the figure below?
(d) Why do syringes and cartridges receive an anti-friction coating on the inside?
Sterility testing (2 points)
(a) Why must sterility testing be carried out in an aseptic process environment?
(b) Why can the direct inoculation method not be used for sterility testing of a sterile antibiotic?
(c) A liquid vaccine is to be tested for sterility according to Ph. Eur. 2.6.1. The filling volume per container is 2.4 ml. What is the minimum volume per container that must be tested for sterility?
a) Why must sterility testing be carried out in an aseptic process environment?
= to ensure the safety of aseptic process & reliability of the results
= to prevent the introduction of contaminants during the process
= because sterility must be verified for each betach to get released
b) Why can the direct inoculation method not be used for sterility testing of a sterile antibiotic?
sterile antibiotic = inhibits the growth of germs (function of antibiotic)
-> if product inhibits the growth of microbes = direct inoculation cannot be used because its not possible to do Positive method suitability test ( = to prove that the product doesn’t inhibit the growth) -> antibiotoc does inhibit growth!
-> other sterility test must be applied
c) 2.4 mL => 1-40mL -> half the contents of ech container but not less tha 1 mL
-> half the content 2.4/ 2 = 1,2 => 1ml per container must be tested
(21) The pipe transition shown (flow direction from left to right) has an unfavourable hygienic design. Draw a transition with an ideal hygienic design. (1 point)
(23) The following illustration shows a CIP/SIP-capable sterile sampling valve. The product chamber is on the left of the picture. How can it be ensured that the sealing point is also reliably cleaned and sterilized? (1 point)
(15) Transfer systems (2 points)
(a) What is meant by mouseholes in barrier systems and for what purpose are they used?
(b) When using RTP systems, how is it ensured that no contaminated air enters the barrier system (see illustration)? Specify two aspects.
(12)
(a) According to GMP Annex 1, every company that manufactures sterile medicinal products requires a CCS. What does the abbreviation CCS stand for?
(b) One of the rules of aseptic working is not to interrupt the laminar air flow. GMP Annex 1 refers to this as the first air principle. Briefly explain why this rule must be followed whenever possible?
(c) Which aseptic working rule should class B personnel follow regarding their gloves?
An infusion solution (large volume parenteral) is to be tested for sterility according to Ph. Eur. 2.6.1. The filling volume per infusion bottle is 500 ml, the batch size is 10.000.
(a) Using the following table from Ph. Eur. 2.6.1, indicate the minimum number of bottles that must be used for sterility testing.
(b) The sterility test is carried out by membrane filtration. Describe the procedure for this test without the subsequent incubation.
a) more than 500 containers (bottles)
2% of 10.000 = 200 -> its too much
whihever is less: minimum 10 bottles for large volume parenterals should be tested
b) Procedure of sterility testing by by membrane filtration
Process: Sample filtration procedure
= should get done under cleanroom, laminar airflow work bench or in isolator
1) 2x empty bottles get connected with an injection needle a hose (Schlauch) through a peristaltic pump with the product bottle
2) Product extraction: with a peristaltic pump the product gets pumped out of the bottle -> product gets transported to the pre-sterilized bottles with a membrane filter
4) product is filtered & empty product bottle gets disconnected
5) a washing solution is placed via hose -> filters get washed to eliminate residuals (esp. of growth inhibiting product)
5) bottles get closed at the bottom
6) Steps for incubation: 2x different nutrient medium get introduced to the bottles to exclude (identify) different bacteria -> incubation for min. 14 days
(8) Ionizing radiation sterilization (1.5 points)
(a) In which case may ionizing radiation sterilization be used for the terminal sterilization of pharmaceutical products?
(b) Name one disadvantage of gamma irradiation compared to X-ray irradiation.
1) Sterility testing (3 points)
(a) How is sterility defined according to Ph. Eur.?
(b) What are the two sterility test types?
(c) An investigational medicinal product should be tested for sterility according to Ph. Eur. 2.6.1. The investigational medicinal product is eye drops. The batch size is 180. Using the following table from Ph. Eur. 2.6.1, state the minimum number of containers to be tested.
a) How is sterility defined according to Ph. Eur.?
= absence of viable micro-organisms
=> defined by a sterility assurance level (SAL) equal to or less than 10^−6
b) Two sterility test types
1) Membrane filtration
2) direct inoculation
c) Minimum number of containers - eye drops 180 batch
=> more than 100 but not more than 500 = min. 10 containers
Because eye drops are usually in single-dose containers (several drops)
(9) Ionizing radiation sterilization (2 points)
(a) A dose of 25 kGy is reached during irradiation. By how many log levels is a population of the yeast S. cerevisiae (D value: 0.5 kGy) reduced?
(b) Name one advantage and one disadvantage of electron irradiation compared to gamma irradiation.
(c) Why is UV-C radiation not suitable for the irradiation of e.g. medicinal products in sealed tubes?
D = 0,5
25/0,5 = 50
= by 50 log levels reduction
eletron irradiation
Advantages:
- No residues
- More gentle than gamma irradiation
- Continuous process, short irradiation time
Disadvantages:
- Limited penetration, but sterilization of products in final packaging possible
- High dose rate
gamma irradiation
- High penetration capability for sterilization in final packaging
- suitable for products with high density % large volume (e.g. whole pallets)
- Rather low dose rate
- Problem of radio active waste
- Not suitable for all materials (cross-linking / degradation reactions of polymers)
c) Why is UV-C radiation not suitable for the irradiation of e.g. medicinal products in sealed tubes?
= because absorbed on surface
= low penetration depth -> no reach of product
(a) Why can sterility testing alone not guarantee the sterility of a batch?
(b) What is examined in the product-specific suitability test?
(c) A liquid vaccine is to be tested for sterility according to Ph. Eur. 2.6.1. The filling volume per container is 1.2 ml. What is the minimum volume per container that must be tested for sterility?
a) Why can sterility testing alone not guarantee the sterility of a batch?
sterility means = absence of viable microorganisms
= because its not possible to test every container without damaging the product/ batch
= in reality only small sample (max. 20 sample size) get taken for testing
=> exponential killing (e^-x) of microorgamisms never reaches zero = never 100 % sterility possible -> D-value never zero
-> sterility has to be assured by the application of a suitably validated production process (geeignetes validiertes Produktionsverfahren)
b) product-specific suitability test
= to prove that the product doesn’t inhibit the growth of the 6 test microorganisms
=> product shouldn’t kill organisms that might me brought in a container during process (e.g. aseptic filling) to detect a contamination
c) minimum volume per container - liquid vaccine 1.2ml
=> liquids 1-40ml = half the contents of each container but not less than 1mL -> half the contents 1.2/2 = 0.6ml
=> 1 ml per container must be testes
What data are needed for validation with biological indicators?
Why is sterile compressed air for counterpressure used and not steam?
1) bioburden N0
2) type of microorganisms -> resistence: D & z-value
-> survival & kill-time
Why is sterile compresses air for counterpressure used and not steam?
= because to generate steam higher °C is needed which raises the pressure: p = p1 + p2 -> container may burst
(6) The autoclave shown is to be used for sterilization at 121 °C. As a starting situation, assume that water of suitable quality is present in the vessel, the items to be sterilized are positioned, the lid is tightly closed and the valve is open. Describe all further steps or processes required until sterilization is complete. Also address the issue of air removal. (3 points)
Describe all further steps or processes required until sterilization is complete. Also address the issue of air removal.
1) water heats up -> generation of steam (with gas burner)
2) steam rises up, reaching the lid -> steam moves down to sterilisation items
3) product is cold -> steam condensates
4) air removal = steam presses the air downwards & out of the system (via open valve)
5) atmopsheric pressure = air is eliminated & temperature can rise to 121 °C -> valve closes when all air is out
(14) Barrier systems (2.5 points)
(a) Why is a safety cabinet considered less safe as an open RABS?
(b) Which RABS type is most comparable to an isolator?
(c) When is overpressure used within isolators?
(d) How can it be tested whether an inert gas atmosphere is maintained in an isolator?
Sterility testing (3 points)
(a) Why are reference germs (Keime) required for sterility testing according to Ph. Eur. 2.6.1?
Name the two purposes for which they are used.
(b) How is the evaluation of the sterility test performed?
(c) A batch of an aseptically filled liquid product (small volume parenteral) consists of 800 disposable syringes. How many syringes must be tested for sterility according to Ph. Eur. 2.6.1 (see table)?
a) Reference germs = test organisms
= are required for sterility testing to ensure the effectiveness of the test results
= for positive control: to verify that the sterilization process is effective in killing or inhibiting the growth of microbes
= to proove that test organisms cannot grow in the nutrient medium or that the product doesn’t inhibit their growth
b) performance evaluation of the sterility test
= visual controll of turbidity after incubation of 2 weeks
c) Table 2.6.1
- more tha 500 containers
- 2% or 20 containers (10 containers for lang volume paranterals) whichever is less => 2% of 800 = 16 is less than 20
Sterility testing
(b) The culture medium plays a central role in sterility testing. Which two microbiological tests must be performed on each batch of culture media?
(c) An infusion solution (large volume parenteral) is to be tested for sterility according to Ph. Eur. 2.6.1. The filling volume per container is 250 ml. Using the following table from Ph. Eur. 2.6.1, indicate the minimum volume per container that must be used for sterility testing
b) microbiological tests of culture media (each batch):
1) Sterility testing = must be verified for each batch
-> incubation for 14 days & no visible microbial growth is allowed
2) Positive growth promotion test (with test microorganisms)
= proove that special test organisms cannot grow in the nutrient medium
c) Table Ph. Eur. 2.6.1
minimum volume per container - infusion solution (large volume parenteral) 250 ml
=> greater than 100ml = 10% of contents of the container (but not less than 20)
10% of 250ml = 25ml per container are used for sterility testing
(22) Static and dynamic seals (2.5 points)
(a) The illustration shows a static seal. When you take a look at the very left side of the seal, what two positive aspects with regard to hygienically designed static seals are fulfilled here?
(b) What is the role of the axial stop in (a)?
(c) What are the two general possibilities to seal the rotational movement of a stirrer shaft in aseptic processing?
(19) Fill and finish
(a) What dosing system is shown in the picture?
(b) Explain the principle of time-pressure dosing.
(c) Why does Annex 1 specify that crimping must be separated from filling and stoppering?
(d) Which test must be carried out on glass ampoules after fusion as a 100% check in addition to the visual inspection?
(e) What is the main advantage of using nested ready-to-fill syringes in tubs compared to using bulk syringes?
(f) How do you call the process for manufacturing infusion or parenteral nutrition bags?
(g) What is used to blow up the containers during the blow fill seal process?
a) (Dual) peristaltic pump
= single-use silicone tube -> tube get sandwiched between rollers (Dialyse-pumpe)
= benefits: single use/ high precision/ quick product change possible
Time-pressure system = gravity & viscosity dependent system
- benefit: ultra-high output/ CIP-SIP compartible
- not for: viscous products & products that are sensitive to silicone tubes
Principle:
- pressure inside vessel must stay constant -> precise monitoring of pressure
- pinched valves are opened & then after defined dosed product valves get closed
- temperature very important (high impact on viscosity) -> influences flow through
Crimping = aluminium cap gets placed on the stopper
• while crimping vials a lot of metal particles get generated -> non-viable particle => therefore, important that crimping is separated from filling area (physical separation)
=> to prevent particle contamination & good air extraction are required
-> closure by fusion, e.g. glass or plastic ampoules should be subject to 100% integrity testing.
bulk syringes = Washing, drying, siliconisation, sterilization necessary before filling
nested ready-to-fill syringes = just opening & filling & closure (ready to use) -> no washing & sterilization etc.
(f) How do you call the process for manufacturing infusion or parenteral nutrition bags? = form- fill- seal production
Blow/ fill/ seal system = purpose-built machine in which, in one continuous operation, containers are formed from a thermoplastics granulate, filled & sealed, all in one production line
Blow mould closes, bottom is welded -> blowing with sterilized compressed air
(16) Barrier systems (3 points)
(a) What is the difference between a passive and active RABS?
(b) Which cleanroom class must be used as the background environment for a RABS?
(c) What are the characteristics of an isolator called "open aseptic"?
(d) How do you perform the leak test of an isolator glove (in addition to the visual inspection)?
• Active RABS = integral HEPA-filtered air supply (HEPA inside)
• Passive RABS = air supply by ceiling mounted HEPA-filters (outside)
Open, active RABS
Open, passive RABS
• Surrounding room classification should be ISO 7 minimum in operation -> surrounding area of RABS = class B
Isolator surrounding = for open isolator class C & for closed isolator class D
Product protection isolators = “Aseptic isolator”
II) Open isolator systems = allows continuous material flow, hermetic seal for decontamination, with unidirectional air flow
= designed for continuous or semi-continuous ingress and/or egress of materials (Ein-/ Austritt) during operations through one or more openings -> mouse holes
= Openings must be engineered (e.g. using continuous overpressure) that no external air/ contaminant get into the isolator
-> has tiny openings (“mouse holes”) with inside overpressure to ensure that no particles get inside
-> min. grade C surrounding (because of holes)
2) Leak testing:
2.1) Pressure decay test = gloves get blowed up with pressure
2.2) airflow test at constant pressure = air gets inside of gloves & measurement of volume that is needed to keep pressure constant => if tight, no additional air needed
(20) Material and surface properties (1.5 points)
(a) An Ra value of 0.8 μm is specified for a surface. What does this mean?
(b) Name a method by which an Ra value of equal to or less than 0.8 μm can be achieved for a stainless steel surface.
(c) A material that comes into contact with a sterile product should be inert to the product. What else must the material be inert to? Specify one aspect.
(20) Specify two aspects that must be taken into account with regard to personnel during visual inspection. (1 point)
1) visual inspection is tiring for eyes -> looking 5 secons & making regular pauses
2) Personnel should follow hygiene practices, to prevent contamination of the products during visual inspection
3) Adequate and good lighting conditions are important for effective visual inspection
4) personnel friendly working condition (temperature/ humidity) are important -> creation of particles through sweating/ low concentration
(21) Media fill (2 points)
(a) During a media fill, the aseptic process defined in the SOP is tested for sterility. What must be considered in addition to the routine process when planning a media fill?
(b) In a media fill, 600 containers are filled with nutrient medium. How many of them are allowed to show germ growth after incubation?
(c) Which special aspects have to be considered for a media fill if a freeze-drying process is part of the procedure to be validated?
a) What must be considered in addition to the routine process when planning a media fill?
Media fill = should imitate as closely as possible the routine aseptic process & include all critical manufacturing steps -> microbial monitoring
= should consider various interventions known to occur during normal production & worst-case situations -> risk analysis
b) In a media fill, 600 containers are filled with nutrient medium. How many of them are allowed to show germ growth after incubation?
= no grotwoth (zero germs) -> otherwise something is not right in production
c) Which special aspects have to be considered for a media fill if a freeze-drying process is part of the procedure to be validated?
Lyophilizates -> partial drying process
- FDA recommends that unsealed containers be exposed to partial evacuation of the chamber that simulates the process.
- Vials should not be frozen, and all precautions should be taken so that microbial growth get supported (for evaluation)
(2)
For a steam sterilization process, the resulting probability of sterility is P = 0.9999998.
(a) What SAL value is achieved here?
(b) Is the SAL value achieved sufficient for a pharmaceutical steam sterilization process or not? Please justify.
a) SAL + Psterile = 1 = 100%
SAL = probability, that germs may be alive
Psterile = probability of germs that are death
1 - P = 0,00000002 SAL = 2*10^-7
b) Is the SAL value achieved sufficient for a pharmaceutical steam sterilization process or not? Please justify.
-> in pharmacy = aim to achieve SAL level of equal to or less 1*10^-6
= sufficient SAL is not achieved because not more than 1 viable micro-organism in 10^-6 sterilised items of final product should be found
-> of 1 million product containers 1 container is contaminated
=> SAL of 2*10^-7 means that of 10 million product containers 2 containers are contaminated
= is achieved because SAL calculated lower than 10^-6
(15) Name four other approaches, different in terms of protection mechanism, as an addition to the use of HEPA filters, how the open product in cleanroom class A can be protected from contamination?
1) physical protection
- Walls/ Barriers
- Shell concept: surrounding class B
2) aerodynamic protection
- protection via laminar air flow
- pressure cascade between zones
3) Personnel (human factor) = try to avoid human interaction through using automatic system
4) Use materials (incl. API & excipients) with very low bioburden
5) Surfaces in production surrounding = compatible quality/ cleaning/ roughness for desinfection
(3) Sterility testing - SAL
(a) What does the SAL value describe?
(b) What is the probability of sterility achieved if the bioburden before sterilization is 20 cfu/ml in a 5 ml volume and sterilisation results in a 7 log reduction?
a) What does the SAL value describe?
SAL = sterility assurance level = probability, that germs may be alive
SAL of a sterilation process = degree of assurance with which the process makes a population of items sterile
= is expressed as the probability of a non-sterile item in that population
b) What is the probability of sterility achieved if the bioburden before sterilization is 20 cfu/ml in a 5 ml volume and sterilisation results in a 7 log reduction?
(22)
(a) Name two further requirements in addition to easy cleanability & smooth surface for materials in contact with the product with regard to their hygienic design.
(b) What is designed in a hygienically unfavourable way here (see picture)? Name one point of criticism.
Requirements for materials - hygenic design
1) Smooth, easy to clean
2) Corrosion resistant -> stainless steel
3) Abrasion-proof, mechanically stable
4) Stable to aging & not change over time -> rubber may break in peaces
5) Inert (inaktiv) to the product & detergents and disinfectants
b) What is designed in a hygienically unfavourable way here (see picture)? Name one point of criticism.
- Sharp corners (≤ 90°) must be avoided
- surfaces hall always slope to one side with a minimum angle of 3°
(11) Sterile filtration (1.5 points)
(a) What is the typical pore size of filters for sterile filtration?
(b) It is to be examined whether product is lost during sterile filtration, i.e. whether active substance is adsorbed in the filter. What is the procedure for this adsorption test?
a) What is the typical pore size of filters for sterile filtration?
< 0.22 micron (or smaller)
b) It is to be examined whether product is lost during sterile filtration, i.e. whether active substance is adsorbed in the filter. What is the procedure for this adsorption test?
Adsorption test = for sterile filterable products
= checking the amount of product loss; muss be 100% (no loss)
-> check change in concentration through sterile filtration
-> measure concentration of API before & after filtration
-> calculation of recovery = must be 100% no change in concentration
recovery = Wiederfindung: amount of product after & before sterilization = should be 100%
(13)
(a) Which prerequisite must be fulfilled for an aqueous medicinal product to be prepared aseptically at all?
(b) The condition from (a) is given and a drug is to be prepared aseptically. However, it cannot be sterile-filtered.
What is the aseptic production process in this case?
a) Which prerequisite must be fulfilled for an aqueous medicinal product to be prepared aseptically at all?
1) sterile material & equipment
2) class A conditions
3) sterile transport
4) Isolator
5) sterile product
b) The condition from (a) is given and a drug is to be prepared aseptically. However, it cannot be sterile-filtered.
= product is not heat stable at all & cannot be filtered then: use pre-sterilised individual or mixed components & aseptic processing
(23) Hygienic design of plant components (4.5 points)
(a) Draw in a design improvement for the pictured container bottom.
(b) Name two criteria for the design of pipework that are not related to the properties of the material / surfaces.
(c) Sketch / draw the compression of a seal with an axial stop.
(d) In the picture you can see two possible sealings of the longitunial movement of a valve rod. Which one is better with regard to hygienic design / contamination risk? Please briefly justify your choice.
(e) Why is a simple mechanical seal not suitable for sealing agitator shafts?
a) Draw in a design improvement for the pictured container bottom.
• Self-drainable -> have special slope (are not straight)
• Fixed connections prefered to detachable ones
Avoidance of
• dead legs (no flow of media) Ú rule: L < 3 x D
• sharp-edged profile extensions & reductions (no corners)
• large-scale flow around / flow shadow (creates dead leg)
c) Sketch / draw the compression of a seal with an axial stop
d) Which one is better with regard to hygienic design / contamination risk? Please briefly justify your choice.
Sealing of longitudinal movements
a) dynamic sealing
b) better: hyrmetic sealing = because hermetically sealed & no germs can enter
Single mechanical seal
1) has a rotating stirrer shaft & rotating ring
2) in the housing is a fixed static ring -> while rotating these 2 parts sliding on each other & spring presses sliding ring against static ring
principle: 2 rings slide on each other (friction & heat creation)
= is NOT 100% sealed = not 100% safe: contamination get through sealing point
-> cannot be used for aseptic purposes & hazardous media -> used for non-sterile/ non-hazardous product/ pumps
-> microbial contamination can get through sliding point
(2) Sterility testing - SAL
(a) How are the SAL value and the probability of sterility related?
(b) For a batch size of 1,500,000 containers, how many non-sterile containers would be expected if the SAL value present would be 10^-5?
c) What is the acceptance criterion for the SAL value of a pharmaceutical steam sterilization process?
b) 10^-5 = 1 container of 100.000 is non-sterile
1,500,000 containers * 10^-5 = 15 containers are non-sterile
c) acceptance criterion for the SAL value
= min. SAL of 10^-6 for pharmaceuticals
-> in reality pharma companies chieve SAL of at least 10^-12
(4) Sterility testing
The following data on heat-killing of a microorganism at a temperature of 125 °C are available to you:
(a) What D value at 125 °C results from the data?
(b) After what total time at 125 °C is a SAL of ≤ 10^-6 achieved?
(c) Is the F0 value reached at the time point 13 min
- below 13 min,
- exactly 13 min or
- above 13 min?
Please justify your choice.
(3 points)
a) 13 -1 = 12 12/8 (log-levels) = 1,5 D-value
b) D-value = 1.5 -> from 10^8 to 10^-6 = 14 log levels
1.5 * 14 (log levels) = 21 minuten
c) above 13 mins -> F0 at 121.1°C
(5) Sterility testing - z value
(a) A mean z-value of 5.5 °C is usually assumed for vegetative
microorganisms. What does this z value of 5.5 °C indicate?
(b) How are the F0 value and the SAL value related?
a) What does this z value of 5.5 °C indicate?
z-value of 5,5 °C = temperature to reduce cfu of factor of 10 log levels
b) How are the F0 value and the SAL value related?
F0-value = special F-value for 121 °C conditions
-> the higher F-value = the lower SAL-value
(4) Complete the appropriate terms in the fields with a grey background. (3 points)
The F-Value has the unit … minutes
The higher the F value, the … slower is the killing of germs.
The highert the z value, the … more resistant are the germs
The lower the F value, the faster is the SAL value achieved
With increasing temperature the D-value … decreases
The F0 value is always connected with a temperature of … 121.1°C (for 1 min)
The larger the F value, the lower is the SAL value achieved
-> high °C results faster killing & a less D-value
a) For a pharmaceutical steam sterilization process, a SAL value of 1.4 * 10^-6 is achieved. Evaluate whether this process meets the specifications of the EMA.
Note: An evaluation includes a justification. (1.5 points)
(b) What is the probability of sterility after a single germ has been subjected to a process that reduces it by one log level?
a) SAL value of 1.4 * 10^-6 is NOT achieved. = 1.4 container of 1 mio non-sterile
EMA - specifiation: 1* 10^-6 = max 1 container of 1 mio non sterile
= the process doesn’t meet the specification of EMA because 1.4 is higher than 1
b) cfu/ml = 1
1 - log (10) = 0
Probability of sterility = 100%
a) For a sterilization process, the SAL value achieved is 1.8 * 10^-11.
What was the initial bacterial count N0 before sterilization if this SAL value was reached after a reduction by 13 log steps?
b) For a sterilization process, the SAL value is 3 * 10^-2. What is the resulting probability of sterility in percent?
a) SAL 1.8 * 10^-11
Redution: 13 log steps
initial baterial count N0 = SAL + log steps = 1.8 * 10^2
b) Probability in percent:
SAL + Pst = 1
Pst = 1 - SAL = 1 - (3 * 10^2) = 0.97 = 97%
(a) The following z value (temperature coefficient) is given in connection with a steam sterilization process: z = 6 °C. What is the D value at 121 °C if the D value at 109 °C is exactly 30 s?
(c) For a steam sterilization process at 134 °C, an F0 value of 18 min is achieved. How long would you have to sterilize at 121.1 °C to achieve the same lethality?
Formula:
a)
c) Formula cannot be used because no z value given!
(13) A stainless steel format part is needed in an isolator for aseptic filling of a medicinal product.
(1) The format part is loaded into a double-door washing machine in class D and then removed in class C.
(2) There, it is packed in a Tyvek bag and loaded in a double-door autoclave.
(3) The sterilized bag with the format part is removed from the autoclave in the filling room class C.
(4) The bag is placed on the work surface of an isolator and the interior of the closed isolator is biodecontaminated with H2O2 vapour.
(5) The bag is opened and the format part is then installed aseptically in class A.
How would you evaluate the process of preparing and transferring the format part: Are all steps (1) to (5) in order with regard to the requirements of Annex 1 and a possible risk to the medicinal product in Class A? Is there potential for optimization? If yes, which one? (2 points)
(8) z-values
How do the z-values for dry heat differ from those for steam sterilisation? Give a reason for the difference as well. (1 point)
How do the z-values for dry heat differ from those for steam sterilisation? Give a reason for the difference as well.
dry heat = 160° for 125 min
steam sterilization = 121.1°C for 1 min
Effectiveness = dry heat is less effective than steam sterilization at the same temperature because no condensation!
because:
1) microorganisms/ spores are more heat resistant under dry conditions (= D-values are approx. 10 times higher at the same temperature) => z-value 10°C
2) dry air has a lower heat capacity & poorer heat transfer and a higher risk of cold islands
- z-values about twice as high compared to steam sterilization because higher D-value necessary -> less effective/ no condensation
- FH value (instead of F0) - for dry heat sterilization
(7) Dry heat sterilization (1.5 points)
(a) What is meant by compensation time in (dry) heat sterilization processes?
(b) The D value for heat-resistant endotoxins is around 50 °C. To achieve the required 3 log reduction of endotoxins the heat treatment is performed at 280 °C for 3 min. How long would the process duration be at 330 °C to achieve the same effect?
(19) Media fill (3 points)
(a) Media fill is also known as APS. What does APS stand for?
(b) How often must a media fill be performed as part of the introduction of a new product?
(c) An SOP for the aseptic formulation and filling of a liquid product specifies that two excipients and WFI should be added to the active ingredient while stirring gently. How do you take these specifications into account for a media fill?
(d) How long does incubation take after a media fill and how are the incubated containers subsequently analyzed?
(5) What role does the bioburden of the sterilization items play in the design of a sterilization process? (1 point)
What role does the bioburden of the sterilization items play in the design of a sterilization process?
the lower the bioburden that easier is it to guarantee sterility
to minimise the bioburden several precautions are necessary e.g.:
- use of components with low degree of microbial contamination
- microbial monitoring
- row materials with low bioburden
- max. bioburden must be defined (via validation)
reduction of bioburden includes washing/ cleaning/ desinfection & sterilization in the end
(8) Dry heat sterilization (3 points)
(b) In a pharmaceutical plant, a hot air sterilization tunnel is used to sterilize glass cartridges.
In addition to sterilization and depyrogenation, what third purpose is met by the treatment in the hot air sterilization tunnel?
(c) The z value for heat resistant endotoxins is about 46 °C. What does this mean when you compare the required process times at 280 °C and 326 °C?
(d) Why is it important to cool glass containers slowly after hot air treatment?
a) What is meant by compensation time in (dry) heat sterilization processes?
Compensation time = time that is needed to reach 121°C from the
reference measuring point inside chamber & all points of product (even coldest spot)
= important to guarantee everywhere 121 °C
= should be as short as possible for good heat transfer
b) In addition to sterilization and depyrogenation, what third purpose is met by the treatment in the hot air sterilization tunnel?
= siliconization : silicon layer (anti-friction coating) is made up to minimize break loose & gliding forces
-> for cartridges. syringes = have stopper inside, it has to move while injection
-> silicone oil or emulsion is sprayed to the inner sruface of syringe
c)
d) Why is it important to cool glass containers slowly after hot air treatment?
= to prevent thermal stress and potential breakage
= to avoide class breakage (due to possible cracks)
(7) Sterility testing
(a) Describe the venting process of a steam sterilization process using the fractionated vacuum method.
(b) How is container bursting prevented in the hot water spray process?
II) fractioned pre-vacuum process
= for special products with very difficult air elimination
- replay change of vacuum phase & steam introduction
- steam gets introduced to reach desired °C
- fractionated cooling & drying: vacuum & air
= through creation of counterpressure by introducing sterile compressed air into container
Sterility testing(7)
(a) For which sterilization goods is the steam-air mixture process used?
(b) Why does the steam-air mixture process use a steam-air mixture and not pure saturated steam?
a) For which sterilization goods is the steam-air mixture process used?
1) steam/air mixture process
-> higher °C to create steam may have negativ impact on product
steam-air mixture allows better control over temperature. Pure saturated steam can have a high temperature & cause damage on produts. By introducing air into the steam, the overall temperature can be moderated to prevent overheating
addition of air to the steam helps to improve heat penetration and distribution throughout the load. Air is a less efficient conductor of heat compared to steam, and the presence of air pockets can aid in achieving more uniform heat distribution within the sterilization chamber
(10) In the diagram the sequence of an ethylene oxide sterilization is shown.
(a) Why are the vacuum-nitrogen cycles required?
(b) What is the purpose of the dwell period?
(c) Name one advantage of ethylene oxide sterilization over hydrogen peroxide sterilization.
ETO = Extremely flammable, forms explosive mixtures with air (O2)
-> therefore O2 needs to be brought out of chamber by vacuum & for gas sterilization is nitrogen used
-> through pulsation vacuumed input of nitrogen air gets better removed (better displacement)
dwell-period = Sterilization time = exposure time of product under constant pressure & temperature
Advantages - ETO
1) Broad spectrum of activity
2) Excellent penetration capacity
Hydrogen peroxide (H2O2) - Wasserstoffperoxid
• Very strong oxidizing agent
• Broad antimicrobial spectrum
• Penetration capacity limited
• Highly corrosive
(6) Sterility testing
(a) Explain the effect of air pockets within the sterilization load during steam sterilization.
(b) How is the pressure in an autoclave related to the amount of noncondensable gases in the autoclave chamber?
(c) How is venting / air elimination performed in a steam sterilization process that uses the gravity or flow principle?
-> when saturated steam is coming into an autoclave = it reaches a cold product surface & gets condensate because the steam cools quickly down & transfers all its saved energy to the product -> so product heats up
-> condensation = very fast & powerful energy transfer process
-> air prevent steam for reaching the product -> bad sterilization
b) How is the pressure in an autoclave related to the amount of noncondensable gases in the autoclave chamber?
limit < 40ml/kg non-condensable gases
3) Pressure increase: In a closed system the pressure rises up (because of high energy & steam needs more space) so the vaporization curve reaching 212°C and it becomes a saturated steam
-> the lower the pressure, the lower amount of non-condensable gases
= if pressure rises to high, then there is a lot if air left causing overpressure
4) Elimination of air -> by gravity/ flow principle
- In the beginning product is cold & contains a lot of air
- air must be eliminated otherwise steam cannot reach product; no condensation
- air is 1,5 times heavier than the steam = moves down & get pressed out of the system
- it reaches air separator (Z) with a valve -> at 100°C no cold air flows out & valve closes
5) more steam generation -> pressure inside rises to achieve p = 2 bar & 121,1 °C
(11) Dry heat sterilization
In a pharmaceutical plant, a hot air sterilization tunnel is used to sterilize glass cartridges.
(a) In addition to sterilization, what two purposes are met by treatment in the hot air sterilization tunnel?
(b) Give a common operating temperature for the sterilization zone.
(c) Which zone of the hot air sterilization tunnel has the highest ambient pressure?
a) in hot air tunnel:
1) Depyrogenation (while sterilization phase) = to reduce heat resistant endotoxins
2) seliconization = silicon layer is spread / made up to reduce friction (for glass or in syringes)
b) Give a common operating temperature for the sterilization zone.
= 280 - 350°C
c) pressure zones in hot air tunnel:
1) washing zone = +2 Pa (refer to class C pressure)
2) heating zone = +4 Pa (sterilization)
3) cooling zone = +6 Pa (class A conditions) = to make shure that no air is getting inside sterilization tunnel
(9) Ionizing radiation sterilization (1.5 points)
(a) Which type of electromagnetic radiation used for sterilization has the highest frequency?
(b) Name one disadvantage of electron irradiation over gamma irradiation.
(c) In which unit do you specify the dose for radiation sterilization?
24) What is the advantage of electron radiation over gamma radiation?
= gamma irradiation
b) Name one disadvantage of electron irradiation over gamma irradiation.
I) Electron irradiation - Characteristics
• Limited penetration, sterilization in final pakaging possible
• High dose rate
• No residues
• More gentle than gamma irradiation
II) Gamma irradiation - type of ionizing radiation: Annex 12 GMP
Characteristics
• High penetration capability sterilization of products in final packaging
• suitable for products with high density and large volume (e.g. whole pallets)
• Rather low dose rate
• No residues -> Problem of radio active waste
• Not suitable for all materials (cross-linking / degradation reactions of polymers)
Dose - ionizing radiation: Gy = Gray
• Target absorbed dose: ≥ 25 kGy (very high!) for def. SAL
• Unit: 1 Gray = 1 Gy = 1 Joule / kg = 1 Nm/ kg = 1 m^2/ s^2
What is the advantage of electron radiation over gamma radiation?
= no radioactive waste & high dosage rate
(10) Ionizing radiation sterilization (1.5 points)
(a) What is the common mechanism of action of all types of ionizing radiation sterilization?
(b) For pallet sterilization of consumables, a radiation sterilization process with the highest penetration capacity should be selected. Material compatibility is given. Which type of radiation can be considered besides X-ray?
(c) Name an area of application for decontamination with UV radiation.
= induction or damage to the genetic material (DNA or RNA) of microorganisms.
- Ionizing radiation has enough energy to ionize atoms/ molecules , leading to the formation of highly reactive ions / free radicals. These can then interact with and damage the genetic material of microorganisms, making them unable to reproduce & causing their destruction.
Gamma irradiation = High penetration capability sterilization of products in final packaging
= suitable for products with high density & large volume (e.g. whole pallets)
= in healthcare (hospitals) for disinfection of surfaces and air to reduce the risk of healthcare-associated infections
(2) Plot a typical heat killing curve of microorganisms (normal, non-logarithmic scaling of the axes) with a decimal reduction time of 1.5 min. (1.5 points)
(b) According to which mode of action does UV radiation, which is not one of the ionizing types of radiation, kill microorganisms?
(c) Which of the ionizing radiation types does not belong to the electromagnetic radiation?
= if it is NOT possible to sterilize pharmaceuticals by heat or steam, then ionizing radiation sterilization may applied
-> for: pharmaceutical medical products; dry powder products, non-aqueos liquids & semi-solids if the product is NOT temperature stable!
e.g. ophthalmic ointments in tubes
I) UV irradiation - type of NOT- ionizing radiation
• Disinfection of smooth surfaces, water and air
• Strong absorption, therefore low penetration depth
• Damages DNA (maximum at 265 nm) -> kiling of germs
7)
a) Why is there a limit for non-condensable gases for pharmaceutical pure steam?
b) What is the key advantage of using saturated steam compared to superheated steam?
a) pure steam = has special purity & high quality so it can get in contact with pharmaceutical
-> if there is a lot of air or non-condensable gases inside sterilization system the steam cannot reach every area of product
=> results: bad sterilisation quality
- no condensation possible
- bad energy transfer -> needed °C cannot be reached for desired duration
b) saturated steam = condensates quickly -> shorter process time
(super-heates steam = needs to cool down to condensate -> longer)
(12) Ionizing radiation sterilization (1.5 points)
(a) Which two irradiation methods are described in Annex 12 of the GMP guide and are thus applicable for terminal radiation sterilization?
(b) Irradiation is also used for sterile transfer, among other things. Give a specific example of use.
a) Which two irradiation methods are thus applicable for terminal radiation sterilization (Annex 12)?
1) electron irradiation
2) gamma irradiation
b) Irradiation for sterile transfer
= E-beam tunnel -> transfer from class C to class A
(11) Sterile filtration (2 points)
(a) When may sterile filtration be used to sterilize a liquid medicinal product?
(b) As part of the validation of a sterile filtration process the bacterial challengetest (BCT) must be performed. How does it work?
Note: the viability test doesnot need to be addressed.
= if product is not heat stable at all, use sterile filtration
= if steam sterilization is not possible then sterile filtration with microbial retentive filter is the last option
=> must be combined with aseptic processing
with presterilized containers -> only final sterile filtration & filling is done in class A
Bacterial Challenge Test, BCT – biological test for filtration
1) Preparation: performance of Viability test (with reference microorganism Brevundimonas diminuta)
= checking if product has bactericidal effect -> germ killing 1log level in 45mins
-> if product is bactericidal = then it needs to get filtered first & then with adding germ suspension
-> if product isn’t bactericidal = germs get added generating a solution with > 107cfu/cm² filter area
2) Provide filtration process = under worst case conditions (with product, filter elements, contact time, pressure, temperature)-> check the filter retention capacity = validation of retention of germs on membrane filters
3) Checking sterility after filtration = 100 % retention of germs (acceptance criterion) -> no single cell of reference germ should pass the filter!
(10) Gamma irradiation
Gamma irradiation and gassing with ethylene oxide are often used for the sterilisation of entire pallets with consumables.
(a) What is the mechanism of action of gamma irradiation?
(b) Give a reason why gamma irradiation is preferred over electron irradiation for this application.
(c) Name one advantage and one disadvantage of gamma irradiation compared to sterilization with ethylene oxide.
(d) Give a reason why ethylene oxide is used for that application but not hydrogen peroxide.(2.5 points)
= use of high-energy gamma rays to disrupt the DNA or RNA of microorganisms, making them incapable of reproduction
- Generation of Ionizing Radiation: = cobalt-60 creates during radioactive decay high-energy gamma rays
- Penetration of Materials = Gamma rays have high energy and short wavelengths, allowing them to penetrate deeply into materials (sterilization)
- Ionization of Atoms & Molecules = As gamma rays pass through a material, causing ionization of atoms -> results formation of highly reactive ions & free radicals -> killing germs
electron irradiation = Limited penetration, but sterilization of products in final packaging possible
Gamma irradiation = High penetration capability sterilization of products in final packaging -> higher assurance level
Gamma Irradiation
High penetration capability sterilization of products in final packaging & no residuces
Problem of radio active waste
2) Alkylation of nucleic acids, proteins (requires H2O)
3) Excellent penetration capacity
Disadvantages - ETO
1) Carcinogenic
2) Extremely flammable, forms explosive mixtures with air
3) Leaves toxic residues -> not allowed in food sector
(d) Give a reason why ethylene oxide is used for that application but not hydrogen peroxide = sterilisation of entire pallets with consumables.
because: ETO has excellent penetration capacity & broad spetrum of activitity -> better for entire pallets with undefined consumables
hydrogen peroxide
= Very strong oxidizing agent
= Highly corrosive
= Penetration capacity limited
(17) Fill and Finish (4.5 points + 0.5 bonus points for part h)
(a) Why is weigh dosing the preferred method for aseptic filling for very high-priced products?
(b) What dosing system is shown in the picture?
(c) How are glass ampoules sealed after the product has been filled?
(d) Which test must be carried out on all glass ampoules as a 100% check after sealing, regardless of whether the product is classified as a parenteral product or not?
(e) You should plan a crimping process as part of a filling and closure line. Specify two aspects in addition to the supply of sterile caps that must be taken into account for the crimping process.
(f) How is the plastic tube produced in the BFS process?
(g) What is the minimum cleanroom class for filling terminally sterilized products?
(h) Bonus question: What is the name of the illustrated method for transferring tubs to class A, which requires double outer packaging of the tubs? Hint: the method is abbreviated NTT.
(13) Chemical sterilization (1 point)
(a) What is the advantage of gas sterilization with hydrogen peroxide, which operates below the saturation concentration of H2O2 and thus without micro-condensation?
(b) Peracetic acid can also be used for gas sterilization. Give one reason why hydrogen peroxide is preferred over peracetic acid.
(a) advantage of gas sterilization with hydrogen peroxide, which operates below the saturation concentration of H2O2 and thus without micro-condensation:
= faster sterilization because no condensation
b) Give one reason why hydrogen peroxide is preferred over peracetic acid.
Peracetic acid = bad smell
= leaves residues (organic produts) that may get i high dooese toxic or have a negative impact on produt
hydrogen peroxide = doesnt leave any persistent chemical residues
= environment friendly
(11) Sterile filtration
What is the so-called bubble point of a sterile filter and how is it determined? (1.5 points)
What is the so-called bubble point of a sterile filter and how is it determined?
1) Bubble point test (manual application)
= Membrane gets wetted with liquid product & pressure (air) gets applied from one side of the filter to press out the solvents out through the filter
-> Solvent is pressed out of filter with increasing pressure
Bubble Point = pressure at which a continuous discharge of air bubbles can be detected (3 to 5 bar)
= depends on the filter properties
- big leak = lower pressure
- the higher pressure = the smaller the pore size
(a) How can you draw conclusions about the integrity of a sterile filter from the measured value of the bubble point?
(b) How is possible product loss during sterile filtration investigated?
- the higher the bubble point / pressure = the smaller the pore size
- The smaller the average pore size of a filter, the higher the pressure. If there is a leak in the filter the pressure (bubble point) stays low
with Adsorption test = for sterile filterable products
= measure concentration of API before & after filtration
-> calculation of recovery = bust be 100% no change in concentration
Plot a typical heat killing curve of microorganisms with a decimal reduction time of 1.5 min. Start with a bioburden of 10^6 and end the curve after 9 min. Note: Do not forget to label the axes and use logarithmic scale for the y-axis.
(14) In preparation for aseptic filling of a medicinal product, the formulated product is usually first subjected to germ-reducing filtration and then finally to sterile filtration in class A.
(a) Why does the formulation of the product have to be carried out at least in clean room class C, when subsequently two filtration steps are carried out to achieve sterility?
(b) What must be checked on each filter before and after use?
= to ensure a low & controlled bioburden before getting to the filtration
-> according to EMA-guidline = Frequent bioburden limit: ≤ 10 CFU / 100 ml -> to fulfill this clean room conditions necessary
= the lower the bioburden, the slower cake formation during filtration (during dead-end filtration)
= Filter integrity testing– GMP Guidline
= The integrity of the filter must be checked before & after use
-> after usage = easy to perform -> filter can get taken out for a quality control in lab or in production area
-> before usage = is critical because it must be assured that filter doesn’t get contaminated -> 2 possibilities to keep filter sterile:
1) sterilization in autoclave after filter testing -> validation important
2) filter gets installed in production area & gets sterilized in place -> filter testing can be done in place
(8) Dry heat sterilization (2.5 points)
(a) What can be criticized about the loading of this hot air sterilization oven?
(b) For depyrogenation, hot air sterilization tunnels must be operated at over 220 °C according to the pharmacopoeia. The z value for depyrogenation is around 50 °C. What does this mean when you compare the required process time at 320 °C and 220 °C?
load should be putted in right position in the autolave (straight)
air distribution = Air must be able to circulate freely
-> metal sterilization cassettes with holes = product should be placed correctly
= process needs longer at 220°C
D value increse when °C insrease
(7)
(a) For which pharmaceutical products can the hot water spray process be used?
(b) How is the counterpressure built up in the hot water spray process to prevent the product containers from bursting?
(c) In which steam sterilization processes is the Bowie-Dick test used?
a) For which pharmaceutical products can the hot water spray process be used?
Hot water spray = Counter pressure sterilization processes
= for aqueous liquids (infusion) in closed containers
= e.g. for bottles, vials made of glass/ plastics & single-multig bags
= compressed air gets introduced in the chamber to build up the counterpressure
= for saturated steam sterilisation with vacuum processes
= proof if sufficient air removal achieved & steam penetration into product sufficient
(18) Preparation of equipment and primary packaging material
(2 points + 0.5 bonus points for part d)
(a) Why is primary packaging material considered more critical then secondary packaging material?
(b) Name one way in which two silicone tubes can be connected aseptically.
(c) Why are cartridges siliconized on the inside?
(d) At which point in the process does siliconization take place?
= its the first layer of packaging that is in direct contact with pharmaceutical product (e.g. vials/ ampoules/ syringes/ stoppers/ caps)
-> shouldn’t cause recontamination or have negative impact on product -> must be sterile (depyrogenation) / inert/ abrasive proof etc.
Disposal systems: Aseptic connection
Aseptic connection via
1) Sterile connectors/ couplings = presterilized system with membraned (gets taken out) for product transfer
2) Silicone hose connectors = used in class A environment of 2 sterile tubes
3) Sterile welding of thermos plastic tubes = 2 tubes get brought together & closed in a special machine (welder) -> inside machine: tubes get cut & welded to each other (+200°C) -> class A air supply
= siliconisation with PDMS = polydimethylsiloxane
• very important for Syringes & cartridges
= get covered with anti-friction coating (silicone) to minimize break loose & gliding forces
without siliconization stopper doesn’t move at all due to high friction or stopper suddenly moves & stops -> syringe doesn’t work well, no correct dosage (insulin) -> dangerous for patient
break loose force = force that is needed to start movement of a stoper
gliding force = force that is needed to keep stopper moving
1) siliconisation of packaging (syringe/ cartridge) usually before sterilisation (after washing & drying) -> often applied backed on siliconization which has formation of silicon layer during (hot air) sterilization
2) siliconisation of syringe & cartridge stoppers = optionally after sterilization -> stoppers must move to achieve equal distribution of silicone
(10) Gas sterilization (2 points)
(a) What steps are needed to biodecontaminate an isolator with hydrogen peroxide?
(b) Why should condensation of H2O2 be prevented if possible?
VHP = vaporized hydrogen peroxide = surface sterilization process / used for biodecontamination -> decrease in 10log levels
General procedure/phases of VHP-process
= in isolators for decontamination/ for transfer airlocks
1) Dehumidification = same starting humidity is important
= reduce of humidity with dehumidifier because H2O2 vapours & creates moisture
2) Conditioning = H2O2 vaporises & gets in sterilisation chamber = concentration of H2O2 & humidity rises
3) Decontamination = desired H2O2 concentration is reached & kept on the same level through adding more H2O2
-> H2O2 concentration must be below 100% saturation, to avoid condensation on the surfaces
4) Aeration (Belüftung) = vapor (gaseous H2O2) can get easily out of the system (as liquid) enabling aeration with fresh air from outside = shorter process time
1) because drops on surface of product increase corrosion
2) duration process takes much longer if H2O2 gets to vapor
(3)
You have the following data on the heat killing of a micro-organism at a temperature of 121 °C: a germ population is reduced by 6 log levels in a period of 180 s.
(a) What D value at 121 °C results from the data?
(b) After what total time is a SAL of 10^-12 achieved if 10000 germs are present at the beginning?
a) What D value at 121 °C results from the data?
b) After what total time is a SAL of 10^-12 achieved if 10000 germs are present at the beginning?
For a sterilization process, the SAL value is 5 * 10^-7.
(a) What was the initial bacterial count N0 before sterilization if this SAL value was achieved after a reduction of 8 log steps?
(b) What is the probability of sterility in percent associated with the SAL value?
a) What was the initial bacterial count N0 before sterilization if this SAL valuewas achieved after a reduction of 8 log steps?
b) What is the probability of sterility in percent associated with the SAL value?
(12) Sterile filtration (2.5 points)
(b) In preparation for the aseptic filling of a liquid medicinal product, the drug is usually subjected to two filtration steps. The second step is sterile filtration. What is the purpose of the first filtration step?
(c) Sterile filters must be examined for leachables. What are leachables?
bioburden reduction prefilters and/or sterilising grade filters may be used at multiple points during manufacturing process to ensure a low & controlled bioburden before getting to the final sterilising filter
Purposes:
- to prevent the build up of filter cake, first filter takes the main germs load out of product
- if the first filter is having a lack the second filter enables the filtration (safety of process)
- increases the process safety to ensure a sterile product
- separation of the 2 steps is possible, so one is done in class C area & the other in the final filtration
Leachables (if required) = only required if toxic extractables detected
= Substances leached from the filter material in contact with the product under standard process conditions
= substances that can migrate or leach from materials used in the manufacturing process into the final product
-> for sterile filters, leachables refer to any chemicals or substances that may be released from the filter material and contaminate the filtered product.
(14) Sterile filtration (2 points)
(a) How is the bubble point of a sterile filter related to the pore diameter?
(b) In preparation for aseptic filling of a drug product, the drug is usually first subjected to germ-reducing filtration and then finally to sterile filtration in class A. Give two reasons why an additional germ-reducing filtration is carried out.
a) bobble point & pore diameter
bubble point = pressure at which a continuous discharge of air bubbles can be detected
=> the smaller the pore size = the higher the pressure (3-5 bar)
= the higher the bubble point
b) Give two reasons why an additional germ-reducing filtration is carried out.
bioburden reduction prefilters & sterilising grade filters
1) to prevent built up of the filter cake
2) if 1-st filter is having a leak the 2-nd filter enables filtration = increase safety of sterility
( 3) separation in 2 steps possible: one in class C area & one in final step)
(5) Why is "packaging material penetration" an important characteristic that a sterilization medium must fulfil? (1 point)
5) Why is "packaging material penetration" an important characteristic that a sterilization medium must fulfil?
= packaging material must be suitable for sterilization medium
to ensure the efficacy & reliability of the sterilization process
to ensure that every part of packed product get reached by sterilization medium (e.g. steam/ radiation)
for qualified sterilization so that germs can survive
(9) Dry heat tunnel (2 points)
(a) Why are hot air sterilization tunnels usually operated at temperatures of 300 - 350 °C, when according to regulations 160 °C would be sufficient for dry heat sterilization? Give two reasons.
(b) Which zone of a hot air sterilization tunnel has the lowest ambient pressure?
(c) Why is it important that glassware cools slowly after hot air sterilization?
a) Why are hot air sterilization tunnels usually operated at temperatures of 300 - 350 °C?
1) to eliminate pyrogens (depyrogenation)
2) because its a (faster) process resulting better killing & higher sterility assurance
b) Which zone of a hot air sterilization tunnel has the lowest ambient pressure?
= the drying/ preheating zone (class c) = +2Pa
-> highes pressure in cooling zone (after sterilization to ensure sterility)
c) Why is it important that glassware cools slowly after hot air sterilization?
(8) Dry heat sterilization (1.5 points)
Hot air sterilization tunnels are used to transfer washed glass containers to the aseptic filling area.
(a) Name two approaches to prevent particles / microorganisms from entering the aseptic area from the glass washer area in addition to the high temperature.
(b) The EMA specifies a minimum temperature of 160 °C for hot-air sterilization, but the pharmacopoeia requires at least 220 °C for hot-air sterilization tunnels. What is the objective (Ziel) of the higher temperature in the pharmacopoeia?
a) Name two approaches to prevent particles / microorganisms from entering the aseptic area from the glass washer area in addition to the high temperature
1) pressure cascade
2) Forced air circulation with HEPA filtered hot air (laminar airflow (LAF) = to remove particles & to control pressure conditions
b) What is the objective (Ziel) of the higher temperature in the pharmacopoeia?
= to ensure a more effective sterilization process
= to enable depyrogenation which starts above 220 °C (for glass) to reduce heat resistant endotoxins
(4) Sterility testing - z value
The following diagram shows the killing curve of a microorganism (z = 5 °C) at a temperature of 121 °C.
(a) Which D value at 121 °C results from the diagram?
(b) What is the D value at a temperature of 131 °C?
calculate: D-value = mins/ log-levels = 8mins/ 4 log levels = 2 mins
or read in diagram
b) What is the D value at a temperature of 131 °C?
(5) F0 & z-values
(a) By a holding time of 30 seconds at an unknown sterilization temperature, an F0 value of 50 min is achieved. The z-value is 5.2 °C. What is the sterilization temperature of the process?
(b) How is the achieved F0 value of 50 min to be interpreted?
a) = 30 seconds
(4) z-value & D-value
(a) A low bioburden before sterilization is not only important to achieve the shortest possible sterilization time and the highest possible sterility assurance. What other reason is there?
(b) How are the D-value and the z-value related?
Prevention of undesirable By-Products -> metabolic-products of germs may begatively influence the quality of product
low bioburden is advantageous for sterile filtration
-> filter capacity = with high bioburden gets quickly blocked by germs (cake formation)
-> depending on amount of bioburden the filter must be huge enough
= Temperature change in °C, which is required to alter the D value by a factor of 10.
= temperature dependency of D-value
(12) Validation of sterilization processes (2 points)
(b) What is checked during the OQ of a steam sterilizer?
(c) What is the acceptance criterion for the Bacterial Challenge Test (BCT) that must be performed as part of the validation of a sterile filtration process?
Parametric release – Ph.Eur.5.1.1
= can be applied when for the terminal sterilization of pharmaceuticals steam sterilization, dry heat sterilization or ionising radiation are used
-> Precondition: sterilization processes must be validated & be reliable = to get permission by authorities
What is parametric release?
= Release of terminally sterilized products without batch-related sterility testing -> for results of sterility testing 14 days needed
= release of product is based solely on the measurement of physical process parameters -> monitoring of parameters
= possible because standard conditions are defined via regulations & are performed in reality with overkilling phase (far above minimal requirements, reaching low SAL) which guarantees sterilisation safety
-> standard conditions (121 °C / 15 min steam sterilization) represent overkill = killing efficiency sterilization safety
-> But must be approved by the authorities
b) What is checked during the OQ of a steam sterilizer?
OQ = Operational Qualification
= Checking basic functions of equipment within the working area including chamber integrity (pressure & vacuum test)
(c) What is the acceptance criterion for the Bacterial Challenge Test (BCT)?
Checking sterility after filtration = 100 % retention of germs (acceptance criterion) -> no single cell of reference germ should pass the filter!
(17) Transfer systems (2 points)
(a) Describe the principle of a transfer using a decontamination airlock into an isolator during operation.
(b) What transfer system is shown in the picture?
Decontamination airlock (Luftschleuse)/ transfer hatch
= has 2 decontamination airlocks (small chambers) on each side of the isolator that are connected to working area
-> material can get inside & out of the isolator
-> double door system
RTP = Rapid Transfer Port
= for transfer of materials, stoppers, caps into aseptic area while processing
- double door transfer system
- consists of Alpha-port which is installed in the wall of isolator/ RABS
- for transfer a Beta-container/ bag gets connected to Alpha-port
- beta-container fits exactly to alpha-port
Principle
1) Beta container = stainless steel with pipe connector & hydrophobic sterile filter
2) container gets autoclaved with vacuum
3) Connection of Isolator + container -> connected lids get opened from inside for material transfer => lids must be airtight connected -> by rotation 60°
(18) Transfer
(a) Which two transfer technologies are best suited for transferring crimping pliers into an isolator during operation?
(b) E-beam tunnels are one of the technologies used to transfer presterilized prefilled syringes in tubs to the filling area (class A).
How is the ingress of germs from the loading area (insertion of the still film-wrapped tubs) into the filling area (Class A) prevented?
1) RTP = Rapid Transfer Port = double door transfer systems
= dog removable storage/ containers bag -> not directly connected to isolator, connection via alpha port
= for transport of stoppers, caps & Liquids
2) Decontamination airlock
(b) E-beam tunnels: How is the ingress of germs from the loading area (insertion of the still film-wrapped tubs) into the filling area (Class A) prevented?
= through pressure cascade & physical barrier (walls)
(15) Barrier systems (3 points)
(a) What characterizes an open, passive RABS?
(b) What characterizes an open isolator?
(c) When is underpressure used within isolators?
(d) How often / at what frequency should the leak test of an isolator glove be carried out?
- Negative pressure = operation for toxic/ hazardous products
= MUST be done at each batch change before every biodecontamination -> prove if isolator & gloves are tight/ closed
(16) Barrier systems (2 points)
(a) Which RABS type is shown in the figure below and what is the main difference to an isolator?
(b) How often / at what frequency should the leak test of an isolator glove be carried out?
open = air circulation between zones (A to B)
active = HEPA filter Unit inside
= open, active RABS
Main difference to Isolator = surrounding environment ISO 7 which is equivalent to class B -> Isolator has an surrounding environment of C/D
(19) Validation of sterilization processes
Preparation of equipment and primary packaging material (2 points)
(a) What is measured to monitor CIP processes?
(b) Sterile couplings are used for single-use systems. What is their purpose?
(c) In addition to the glass body, which parts of a finished syringe are also siliconized?
= is a Validated automatic cleaning without disassembly
-> Mobile or fixed systems with connection to SIP
Monitoring = measurement of Conductivity (possibly also pH / TOC = total org. carbon) in the effluent (Abwasser)
Conductivity =property of water to conduct electrical current. Is influenced by the concentration of ions in the water.
= used as an indicator of overall water quality.
High conductivity = high concentration of dissolved ions in water
= may suggest the presence of contaminants or changes in the composition of water.
Sterile connectors/ couplings = presterilized system with membraned (gets taken out) for product transfer
= for Hose connection with sterile couplings (under class A air) = disposable system for product transfer in class A
- tiny hole in the wall between class C & class A (overpressure)
- tube can be fixed in this hole & get connected with sterile coupling
- syringe plunger & needle & stopper
(16) Barrier systems
(a) From what can it be determined that the RABS shown is an open, active RABS?
(b) Which GMP clean room class is required as the background environment if aseptic production takes place in this RABS?
open = air circulation in & outside chamber (from A to B class)
active = HEPA Filter inside
= class B environment (for RABS)
(17) Leak test isolator glove
What is the leak test procedure for an isolator glove (in addition to the visual inspection)? (1 point)
What is the leak test procedure for an isolator glove?
2.2) airflow test at constant pressure = air gets inside of gloves & measurement of volume that is needed to keep pressure constant -> if tight, no additional air needed
(17) Barrier systems (4 points)
(a) What is the meaning of the abbreviation RABS?
(b) What characterizes an active, closed RABS?
= Restricted Access Barrier System
= always for aseptic purposes = + over-pressure inside (protection of product)
(17) Preparation of equipment and primary packaging material (2 points)
(a) Which medium is used in a CIP process for the final rinse of a format part that is later used in an aseptic process?
(b) Name one way in which two silicone tubes can be aseptically connected.
(c) What process takes place for cartridges after washing before treatment in the hot air sterilization tunnel?
(d) What is the final process step of an automated stopper treatment?
->Mobile or fixed systems with connection to SIP
CIP – steps = 2 water qualities required
1) Pre-rinse with purified water (pre-rinse)
2) If necessary, wash cycle with (alkaline) detergent
3) If necessary, rinse with purified water
4) Rinsing with water for injection (final rinse)
5) Drying -> otherwise can cause dilution of product
= aseptic connection via:
Sterile welding of thermos plastic tubes = 2 tubes get brought together & closed in a special machine (welder) -> inside machine: tubes get cut & welded to each other -> class A air supply
= siliconization
Capping/ Crimping = after placing a stopper on a vial, capping must be perform to fully seal the vial
= ONLY after this step container is considered ready & sealed -> can leave class A
(18) Preparation of equipment and primary packaging material (1.5 points)
(a) What is the riboflavin test used for?
(b) Which medium is used for the final rinse in the washing process of primary packaging materials?
(c) Why are syringes and cartridges siliconized?
a) What is the riboflavin test used for?
= for cleaning validation -> validation of CIP
b) Which medium is used for the final rinse in the washing process of primary packaging materials?
= water for injection (WFI) due to contact with product
c) Why are syringes and cartridges siliconized?
= to reduce friction & to chieve easy/ precised application (easier stoppes movement)
(20) Barrier systems (2.5 points)
(a) RABS and isolators are operated by glove/sleeve systems. What checks/tests must be performed on the gloves before usage?
(b) Describe the principle of a transfer into an isolator during operation using a decontamination airlock.
(c) Indicate one way in which a continuous transfer of material can be made out of an isolator?
a) What checks/tests must be performed on the gloves before usage?
1) Visual inspection of the gloves -> if leak detected no further test needed
b) principle of a transfer into an isolator during operation using a decontamination airlock.
3.1) Decontamination airlock/ transfer hatch
airlock = Luftschleuse
c) Indicate one way in which a continuous transfer of material can be made out of an isolator?
Open isolator systems = allows continuous material flow, hermetic seal for decontamination, with unidirectional air flow
Mousehole – transfer system
= for continuous material transfer between different pressure zones within the isolator or out of isolator
-> Pressure tight closure for decontamination
-> fit exactly to the maximum dimension of the transported material (e.g. vials)
Active mousehole = with suction (additional protection) Ò overpressure cascade inside isolator
(18) Preparation of equipment and primary packaging material (2.5 points)
(a) In CIP processes, it is important that no spray shadows occur. What is meant by this?
(b) Stoppers that will later be used in syringes in an aseptic process are to be prepared. Outline the process steps of automatic stopper treatment (excluding transfer and storage processes).
(c) What process takes place for syringes after final rinsing and blow-out with compressed air before treatment in the hot air sterilization tunnel?
"spray shadows" = areas or surfaces within a processing system that are not effectively reached or cleaned by the spray nozzles during the CIP cycle.
Automatic stopper treatment
1) Washing & cleaning = to remove any contaminants, particles, or residues.
2) Rinsing = After cleaning, the stoppers undergo rinsing to eliminate any remaining cleaning agents or impurities
3) Drying = to remove moisture
4) Sterilization = autoclaving, gamma irradiation
5) Siliconization = application of a thin layer of silicone oil to the stopper surfaces. This is done to facilitate the movement of the stoppers within the vial or syrange and reduce frction & enebale correct dosing
6) Inspection and Quality Control = to ensure that quality: visual inspection for defects, dimensional checks, and testing for sterility.
7) insert stopper in syringe (with pipe or vacuum)
(c) What process takes place for syringes after final rinsing and blow-out with compressed air before treatment in the hot air sterilization tunnel? = siliconization
(20) Aseptic filling (2 points)
(a) What is controlled fully automatically during aseptic filling? Name two aspects.
(b) Why are cartridges always filled to the top?
(c) What is the name of the illustrated dosing system?
1) Air circulation/ distribution & pressure cascades -> temperature
2) Filling weight
Filling Process according of cartridges to bulk syringes, but
1) Stopper first
2) Filling to the edge (100 %) to exclude air inclusions /bubbles
3) Closure with stopper and flanged cap or special closure system
c) Rotary piston pump = Drehkolbenpumpe: for temperature sensitive product
= used for: viscous products & for high precision filling (low volumes)
-> not for: metal sensitive products & quick product changes
- cylinder (no valve) & a piston that moves up/ down
- groove of upper part can rotate
- product get inside, groove rotates (180°C) & get filled to final container
(18) Aseptic filling (2.5 points)
(a) Name two dosing systems besides the peristaltic pump that are used for aseptic filling.
(b) What is the purpose of a stopper sorting bowl? Name two aspects.
(c) What needs to be considered when placing a lyophilization stopper following aseptic filling?
1) Rotary piston pump = Drehkolbenpumpe: for temperature sensitive product
2) Rolling diaphragm pump = multi-use or single use pumps
- intake port open & product gets inside, piston moves inside
- discharge port opens & product gets out, piston moves out
- diaphragm inside housing covers completely the piston,
no connection to outside (closed system)
(b) What is the purpose of a stopper sorting bowl? Name two aspects
Feeding of stoppers (Zufuhr von Stopfen) = Singularization (Vereinzelung) & correct stopper alignment with (vibratory) sorting bowl
- inside barrier system get transported to a vibratory sorting bowl (is rotation & vibrating)
- only stoppers that are located in right direct/ position are get transferred for capping
Lyophilization = performed in free dryer under class A conditions
-> because: - vials have special lyophilization stopper (with opening), so the solvent/ vapor can get out
- opening in stopper = risk of contamination -> class A surrounding & CIP/SIP capable (steam sterilization/ H2O2)
(19) Aseptic filling
An infusion bottle containing 1 liter of 0.9 % saline solution is to be produced using the blow-fill-seal process.
Specify all production steps starting from extrusion to the finished, sterile product. (2.5 points + up to 1 bonus point)
blow-fill-seal process:
Cyclic process (BFS)
1) Extrusion: Melting & homogenization of plastic granules in an extruder -> at 170°C get automatically sterilized
2) Pressing the liquid plastic through a ring nozzle -> a hose (schlauch) is formed
3) Extrusion & cutting hose below the nozzle -> most critical
4) Blow mould closes, bottom is welded -> blowing with sterilized compressed air
5) Inflating to a container with sterile compressed air (optional vacuum in the blow mould)
6) Filling with the product via the mandrel (Dorn)
7) Seal: Lifting of the mandrel, closure of the head mold & formation of the head closure (with vacuum)
8) Opening of the blow mould
9) Visual inspection for integrity necessary -> if plastic is to cold while forming because it creates holes -> bubble test
You want to transfer forceps into an isolator in which a product is currently being aseptically filled.
A beta container is to be used for the transfer.
Describe the process for transferring the forceps. (2 points)
A beta container is to be used for the transfer. Describe the process for transferring the forceps
1) Rapid Transfer Port = RTP
- consists of an Alpha-port which is installed in the wall of isolator or RABS
-> beta-container fits exactly to alpha-port
1) Beta container = stainless steel with pipe connector & hydrophobic sterile filter = forceps are put in the beta container
2) container (with forceps inside) gets autoclaved with vacuum
3) Connection of Isolator + container -> connected lids get opened from inside for material transfer = must be air-tight to avoid decontamination
-> lids must be airtight connected ->opening of lid only possible when fully connected-> by rotation 60°
4) forceps can be transfered into isolator & there used
-> higher pressure inside transfer pressure for product protection
(21) Blow fill seal (1.5 points)
(a) Which step of the blow-fill-seal process is the most critical step for the integrity of the container? In addition, provide a brief justification.
(b) Name one advantage of the blow-fill-seal process over traditional filling in glass containers besides the exclusion of glass breakage
Critical:
• sealing step = not good sealed container is a high risk of contamination
= is used for aseptic production
Advantage: BFS gives very high process & product safety (no transport routes, sterilization of vessels)
(24) Form fill seal / blow fill seal (1.5 points)
(a) What primary packaging materials are created in form-fill-seal processes?
(b) What is used in the BFS process as a supplement in the "Blow" step when very small containers are filled or a more complex container geometry is realized?
= sealed pouches & infusion bags
-> to form small or complex structure containers always vacuum is needed!
(22) A parenteral product was aseptically filled into glass vials and sealed with a stopper and crimp cap. The product is then visually inspected.
What is checked during this inspection process? Name two points.
(1 point)
What is checked during this visual inspection process? Name two points.
Inspection of:
1) visible particles in product
2) container integrity
3) correct crimping
4) other defects/ deviatiations
(23) Media fill (3 points)
(a) In which documents is the execution of a media fill specified?
(b) How often / with what frequency must a media fill be repeated after successful initial validation?
(c) An SOP for aseptic formulation and filling of a product requires that the product is freeze-dried and nitrogen is applied for inertisation before closing the stopper after freeze-drying. How do you take these process steps into account for a media fill?
= Validation plan
Frequency - EU GMP Annex 1
-> APS should be performed as initial validation = including 3 successful APS that cover all working shifts when significant modification in process occurs = again initial validation is required
-> periodic re-validation = APS should be repeated twice a year (2x/ year) per shift & process
-> Every employee who is routinely involved in aseptic processes must participate in 1x successful APS min. 1x/ year with an appropriate job profile.
c) An SOP for aseptic formulation and filling of a product requires that the product is freeze-dried and nitrogen is applied for inertisation before closing the stopper after freeze-drying. How do you take these process steps into account for a media fill?
• Lyophilizates -> partial drying process
(20) Media fill (2.5 points)
(a) What is the purpose of a media fill?
(b) A media fill plan includes extended microbiological environmental monitoring during a media fill compared to routine production. How do you evaluate this?
(c) How is a media fill evaluated after incubation?
a) What is the purpose of a media fill?
Medial fill – EU GMP Annex 1
= filling of a nutrient medium instead of product for aseptic process simulation
= to verify that process has high capability in assuring product sterility
= all aseptic operations of process must be included
-> process simulation test = should imitate as closely as possible the routine aseptic manufacturing process & include all the critical subsequent manufacturing steps -> microbial monitoring
b) A media fill plan includes extended microbiological environmental monitoring during a media fill compared to routine production.
How do you evaluate this?
Biological indicator
Evaluation = Visual inspection for growth by personnel, germ identification/turbidity -> Objective: zero growth
c) How is a media fill evaluated after incubation?
= visual inspection & microscope
= no turbidity
(21) Media Fill (2.5 points)
(a) What should be considered in a media fill with regard to personnel besides going for the maximum number of persons involved in the routine process? Name one additional point.
(b) An SOP for aseptic formulation and filling of a liquid product states that two excipients and WFI are added to the active ingredient for formulation, followed by stirring at 300 rpm for 30 min ± 10 min. How do you take these specifications into account for a media fill?
(c) Which number is missing in the following extract from Annex 1?
“Process simulation tests should be performed as initial validation with ____ consecutive satisfactory simulation tests”.
a) What should be considered in a media fill with regard to personnel besides going for the maximum number of persons involved in the routine process? Name one additional point.
b) An SOP for aseptic formulation and filling of a liquid product states that two excipients and WFI are added to the active ingredient for formulation, followed by stirring at 300 rpm for 30 min ± 10 min. How do you take these specifications into account for a media fill?
c) Which number is missing in the following extract from Annex 1?
(20) Media fill (3 points)
(a) What should be considered during a media fill with regard to personnel? Name two points.
(b) An SOP for aseptic formulation and filling of a liquid product specifies stirring for three hours as part of the aseptic batch formulation and storing the product at -20°C after filling into syringes. How do you take these specifications into account for a media fill?
(26) Media fill (3 points)
(a) List two worst-case situations that could be considered in a media fill of an aseptic filling.
(b) An SOP for aseptic formulation and filling of a liquid product specifies that for formulation two excipients are added to the active ingredient while stirring. The product is then sterile filtered and aseptically filled. After filling, the product is stored at -80°C. How do you take these process steps into account for a media fill?
(24)
(a) How must surfaces be designed so that they can be certified as hygienic? Name two different aspects in addition to a suitable Ra value.
(b) Which stainless steel surface finishing process is used if an optimum surface finish is to be achieved?
1) smooth, no scratches
2) chemical (corrosion) resistant & inert
3) not attracted for particles -> not 90°C surfaces
= mechanical polishung
(25) Name two positive and additionally one negative aspect regarding the compression of the seal shown.
a) (25) Name two positive and additionally one negative aspect regarding the compression of the seal shown.
negative = huge gap -> sealing not safe
positive = enough expansion space for sealing (get pressed) & rounded contact of seal to metall
(26)
(a) Why is there the rule L < 3 * D in piping systems or what does this rule say?
(b) What is the most important criterion for the design of vessels / tanks that is definitely fulfilled for the reactor shown below?
a) 3-D rule: L < 3 x D
L = length of the dead leg from the pipe wall
D = diameter of the pipe (connection piece)
In pipes for aseptic processing ( e.g. biorechtor pipes) dead ends & dead legs should be avoided because there is no flow of media possible or media may assemble there. Pipes must be designed to be self-drainable.
For a good hygenic design (if connection piece cannot be avoided) its length has to be lower than the 3x time diamater.
b)
(27) What is the difference between a double mechanical seal and a single mechanical seal? (1 point)
What is the difference between a double mechanical seal and a single mechanical seal? (1 point)
= primary difference between a double mechanical seal and a single mechanical seal lies in the number of seal faces and the barrier fluid used for sealing
(22) Design of a piping systems (2 points)
(23) Hygienic design of plant components (3 points)
(a) What is the name of the static connection shown in the figure?
(b) The figure below shows a hygienic static connection with front-flush sealing. One of the requirements for hygienic static seals is that there should be the maximum contact pressure at the sealing point. Is this requirement fulfilled here? Please give a brief justification for your answer.
(23) Hygienic design of plant components
(c) In the figure below a sterile sampling valve with bellows coating is shown. Explain one aspect that may be critical to successful cleaning of all valve internal surfaces?
(d) How can rotational movements of stirrer shafts be hermetically sealed?
(28) Hygienic design of plant components (3 points)
(a) The following figure shows a tank with an unfavorable hygienic design. Indicate what needs to be improved for the two marked points.
(22) Aseptic filling (2.5 points)
(a) Which routes can be used to transfer the product to be filled to the filling area (class A)? Name two possibilities.
(b) Name one advantage that peristaltic pumps offer over rotary piston pumps in aseptic filling.
(c) Explain the general principle of time-pressure dosing.
a) Which routes can be used to transfer the product to be filled to the filling area (class A)? Name two possibilities.
1) from class D/C to B the to A = e.g. RABS/ decontamination airlock
2) from class C to A -> inside sterilization tunnel (via via conveyor belt)
(23) Hygienic design of plant components (4 points)
(a) Which two aspects should be improved in this reducing piece? Note: the arrow indicates the direction of flow.
(b) Draw the compression of a front-flush seal with axial stop and label everything.
a) two aspects should be improved in this reducing piece?
sharp edges should be rounded
no engle (alpha); should be a straight area for liquid to flow
b) front-flush seal with axial stop
(c) The housing of which valve type is shown here?
(d) In the figure below you can see a valve for sterile sampling. The product chamber is on the left of the picture. How can it be ensured that the sealing point is also reliably cleaned and sterilised?
c) the housing of diaphragm valve
d) How can it be ensured that the sealing point is also reliably cleaned and sterilised?
= due to CIP/SIP connection
= valve can get opened for cleaning and sterilisation
(15)
State two principles of aseptic work in addition to slow, controlled movements, and briefly indicate in each case what the consequences of not following the principle would be for the production environment or process.
State two principles of aseptic work. What the consequences of not following the principle would be for the production environment or process.
Principles of aseptic working
1) Do not work above the product = risk of product contamination & risk for inhalation of substances for personnel
2) Keep sterile equipment under Class A conditions & protect
it from contamination; exchange it in case of contamination = risk of product contamination/ risk of cross-contamination/ if toxic subsatnce risk for personnel
(21) Preparation of equipment and primary packaging material (2.5 points)
(b) Which test is performed in the pharmaceutical sector for the validation of CIP processes?
(c) When in the course of the process does siliconization of glass syringes take place?
(d) Name one disadvantage that oily siliconization has compared to baked-on siliconization.
(e) For which pharmaceutical products is the final drying of the stoppers after automatic stopper treatment of particular importance?
Validation = Riboflavin test (at least 2 times): flush is fluorescence (green) substance & good water soluble
1) contamination of the inner surfaces with a riboflavin solution
2) drying
3) CIP verification of successful cleaning with UV lamp
-> no green fluorescence should be visible! otherwise dirt left
= before sterilization -> baked-on siliconization preferred & to reduce friction while transport/ sterilization
d) Name one disadvantage that oily siliconization has compared to baked-on siliconization.
oily siliconization = thick layer -> may cause mechanicall problems e.g. in syrange dur to limited space
-> backed-on silionization = due to treatment by heat (during sterilization) mire robust silocon layer results, lower risk of silicone migration
e) For which pharmaceutical products is the final drying of the stoppers after automatic stopper treatment of particular importance?
Lyophilized (Freeze-Dried) Products = are highly sensitive to moisture
= Dual-chamber technology - syringes
• Chamber 1 = lyophilized drug
• Chamber 2 = diluent for reconstitution
-> Used because: dilution of drug is easier,
safer & higher dosage precision
(6) A suitable sterilization process is to be selected for the sterilization of a new aqueous pharmaceutical product. The product is sterile-filterable. Other sterilization processes have not yet been tested for their suitability. Describe how, following the guidelines of the EMA, the selection of the process has to be done. (2 points)
Describe how, following the guidelines of the EMA, the selection of the process has to be done.
EMA: choice of sterilization method should be justified
-> according to decision tree = sterile filterable aquous product is NOT heat stable => normal steam sterilization not possible
1) product not heat stable = can be sterilised with F0-value > 8mins
-> lower °C & longer time to protect product
2) if product is not heat stable at all = use sterile filtration
3) product is sterile filterable (class A conditions) = apply combination of aseptic processing & filling
(b) How can a longitudinal movement of a valve rod be hermetically sealed. Name one possibility.
(c) Mark the location within the sampling valve that would be most difficult to clean in a CIP procedure (besides the sealing point) and justify your choice
(d) What is an alternative to the magnetic coupling for hygienic sealing of rotational movements that require high forces?
(10) autoclave
(a) Which principle is used for air removal in the autoclave shown?
(b) For what kind of sterilization items are counterpressure autoclaves used and how is the counterpressure generated?
= via air separator (in the bottom of system)
-> air is 1,5 times heavier than steam = it moves down & get pressed out of system
-> valve of air separator is open so air gets out
-> temeprature inside autoclave rises & get detected via temperature measurement -> at 100°C no cold air flow out & valve closes
b) counterpressure in autoclaves
counterpressure autoclaves are used for = aqueoues liquids (e.g. infusions) in closed containers -> air elimination not possible so counterpressure necessary
-> e.g. for bottles/ ampoules/ contect lenses…
generation of counterpressure = additional pressure gets introduced in the autoclave chamber with sterile compressed air
(17) Terminally sterilised products
Sterile drugs can be manufactured either by terminal sterilization or aseptic processing. In this context, complete the missing information in each of the following excerpts from Annex 1 of the GMP guide. (1.5 points)
Terminally sterilised products:
28. Preparation of components and most products should be done in at least a grade D environment […].
29. Filling of products for terminal sterilisation should be carried out in at least a grade C environment.
Aseptic preparation:
32. Preparation of solutions which are to be sterile filtered during the process should be done in a grade C environment
33. Handling and filling of aseptically prepared products should be done in a grade A environment […] with class B environment
(18)
Annex 1 of the GMP guideline contains the following remark on the purpose of cleanroom class B: "For aseptic preparation and filling, this is the background environment for the grade A zone."
When is a Class B cleanroom not required?
List two possibilities.
1) for terminally sterilised products = classes A/C/D required
2) for aseptic preparations = classes A/C/D required
(25) Annex 1 of the GMP guide specifies: „124. Filled containers of parenteral products should be inspected individually for extraneous contamination or other defects.“
What is the procedure for testing if it is done manually?
= visual inspection by pesonnel
1) filling weight
2) particles
(19) Barrier systems (2 points)
(a) Which RABS type is shown in the figure?
(b) Give two differences between a product protection isolator and a personal protection isolator (for handling a non-sterile toxic product).
a) open active RABS
b) differences between a product protection isolator and a personal protection isolator (for handling a non-sterile toxic product).
product protection isolator
1) protection of product -> operation at overpressure +
2) working with gloves
3) Fully sterilizable (H2O2 decontamination)
personal protection isolator
1) protection of personnel
- Negative pressure = operation for non-aseptic products
+ Overpressure = for aseptic/ toxic products
2) working with special manipulators (NO gloves!)
3) Spray cleaning (CIP) = manual spray cleaning from outside by personnel
4) exhaust air protected = air gets filtered bevor leaving isolator
(23) Glass vials
(a) A product is filled aseptically. Glass vials serve as the primary packaging material. Rubber stoppers and aluminium caps are used as the closure system. At what point in the process are the containers considered to be completely sealed?
(b) Indicate one way the stopper can be placed in syringes without pushing product out of the syringe.
(27) (a) A material in contact with the product should be inert to the product. Explain why this requirement is very important in terms of patient safety?
(b) Which parameter is used to describe the surface finish / roughness of a material?
(c) Why are scratches/crevices to be avoided on a surface in contact with the product?
(a) A material in contact with the product should be inert to the product. Explain why this requirement is very important in terms of patient safety?
= Inert (inaktiv) to the product & detergents and disinfectants to avoid interaction of material & product -> proterties & quality of product my change/ cause toxic reactions -> can be dangerous for patients
= for recure storage/ ling term stability
Roughness = Ra-value = gets measured perpendicular to grinding direction
-> Reference value: Ra ≤ 0.8 μm for surfaces in contact with the product
=> ideal: Ra ≤ 0.5 μm
-> For stainless steel surfaces a Ra-value of ≤ 0.8 μm achieved by mechanical polishing or machinings is recommended
-> most microbes are around 1 μm so they don’t fit in a roughness profile of ≤ 0.8 μm
= germs can accumulate in the rough profile (scratch)
-> difficult to clean
(29) Sealing of rotational movements (1 point)
(a) What is the main advantage of a magnetic coupling compared to a double mechanical seal?
(b) In addition to removing possible contamination, what purpose does the barrier medium serve in a double mechanical seal?
(5)
(a) By which process is most of the thermal energy transferred to the sterilisation load during steam sterilisation?
(b) DIN 58950 specifies for pure steam that the superheat must be below 5°C. Why is this specification important?
= saturated steam
(6)
(a) Draw the pressure curve of a steam sterilisation process using fractionated vacuum for air elimination and a single vacuum phase for drying.
(b) How can sufficient air removal be verified / checked? Name one possible way to do this.
= Bowie Dick test
(7) Glass bottles with WFI are to be terminally heat-sterilised by a hot water spray process. Describe the basic principle of the process, including how to prevent the glass bottles from bursting. (2 points)
Hot water spray
(9) Hot air sterilization tunnel (1.5 points)
(a) Hot air sterilization tunnels are a transfer system for primary packaging materials. Between which cleanroom classes does the transfer take place?
(b) Why is there the highest pressure in the cooling zone?
a) sterilization tunnel: Between which cleanroom classes does the transfer take place?
= C to A -> because in claass C washing of items is taking place
b) Why is there the highest pressure in the cooling zone?
= to avoid entrance of air inside or particles & to prevent recontamination of sterilized products
(13) A buffer is required for the aseptic formulation of a medicinal product which is not finally sterile-filterable. The buffer is prepared in cleanroom class D and subjected to germ-reducing filtration. For aseptic formulation in cleanroom class A, the sterile active substance is added directly to this buffer. Which two points are incorrect here? (2 points)
A buffer is required for the aseptic formulation of a medicinal product which is not finally sterile-filterable. The buffer is prepared in cleanroom class D and subjected to germ-reducing filtration. For aseptic formulation in cleanroom class A, the sterile active substance is added directly to this buffer. Which two points are incorrect here?
(14) State one activity/step in the aseptic manufacture of a pharmaceutical product carried out in clean room class B. (1 point)
State one activity/step in the aseptic manufacture of a pharmaceutical product carried out in clean room class B.
Double-door / pass-through autoclaves
Usage: - for manufacturing sterile pharmaceuticals -> aseptic production
- as transfer system
- always has 2 doors: loading from the dirty side & unload from clean side
- installed between class C (loading) & class B (unloading sterile products)
-> high assurance that no contamination in class B (only 1 door opens at a time)
(a) Why is one of the principles of aseptic working that (as far as possible) no work should be done over the open product?
(b) During the media fill experiment, sterile forceps were made unsterile by touching a potentially contaminated surface, but were still used for handling sterile stoppers. Which principle of aseptic working was violated?
a) Why is one of the principles of aseptic working that (as far as possible) no work should be done over the open product?
to avoide contamination of product —> product protection
for self-protection of personnel -> not to inhale substances
b) Which principle of aseptic working was violated?
• Only touch sterile materials with sterile equipment
• Keep sterile equipment under Class A conditions & protect it from contamination; exchange it in case of contamination
(17)
(a) Describe the way to accomplish the transfer of sterilized rubber stoppers, which are present in a beta bag, into an isolator during operation.
(b) The stoppers are used to close vials. For this purpose, the stoppers are singularised and placed on the vials by means of pick & place. What is the principle of pick & place?
(19) Fill and finish (4 points)
(a) What is shown in the figure below?
(b) Why is fill by weight preferred for high-priced medicinal products?
(c) Why are two-hose and not single-hose peristaltic pumps usually used?
(d) In which cleanroom class must freeze-drying take place after aseptic filling?
(e) Why does Annex 1 specify that crimping must be separated from filling and stoppering?
(f) How are ampoules closed?
(g) How do you call the process for manufacturing infusion or parenteral nutrition bags?
(h) How is the plastic hose produced in the blow fill seal process?
20) Which two data / parameters / documents must not be taken into account when releasing an aseptically manufactured product batch? (1 point)
Which two data / parameters / documents must not be taken into account when releasing an aseptically manufactured product batch?
site master file
fulfillment of product specofocations
For batch release must be taken into account:
1) Manufacturing documentation
2) In-process & final product controls = Sterility of product: all quality controls must be approved!
3) Controlling all Monitoring data (environment, personnel, WFI, etc.)
4) check deviations (Abweichungen): e.g. interventions, shutdown, equipment malfunction, CAPA
(a) Materials in contact with a pharmaceutical product should be smooth (low Ra value), easy to clean and inert. State two further requirements in addition to that.
(b) What is the guideline value for the Ra value of surfaces in contact with products in the pharmaceutical sector?
a) Materials in contact with a pharmaceutical product should be smooth (low Ra value), easy to clean and inert. State two further requirements in addition to that.
Requirements for materials
b) What is the guideline value for the Ra value of surfaces in contact with products in the pharmaceutical sector?
Many sterilization items that are steam sterilized are difficult to vent / remove air from.
(a) Name a sterilization item that is difficult to vent other than textiles.
(b) How can sufficient air removal be verified / checked? Name two possible ways to do this.
(a) Describe how the air removal is done in the autoclave shown below?
(b) Why does the temperature measurement take place at the bottom of the system (see illustration)?
a) Describe how the air removal is done in the autoclave shown below?
b) Why does the temperature measurement take place at the bottom of the system (see illustration)?
(8) heat sterilization processes
In heat sterilization processes, the aim is to achieve the shortest possible compensation time. Illustrate the compensation time graphically.
Note: Do not forget to label the axes. (2 points)
(11) Describe the steps that are needed to biodecontaminate an isolator with hydrogen peroxide
(VHP process). (2 points)
Describe the steps that are needed to biodecontaminate an isolator with hydrogen peroxide (VHP process).
VHP = vaporized hydrogen peroxide
= surface sterilization process -> decrease in 10log levels
= used for biodecontamination
= reduce of humidity with dehumidified because H2O2 vapours & creates moisture
2) Conditioning = H2O2 vaporises & gets in sterilisation chamber
= concentration of H2O2 & humidity rises
4) Aeration (Belüftung) = vapor (gaseous H2O2) can get easily out of the system (as liquid) enabling aeration with fresh air from outside
-> shorter process time
Why is it good to avoid condensation of H2O2?
(13) What should be considered when validating a steam sterilization process with regard to loading? (1 point)
(14) (a) A company wants to produce a new sterile product via aseptic processing, but without any robust data on the sterilizability of the product. You do not agree with this and suggest another procedure to the company. Which one?
(b) What is the minimum cleanroom grade for the preparation of a medicinal product that is later filled aseptically?
(15) Silicone tubes are used for the aseptic filling of a medicinal product.
[1] These are inserted into a double-door washing machine in a D zone, washed and dried therein and then removed to class C.
[2] There, the tubes are packed in a Tyvek bag, sterilized in an autoclave and removed again in class C. The bag is then placed in a disinfected material transfer hatch in class C and removed from it in class B.
[3] In class B, the transfer takes place into a previously decontaminated RABS. The Tyvek bag is placed in class A via a small opening in the window of the RABS.
[4] Finally, with the doors of the RABS closed, the opening of the Tyvek bag and the installation of the tubing are carried out via glove ports.
How do you evaluate the process? Are all steps (1 to 4) in compliance with the requirements of annex 1 and a minimal contamination risk for the drug processed in class A? Is there potential for optimization? If yes, which?
(3 points + 1 bonus point)
Nr. 1) is right according to Annex 1
Nr. 2) is wrong:
- for class B use pass through autoclave
- make biodecontamination
- the use of 2 Tyvek bags is needed for this process
Nr. 3) is wrong: small opening is not good in RABS
Nr. 4) is right according to Annex 1
(16)
(a) Give two reasons why, in critical cleanroom classes, cleanroom personnel should make only slow, controlled movements?
(b) Name two more basic rules of aseptic working.
a) Give two reasons why, in critical cleanroom classes, cleanroom personnel should make only slow, controlled movements
1) to avoid additional particle generation & distribution
2) to avoid interruption of lamina airflow & creation of air turbulances
3) to reduce risk of product contamination
• Always perform aseptic procedures from the side, do not work above the product
• Do not speak in critical areas if possible
• Disinfect sterile gloves regularly or change them
8)
8) The larger the F value (holding time), the slower is the SAL value achieved
With a D value of 1 min, an F value of 5 min results in 5 log levels of germ reduction.
(19) Fill and finish (4.5 points)
(a) What is checked during aseptic filling as part of an in-process control? Name one aspect.
(b) Name one advantage that rotary piston pumps can offer over peristaltic pumps in aseptic filling.
(c) In which cleanroom class must freeze-drying take place after aseptic filling?
(d) What is part of the aseptic filling process for oxidation-sensitive products?
(e) Which stopper placement method is shown here?
(f) What is the main advantage of using nested ready-to-fill syringes in tubs over bulk syringes?
(g) What are the two options for forming the plastic container in blow-fill-seal processes?
(h) What must be checked for all finished BFS containers, just as for ampoules, regardless of whether the product contained is administered parenterally?
(21)
(a) The surface quality is usually described by the Ra value. What is the Ra value or does the Ra value express?
(b) A material in contact with a sterile product should be abrasion-proof. Why?
a) The surface quality is usually described by the Ra value. What is the Ra value or does the Ra value express?
= Reference value: Ra ≤ 0.8 μm for surfaces in contact with the product
b) A material in contact with a sterile product should be abrasion-proof. Why?
1) for prevention of Particle Generation & crosscontamination
2) for maintaining Product Integrity: Abrasion can cause physical damage to materials, leading to cracks, scratches
3) to enhance Long-Term Durability: Sterile products are often intended for use over an extended period. Abrasion-proof materials contribute to the long-term durability to maintain the quality of product
(22) Why are metal-to-metal contact surfaces problematic in terms of hygienic design? (1 point)
Direct metal-to-metal joints shall not be used other than welding
= may harbour soil, liquids and could corrode
= create huge gap because metal is not flexible,
small pieces can break through friction & be dangerous for product
Why are hot air sterilization tunnels dsigned in such a way that they become slightly wider in the course of the sterilization zone?
= because of elongation of glass containers that get wider through heating -> containers expand
-> otherwise they may get truck & glass can break
-> glass breaking is a very huge problem because its very difficult to clean it up in the tunnel
Question: In preparation of aseptic filling of a drug product, the drug is usually first subjected to germ-reducing filtration
& then finally to sterile filtration in class A. Give reasons why an additional germ-reducing filtration is carried out.
(a) The illustration shows a hygienically designed static seal. For the positions marked 1 and 2, please indicate which positive aspects regarding hygienic design have been realised here. Please note: For position 2 it is already specified that it is a front-flush seal. This is to be supplemented by another aspect.
(15) Validation of sterilization processes (1.5 points)
(a) List four different non-economic requirements that should be included in a user requirement specification (URS) for an autoclave.
(b) What must be considered in the PQ of a steam sterilization process with regard to loading - in addition to the loading schemes of routine production?
(a) List four different non-economic requirements that should be included in a user requirement specification (URS) for an autoclave
1) Underlying regulatory requirements for the system
· Standards to be complied with, guidelines
2) Location / Environment / Dimensioning
· Place of installation/ Environment / zone requirements
· Constructional conditions, e.g. available area, floor load
· Dimensions (chamber volume)
3) Process
· Procedure / operating mode (e.g. FRVV)
· Special requirements for the process (e.g. accelerated cooling)
· Design / Type, (cylindrical/rectangular; horizontal/vertical)
· Working range (pressure / temperature)
· Performance / energy consumption / energy efficiency
4) Monitoring / Measurement technology
· Sensors/ Accuracy/ Additional features
5) PQ = Performance Qualification = Verification of sterilization success based on relevant loads -> actual process validation
= provide evidence that equipment works according to specified criteria (reliable sterilization)
= include physical & biological tests
Requirements for PQ (Verification of sterilization process)
Inspections must verify that:
1) the documentation confirms a successful IQ & OQ -> IQ/ OQ & cycle development should be successfully finished & working
2) the test load of the sterilizer contains products which are routinely handled -> loading patterns must be defined and [...] represent product families which present the greatest difficulty for the sterilisation -> include worst case conditions
3) the packaging system must be identical to the packaging used for routine production
4) the preconditioning = storage before sterilization must correspond to criteria
5) the loading configuration & amount of load has to bedefined -> size and/or the mass of the sterilizer load
(12) Validation of sterilization processes (2.5 points)
(b) What should be considered when validating a steam sterilisation process with regard to loading?
(c) What is the general name of test strips with bacterial spores used to validate sterilization procedures?
b) What should be considered when validating a steam sterilisation process with regard to loading?
= worst case conditions -> coltest spot
Cycle development = based on relevant loads -> Optimization of steam penetration, venting etc. for different loads
= establishment of sterilization cycles for routine manufacturing for different loads
PQ = Performance Qualification = Verification of sterilization success based on relevant loads -> actual process validation
2) the test load of the sterilizer contains products which are routinely handled -> loading patterns must be defined
and [...] represent product families which present the greatest difficulty for the sterilisation -> include worst case conditions
c) What is the general name of test strips with bacterial spores used to validate sterilization procedures?
The general name for test strips with bacterial spores used to validate sterilization procedures is "biological indicators" or "biological indicator strips”
These strips contain a known number of highly resistant bacterial spores, such as Bacillus subtilis or Geobacillus stearothermophilus. Biological indicators are employed to assess the efficacy of sterilization processes, for steam (autoclaving), ethylene oxide gas, hydrogen peroxide gas et.
(a) What is checked during the OQ of a steam sterilizer?
(b) What is the general name of test strips with bacterial spores used to validate sterilization procedures?
(c) What should not be included in the URS of a steam sterilizer? Mark with a cross.
a) What is checked during the OQ of a steam sterilizer?
OQ = Operational Qualification = Checking basic functions of equipment within the working area including chamber integrity
-> checking if:
1) sterilization possible
2) are required °C possible to achieve
3) chamber integrity -> possible with vacuum
b) What is the general name of test strips with bacterial spores used to validate sterilization procedures?
= microbial tests
c) DQ = Design Qualification = Determination of requirements for equipment
URS = user requirements specifications & functional specification document
(a) During the DQ you have to select a steam sterilization process for the sterilization of 2 m long silicone tubes used for aseptic filling. Which process do you choose? Give a justification for your choice.
(b) What is the name of the document in which the user demands on a sterilization system are recorded as part of the DQ?
(c) What is checked during thermoelectric testing with an empty sterilization chamber as part of the PQ of a steam sterilization process?
DQ = Design Qualification = Determination of requirements for the equipment ->(URS = user requirements specifications & functional specification document)
Ionizing radiation sterilization -> E-beam tunnel
= its applicabel for tube & E-beam tunnel is long, has enough space for 2 m logn tube
1) Thermo-electric test with empty chamber
= temperature and pressure sensors get distributed inside empty chamber to check uniformity of parameters (°C/ ∆p)
-> minimal deviations (+/- 2°C; +/-10 kPa) allowed -> even coldest spot must fulfil criteria
-> must be performed for minimum & maximum cycle times and temperatures
(9) F0 value
For a steam sterilization process at 125°C, an F0 value of 30 min is achieved. How is the achieved F0 value of 30 min to be interpreted?
9)
It means that the steam sterilization process for 30min at 125°C reaches an equal lethality as the process for 74 mins at 121.1°C.
(3) F0 & D value
a) You shall define the minimum duration of a pharmaceutical steamsterilization procedure. The D value of the microorganisms present in the material to be sterilized at 121 °C is given as D = 1.25 min. What is the minimum time you need to sterilize at 121 °C if the bioburden before sterilization is N0 = 10^6?
(b) What is the F value at 121 °C for the data in (a)?
F0 & D-value
In preparation for aseptic filling of a drug product, the product formulation is usually first filtered in a germ-reducing step and then finally sterile filtered in class A.
(a) What would be the consequence if the product formulation was not carried out in at least class C, as required, but in class D? Provide an explanation.
(a) What would be the consequence if the product formulation was not carried out in at least class C, as required, but in class D?
Provide an explanation.
1) higher risk of product contamination of particles out of air or materials (“dirty” environment)
2) Impact on product quality -> may have unwanted particles -> danger for patient
3) Risk of bad sterility -> not sterile enough -> no release of product (additional costs)
b) What must be checked on each filter before and after use?
-> before usage = is critical because it must be assured that filter doesn’t get contaminated
2 possibilities to keep filter sterile before usage:
1) sterilization in autoclave after filter testing Ò validation important
2) filter gets installed in production area & gets sterilized in place Ò filter testing can be done in place
(4) F0 & D value
(a) How are the F-value and the D-value related?
(b) For a given z-value of 10 °C, how does the D-value change if steam sterilisation is carried out at 120 °C (increased pressure, e.g. pressure cooker) instead of 100 °C (atmospheric pressure)?
(c) For a steam sterilisation process, the F0 value is 30 min. How can this value be interpreted?
F-value = Effectiveness of a sterilization process:
time (holding time) at a temperature T in minutes to reduce the existing population (N0) with the specific D value by the desired number of powers often/ log10 levels (n) to a final value (N)
= if processing time (F-value)
= D-value decreases for factor 10^-2
F0 = 1 min means the lethality of a method that works for 1 minute at 121.1 °C (250 °F) and a z-value of 10 °C (18 °F) or an equivalent temperature-time combination
F0 value of 30min = means the same lethality of a method that works for 1 minute at 121.1°C
(16) Give two reasons why, in critical cleanroom classes, cleanroom personnel should make only slow, controlled movements? (1 point)
Why, in critical cleanroom classes, cleanroom personnel should make only slow, controlled movements?
at high clean room classes are high requirement for aseptic working
slow controlled movements to avoid:
1) interruption of unidirectional flow or creation of air tubulances
2) contamination of products or spread out additional particles
(14)
In preparation for aseptic filling of a drug product, the formulation is usually first subjected to germ-reducing filtration and then finally to sterile filtration in class A.
(a) What is the minimum cleanroom class in which the preparation must take place?
(b) Give two reasons why additional germ-reducing filtration is carried out.
= Class C
b) Give two reasons why additional germ-reducing filtration is carried out.
bioburden reduction prefilters and sterilising grade filters
1) to prevent the build up of filter cake, first filter takes the main germs load out of product
2) if the first filter is having a lack the second filter enables the filtration (safety of process)
3) increases the process safety to ensure a sterile product
4) separation of the 2 steps is possible, so one is done in class C area & the other in the final filtration
A silicone tubing shall be used for the aseptic filling of a drug. The preparation of the silicone tubing consists of the following steps:
[1] In class D it is placed in a double-door washing machine, washed therein, and then removed to class C.
[2] In class C the hoses are double-wrapped in tyvek bags and loaded into a pass-through autoclave.
[3] The removal of the sterilized hoses takes place in class B.
[4] The hoses are then transferred to a previously decontaminated RABS. During transfer, the door of the RABS is briefly opened and the outer tyvek bag is removed.
[5] Finally, with the doors closed, glove ports are used to open the inner tyvek bag and install the tubing.
How do you evaluate the process?
Are all steps (1 to 5) in compliance with the requirements of annex 1 and a minimal contamination risk for the drug processed in class A?
Is there potential for optimization? If yes, which?
= the process doesnt fulfill the criteria of aseptic preparation
= No are not, high contamination risk through wrong unpacking in wrong zone
[1] ist right
[2] & [3] = tubs get unpacked/ opened (removal of second packaging tyvek bag) in class C -> surface sterilization from C to B is correct
[4] door of RABS shouldn't get opened -> high contamination risk (class A inside) & unpackeging of second packaging layer has to be in class C
[5] doors shouldn't open (transporttation via decontamiantion airlock or RTP better) - inner tyvek bag better to open in class B area & transfer sterile tub to class A for final filling/ processing
Main steps:
I) Class C/D: 1) tubs get mechanically unpacked/ opened -> HEPA filtered air
2) tubs get introduced into E-beam tunnel -> through airlock doors
3) tubs get transferred to irradiation zone -> surface sterilisation
II) Class B: 4) tubs get transported trough class B zone to class A
III) Class A: 5) filling zone -> presterilized syringes/ vials get filled
(10) Ethylene oxide sterilization (2 points + 0.5 bonus points)
Cartons of individually wrapped disposable syringes (stacked on a pallet) are to be sterilized using ethylene oxide.
Describe the complete sequence of the procedure.
Describe the complete sequence of sterilization using ethylene oxide.
= generally, only acceptable if no other method of sterilisation is possible” -> use in food sector prohibited
= in pharma limits of dosage are defined due to toxic residues
Process sequence (negative pressure process)
= runs under vacuum because of 2 purposes:
I) test of integrity of sterilization chamber
II) to take out air because it forms explosive mixture with ETO
1) Conditioning outside the sterilization chamber
= ETO gets brought to defined humidity outside chamber 55-85%
2) Transfer of the sterilization items into the chamber
3) Vacuuming & injection of nitrogen, then moistening the chamber with steam = air gets succeed out with vacuum & nitrogen gets inside via pulsation repeats
4) Injection of ethylene oxide (ETO)
5) Sterilization time / dwell period = exposure time
6) Washing steps: vacuuming and introduction of inert gas (N2) + catalytic conversion of ETO to CO2 + H2O = conversion important because ETO is dangerous to nature
7) Venting (heated) = entlüften = ventilation to reach limits of residues defined in EMA guideline
-> take few days to take out all ETO & ventilation systems must have high qualities
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