Why Validation is Important

Validation is a concept that has been evolving continuously since its first formal appearance in United States in 1978. The concept of validation has expanded through the years to encompass a wide range of activities which should take place at the conclusion of product development and at the beginning of commercial production. Validation is confirmation by examination and provision of objective evidence that the particular requirements for a specified intended use are fulfilled.

We all love validation!

Objective Measures

Validation is the overall expression for a sequence of activities in order to demonstrate and document that a specific product can be reliably manufactured by the designed process, usually, depending on the complexity of today’s pharmaceutical products, the manufacturer must ensure. Quality cannot be adequately assured merely by in-process and finished product inspection or testing so the firms should employ objective measures (e.g. validation) wherever feasible and meaningful to achieve adequate assurance.
Today we have different definitions of validation, which are as follows-

• Establishing documented evidence which provides a high degree of assurance that a specific process will consistently produce a product meeting its pre-determined specifications and quality characteristics.
• The collection and evaluation of data, from the process design stage throughout production, which establishes scientific evidence that a process is capable of consistently delivering quality products.
• Validation is a process by which a procedure is evaluated to determine its efficacy and reliability for forensic casework analysis.

Why Validation is Important

The principles – Quality, Safety and Effectiveness must be designed and built in to the product, quality cannot be inspected or tested in the finished products and each step of the manufacturing process must be controlled to maximize the probability that the finished product meets all quality and design specifications. Now let me explain the specific importance of the validation – it is the concept detailed in quality guidelines of Product Lifecycle and with the help of which we can do the following:

• Determine the process parameters and necessary controls.
• To confirm the process design as capable of reproducible commercial manufacturing.
• Risk/Worst Case assessment. What is Worst Case? It is a set of conditions encompassing upper and lower limits and circumstances, including those within standard operating procedures, which pose the greatest change of process or product failure when compared to the ideal conditions.
• To provide ongoing assurance that the process remains in a state of control during routine production through quality procedures and continuous improvement initiatives.
• Quantitatively determine the variability of a process and its control.
• The variability within and between batches can be evaluated to determine the inner and intra-batch variability.
• Greater scrutiny of the process performance for development and deployment of process controls.
• Scientific study performed prior to implementing a change to a process can support the implementation of a change without revalidation.
• Safeguard and process against sources of variation which may not have been identified during the original process development.
• The most compelling reason to optimize and validate pharmaceutical productions and supporting processes and cost reduction.
• Control point in the context of preventive maintenance.
• Investigate deviations if any from established parameters.

Conclusion

Validation allows us to focus on our everyday business operations of making and selling quality products that also comply with regulatory requirements such as the FDA, Schedule M, etc. The industry which has adopted a lifecycle approach to the product development, validation and modern risk analysis tools can control critical process parameters. The companies can create a new standard of industry best practice by embracing the ability of validation practices which will lead in technological revolution.

Critical Parameters Affecting Process Validation

Validation is an integral part of quality assurance; it involves systematic study of systems, facilities and processes aimed at determining whether they perform their intended functions adequately and consistently as specified. Validation in itself does not improve processes but confirms that the processes have been properly developed and are under control. Adequate validation is beneficial to the manufacturer in many ways – It deepens the understanding of processes; decreases the risk of preventing problems, defect costs, regulatory non compliances and thus assures the smooth running of the process.

Process Validation is key to a robust manufacturing process

Process validation involves a series of activities taking place over the lifecycle of product and process. Validation requires a meticulous preparation and careful planning of the various steps in the process. All work involved should be carried out in a structured way according to formally authorized standardized working procedures.

What are the Critical Parameters affecting Process Validation?

The critical parameters should normally be identified during the development stage or from historical data or during manufacturing and process control. Process validation involves three stages and now will identify the critical parameters in these stages.

Stage One: Process Design

Process design is the activity of defining the commercial manufacturing process. The goal of this stage is to design a process suitable for routine commercial manufacturing that can consistently deliver a product that meets its critical quality attributes. A product development activity provides key inputs to the design stage, such as the intended dosage form, the quality attributes, and a general manufacturing pathway. The functionality and limitations of commercial manufacturing equipment should be considered, also contributions of variability by different component lots, production operators, environmental conditions and measurement systems in the production setting.
Designing an efficient process with an effective process control approach is dependent on the process knowledge. Use of risk analysis tools to screen potential variables for Design of Experiment (DOE) studies to minimize the total number of experiments. The results of DOE studies can provide justification for establishing ranges of incoming component quality, equipment parameters and in-process material quality attributes. Manufactures should document the variables studied for a unit operation and the rationale for those variables identified as significant. This information is useful during the process qualification and continued process verification stages, including the design is revised or strategy for control is refined.
Process control addresses the variability to assure quality of the product. Controls can consists of material analysis and equipment monitoring at significant processing points designed to assure that the operation remains on target and in control with respect to output quality. Timely analysis, control and adjust the processing conditions so that the output remains constant.

Stage Two: Process Qualification

During this stage, the process design is confirmed as being capable of reproducible commercial manufacturing. It confirms that all established limits of the critical parameters are valid and that satisfactory products can be produced even under worst case condition. This stage has following elements –Qualification of Utilities and Equipment.
Installation Qualification is an essential step preceding the Process Validation exercise which is normally executed by Engineering group. The installation of equipment should follow well defined plans which is developed and finalized following progression through a number of design stages. This stage of validation includes examination of Equipment Design, Determination of Calibration, Maintenance and Adjustment Requirements.
Consider the following Equipment Calibration Requirements
1. Confirmation of calibration of calibrating equipment with reference to the appropriate national standard.
2. Calibration of measuring devices utilized in the Operational Qualification stage.
3. Identification of calibration requirements for measuring devices for the future use of the equipment. At the Installation Qualification stage the company should document preventive maintenance requirements for installed equipment.
Operational Qualification is an exercise oriented to engineering function referred as commissioning. It is important stage to assure all operational test data conform with pre-determined acceptance criteria and manufacturer should develop draft standard operating procedures for the equipment, service operation, cleaning activities, maintenance requirements and calibration schedules.
The critical operating parameters for the equipment or the plant should be identified at the Operational Qualification stage. Critical variables should incorporate specific details and tests that have been developed. The completion of a successful Operational Qualification should include the finalization of operating procedures and operator instructions documentation for the equipment.
Performance Qualification combines the actual facility, utilities, equipment, trained personnel, control procedures and components to produce commercial batches. Performance qualification will have a higher level of sampling, additional testing and greater scrutiny of process performance. The level of monitoring and testing should be sufficient to confirm uniform product quality throughout the batch during process.

Stage Three: Continued Process Verification

Continually assure that the process remains in a state of control during commercial manufacturing. A system or systems for detecting unplanned departures from the process as designed is essential. The following points to be considered in Continued Process Verification.
Collection and evaluation of information and data about the performance of the process will allow detection of process drift. Evaluation should determine whether action must be taken to prevent the process from drifting out of control.
An ongoing program to collect and analyze product and process data that relate to product quality must be established to verify the critical quality attributes are being controlled throughout the process.
Process variation also can be detected by assessment of defect complaints, out of specifications finding, process deviation reports, process yield variations, batch records, incoming raw material records and adverse event reports.
Operator’s errors should be tracked to measure the quality of the training program.
Maintenance of the facility, utilities and equipment is an important aspect of ensuring that a process remains in control.

Conclusion

Process validation is a mean of ensuring and documenting that the processes are capable of producing a finished product of the required quality consistently and should cover all the critical elements of the manufacturing process. The process design stage and the process qualification stage should have as a focus the measurement system and control loop establishing scientific evidence that the process is reproducible and will consistently deliver quality products.
Good process design and development should anticipate significant sources of variability and establish appropriate detection, control, appropriate alert and action limits. Process variability should be periodically assessed. It is the responsibility of the manufacturer to judge and provide evidence of a high degree of assurance in its manufacturing process.

References

• Guidance for Industry Process Validation: General Principles and Practices – US Dept. of Health and Human Services, Food and Drug Administration. Nov. 2008 Current Good Manufacturing Practices.
• ANNEX 15. Validation Master Plan, Design Qualification, Installation and Operational Qualification, Non Sterile Process Validation, Cleaning Validation. 17th Sep. 1999.

Good Documentation Practices (GDP) are Critical to Success!

Good Documentation Practices (GDP) are critical to the success of any operation or project within a regulated industry. Deployed [usually] via a Document Management Plan in accordance with Standard Operating Procedures (SOPs), GDP is cascaded through an organisation to enable consist, correct entries being made on and to documentation.

GDP requires a consistent approach

There is a more than one way to skin a cat, one might say so there is certainly more than one way to work with documents and for this reason alone GDP is critical. For example, you have a group of operators making up a batch of drugs on a rotating shift basis – they’re all completing the relevant batch records whilst adhering to the SOPs governing the make-up; the drugs are made [probably correctly], however upon QA review the specialist doesn’t fully understand some of the entries in the batch record.
There are blank spaces in some, date formats are different (EU vs US); felt pens, highlighters and biros are used to make the entries and mistakes are left scribbled out, ripped off or just left. How can the company stand over the integrity of the drugs when their own internal QA specialists can’t understand the batch records?
Answer: They can’t.
What next? Drugs in the bin, poor documentation equals poor assurance and that means lost revenue at best and lost customers or patient risk at worst!

How can we improve?

Remedy: Good Documentation Practices. If everyone is utilising the same set of documentation rules, whether this be the people making drugs or the people checking HVAC (Heating, Ventilation and Air Conditioning) logbooks the message is clear. Follow the procedures or your work won’t be acceptable. If everyone handles mistakes with a single-strikethrough and initials and dates modifications, paginates in the form x of y; everything is consistent. Everyone can understand and QA reviews can be successful leading to getting product out the door.

Have clear training requirements

Good Manufacturing Practices (GMP) are fundamental to the success of drug manufacturing; GDP is fundamental to GMP. But how is this achieved? Training. Plain and simple. Training all personnel as to the criticality of GDP is essential. Show them how it is done, show them how is shouldn’t be done – make sure each person has a curricula detailing their training requirements and make sure training is fully documented and in cases of great importance such as GDP make sure there is an assessment so that the trainee can be verified to have understood the course.
Training doesn’t have to take long, it just has to be right. 30 to 60 minutes is sufficient to train people in the use of GDP – but make sure a robust document management plan is in existance first, use this as the driver to push out the importance of GDP, ensure that all documentation eventualities are addressed from labelling and cross referencing attachments to the usage of tip-ex (or rather not to ever use tip-ex – this is simply forbidden).
Work instructions are SOPs and will implement the requirements of the document management plans so that GDP simply can’t go un-noticed, after all if people are trained in procedures that they aren’t following properly this is going against the grain of the job brief as well as moving the business out of compliance in certain areas, which of course is completely unnacceptable, leaving the offenders open to disciplinary action.

GDP is here to stay

Clearly and simply, or rather to be blunt – GDP is here to stay and once proper procedures have been established, users trained and the wheels are in motion there is no turning back. QA will reject illegal entries and users will simply just apply GDP, well it is better than the sack. Once these have been implemented it will be quite difficult to NOT understand the point whilst at the same time people won’t be able to wonder why or how they were even there before GDP.

When is GDP applicable?

When is GDP applicable? Officially, when completing documentation that supports the manufacture of drug products or offical materials, the storage, holding and distribution of goods are within the remit of GDP to name but a few. Professionally, it is good practice to apply GDP always – even when handling documents relating to test and development systems. If nothing else, GDP is as much part of Good Engineering Practice (GEP) as it is relevant to GMP, if in doubt apply GDP. At least you’ll sleep safely at night without worrying about noise in the night been your boss at the door.

GDP all the way!

Enforcement. The Document Management Plan should detail how these documents should be verified to be correct. We’ve already mentioned QA reviewing batch records but they don’t necessarily get to see everything and to this end documents that are deemed to fall under the GDP umbrella should be audited to verify that people completing GDP entries are doing so correctly; this is standard practice and is a useful exercise, after all it is better identifying and resolving such issues sooner than later; the consequences of the FDA or EMEA findings faults during their own audits doesn’t bear thinking about!
So, GDP or not GDP? I think I’m safely on the GDP side of the fence, how about you? Plan to succeed with GDP and ensure your systems are set up properly. Otherwise you’ve simply failed by virtue of failing to plan!! Don’t delay, write your document management plan today!

Cleaning Validation Methods

leaning can be defined as the removal of residues from previous batch, other residues, and traces of cleaning agents. There are several mechanisms associated with cleaning of equipment. The mechanisms involved can be mechanical action, chemical action between the residues and the cleaning agent. The selection of cleaning agent and mechanism involved in cleaning is largely dependant on the process residue to be cleaned.

Cleaning Validation Mechanisms

Cleaning Mechanisms

The cleaning mechanism totally depends on the selection of cleaning agent and type of residue to be cleaned. Following can be the one of the methods involved in cleaning of residues,

• Dissolution
• Saphonification
• Wetting
• Emulsifying

 

Many cleaning compound agents perform several functions at once. Butyl, for instance, can serve as a wetting or surface tension reducing agent as well as a solubilizing agent. It also can contribute to emulsifying capabilities when combined with anionic surfactants or soaps (alkali-metal salts of carboxylic acids).

Dissolution

Dissolution is the process by which a solid or liquid forms a homogeneous mixture with a solvent or solution. This can be explained as a breakdown of the crystals into individual ions, atoms or molecules and their transport into the solvent
The mechanism involved in this type of cleaning is solubility of the residue in the cleaning agent or solvent. The monobasic buffers i.e. sodium chloride are soluble in cool and hot WFI. Ethylene glycol butyl ether is soluble in water as well as oil is also used in solubilizing agent. Chelating agents and builders are added to the formula to keep water hardness from interfering with the cleaning process.
Rate of dissolution is depend on,

• Nature of solvent or residue to be dissolved
• Temperature of solvent
• Presence of mixing
• Area of contact
• Presence of inhibitors

Saphonification

Saponification can be defined as “hydration reaction where free hydroxide breaks the ester bonds between the fatty acids and glycerol of a tri-glyceride, resulting in free fatty acids and glycerol”, which are each soluble in aqueous solutions.
This process specifically involves the chemical degradation of lipids, which are not freely soluble in aqueous solutions. Heat treated lipid residues are difficult to remove than non-heat residues due to polymerization.
Saphonification plays a critical role in cleaning lipids which are present in the areas of process involving cell growth and cell processing i.e. Bacterial fermentation, Cell disruption process

Wetting

Wetting can be defined as a process “involves the lowering of the surface tension of the cleaning solution, thus allowing it to better penetrate residues that are adhered to equipment and piping surfaces”. Wetting agents, or surfactants, are often used in relatively small amounts and they can substantially reduce the quantities of cleaning agent (in this case, alkali) required for residue removal.
Advantages of Wetting

• Lowers the surface tension of the cleaning solution
• Allow better penetrate residues which are adhered to equipment
• Used in small amount
• Sticky residues which are hydrophobic in nature get easily removed

Water acts as a solvent that breaks up soil particles after the surfactants reduce the surface tension and allow the water to penetrate soil (water is commonly referred to as “the universal solvent”).

Emulsifying

Emulsifying and suspending agents are often used to keep residues from precipitating by providing “hydrophobic groups” onto which hydrophobic areas of residues can associate, thus preventing them from associating with other residues and forming larger particles which are likely to leave solution. These agents also typically have “hydrophilic groups” which keep them very soluble in aqueous solutions of moderate to high ionic concentrations. Emulsifiers increase the capacity of a cleaner to emulsify non-soluble compounds in the cleaner. i.e. anionic soap surfactants, cationic surfactants, neutral surfactants
Advantages of Emulsifying agents,

• Prevent association of residues
• Allow the residue to precipitate and not allow thdse residue to redeposit on surface

Cleaning In Place (CIP) Vs Cleaning Out of Place (COP)

The basic regulatory requirement is to provide pharmaceutical products of highest quality to the patients. A cleaning problem can have various consequences to health, economics, environment and regulatory approvals. Absence of good cleaning, leads to contaminated drugs, which pose risks for patients as well as manufacturing personnel. Regulatory risk includes possible removal of authorization. Economical risk involves production delays and stock shortages or losses but also manifest as public relations problems for a company.
There are four main cleaning processes used in regulated environments.  These include:

• Cleaning-in Place (CIP)
• Cleaning-out-of-place (COP)
• Manual Cleaning
• Immersion Cleaning

So whats the difference between all of these cleaning techniques?

Cleaning In Place (CIP)

Cleaning in place can be described as the cleaning of equipment and vessels at the same place without movement of them to a different place. The cleaning agents can be transferred to the vessel or equipment types either thorough fixed piping or flexible hoses.
The CIP process can consist of the following elements:

• Supply pump
• Return pump
• Heat exchanger with Black/Plant steam supply
• Chemical tanks i.e Acid, Alkali tanks
• Supply Pressure gauge or transmitter
• Supply temperature sensors
• Conductivity meter with sensor

Cleaning Out of Place (COP)

Cleaning Out of Place is defined as a method of cleaning equipment items by removing them from their operational area and taking them to a designated cleaning station for cleaning. It requires dismantling an apparatus, washing it in a central washing area using an automated system, and checking it at reassembly.

Manual Cleaning

Manual cleaning is the universal practise among the pharma and biopharma industries. The design, configuration and construction of equipment or the whole equipment which necessities the manual cleaning for the piece of equipment. The efficiency of the manual cleaning accomplished by training the cleaning operators, ensuring exact method of cleaning in the manual cleaning SOP, validating the method from different operators and verifying the procedure with interval of time.
The manual cleaning is dependent on,

• Concentration of detergent used
• Temperature of washing liquid

Immersion Cleaning

This is the type of cleaning in which the parts to be cleaned are placed in the cleaning solutions to come in contact with the entire surface of the parts.
Immersion cleaning is preferred for parts that must be placed in baskets and for processes requiring a long soaking time because of the type of contamination to be removed or the shape of the parts to be cleaned.
It is the most effective method, even if not the fastest one, and can be used with any type of cleaner for any process, heated or at room temperature. Immersion washers can be portable or stationary; single or multi-compartment; and are available with a variety of options, controls and valve configurations including CIP capability.
The important aspects during design of immersion washer should be

• To minimize cycle time
• Lower chemical usage
• Reduce water and utility costs

Performance for immersion cleaning can be improved by moving the parts within the liquid or with agitation of the liquid, mechanically or with the addition of ultrasonic energy.

Cleaning Validation Forum

If you would like to learn more about CIP or COP please read the following threads:

• Criteria of determination of cleaning of residue
• Post Cleaning Rinsing
• Cleaning Validation Conclusion Report
• Cleaning Validation Calculation

Process Robustness in Pharmaceutical Manufacturing

The objective of this study is to unify understanding of the current concepts of process robustness and application of robustness principles to non-sterile solid dosage form manufacturing. Process robustness activities start at the earliest stages of process design and continue throughout the life of the product, it suggests greater process certainty in terms of yields, cycle times and level of discards.

Process Robustness in Pharmaceutical Manufacturing

An assessment of process robustness can be useful in risk assessment, reduction, potentially be used to support future manufacturing and process optimization. Robustness cannot be tested into a product; rather it must be incorporated into the design and development of the product. Performance of the product and process must be monitored throughout scale up, introduction and routine manufacturing to ensure robustness is maintained.

Principles Of Process Robustness

Definition of Robustness –

“The ability of a process to demonstrate acceptable quality and performance while tolerating variability.”

Process performance and variability may be managed through the choice of manufacturing technology. Well designed processes reduce the potential for human mistakes, thereby contributing to increased robustness. During product and process development both the inputs and outputs of the process are studied to determine the critical parameters and attributes for the process, the tolerances for those parameters and how best to control them. Critical Quality Attributes, Process Parameters, Process Capability, Manufacturing and Process Control Technologies and Quality System Infrastructure are referred as Manufacturing Science underlying a product and process. Principles of process robustness are as follows –
(A) Critical Quality Attributes (CQAs) – The identified measured attributes that are deemed critical to ensure the quality requirements – intended purity, efficacy and safety of an intermediate or final product, termed as Critical Quality Attributes.
(B) Critical Process Parameters (CPPs) – Is a process input that, when varied beyond a limited range has a direct and significant influence on a Critical Quality Attribute. It is important to distinguish between parameters that affect critical quality attributes and parameters that affect efficiency, yield, worker safety or other business objectives. Most processes are required to report an overall yield from bulk to semi-finished or finished product. It is important to have an understanding of the impact of raw materials, manufacturing equipment control, degree of automation or prescriptive procedure necessary to assure adequate control.
(C) Normal Operating Range (NOR) and Proven Acceptable Range (PAR) – In developing the manufacturing science a body of experimental data is obtained and the initially selected parameter tolerances are confirmed or adjusted to reflect the data. This becomes the Proven Acceptable Range for the parameter, and within the PAR an operating range is set based on the Normal Operating Range for the given parameter. In a robust process, critical process parameters have been identified and characterized so the process can be controlled within defined limits for those CPPs. A process that operates consistently in a narrow NOR demonstrates low process variability and good process control. The ability to operate in NOR is a function of the process equipment, defined process controls and process capability.
(D) Variability: Source and Control – Typical sources of variability includes process equipment capabilities, calibration limits, testing method variability, raw materials, human factors for non automated processes, sampling variability and environment factors within the plant facility.
(E) Setting Tolerance Limits – Upper and lower tolerances around a midpoint within the PAR of a parameter should be established to provide acceptable attributes. The defined limits should be practical and selected to accommodate the expected variability of parameters while confirming to the quality attribute acceptance criteria.

Development Of A Robust Process

A systematic team-based approach to development is one way to gain process understanding and to ensure that a robust process is developed. The following are the steps for the development of a robust process –
(1) Form the Team – Development of a robust process should involve a team of technical experts from R&D, technology transfer, manufacturing, statistical science and other appropriate disciplines. This team approach to jointly develop the dosage form eliminates the virtual walls between functions, improves collaboration and allows early alignment around technical decisions leading to a more robust product. This team should be formed before optimization and scale-up.
(2) Define the Process – A typical process consists of a series of unit operations. Before the team can proceed with development of a robust process they must agree on the unit operations they are studying and define the process parameters and attributes. Defining the process is to list all possible product attributes and agree on potential Critical Quality Attributes. The final step in defining the process is determining process parameters. Categorizations of parameters to consider are materials, methods, machines, people, measurement and environment.
(3) Prioritize Experiments – It is recommended that the team initially use a structured analysis method such as a prioritization matrix to identify and prioritize both process parameters and attributes for further study. A ranking of parameters of importance is calculated by considering the expected impact of a parameter on attributes as well as the relative importance of the attributes.
(4) Analyze Measurement Capability – The analysis of a process cannot be meaningful unless the measuring instrument used to collect data is both repeatable and reproducible. Analysis should be performed to assess the capability of the measurement system for both parameters and attributes. Measurement tools and techniques should be of the appropriate precision over the range of interest for each parameter and attribute.
(5) Identify Functional Relationship Between Parameters and Attributes – The functional relationships can be identified through many different ways, including computational approaches, simulations or experimental approaches. Design of Experiments is the recommended approach because of the ability to find and quantitate interaction effects of different parameters. Properly designed experiments can help maximize scientific insights while minimizing resources because of the following –

• The time spent on planning experiments in advance can reduce the need for additional experiments.
• Fewer studies are required and each study is more comprehensive.
• Multiple factors are varied simultaneously.

(6) Confirm Critical Quality Attributes and Critical Process Parameters – After a sufficient amount of process understanding is gained, it is possible to confirm the Critical Quality Attributes previously identified. Critical Process Parameters are typically identified using the functional relationships from step 5 (Identify Functional Relationship Between Parameters and Attributes).

Conclusion

The pharmaceutical manufacturers should implement robust manufacturing processes that reliably produce pharmaceuticals of high quality and that accommodate process change to support continuous process improvement. Creating a system that facilitates increased process understanding and leads to process robustness benefits the manufacturer through quality improvements and cost reduction. The goal of a well characterized product development effort is to transfer a robust process which can demonstrate, with a high level of assurance, to consistently produce product meeting pre-determined quality criteria when operated within the defined boundaries.

References

• PQRI Workgroup Members. Process Robustness – A PQRI White Paper. Pharmaceutical Engineering The Official Magazine of ISPE November/December 2006; Available from: http://www.06ND-online_Glodek-PQRI.pdf
• Taguchi G., Y. Wu., A. Wu. Taguchi Methods for Robust Design. American Society of Mechanical Engineers 2000.
• Johnson D. B., Bogle I. D. L. A Methodology for the Robust Evaluation of Pharmaceutical Processes under Uncertainty. Chem. Papers 54 (6a) 398-405 (2000).
• Innovation and Continuous Improvement in Pharmaceutical Manufacturing (Pharmaceutical CGMPs for the 21st Century) The PAT Team and Manufacturing Science Working Group Report: A Summary of Learning, Contributions and Proposed Next Steps for moving towards the “desired State” of Pharmaceutical Manufacturing in the 21st Century. Available from: http://www.2004-4080b1_01_manufSciWP.pdf.

Validation Process in Pharmaceutical Industry

Validation is a method to keep a check on the specific process, whether the ongoing process is able to meet the desired requirements. The definition of Validation as given by GMP is  "Establishing documented evidence which provides a high degree of assurance that a specific process will consistently produce a product meeting its pre-determined specifications and quality attributes.”

Validation documentation includes analytical information, reports determining development, formulae which are used in the manufacturing process, standard operating procedures, development reports. Documentation also provides with the information for the currently running process. Activities performed under Validation will incorporate a level of Impact Assessment to ensure that systems, services and products directly influenced by the testing have been identified.

Validation process is conducted in various ways, some of them are:
Prospective validation
Under this kind of validation, a documented evidence is made defining, that a process will do, what it is supposed to do. And this specification is based upon a pre planned series of scientific tests as defined in the validation plan.

Concurrent validation
This kind of validation comes in a picture when an existing process is in a state of control because of various tests applied on samples throughout a process and when, same can be shown. For the documentation to be presented, all data is collected with the proper implementation of the process. Moreover, collecting of data continuous till sufficient information is available to demonstrate process reproducibility.

Retrospective validation
Finally Retrospective validation is documented, which is actually based on review and analysis of historical data. This particular validation defines, that a process does what it purports to do

Concept of Process Validation For Pharmaceutical Industry

Concept of Validation
According to GMP definition Validation is "Establishing documented evidence which provides a high degree of assurance that a specific process will consistently produce a product meeting its pre-determined specifications and quality attributes."
Appropriate and complete documentation is recognized as being crucial to the validation effort. Standard Operating Procedures (SOPs), manufacturing formulae, detailed batch documentation, change control systems, investigational reporting systems, analytical documentation, development reports, validation protocols and reports are integral components of the validation philosophy. The validation documentation provides a source of information for the ongoing operation of the facility and is a resource that is used in subsequent process development or modification activities.
All validation activities will incorporate a level of Impact Assessment to ensure that systems, services and products directly influenced by the testing have been identified.
A revalidation program should be implemented based on routine equipment revalidation requirements and on the Change Control Policy.
Types of Validation
Prospective validation
Establishing documented evidence that a piece of equipment/process or system will do what it purports to do, based upon a pre-planned series of scientific tests as defined in the Validation Plan.
Concurrent validation
Is employed when an existing process can be shown to be in a state of control by applying tests on samples at strategic points throughout a process; and at the end of the process. All data is collected concurrently with the implementation of the process until sufficient information is available to demonstrate process reproducibility.
Retrospective validation
Establishing documented evidence that a process does what it purports to do, based on review and analysis of historical data.
Design Qualification (DQ)
The intent of the DQ is met during the design and commissioning process by a number of mechanisms, which include:
- Generation of User Requirement Specifications
- Verification that design meets relevant user requirement specifications.
- Supplier Assessment /Audits
- Challenge of the design by GMP review audits
- Product Quality Impact Assessment
- Specifying Validation documentation requirements from equipment suppliers
- Agreement with suppliers on the performance objectives
- Factory Acceptance Testing (FAT), Site Acceptance Testing (SAT) & commissioning procedures
- Defining construction and installation documentation to assist with Installation Qualification (IQ).
Installation Qualification (IQ)
IQ provides documented evidence that the equipment or system has been developed, supplied and installed in accordance with design drawings, the supplier's recommendations and In-house requirements. Furthermore, IQ ensures that a record of the principal features of the equipment or system, as installed, is available and that it is supported by sufficient adequate documentation to enable satisfactory operation, maintenance and change control to be implemented.
Operational Qualification (OQ)
OQ provides documented evidence that the equipment operates as intended throughout the specified design, operational or approved acceptance range of the equipment, as applicable. In cases where process steps are tested, a suitable placebo batch will be used to demonstrate equipment functionality.
All new equipment should be fully commissioned prior to commencing OQ to ensure that as a minimum the equipment is safe to operate, all mechanical assembly and pre-qualification checks have been completed, that the equipment is fully functional and that documentation is complete.
Performance Qualification (PQ)
The purpose of PQ is to provide documented evidence that the equipment can consistently achieve and maintain its performance specifications over a prolonged operating period at a defined operating point to produce a product of pre-determined quality. The performance specification will reference process parameters, in-process and product specifications. PQ requires three product batches to meet all acceptance criteria for in-process and product testing. For utility systems, PQ requires the utility medium to meet all specifications over a prolonged sampling period.
The PQ documentation should reference standard manufacturing procedures and batch records and describe the methodology of sampling and testing to be used.
What Gets Validated
General
All process steps, production equipment, systems and environment, directly used for the manufacture of sterile and non sterile products must be formally validated.
All major packaging equipment and processes should be validated. This validation is less comprehensive.
All ancillary systems that do not directly impact on product quality should be qualified by means of a technical documentation of the extent of the system and how it operates.
Facility
- Manufacturing Area Design.
- Personnel and material flow etc.
Process and Equipment Design
Process steps and equipment description. i.e. Dispensing, Formulating, Packaging, Equipment washing
and cleaning. etc
Utility Systems Design
Raw/purified steam, Purified water, Compressed Air, Air conditioning system, Vacuum, Power supply, Lighting, Cooling water, Waste etc
Computerized Systems Design
Information system, Laboratory automated equipments, Manufacturing automated equipments, Electronic records etc
Cleaning Validation (CV)
CV provides documented evidence that a cleaning procedure is effective in reducing to pre-defined maximum allowable limits, all chemical and microbiological contamination from an item of equipment or a manufacturing area following processing. The means of evaluating the effectiveness of cleaning involves sampling cleaned and sanitized surfaces and verifying the level of product residues, cleaning residues and bacterial contamination.
The term CV is to be used to describe the analytical investigation of a cleaning procedure or cycle. The validation protocols should reference background documentation relating to the rationale for "worst case" testing, where this is proposed. It should also explain the development of the acceptance criteria, including chemical and microbial specifications, limits of detection and the selection of sampling methods.
Method Validation (MV)
MV provides documented evidence that internally developed test methods are accurate, robust, effective, reproducible and repeatable. The validation protocols should reference background documentation relating to the rationale for the determination of limits of detection and method sensitivity.
Computer Validation
Computer Validation provides documented evidence to assure systems will consistently function according to their pre-determined specifications and quality attributes, throughout their lifecycle. Important aspects of this validation approach are the formal management of design (through a specification process); system-quality (through systematic review and testing); risk (through identification and assessment of novelty and critical functionality) and lifecycle (through sustained change control).
Where equipment is controlled by embedded computer systems, elements of computer validation may be performed as part of the equipment IQ and OQ protocols.

General process, cleaning and methodology validation concepts are described in this article with a special view to pharmaceutical industry

Definition of Validation Protocol

By Katina Blue, eHow Contributor

updated: September 11, 2010

I want to do this! What's This?

Validation protocol ensures that specific procedures are followed correctly.

business image by peter Hires Images from Fotolia.com

Validation protocol is a means of testing a process to ensure its effectiveness or validity. It helps businesses and other organizations reach specific goals through tests and examinations.

Purpose

1. Validation protocol helps determine whether certain rules or procedures are being done correctly. It employs specific instructions or guidelines that must be followed to accomplish an end result.

Example

2. Businesses use validation protocol for various reasons, including evaluating the final construction of a product. For instance, an auto manufacturing plant could devise certain guidelines and tests to measure how well employees are assembling parts. The tests would evaluate how the part is constructed from beginning to end based on the given set of rules. The use of tests to measure the success of an outcome, such as a properly crafted auto part, is a form of validation protocol.

Benefits

3. Validation protocol is useful for businesses, universities, hospitals and any other entities that need to attain specific goals. It helps companies and individuals assess if products are created according to standards. It also aids in ensuring that the desired outcome of a procedure is achieved.

Definition of Process Validation

Process validation is important to a wide range of industries, particularly that of pharmaceutical production. Without process validation, manufactured products would be both inconsistent and substandard.

Definition

1. Process validation is a procedure used to ensure that a certain process produces consistent, desirable results over a period of time. It guarantees that a given method for making products allows them to meet predetermined standards in a cost-effective way.

Areas Inspected

2. When a process undergoes validation, inspectors begin by checking to see that the equipment used in the process operates properly. Then, they check the results to make sure they fully meet quality standards. Finally, they observe the process over a period of time to ensure that the results are consistent.

Results

3. After observing these three areas, professionals compare the results to see if they match up with a predetermined set of standards. The process is then either validated or adjusted to make it fit the company's requirements.

Pharmaceuticals

4. Process validation is especially important to the pharmaceutical industry, because drug companies must be certain that their products are consistently identical. Because of this, the FDA strictly regulates process validation in this area.

Microbiological Aspects of Process Validation

EtO Sterilization: Microbiological Aspects of Process Validation

Deliberate decision making during the structuring of microbial challenges, product loads, and biological indicators can provide a validation process for EtO sterilization that ensures accuracy, the absence of microbes, and a smooth testing procedure.

Susan Edel Satter and Paul J. Sordellini

A companion to this article, EtO Sterilization: Principles of Process Design, discussed the components of each phase of two 100% EtO with nitrogen processes, focusing on the engineering aspects of designing EtO cycles.¹ This article focuses on various approaches to medical device sterilization cycle validation from a microbiological standpoint. The discussion assumes that the first two stages of the validation—the commissioning and the physical performance qualification of the sterilization chamber—have been successfully completed. Therefore, this discussion deals solely with the third stage of EtO validation: the microbiological performance qualification (MPQ). The topics that are discussed include how to choose the appropriate microbial challenge for an EtO sterilization process, approach to developing the EtO cycle, product load, placement of biological indicators in/on the product load, method for determining cycle lethality, and calculations to determine the D-value.

Information is also given on the documentation for the report on validation and certification of the process, and revalidation is discussed briefly. Some suggestions exceed the current requirements presented in international standards, but they can enhance a validation process, resulting in a more thorough and accurate study.

International standards ISO 9001 and 9002 present the quality system requirements for the design, development, production, installation, and servicing of healthcare products. The ISO 9000 series treats medical device sterilization as a special manufacturing process because the results cannot be verified by inspecting and testing 100% of the product at the conclusion of the cycle. Sterilization processes must, therefore, be assessed for special considerations, validated prior to use (or during use in certain situations), and routinely monitored.

To design an effective validation and routine control program for a sterilization process, the bioburden on the product and packaging must be considered. Bioburden is defined as "the population of viable microorganisms on a product and/or a package" and is characterized in terms of number, identity, and resistance. A validated test method must demonstrate that it can consistently and adequately remove the bioburden from the product and packaging. There are various bioburden test methods and associated validation procedures from which to choose.²

Many factors can contribute to the bioburden on product and packaging, including the origin of the raw materials and components, transit and storage conditions, and the manufacturing environment in which the finished products are assembled and packaged. European standards place particular emphasis on controlling the processes used to manufacture sterile products.³

Sterility is defined as the "state of being free from viable microorganisms."⁴ Microbial death relating to the gaseous sterilization of healthcare products is an exponential function typically defined as the probability of a nonsterile item existing per a given number of units in a batch. This probability, called the sterility assurance level (SAL), defines the "probability of a viable microorganism being present on a product unit after sterilization."⁴ While sterilization can reduce the bioburden on a given product to a very low number, that probability can never be reduced to zero. Therefore, in order to achieve the desired bioburden levels, it is critical to design a validation program that provides a high degree of confidence for consistent sterilization.

DETERMINING THE APPROPRIATE MICROBIAL CHALLENGE

A biological indicator (BI) is an "inoculated carrier contained within its primary pack providing a known resistance to the relevant process."⁴ There are many different types of BIs but the most common include:

• Commercial units supplied with the manufacturer's certification, such as bacterial spore strips individually packaged in glassine envelopes or pieces of filter paper impregnated with a certified population of a challenge organism that has a known resistance to EtO, such as the spores of Bacillus subtilis var. niger.
• A liquid bacterial spore suspension commercially sold and certified by the manufacturer. The suspension is placed on or in the product, which is then referred to as inoculated product.
• A liquid bacterial spore suspension made by a device manufacturer using a commercial, certified strain of a bacterium.
• Strains of microorganisms that have been isolated from a device manufacturing facility. These strains represent the most resistant organisms found in or on the devices and are typically used for the combined BI/bioburden method of validation.

It is crucial to ensure that the type of BI used to validate or routinely monitor a given sterilization process is the most appropriate indicator for that process. In addition to identity, quantitation, resistance, storage, general directions for use, and disposal conditions, the manufacturers of BIs are required to provide information regarding the optimal culturing conditions, such as temperature and type of growth media.

Irrespective of which BI is chosen, the methods used to recover the challenge organism must be validated. This recovery is expressed in terms of the percent recovery of the original inoculum. These recovery studies can be especially challenging when using liquid spore suspensions because of the interaction between the suspension and the material onto which it is inoculated. The material substrate can alter the resistance characteristics of the inoculum because of such anomalies as spore clumping or the physical sheltering of spores in certain sites within the product.

The goal is to kill the microbes, which means disabling their ability to reproduce even in their most favorable growth conditions as described by their manufacturer. There have been records of seemingly killed microbes that regenerated when conditions become favorable.⁵ The user must validate that the incubation time under the prescribed conditions is sufficient to recover delayed growth of the organisms after exposure to a given sterilization process. For routine processing, this time period is typically 7 days unless validated for a shorter time period in accordance with current national requirements.⁶ In such cases, periodic checks should be run to confirm that the shorter time period yields equivalent recoveries to those obtained from the longer incubation period. It is also important to ensure that the incubation time is sufficient to recover growth from injured organisms exposed to sublethal cycles. In some cases, this may mean using a 14-day incubation period. This incubation period is also required by the U.S. Pharmacopeia for product sterility testing.⁷

Just as the BI must provide a defined resistance to a specified process, it is necessary to prove that the inherent bioburden on the product does not have a greater resistance than the BI. Characterizing bioburden involves quantitation, identity, and resistance of the bioburden. Several methods can be employed to determine which BI is appropriate for a specific situation.

• Challenge organisms that make up BIs typically resist a particular sterilization process far more than do common bioburden organisms. The resistance of the bioburden cannot be adequately evaluated by quantitation only, yet it must be determined if the products to be processed might contain organisms that are more resistant than the challenge organism. After the microbial identifications and quantitation of the bioburden have been completed and analyzed, comparative resistance determinations of the most resistant bioburden component or product must be calculated. A literature review is also required.
• When microbial identifications of the bioburden are not performed, at least one sublethal cycle should be run to compare the relative inactivation rates of the bioburden with that of the challenge organism. Product sterility testing, after exposure to at least one sublethal cycle under appropriate experimental conditions, can ensure that the product's bioburden is not more resistant than the challenge organism.
• If the quantity, identity, and resistance of the product's bioburden are known, it might be possible to validate and routinely monitor the sterilization process by combining BI and bioburden methods. It must be demonstrated that the BI's degree of challenge to the sterilization process is adequate to ensure that the process will attain the desired SAL for the bioburden. Combining the BI and bioburden methods to determine the appropriateness of the BI can be time-consuming and result in additional testing costs. However, the required sterilization parameters can be more accurately determined, which can result in reduced processing time and reduced exposure to the sterilant.

BIs can be configured in many different ways depending on the cycle development method chosen.

Inoculated Product. The actual product, configured and packaged as it is intended to be sold, can be inoculated with spores of a microorganism such as Bacillus subtilis var. niger. Direct inoculation usually uses spores suspended in liquid, then placed on the product and dried. The product's surface characteristics will affect the distribution of spores and may lead to a difference in resistance behavior compared with other challenge systems.⁴ It is, therefore, important to achieve an even distribution of spores on the product's surface. Indirect inoculation involves placement of a carrier, such as filter paper that has been impregnated with spores, in the product or its package.

Inoculated Unit. A carrier, such as a filter paper strip or disk, can be inoculated with a population of a resistant organism, such as Bacillus subtilis var. niger, that has been extensively characterized and certified by the manufacturer. The resistance of this inoculated carrier must be compared to that of the inherent bioburden of the product being validated or the equivalent simulated product. An inoculated unit is usually used when there is the potential that the bioburden on the product is more resistant than the indicator organism and is required for the combined bioburden and BI cycle development method.

Inoculated Simulated Product. A simulated product that comprises the most difficult to sterilize portions of a device or that configuratively represents a device family can also be directly or indirectly inoculated. This simulated product must present the greatest challenge to the process in order to be considered an adequate microbial challenge. Each unit must contain a certified inoculum either in liquid form or on a carrier.

Natural Product. The inherent bioburden on the product can also be used as the microbial challenge during validation and for routine monitoring when the absolute bioburden method is employed for cycle development (see "EtO Cycle Development Approaches", below).

All validation methods for EtO sterilization require that the BI used for validation and to monitor routine cycles must be more resistant than the bioburden of the product and be placed in a location that is more difficult to sterilize. Comparative resistance testing is an effective means of selecting the BI and its location in the product that presents the greatest challenge to the sterilization process. Such an assessment should be made prior to validation as part of determining the appropriateness of the BI. These studies are usually carried out in small chambers that are capable of delivering rapid ramp rates, e.g., the times required to achieve specific pressure set points.

Products should be exposed to cycles in which the only variable is the gas exposure time period. The data obtained from this testing can be used to justify the choice of a specific actual or simulated product to inoculate and use for the BI. If the design of the product is such that a BI unit cannot be placed in the part of the device that is the most difficult to sterilize, the product should be inoculated with a liquid spore suspension to provide a known number of viable spores. The spore suspension, materials, and techniques used should comply with ISO 11138, parts 1 and 2.^8,9

Many device manufacturers include an additional objective in their validation plan that involves the use of external BI monitoring systems. Often referred to as process challenge devices (PCDs), they assess the lethality of the EtO process after the cycle has been designed. The PCDs are geometrically distributed around the load rather than in internal locations in the case cartons. Direct comparisons can then be made between the sterility test data obtained from these external PCDs and the BIs placed in internal locations. A PCD must be shown through comparative resistance studies to provide more of a challenge to the process when it is placed in external locations in the load than do the the BIs placed in internal locations. They usually, therefore, bear no resemblance to the product. Examples of external PCDs are spore strips double-packaged in plastic bags, in sealed plastic tubing, or in syringes. There are also commercially available PCDs that are sold as ready-to-use packaged systems. It is advisable during the validation studies to evaluate different PCD configurations during the comparative resistance studies to determine the best candidate. To monitor routine sterilization cycles, it must be shown at the time of validation that the PCDs in the external locations comply with the same requirements for resistance to sterilization.

ETO CYCLE DEVELOPMENT APPROACHES

There are three basic approaches to developing EtO sterilization cycles—the overkill method, the combined bioburden and BI method, and the absolute bioburden method.

The overkill method is probably the most widely used because it is relatively easy to use and it results in a robust SAL. The method ensures that the sterilization process will inactivate a specific number of microorganism spores known to be resistant to the EtO sterilization process. The organism most commonly used to monitor the overkill process is Bacillus subtilis var. niger. A certified preparation consisting of a stated population of Bacillus subtilis var. niger spores is inactivated through exposure to specific cycle parameters that have been assessed to be significantly higher than those required to kill the inherent bioburden on the product. The parameters are increased on a routine basis to provide the desired SAL (see "Methods for Determining Cycle Lethality," below).

The combined bioburden and BI method is used when the two are equally resistant. This method requires routine bioburden and BI testing in addition to a considerable amount of routine sterility testing to develop a cycle that will inactivate the BI challenge population. The BI must be sufficiently resistant to ensure that the EtO process will deliver the desired SAL relative to the bioburden on the product.

The absolute bioburden method is used less frequently in cycle development because it requires extensive testing in both the development phase and routine processing. However, it must be used when the product's bioburden is more resistant than the BI. Such bioburden resistance to the EtO process can be caused by any number of factors, such as the configuration of the product, the quantity or location of the microorganisms, or the bioburden's intrinsic resistance. Since the bioburden on the product constitutes the essential microbial challenge for the process, the bioburden test method must be validated and strictly controlled. The resistant microorganisms are screened through bioburden testing and may be isolated and propagated for use in cycle development studies. One negative of this method is that the microorganisms' resistance can change as a result of how they are cultured, which can adversely affect the results of the cycle development studies. The absolute bioburden method also requires extensive controls of the manufacturing environment in addition to routine product bioburden monitoring and resistance studies.

DETERMINING THE APPROPRIATE PRODUCT LOAD CHALLENGE

Microbiological performance qualification (MPQ) should be performed using specified products and packaging configured in the same manner in which they will be routinely sterilized. For the cycle to be accurate, the product load must represent the greatest challenge intended for future routine sterilization. If a device manufacturer intends to use multiple load configurations on an ongoing basis, the densest configuration should be used for the MPQ.

Each type of configuration must be documented in terms of the number of product units per case, the number of cases per pallet, the stacking patterns on the pallet, and the density. This documentation should be included with the validation data. Some testing should also be conducted on the least dense configuration, which, theoretically, presents less of a challenge to the process. This testing can be as simple as placing thermocouples throughout the least dense load on a routine cycle and comparing the temperature distribution with that of the densest load. In other cases, additional microbial challenge studies might be required. Changes in the product load must be evaluated carefully because seemingly innocuous changes, such as changing the shrink wrap or corrugate on the load, can have a significant effect on the cycle's efficacy from the perspective of product sterilization.

BI PLACEMENT IN THE PRODUCT LOAD

After the product load challenge has been identified, the BI positioning and placement can be determined. BIs should be distributed throughout the product load and, as much as possible, in the same orientation (e.g., vertical). The placement must include those locations that are considered to present the greatest challenge to the process and can be the same as those used for temperature monitoring. The ANSI/AAMI/ISO 11135-1994 standard suggests placing two BIs at each location with a temperature-monitoring device in order to obtain additional information on process efficacy. It also provides the following recommendation for the number of BIs to be included in each validation cycle:

• At least 20 BIs for usable chamber volumes up to 5 m³.
• Increase the number of BIs by two for every additional 1 m³ of usable sterilizer chamber volume between 5 and 10 m³.
• Increase the number of BIs by two more for every additional 2 m³ of usable sterilizer chamber volume above 10 m³.

The AAMI technical information report "Contract Sterilization for Ethylene Oxide" can provide additional information on the number of BIs and monitoring devices recommended based on product load volume.¹⁰

METHODS FOR DETERMINING CYCLE LETHALITY

Results obtained from commissioning and physical performance qualification and monitoring devices should be used to identify critical features of the equipment or process that can be investigated during the MPQ. For example, it is critical that the sterilant injection time is consistent among the MPQ cycles to ensure a uniform delivery from one cycle to the next. Even minor changes in the sterilant injection time can result in significant differences in lethality.

The MPQ should be performed in the industrial chamber that will also be used for routine processing unless equivalency can be demonstrated between the industrial chamber and whatever chamber is used for the qualification. Maintaining the precise and consistent delivery of the sterilization cycle parameters is more difficult to accomplish in large industrial chambers than in small test chambers. It is also important to conduct these studies using the actual product load intended for routine sterilization. Hence, these studies are usually conducted in large industrial chambers rather than in small test chambers.

The MPQ can be performed by determining the lethality of the cycle on the basis of the number of D-values applied. The D-value is defined as "the time required to reduce a specific microbial population by 90% or one logarithm."⁴ The survivor curve construction or fraction-negative methods (described below) may be used as outlined in current standards.⁴ Another means of evaluating the MPQ is the half-cycle method, based on the number of times required to completely inactivate the BI microorganisms with an added margin of safety. The ultimate objective of each method is to determine the full cycle to which the product load must be exposed.

Survivor Curve Construction Method. The survivor curve construction method involves the direct enumeration of survivors in terms of colony forming units (CFUs) recovered after exposure to graded amounts of the sterilization cycle. A CFU is defined as "a visible outgrowth of a population of organisms arising from a single or multiple cells."² A minimum of five cycles should be run, each using different graded time exposures to EtO.⁴ The parameters used, with the exception of the gas exposure time, must be kept consistent. The first cycle is a time zero study in which the initial CFU survival count of the BI is determined by exposing the BIs to all stages of the process, including preconditioning if used, prior to the EtO injection phase of the cycle. All BIs should survive because they will not be exposed to the sterilant.

After each of the four or more additional cycles, all employing different gas exposure time periods, the number of BIs that survive the processes are counted. The BIs should be removed from the chamber and the load as soon as possible within the confines of worker exposure policies. The BIs should also be tested as soon as possible after being removed to reduce their exposure to EtO residuals, which can affect BI survival rates. In all cases, the time intervals between when the load is removed from the chamber, when the BIs are removed from the load, and when they are subjected to the enumeration process must be consistent among the cycles. Ideally, a final enumeration of the BIs from one cycle should be obtained before the next cycle is initiated to more accurately assess the exposure time to use. This is not always feasible because the enumeration process can take days or weeks to complete.

The number of BIs used in each of the cycles in the study should be statistically significant to ensure obtaining dependable data. The number can also be based on the size of the chamber or the size of the product load. The data acquired in the study are used to calculate an EtO exposure time and the minimum process parameters expected to elicit a specific probability of survival of the challenge organism expressed in CFUs (see "The Full Cycle," below). Theoretically, because all other process parameters are the same, the statistical evaluation of the data should result in a plotted survivor curve or regression analysis that demonstrates a consistent relationship between the EtO gas exposure time and the number of survivors (positive BIs). Unfortunately, this is not always the case. In addition to the issue of consistency between the BIs, the integrity of the data is dependent on the consistent delivery of the process parameters for each cycle, including precise control of gas injection times that can be considered part of the gas exposure phase for the purposes of this study. Controlling the gas injection time in large industrial EtO chambers necessitates ensuring that the head space pressure in the gas tanks is consistent from one cycle to another and that the gas delivery lines are uniformly either full or purged of gas. A few minutes difference in gas inject time can significantly change the results in this study. Other variables to consider include the temperature of the gas volatilizer, evacuation rates and times, gas makeups, and air exchanges.

If the survivor curve study is conducted in an industrial chamber, the data should elicit an accurate probability of survival of a specific challenge organism from which routine cycle parameters can be determined. The BI should be an inoculated carrier. For example, if the BI is a spore strip that contains a population of at least one million (10⁶) spores of Bacillus subtilis var. niger and the construction of the survivor curves demonstrates that total kill was obtained at 2 hours of gas exposure time, the full cycle used to routinely sterilize the product to an SAL of 10^–6 could employ 4 hours of gas exposure. To achieve a smaller SAL, for example 10^–3, would require adding the appropriate additional gas exposure time to the original 2 hours.

The BI testing process should be validated prior to initiation of the survivor curve study to obtain an acceptable and documented recovery method. The process begins with macerating the BIs in sterile water. The suspension is evaluated by plating specific dilution aliquots of serial dilutions of the suspension onto a selected agar medium and counting the number of CFUs after incubation. The number and extent of dilutions needed will be based on the duration of gas exposure and concentration in relation to the number of BI organisms. In other words, more dilutions are required when more survivors are expected.

A statistical analysis made from each cycle of survival data should show the log₁₀ of the surviving population plotted against EtO exposure time intervals. The best-fit rectilinear curve through the data can be drawn or determined by regression analysis using the method of least squares.⁸

The survivor curve method is complicated by the number of serial dilutions that must be prepared and the quality of the dilutions being dependent on the skill of the person performing the test as well as the precision of the equipment used. Only trained personnel who can adequately practice aseptic technique should conduct this test. Calibrated pipettes and dilution controls help ensure the test's accuracy. The dilutions should also be chosen to yield counts between 30 and 300 CFUs. It is generally assumed that numbers ranging from 30 to 100 CFUs should be used because it is thought that higher numbers of CFUs per plate could result in inaccurately low counts and that numbers lower than 10 CFUs per plate could give unreliable counts. For practical purposes, counts between 30 and 300 are generally acceptable.

Fraction-Negative Method. The fraction-negative method also involves exposing BIs to multiple cycles of graded exposures to EtO. The differences between the two methods are the number of cycles recommended and the number used to enumerate the survivors (in terms of the positive BIs).⁴ A minimum of seven cycles should be employed in this study, each utilizing different gas exposure time periods. These seven cycles should elicit the following survivor data:

• At least one sample set that elicits all survivors (growth in all BIs tested).
• At least four sample sets that elicit fractional data, i.e., a fraction of the BIs in each set demonstrates growth or survival.
• At least two sample sets in which there is neither growth nor survivors.

The method used to enumerate the number of positive BIs is more straightforward than in the survivor curve method. In the fraction-negative method, the BIs are immersed directly into the appropriate media and incubated. Results are recorded in terms of the total number of BIs demonstrating growth and the total number eliciting no growth for each set of test samples.

METHODS FOR DETERMINING CYCLE LETHALITY

Calculating D-Values. The D-value corresponding to the survivor curve construction results is determined by either reading it from a graph or calculating from the data the relevant time interval for reducing the count by one log (Table I). The performance of the fraction-negative method will generate data that are then utilized to calculate a D-value using a method described by Pflug and Holcomb.¹¹ This method is also referred to as the full or generic Spearman-Karber procedure. It does not require the use of the same number of replicates nor the same gas exposure time intervals because it involves spore strips rather than counting CFUs.

Time of Exposure to Sterilant (t)	Number of Test Samples Exposed (n)	Number of Test Samples Showing No Growth (r)
t1	n1	r1
t2	n2	r2
t3	n3	r3
t4	n4	r4
t5	n5	r5
t6	n6	r6
t7	n7	r7

Table I. Variables used in calculating D-values during MPQ.

In Table I, t₁ represents the shortest exposure time to sterilant, and all test samples run through this short time frame should show growth. The variables t₂ to t₅ correspond to increasing exposure times in the fraction-negative area, otherwise known as the quantal region. Exposure times t₆ and t₇ should represent tests in which none of the tests show growth.

In the following equations, r₁ is the number of test samples out of the number exposed (n_i) that show no growth at exposure time t₁. For each period of exposure to sterilant t₁ to t₆, the factors x and y are calculated as shown:

At t_i, all test samples show growth, therefore,

From the calculated values of x_i and y_i above, the value µ_I (mean time to attain no growth) can be calculated for each period of exposure (t₁ to t₆) as follows:

The mean time to attain no growth from all of the test samples, µ, can be calculated from the sum of µ_I for each time of exposure t₁ to t₆:

Where the interval between exposure times (d) is constant and the same number of test samples (n) is used at each exposure time, the mean to attain no growth (µ) can be calculated from the equation:

The mean D-value (D) can be calculated from the equation:

where N₀ = the initial number of challenge organisms per test sample.

For the purposes of calculating the sterilization period using the D_calc method below, the upper 95% confidence level for D should be used. This can be calculated from the equation:

where V is derived as follows:

In addition to the method described above, there are three other commonly used means of calculating D-values. The limited Spearman-Karber procedure requires the same number of replicates for each exposure time and equal intervals between the various gas exposure times. For example, the exposure times of 9, 12, 15, and 18 minutes provide an arithmetic series of 3-minute time intervals.

The Stumbo-Murphy-Cochran procedure requires one result in the fraction-negative range, consisting of time (t), the number of units negative for growth (r) and the number of replicates (n) at each exposure time as well as the initial number of microorganisms per replicate (N₀). The D-value for each exposure time is calculated from the following equation. The confidence limits of the mean may be calculated by conventional procedures using natural logs.

The factors in the equation represent the following:

U = exposure interval.

N₀ = initial number of organisms per replicate carrier unit.

N_U = ln (n/r).

n = total number of replicate units at exposure U.

r = number of units negative for growth at U.

The D-values are averaged to obtain the overall D-value for the experiment.

The limited Stumbo-Murphy-Cochran procedure requires only one exposure time in the fraction-negative range.

It is important to choose the most appropriate procedure of the three described to calculate the D-value and its upper confidence limit. Certain methods have limitations. For example, Graham and Boris have noted that a confidence interval cannot always be determined from the Stumbo-Murphy-Cochran procedure.¹² Shintani et al. have observed that the Stumbo-Murphy-Cochran procedure can yield invalid results if r is close to n.¹³ They also state, however, that this procedure may be superior in situations in which the sample population is at least 50, r is at least 1, and r/n is less than 0.9.

Half-Cycle Method. The half-cycle method determines the minimum time a specific product load must be exposed to an EtO process to guarantee that no survivors exist from the BIs used to monitor the cycle's efficacy. The cycle chosen is based not only on the elements discussed in the companion to this article but also on the microbiological data obtained prior to determining the parameters for the half-cycle.¹ The minimum ranges of the half-cycle are based on this predetermined microbiological data, whereas the maximum ranges are dictated by the product and packaging tolerances established through functional testing.

If the half-cycle method is chosen, an additional cycle should be performed, preferably before the first half-cycle is conducted. In some cases, other proportionate cycles, referred to as sublethal or fractional cycles, may be used. The gas exposure time is shortened in order to obtain survivors from the BIs used to monitor the cycles. The process parameters in a sublethal cycle are the same as those in the half-cycle with the exception of the gas exposure time period. Such cycles are conducted to demonstrate that positive BIs can be recovered and to prove the adequacy of the BI, the sterility test method, and associated equipment and materials. The sublethal cycle is not required if either the survivor curve construction or fraction-negative method is used because these methods require positive sterility test results as part of the study.

A second objective in conducting a sublethal cycle is to attempt to demonstrate that the resistance of the product's bioburden is less than that of the BI. This is accomplished by using actual product test samples that have been manufactured, packaged, and handled in the same manner intended for routine production. It is imperative that these test samples were made in accordance with routine manufacturing procedures because the bioburden on the product must represent what the product would normally contain. Sometimes using a number of different product samples can yield useful information when there is concern about not only the quantity of bioburden but also where it is located. If a number of different samples have been sterility tested, those with the highest bioburden should be used for validation. After the completion of the sublethal cycle, the product samples are sterility tested. Since the objective of the test is to demonstrate sterility of the bioburden, the data can be compared to the positive sterility test results obtained from the BIs to demonstrate that the resistance of the product's bioburden is less than that of the BI.

A third objective of the sublethal cycle study is to determine that a PCD used as an external monitoring device for routine sterilization cycles yields more positive sterility tests than an internal BI does. It has been suggested that it is acceptable to obtain positive sterility test results from those external PCDs exposed to the half-cycles if previous comparative resistance studies have proven that they are as resistant or more resistant than the internal product BI and if no positive sterility test results are obtained from the BIs placed in internal load locations. If total kill is obtained from all PCDs on all half-cycles, it can be assumed that the resulting validated full cycle will provide an additional safety factor that goes beyond the minimum requirements.

Typically, the sterilization efficacy of the first half-cycle is determined before any subsequent cycles are conducted. If the first half-cycle experiment elicits all negative BIs, two additional half-cycles are conducted to confirm the data. All three half-cycles must employ the same parameters to demonstrate reproducibility and reliability. All three half-cycles must demonstrate the ability to elicit all negative BIs. If a SAL of 10^–6 is required, a gas exposure time that is at least double that used in the half-cycle becomes the minimum gas exposure time used routinely in the full cycle.

THE FULL CYCLE

The data obtained from the survivor curve construction method, the fraction-negative method, or the half-cycle method are used to design the full cycle that will be used to routinely sterilize the product. The full cycle must be capable of reliably demonstrating a required SAL that consists of the minimum time to obtain all negative BIs or CFUs with an additional margin of safety. Normally this can be expressed as 10^–n where n is the cumulative probabilities of the log minimum time to sterilize and the log margin of safety. For example, if the half-cycle method was chosen to validate a given sterilization cycle with a required SAL of 10^–6, the half-cycle must demonstrate the ability to ensure that there are no survivors from BIs that have a certified population of 10⁶. Since the BIs have a greater resistance than the product bioburden, it can be concluded that the time is sufficient to achieve product sterility. However, adding an equivalent sterilization time period increases the margin of safety. The additional 6 log reduction in the population (10^–6) is theoretical and is obtained by doubling the half-cycle gas exposure time. In other words, if the half-cycle gas exposure time period was 2 hours, the full-cycle gas exposure time would be a minimum of 4 hours.

The processing ranges established for the critical parameters in the full cycle should be based on the ranges determined in the half-cycle. The minimum process parameters in the full cycle, such as temperature, sterilant concentration, pressure, and humidity, may also be greater than those used in the half-cycle to ensure greater lethality with the full cycle. The first full cycle should be included in the validation to complete the profile of the product load. The same type and amount of monitoring should be conducted during the first full cycle as was conducted on each of the half-cycles. Comparisons of temperature distribution throughout the chamber and the load can justify a determination of reliability and repeatability for the process. These data can also be useful for determining the significance of any minor changes that are made to the load.

REPORT ON VALIDATION AND CERTIFICATION

The documentation on the validation and certification should include but not be limited to the following:

• References to the maintenance and calibration procedures for the processing equipment.
• Specifications for the EtO chamber.
• References to the commissioning data.
• An indication that all gauges, monitoring devices, etc., were calibrated prior to initiation of and at the conclusion of the validation.
• The validation protocol.
• Comparative resistance test data and reference to the protocol.
• A complete description of the products used as test samples, including packaging.
• BI certification and all data, according to current standards, required in the labeling.
• The physical and biological records from all of the validation cycles.
• Placement diagrams and data for the monitoring devices (including temperature, gas concentration, and relative humidity).
• Documentation on and verification of the positioning and location where the BIs and product test samples were placed on the product load.
• BI population determination test results and statement of retention samples.
• All biological test data associated with the validation.
• EtO residual test data.
• References to the test procedures used during the validation.
• Documentation of operating procedures, including process control limits.
• A test report summarizing and analyzing the validation data.

ANNUAL REVALIDATION

An annual revalidation of the process should be conducted to verify the integrity of the original validation data. It may be advisable to conduct this revalidation within 13 months of the initial validation and each year thereafter until sufficient data are obtained to allow that time period to be extended. Afterwards, revalidation should be conducted at least every 2 years.

Data from each revalidation should be compared with that from the original validation and any subsequent revalidation to confirm that the original performance specification remains valid. A revalidation consists of a thorough review of all the factors that could affect the sterilization process, including changes to the product or its packaging, the manufacturing methods and facility, and component and materials suppliers. Product bioburden test data should be analyzed and trends noted to ensure that there have been no substantial unacceptable changes in quantity or characterization. The testing conducted of the preconditioning room, the sterilization chamber and ancillary equipment, and the aeration room and equipment should be compared with the data obtained during the original validation as well as to the commissioning data. All associated programs, such as preventive maintenance and calibration, should be reviewed for effectiveness.

CONCLUSION

A well-designed and scientifically sound MPQ helps ensure that the process is safe, efficacious, and efficient. Performed well, the MPQ should deliver the required SAL for the product in an economical process and should enable the process to continue without having to be repeated. It should also prevent product reprocessing and delays in product release to the marketplace. A knowledgeable approach that considers all of the various aspects of the MPQ will ensure the success of the process.

ACKNOWLEDGMENT

The authors would like to express their gratitude to Aubrey S. Outshoorn, PhD, of the United States Pharmacopeial Convention Inc. for his contributions to this article.

REFERENCES

1. PJ Sordellini and SE Satter, "EtO Sterilization: Principles of Process Design," Medical Device & Diagnostic Industry 20, no. 12 (1998): 47–59.

2. "Sterilization of Medical Devices—Microbiological Methods—Part 1: Estimation of Population of Microorganisms on Products," ANSI/AAMI/ISO Standard 11737-1 (Arlington, VA: Association for the Advancement of Medical Instrumentation, 1995).

3. "Guidance on the Application of EN 29001 and EN 46001 and of EN 29002 and EN 46002 for Non-Active Medical Devices," BS/EN 724 (London: British Standards Institution, 1995).

4. "Medical Devices—Validation and Routine Control of Ethylene Oxide Sterilization," ANSI/AAMI/ISO Standard 11135 (Arlington, VA: Association for the Advancement of Medical Instrumentation, 1994).

5. EW Nester et al., Microbiology, 2nd ed. (Chicago: Holt, Rinehart, and Winston, 1978).

6. The Center for Devices and Radiological Health, FDA Guide for Validation of Biological Indicator Incubation Time (Rockville, MD, FDA, 1985).

7. United States Pharmacopeia 23, 8th supp., monograph <71>, 1998.

8. "Sterilization of Health Care Products—Biological Indicators—Part 1: General Requirements," ANSI/AAMI/ISO Standard 11138-1 (Arlington, VA: Association for the Advancement of Medical Instrumentation, 1994).

9. "Sterilization of Health Care Products—Biological Indicators—Part 2: Biological Indicators for Ethylene Oxide Sterilization," ANSI/AAMI/ ISO Standard 11138-2 (Arlington, VA: Association for the Advancement of Medical Instrumentation, 1994).

10. "Contract Sterilization for Ethylene Oxide," AAMI TIR no. 14-1997 (Arlington, VA: Association for the Advancement of Medical Instrumentation, 1997).

11. IJ Pflug and RG Holcomb, "Principles of Thermal Destruction of Micro-Organisms," in Disinfection, Sterilization and Preservation, ed. SS Block, 3rd ed. (Philadelphia: Lea and Febiger, 1983).

12. GS Graham and CA Boris, "Chemical and Biological Indicators," in Sterilization Technology: A Practical Guide for Manufacturers and Users of Health Care Products (New York: Van Nostrand Reinhold, 1993).

13. H Shintani et al., "Comparison of D₁₀-Value Accuracy by the Limited Spearman-Karber Procedure (LSKP), the Stumbo-Murphy-Cochran Procedure (SMCP), and the Survival-Curve Method (EN)," Biomedical Instrumentation Technology 29, no. 2 (1995): 113–124.