A Practical Approach toCommissioning and Qualification -A Symbiotic Relationshipby Timothy D. Blackburn, PE
This articleoffers apracticalapproach tocommissioningand a method ofsynergy withqualification.Practicalapplications areexplored, aswell as the useof EnhancedCommissioningDocumentationto minimize thequalificationeffort.
An initial response to the recommenda-tion to perform commissioning is thatit is just an additional step – anotherroadblock to engineering success and
something repeated during qualification. How-ever, effective commissioning supports engi-neering and qualification success. This articleaddresses efficient commissioning techniquesand synergizing with qualification. Examplespresented are not all definitive, and documen-tation may exceed or not include certain ele-ments – commissioning (and qualification) mustbe structured for the project.
Commissioning StreamlinesQualification
Effective commissioning results in a focusedand first-time-success validation effort. Thereare many ways commissioning can benefit quali-fication – reduce costs (but don’t overstate), aless rigorous documentation regimen (exceptfor enhanced commissioning requirements),tests are closer to the source (suppliers, con-tractors, etc.) and therefore are often moremeaningful, debugging/trouble shooting is mini-mized during qualification, faster qualifica-tion, catch problems qualification might miss,
better schedule attainment, and better project quality attain-ment, better customer satisfaction (when they finally realizethe value of commissioning).
There are good reasons formal commissioning is needed,many of which are directly related to more efficient qualifica-tion. The following are a few examples:
1. Ratcheting Validation Costs - each project has the ten-dency to “one up” the previous one and qualificationsuccess may be graded by the weight of the paper gener-ated.
2. Validation, a Debugging Exercise - due to a lack of propercommissioning, problems may be discovered during quali-fication that add cost, schedule duration, and undo stress.Validation should be a one-shot exercise and successfullycompleted as much as possible on the first try.
3. Overly Extensive Validation, Undue Lifecycle Burden -there is a tendency to over-qualify due to a lack of confi-dence in the installation (actually due to a lack of adequatecommissioning), which not only adds initial cost, butunnecessary lifecycle maintenance of a validated state.This over-qualification may extend to areas not associatedwith product quality, and is not necessary when effectivecommissioning is applied.
4. Repeating Informal Commissioning Activities - Mostprojects include some level of Commissioning, which areoften repeated during qualification.
Validation/Commissioning: The DistinctionsIt is important to understand the definitions of Validation,Qualification, and Commissioning to determine the distinc-tion and how they can effectively work together. First, valida-tion is defined as “establishing documented evidence whichprovides a high degree of assurance that a specific processwill consistently produce a product meeting its pre-deter-mined specifications and quality attributes.”1 Qualification isa subset of validation which includes IQ/OQ/PQ, and isdefined as “The documented verification that all aspects of afacility, utility, or equipment that can affect product quality...adhere to approved specifications” (Installation Qualifica-tion or IQ) ... operate as intended throughout all anticipatedranges (Operational Qualification or OQ) ... perform as in-tended meeting predetermined acceptance criteria”2 (i.e.,over time Performance Qualification or PQ).”
Commissioning is defined as “a well planned, documented,and managed engineering approach to the start-up andturnover of facilities, systems, and equipment to the end-userthat results in a safe and functional environment that meetsestablished design requirements and stakeholder expecta-tions.”3 In other words, commissioning verifies what wasspecified was installed, that it functions properly, and it wassuccessfully turned over to the user, and reasonably ensuresqualification success (avoid qualification becoming a trouble-shooting exercise). For cGMP, formal commissioning pro-
vides necessary documentation to verify and record commis-sioning was done and supports qualification documentation.
Note the distinction between the two definitions. Thevalidation/qualification definition emphasizes product; thecommissioning definition emphasizes equipment. Validation/qualification is primarily concerned with and verifying as-pects that could affect product quality. Commissioning isconcerned with Good Engineering Practice (GEP) and quali-fication success, and is an equipment/system/facility focus.When commissioning is properly implemented, qualificationcan focus on what is important – aspects that could affectproduct quality. Defining qualification and commissioningearly in a project also allows commissioning to emphasizedirect impact elements to ensure qualification success.
The “W” ModelCommissioning supports qualification relationally; for ex-ample, inspection activities support and are similar to IQ,and testing activities support and are related to OQ/PQ.Factory Acceptance Tests (FATs) and Site Acceptance Tests(SATs) support and are similar to the overall qualificationeffort. Figure 2, a “W” Model, illustrates the relationshipbetween design, commissioning, and qualification. This issimilar to the familiar “V” model, except a center portion isadded to illustrate the commissioning relationship. The pri-mary User Requirement Specification (URS) or similar docu-ment defines the high level, low detail fundamental require-ments of the project. Certain Commissioning FunctionalityTests should verify the URS was complied with, which leadsto PQ. Commissioning testing activities also should suffi-ciently verify the installation complies with the FunctionalRequirement Specification (a somewhat more detailed docu-ment than the URS), which leads to OQ. Commissioninginspection activities should sufficiently address the detailedspec, which leads to IQ. FAT/SAT documents may includemost commissioning testing/inspection elements for someprojects, and therefore be relational to all the design docu-ments and lead in to related qualification.
InVEST Wisely in CommissioningWhen establishing commissioning requirements, it is impor-tant to remain focused on common sense objectives to makethe effort meaningful and cost effective. The acrostic “InVEST”is helpful in establishing the focus:
• Integrate: integrate commissioning with qualification.Don’t automatically do things twice.
• Verify: does the commissioning activity adequately verifythe equipment or system is what was specified and worksas it should?
• Ensure Qualification Success: does the commissioningeffort sufficiently ensure qualification will be successful –first time?
• Sensible: do enough, but don’t over do it.• Traceable: document it. Remember the saying, “if you
Before developing commissioning documentation, establishthe extent of commissioning needed, and design efficient andeffective commissioning around the needs of the project(hopefully as expressed in a well-written URS/FRS.) Effec-tive commissioning documentation defines the commission-ing process (with signatory approval when needed), definessetting to work verifications, inspections, and tests; mayconfirm training completion (the project is not complete untilusers know how to use it); and may confirm documentationturnover (the project is not complete until drawings, specs,and O&M manuals are turned over to record/as-built condi-tion and enable users to operate/maintain).
Typical Commissioning Documents may include the fol-lowing, depending on project complexity - Figure 1.
• Overall Commissioning Plan - for large and more complexprojects - this is a master plan for commissioning when theapproach needs preplanning and structure. On smallerprojects/single equipment, consider relying on StandardOperating Procedure (SOP) requirements rather than aseparate overall Plan.
• Pre-Commissioning: includes Factory Acceptance Test(FAT), Site Acceptance Test (SAT), and possibly otherearly inspection/test activities. These are usually struc-tured for individual systems, and can be included in or
required by the commissioning plan. These could be standalone for individual equipment/systems, and/or includeessential elements of the commissioning test/inspectionplans.
• Commissioning Test and Inspection Plans: these could bestand alone for individual equipment/systems. These alsomay supplement areas not covered by FATs/SATs. Fur-ther, self-contained commissioning checklists can be usedfor simple/small work. Don’t create unnecessary volumesof documentation.
Enhanced CommissioningCertain commissioning activities need not be repeated dur-ing qualification. It is possible to do commissioning activitiesthat satisfy elements of qualification. This is called “En-hanced Commissioning.” Documentation created by enhancedcommissioning is considered sufficient for a related qualifica-tion aspect and not repeated during qualification. Enhanceddocumentation may require more extensive and/or a morerigorous test/inspection regimen, as well as additional signa-tures. Essentially, enhanced documentation must satisfy allthe requirements of qualification documentation.
Note that commissioning never replaces qualification fordirect impact systems. The commissioning process can coveronly elements of qualification, and is not a substitute. Quali-fication should link back to properly documented enhancedelements. Consider the impact of change control (formal orproject) that could affect decisions as to when to use enhancedcommissioning.
Factory Acceptance Tests (FATs) and Site AcceptanceTests (SATs) may include enhanced elements. However, becareful when using FAT especially for enhanced becausechanges may be made at the factory in an uncontrolledsetting that affects other outcomes.
FAT/SAT ConsiderationsFor many projects (especially single equipment), the SATmay constitute the majority of the commissioning activities.When FATs (usually a business decision) are provided, SATscan have a reduced regimen; however, this must be carefullythought out when enhanced elements are included.
Typical FAT/SAT considerations may include the follow-ing, many of which are good candidates for enhanced classi-fication. (Note: prime potential candidates to include en-hanced documentation are noted by (E)).
• functionality - operate equipment/system during testing(E)
• alarms and safeties• PLC/control thorough checkout/challenge (E)• utilities (E)• maintenance needs• calibration (E)• labeling• training and turnover (E)
Commissioning Test PlansCommissioning test plans may be needed to supplementSATs and to commission in an integrated setting, manyelements of which may be good candidates for enhanceddesignation. This is not to be confused with a commissioningplan, which is the umbrella or overall document. First, thefollowing are Inspection (Supporting IQ) questions that mustbe answered as applicable and included in a commissioningtest plan:
• Was specified equipment/systems installed? (E)• Installed correctly?• Proper utilities? (E)• Appropriate human interface?• Safety/environmental/ergonomics?• Documentation (user manuals) and other closeout needs
completed? (E)• Training of user personnel completed? (E)
The commissioning test plan also includes Testing Consider-ations that support OQ, which may answer the followingquestions as applicable:
• Does the equipment or system perform as specified? (E)• Does it deliver URS/FRS or Basis of Design (BOD) require-
ments (or other acceptance criteria)? (E)• Does it operate safely and produce safe results? (E)• Does it properly function in an integrated setting? (E)• Calibration (E)
Self-contained commissioning checklists are useful for smallprojects where commissioning plans and test plans are notwarranted. These are useful for small work where the com-plete commissioning exercise can be accomplished on a suc-cinct document. Again, InVEST wisely – don’t do more thanis needed. These checklists can be enhanced, and may includethe following:
• verify item specified was installed (E)• utility connection (E)• functionality checkout (E)• verify calibration completed (E)• verify closeout documentation completed (E)• verify training or orientation completed (E)• CMMS entry (E)• other internal requirements (E)
Impact AssessmentsBefore drafting the commissioning or final qualification docu-mentation, it is essential to perform an impact assessment.This process is well defined in ISPE materials. An impactassessment is crucial because it enables qualification andcommissioning to focus on what is important. This focus alsoallows commissioning to minimize qualification while sup-porting its success. Qualification is minimized both by breadthof coverage, and benefits from commissioning enhanced docu-mentation. Only cGMP direct impact equipment/systems re-
quire validation, and other aspects (indirect impact and noimpact) can be commissioned in accordance to Good Engineer-ing Practice in lieu of an overstated qualification protocol.
SMART Commissioning and QualificationAcceptance Criteria/Ranges
Also important in synergizing Commissioning and Qualifica-tion and increasing the likelihood of success in both is toassign SMART acceptance criteria. The acrostic SMART is asfollows:
• Sensible: be practical in assigning validated ranges. Is therange really needed to ensure product quality? What doesthe product really require? Can the equipment deliver thisrange consistently? Do the ranges also meet business/payback objectives?
• Maintainable: will the range be maintained over time?
• Accurate: is the range measurable? Are realistic toler-ances considered? Can equipment consistently meet thistarget?
• Range: is a reasonable range assigned? Rarely can pointvalues be maintained. Design values must be well withinvalidated ranges to minimize nuisance alarms and qualityintervention.
• Traceable: has/can the attainment of the range be verifiedand documented? Can it be verified later?
ISPE Baseline® Guides present design, normal operating,and operating ranges. (See Figure 3 for a graphical illustra-
tion.) Design is the value to which the equipment or system isdesigned. Normal operating is the range, wider than design,at which a pre-alert could occur for maintenance notification– this could be the commissioned range. Even wider is theoperating or validated acceptance range. It is crucial to havea less stringent validated (operating) range than the commis-sioned (normal operating) range, both of which should be lessstringent than the design range or value. For example, if thedesired validated (operating) range of a filler may be 300 vialsor bottles per minute, the commissioned (normal operatingrange) might be 320, and the design range 340. If the operat-ing range was set at the design value or range, occasionalfailures would likely occur. (For this example, don’t forget toalso check at the lower speed during commissioning – someequipment may not operate properly at slow speeds.) Buffersshould be provided. Remember, once operating or validatedranges are assigned, there could be a quality interventionrequired when there are excursions – obviously, this shouldbe avoided. Ideally, acceptance criteria should be determinedearly, and be a part of the FRS against which final commis-sioning and qualification documents are drafted.
Specific ExamplesThus far, this article has argued the need for commissioning,the need to InVEST wisely and set SMART acceptancecriteria, and use enhanced commissioning documentation inthe qualification effort. The remainder of the article willcover examples of typical commissioning considerations andapproaches for GMP technology and GMP utility systems.Obviously, any application could differ, requiring more orless of the listed considerations.
Technology systems include computer/control systems,packaging/fill, and process/manufacturing. Typical cGMP
direct impact utilities could include HVAC, purified (or WFI)water, compressed air, and others (site/product specific). Asbefore, prime potential candidates for enhanced elements aremarked with (E). URS/FRS (or acceptance criteria) elementsof commissioning verification are indicated, as well as pos-sible commissioning vehicles, i.e., documents. Given thecomplexity of the various systems or with some combinationsof systems, overall commissioning plans also should be con-sidered where needed.
Computer/Controls• URS/FRS elements or acceptance criteria commissioning
verification- hardware/software verification and testing (E)- security (E)- Part 11 issues (E)- functionality/challenge (E)- alarms (E)- trends (E)- data verification and integrity (E)- human interface/graphics (E)- backup (E)- input/output verification (E)
• include verification of items being controlled - somewhere(E)
• commissioning vehicle: most commissioning activities (in-spections/tests) can be captured in FAT/SAT (E)
Packaging/Fill• URS/FRS elements or acceptance criteria commissioning
verification- verify specified equipment installed (E)- utility connections (E)- instrumentation/calibration (E)- controls interface (E)- proper installation/alignment (E)- materials of fabrication (E)- safeties/ergonomics- additional for sterile (E)- run product
~ line speeds (E)~ labeling (E)~ tolerances (E)~ proper product encapsulation (E)~ finish form acceptance criteria (E)~ cartoning
• commissioning vehicle- most commissioning activities (inspections/tests) may
be captured in FAT/SAT (E)- supplement with commissioning test plans (E)- great opportunity for qualification synergy (E)
Process/Manufacturing• URS/FRS elements or acceptance criteria commissioning
• commissioning vehicle: commissioning plan, commission-ing test plans, and FAT/SAT on individual major equip-ment when needed. If project essentially consists of asingle equipment, FAT/SAT could satisfy most of (if notall) the commissioning test/inspection activities. (E)
HVAC• BOD/URS/FRS elements or acceptance criteria commis-
• BOD/URS/FRS elements or acceptance criteria commis-sioning verification- viable and non-viable particle counts (E)- moisture (dew point) (E)- flow rate/Pressure (E)- oil free? (E)
• commissioning vehicles- pre-commissioning: SATs of major equipment (E)- commissioning test plan (E)
~ challenge installed system to meet acceptance crite-ria, alarms, safeties, automatic operation, etc.
~ trends~ inspection activities
SummaryCommissioning documentation and qualification are symbi-otic when properly applied. Qualification helps define what isimportant for commissioning to emphasize, while commis-sioning minimizes the validation effort and supports itssuccess. Remember to “InVEST” wisely (integrate commis-sioning with qualification, verify, ensure qualification suc-cess, sensible, traceable/document it), and set SMART accep-tance criteria in the beginning (sensible, maintainable, accu-rate, range, traceable). To get more information, see varioustrade organizations (ASHRAE, etc.). Tried and tested GEPapproaches and documents are available, and translate eas-ily into documented GEP commissioning and enhanced com-missioning. Of course, ISPE has many publications avail-able, including the “Commissioning and Qualification”Baseline® Guide. But mostly, learn by doing.
References1. FDA Guidelines on General Process Validation, May 1987.2. ISPE Baseline® Pharmaceutical Engineering Guides for
New and Renovated Facilities - Volume 5: Commissioningand Qualification, Glossary.
About the AuthorTimothy D. Blackburn holds an MBA fromthe Kenan-Flagler Business School (UNC-Chapel Hill) and a Bachelors degree from theWilliam States Lee College of Engineering(UNC-Charlotte) Summa Cum Laude. He isa registered Professional Engineer in VA,PA, and NC with more than 18 years ofexperience, and is a published author and
conference speaker. He is currently the Manager of Engineer-ing for Wyeth Pharmaceuticals in Richmond VA; however, allopinions expressed in the article are the author’s. Further,the author implies no warranty regarding any of the contentsof the article. Blackburn can be contacted by tel: 1/804-257-3609 or by email: [email protected].
Wyeth Pharmaceuticals Inc., PO Box 26609, Richmond,VA 23261.
PAT, HACCP, and Six Sigma -Making Sense of it Allby George R. Johnson
This articleexplores thepotential impactof PAT, HACCP,and Six Sigmaon the LifeSciencesIndustry.
Introduction
The FDA, pharmaceutical and life sci-ences industry have recently focusedon a framework known as Process Ana-lytical Technology (PAT) with the FDA’s
publication of the “Guidance for Industry, PAT– A framework for Innovative PharmaceuticalManufacturing and Quality Assurance.” Thisguidance is part of the FDA’s GMPs for the 21st
Century initiative. PAT focuses specifically onthe growing need to “employ innovation, cut-ting edge scientific and engineering knowl-edge, along with the best principles of qualitymanagement to respond to the challenges ofnew discoveries (e.g. novel drugs andnanotechnology) and ways of doing business(e.g., individualized therapy, genetically tai-lored treatment).” Primarily due to significantconsumer risks inherent in rapid changes andthe associated innovation, the FDA and the lifesciences industry have been hesitant to imple-ment new technologies and approaches untilthey have been accepted as a standard practicein the industry. This “chicken and the egg”cycle imposes restrictions that have impededthe implementation of technology or processimprovements in the past, even when thesenew technologies and improvements offer sig-nificant promise.
In the current business environment, thesynergy of Hazard Analysis and Critical Con-trol Point (HACCP) and Six Sigma with a PATstrategy offers an opportunity to mitigate riskwhile implementing improvements to a pro-cess. This article will explore the potentialimpact of these three methodologies on the lifesciences industry as well as their synergisticability to realize the promise of PAT whileminimizing potential risks to the customer.
What is HACCP?In the late 1950s, the National Aeronauticsand Space Administration (NASA) recognizedthat special foods would need to be developedfor space travel. During this effort, one crite-rion that was identified was the need to ensurethat the food products were absolutely safe tominimize the potentially devastating impacton future astronauts. At that time, food qual-ity and safety were assured primarily throughinspection after the fact. NASA was not com-fortable that this strategy provided the appro-priate level of safety and asked the primaryvendor (Pillsbury Company) to design a sys-tem for assuring food safety using a processcontrol focus. As a result of this initiative,Pillsbury developed the original Hazard Analy-sis and Critical Control Point (HACCP) sys-tem.
In order to implement a HACCP system, thefollowing five preliminary tasks need to becompleted:
1. Identify and assemble the HACCP Team -the team membership should include repre-sentatives from the manufacturing processas well as the technical staff. This step alsorequires that the team (and other appropri-ate people) receive training from an accred-ited HACCP course provider.
2. Describe the product and its distribution -this includes the common name of the prod-uct, processing methods, and distributionrequirements (shelf life, heat, humidity etc).
3. Describe the intended use and consumers ofthe products - this includes the expectedapplication/dosage as well as consideration
for the likely end user of the product. Identification shouldbe made of any potential end user groups that may haveunique risk factors that preclude the safe use of theproduct.
4. Develop a flow diagram that describes the process - theflow diagram should detail each process step under controlof the company (receiving to shipping) as well as who/whatgroup has the primary responsibility for each step.
5. Verify the accuracy of the flow diagram - this includes aphysical “walk through.”
Once the preliminary tasks have been completed, the follow-ing seven basic HACCP system components can be imple-mented:
1. Analyze hazards - potential hazards and potential mea-sures to control those hazards are identified. Note that aclear distinction should be made between those consider-ations that may be quality related as opposed to thoseconcerns that are hazards (life or health threatening).
2. Identify Critical Control Points (CCP) - these are points ina product’s production process at which a potential hazardcan be controlled or eliminated. If multiple control pointshave been implemented that are associated with thecontrol or elimination of a specific hazard, the final assur-ance point in the process flow becomes the Critical ControlPoint for that hazard.
3. Establish preventative measures with critical limits foreach CCP - critical limits can be either extreme limits(temperature no greater than 140°F) or max/min (5 +/-1%).
4. Establish procedures to monitor the preventative mea-sures at each CCP.
5. Establish corrective action to be taken when monitoringshows that a CCP critical limit has not been met.
6. Establish procedures to verify that the system is workingproperly - this often is an oversight or audit program aswell as measurement systems analysis/validation.
7. Establish effective record keeping documenting the HACCPsystem - this would include records of hazards and control
methods, the monitoring of CCP measures, and actiontaken to correct potential problems.
An application example for a CCP could be in the manage-ment of raw materials. Material contamination by microbio-logical, chemical, or physical hazards as well as the associ-ated purity could result in significant risks to the consumer.The primary questions here are:
1. Is there a significant hazard that could be associated withthe raw material?
2. Will this hazard be processed out during the manufactureof the product?
3. Is there a cross-contamination risk to the facility or toother products?
Depending on the answers to the questions above, the man-ner in which the material is certified, received, handled, orstored might be CCPs. The presence of these CCPs as well asthe capabilities required by the appropriate prerequisiteprogram (cGMP, etc.) protect the process from significantrisks associated with vendor management and material sourc-ing concerns, while allowing a significant degree of flexibility.
What is Six Sigma?Six Sigma (a federally registered trademark), is a methodol-ogy that was originally developed by Motorola during theearly 1980s to drive improvement throughout their variousoperations. While its origins had a heavy manufacturingfocus, Six Sigma implementation has since spread to a broadlydiverse number of industries and has become known through-out the world as a powerful framework that can be used tooptimize business processes and link the results to thebottom line. Any processes that require improvement to ahighly efficient state of near-zero defects have proven to besuccessful Six Sigma project opportunities.
Six Sigma achieves this amazing result by the methodicalidentification and control of process factors known as KeyProcess Input Variables or KPIVs. In the terminology of SixSigma, this is often referred to as “Y is a function of x” orY=f(x) where Y is the Key Process Output Variable(s) or KPIVand the x factors are the KPIVs. Y=f(x) focuses on theachievement of a thorough understanding of and subsequentcontrol of the process rather than monitoring for defects afterthe fact.
“While its origins had a heavy manufacturing focus, Six Sigma implementationhas since spread to a broadly diverse number of industries and has become known
throughout the world as a powerful framework that can be used to optimize businessprocesses and link the results to the bottom line.”
The most common framework for Six Sigma project activi-ties is known as Define, Measure, Analyze, Improve, andControl (DMAIC). Each one of these steps has a clear purposeand data driven focus which synergistically unleashes thepower of Six Sigma. These are simply:
Define - select process areas that are critical to the businessand need to be improved.
Measure - identify process KPOVs and target improvementvalues. Conduct a Measurement Systems Analysis to ensurethat you have an effective means to measure the KPOVs.
Analyze - map the value stream of the process and identifyKPIVs.
Improve - determine the KPIV values that optimize theassociated KPOVs.
Control - determine and implement the most effective meansto control the KPIVs.
The power of this framework follows a very logical progres-sion and activity flow for quality improvement activities. TheDefine stage ensures that the use of Six Sigma trainedresources are applied to an area of the business that will yieldthe maximum benefit and ensures that the project is wellscoped.
The Measure stage ensures that we have a metric that willbe a good measure of the improvement activity and that themeasurement system is valid. The purpose of the Analysisstage is to understand what issues drive the improvementopportunity. Without this understanding and the associatedvalidation activities, improvement activities become littlemore than ‘best guess’ implementations which potentiallyhave no improvement impact. The Improve stage uses whatwas learned during the analysis stage to design processimprovements that have a high probability of success. Thepurpose of the last stage (Control) is to lock the improvementsin place. Each stage neatly builds on the foundation laid bythe earlier work.
Six Sigma uses a number of well defined roles and respon-sibilities within its framework. They are:
Master Black Belt - mentor and trainer of Black Belts andoften associated with working on strategic cross functionalimprovement opportunities.
Black Belt - mentor and trainer of Green Belts. The BlackBelt is often associated with major business improvementopportunities.
Green Belt - a part time improvement resource who oftenworks projects to improve areas within their daily scope ofwork. The Green Belt is the primary driver of the shift to a SixSigma culture.
Champion - a senior business leader who supports a SixSigma project effort. Champions review activities and ensureappropriate resource availability as needed to ensure theproject is a success.
Six Sigma easily lends itself to any number of “ad hoc”potential improvement issues within the operation. Applica-tion and focus differ widely, determined predominately by theneeds of an individual business. Any area within the opera-tion that is not optimal or is critical for long term success isa good potential Six Sigma project opportunity.
Pieces of the Puzzle -PAT, HACCP, and Six Sigma
PAT, HACCP, and Six Sigma have a powerful synergy thatholds great promise for those companies that specialize in thelife sciences – pharmaceuticals, medical devices, and nutra-ceuticals. The following discussion will demonstrate how thevarious pieces can be structured to achieve this promise.
PAT offers the vision that cutting edge technology, tools,and approaches as well as a process based focus is the path tohigh operational efficiency and high quality products. Whilelong on vision, PAT as a stand alone lacks the detail necessaryto show how this can be achieved in a dynamic industry wherefrequent change often raises the specter of potential signifi-cant risk to the customer.
HACCP offers a proven risk mitigation strategy and ap-proach that is specifically focused on ensuring that thecustomer is protected from any hazards created by variabilityor changes in the process or incoming materials. It focuses onthe few CCPs necessary to achieve this objective. Its weak-ness as a stand alone is its heavy reliance on research ofknown hazards (biological, chemical, or physical) during thehazard analysis. This presents a significant potential prob-lem in those processes which are “one of a kind” in the riskspresented (formulation and/or unique customer application).
Six Sigma offers a detailed and time proven structure forimprovement which has been demonstrated to be applicableto a broad range of industries and applications. Its weaknessis that while Six Sigma has a very capable tool set andstructure, it needs to be clearly linked to the needs of thebusiness to be effective. Without this focus, program benefitstend to be local in nature and as such miss much of theopportunity that the program offers.
Making Sense of it AllSo how does this all work? First, it’s worth noting that allthree structures (PAT, HACCP, and Six Sigma) start with anassumption that the fundamental processes are consistentwith the application of current Good Manufacturing Prac-tices (cGMPs). This foundation is critical to ensuring that thefundamental safeguards and associated documentation arein place to demonstrate that production practices are as freefrom error as possible and produce a product that is effective,high quality, and absolutely safe for the consumer. Once thisfoundation has been established, the process is assumed to bewell managed and stable. At some point, changes in technol-
ogy or capability will create pressure to change the originalprocess. It is at this point that all three structures (PAT,HACCP, and Six Sigma) can be utilized to minimize theadverse impact of the change to the business while assuringan absolutely safe product for the consumer.
PAT takes a philosophical center stage during this effort.All changes must be done with a full understanding of therisks and potential factor interactions introduced by theproposed changes. Six Sigma helps during this effort throughits focus on real time control of output characteristics (KPOVs)by real time (ideally) control of process and input character-istics (KPIVs). Risk is significantly mitigated by the estab-lishment and control of HACCP Critical Control Points in theprocess. These control points act as stage gates, ensuring thatany process changes that occur prior to the CCP do not impactthe product in a way that would create a hazard if passed onto the consumer.
Utilizing all three structures has the potential of enablingprocess capability updates to be implemented relativelyquickly and without significant risk to the customer.
None of these methodologies have been recommended noradopted by the FDA as of this date. These methodologies canadhere to the PAT guidelines when instituted using theproper metrics, measurements, and critical analysis.
References1. Guidance for Industry: PAT - A Framework for Innovative
Pharmaceutical Manufacturing and Quality Assurance.PAT Web page at www.fda.gov/cder/OPS/PAT.htm., Au-gust 2003.
2. Mathis, Nancy, “PAT - Quality by Design” PharmaceuticalFormulation and Quality, March/April 2004.
3. George, Michael L., “Lean Six Sigma for Service – Hot toUse Lean Speed and Six Sigma Quality to Improve Ser-vices and Transactions,” McGraw-Hill, New York, 2003.
About the AuthorGeorge R. Johnson is the Director of Qual-ity for APV Products (an Invensys Company)in Foxboro, Massachusetts. He is a graduateof the US Naval Academy (Annapolis) with aMS in quality from Anna Maria College anda MA in marine affairs from the University ofRhode Island. He is a Six Sigma CertifiedMaster Black Belt as well as an ASQ Certi-
From Good Manufacturing Practice toGood Manufacturing Performanceby Professor Roger S. Benson, FREng andJim D.J. McCabe
This article wasadapted from apresentation atthe 2003 ISPEUK AffiliateAnnual Seminar,Manchester. Itdemonstrateshow thepharmaceuticalindustry iscapable ofimprovedmanufacturingperformance.
Table A. Typicalbenchmark data.
Measure Pharmaceutical Industry A Winning A World Class FactoryPharmaceutical Factory
Stock turn 3 to 5 14 50OTIF 60% to 80% 97.4% 99.6%RFT 85% to 95% 96.0% 99.4%CpK 1 to 2 3.5 3.2OEE 30.0% 74.0% 92.0%Cycle time (hrs) 720 48 8Safety/100,000 hrs 0.100 0.050 0.001
Introduction
The pharmaceutical industry has a con-tinuous record of growth, innovation,and profitability. Operations are con-trolled through the principles of Good
Manufacturing Practice (GMP) and regulatedby the US Food and Drug Administration (FDA)and national regulatory bodies such as theMedicines and Healthcare Regulatory Author-ity (MHRA) in the UK.
However, the industry is faced with thefollowing pressure to change:1
• stock market demanding continuation ofhistoric growth and profitability
• reduced numbers of new chemical entitiescompared to increasing research and devel-opment costs
• pressures from healthcare providers to re-duce the cost of life-saving medicines
• drive to increase access to life-saving medi-cines in developing countries
• growth of generic competition
One factor arising from these pressures is arequirement to improve manufacturing perfor-mance. The FDA in particular recognizes thesepressures and has publicly stated its willing-
ness to support the challenge of improvingcompetitiveness in the pharmaceutical indus-try.2 This combination of competitive pressureand supportive regulatory environment cre-ates the necessary conditions for change to takeplace. Is the pharmaceutical industry ready tostep up to the plate?
This article will argue that in many ways itis. Good manufacturing performance is sus-tained by good manufacturing practices. Com-panies that meet GMP requirements have anexcellent foundation to develop and adopt inno-vative solutions to maintain product qualityand improve manufacturing performance.
BenchmarkingBenchmarking is a process where your perfor-mance is judged against the best in the world.That is not to say that your factory or even thepharmaceutical industry has to be the best inthe world, but it is important to know what thebest is and to have made a conscious decision tooperate at a different level of performance. Inmany ways, good manufacturing performanceis like the decathlon in the Olympics. You don’thave to win every event to win the decathlon,but you need to win some events and be aboveaverage in the other events.
“Benchmarking is the process of continu-ously measuring and comparing one’sbusiness performance against comparable
processes in leading organizations to obtain informa-tion that will help the organization identify and imple-ment improvements.”3
It is important to note that benchmarking must be a struc-tured process to ensure consistency of definitions and validityof benchmarks. The process must look beyond the perfor-mance measures. It is not sufficient to know that a world-class stock turn is 50 – the process must identify what world-class companies do differently to achieve that level of perfor-mance. An external focus is essential to learn from otherindustries. Why can the semiconductor industry achieveOverall Equipment Effectiveness (OEE) measures in excessof 85% when the pharmaceutical industry typically operatesbelow 50% OEE? It is not enough to benchmark within yourown industry – learn from the best, whatever their industry.Finally, benchmarking is not a competition. Use it to find outwhere to improve and how to improve. How far and how fastyou can improve your own performance is much more impor-tant than your overall position.
This is illustrated by some real benchmarks from compa-nies that are FDA regulated. Consider the following defini-tions:
Stockturn - This is the total turnover of the site at manufac-turing price divided by all the stock on the site on the samebasis. The stock includes finished goods, work in progress,and purchased raw materials.
OTIF - On Time In Full delivery. This is the percentage oforders that are satisfied On Time In Full with zero defects.Note: if there is one defect in an order, the OTIF is zeropercent.
RFT - Right First Time. This is a percentage of the productsat the point of manufacture that are delivered right first timewith no defects. Any recycling, blending, rework of documen-tation, or laboratory testing or other adjustments are ex-cluded from the Right First Time figure.
CPK - Is a statistical process control measure of the variabil-ity of the product. A six-sigma figure corresponds to only fourdefects per million products, while a two-sigma figure corre-
sponds to 308,000 defects per million products. It is measuredon a logarithmic scale.
OEE - Overall Equipment Effectiveness. This measures howeffectively the manufacturing equipment is used. It is aproduct of the product rate multiplied by the quality ratemultiplied by the plant availability. A figure of 100% impliesthat the plant is running flat out every hour of the day makingperfect product. A figure of 10% implies that the plant couldachieve ten times the output that it currently achieves.
Cycle Time Hours - This is the total time from commencingmanufacture to delivering products to the customer which inmany cases is the factory warehouse.
Safety per 100,000 Hours - This is the number of reportableaccidents, greater or equal to three days absence per hundredthousand working hours.
Table A presents the figures for three typical operations. Thisfirst column is for figures for the pharmaceutical industry inthe UK that have been established and developed over sev-eral years from benchmarking discussions with a wide rangeof manufacturers.4
The second column is an award-winning pharmaceuticalmanufacturer that manufactures over the counter drugs,prescription drugs, and injectables. Last year, it was a winnerin the UK Awards for Manufacturing.
The world-class plant is in fact a food plant supplyingsupermarkets and grocery stores. It may be argued that foodis different than pharmaceuticals. However, food manufac-ture also is regulated under the principles of GMP. Consumerprotection and product safety is no less a concern for a foodmanufacturer than it is for a pharmaceutical company. Youwill note from the figures that there is a significant differ-ence.
Consider, for example, the stockturn where the pharma-ceutical industry average is between three and five, butalready there is a pharmaceutical manufacturer achieving14. World-class is 50. If all of the world pharmaceuticalindustry (estimated annual turnover $290 billion) was tomove from its current average to that of the award-winningfactory, the cash released would be in the order of $76 billion
and if it were to move to the world class condition, it wouldrelease a further $15 billion! While these are one-off releasesof cash, they are extremely significant, and would havedramatic effects on both the short-term and long-term profit-ability of the companies.
Stock turn is a powerful measure of manufacturing perfor-mance. High stocks allow operations a buffer to insulate themagainst poor performance. Only excellence in manufacturingwill deliver a high stock turn. It is not something the accoun-tants can adjust.
Comparison of the OEE measures would suggest thatbetween the industry average of 30% of the equipment beingused to its full potential and a world class figure of 92%, theindustry could increase the output of the presentassets by more than a factor of three with minimumcapital investment.
It may be argued that since the capital has been invested,this does not represent a saving, but there is either thepotential to rationalize the asset base or to use alreadyinvested capital to rapidly introduce new products to themarket.
Safety is an excellent measure of manufacturing perfor-mance. Experience from other industries indicates there is adirect correlation between excellent manufacturing and ex-cellence in safety.3
The other measures speak for themselves. Given thenature of the industry, one would expect a high OTIF, theRight First Time to be good, and the CPK to be running in thearea of 4 - 6, not in the area of 2 - 3. Similarly, for manyformulation and packaging operations, one could expect thecycle time to be measured in less than 24 hours rather thanseveral days.
Creating Good Manufacturing Performance -the Virtuous Cycle
The evidence would suggest that in terms of manufacturing,sections of the pharmaceutical industry are in what we wouldcall a destructive cycle - Figure 1.
Many pharmaceutical processes are complex and incom-pletely understood. Processing parameter ranges are oftenestablished empirically based on what has worked in thelaboratory and confirmed during process validation. It isdifficult to predict how the process will respond to variabilityin raw materials or control variables. This lack of predictivecontrol can result in low CpK. Out of Specification (OOS)investigations may be triggered by deviations outside theallowed parameter ranges when such deviations have noimpact on product quality, increasing the cost of compliance.
Without good process understanding, new product intro-ductions can be delayed due to incomplete technology trans-fer between development and manufacturing. If the processis understood, resources can be directed at the critical controlpoints, those parameters that have a demonstrated directimpact on product quality, in order to introduce a more robustand capable manufacturing process.
If the manufacturing process cannot be relied upon toproduce product when it is needed, high levels of safety stocks
are required to meet customer commitments. This results inlow stock turns.
Such facilities are often characterized by high levels offirefighting with manufacturing seen as a problem ratherthan a driver for competitive advantage. Absenteeism (mea-sured by percentage working days lost through absenteeism)or high levels of staff turnover are indicators of low moraleand high levels of workplace stress. A culture of firefightingand pressure to meet production targets is often accompaniedby reductions in EHS performance.
Few companies will find themselves locked fully into thedestructive cycle, but certain of these observations may bepresent to some extent in many pharmaceutical operations.
The priority is to move to the virtuous cycle - Figure 2.Note: this takes time and continuous focus over a number ofyears.
In the virtuous cycle, good manufacturing performancedelivers higher profitability from lower stocks with quickerspeed to market and equipment which is more highly used.
Practices Drive PerformanceGood manufacturing performance is driven by good manufac-turing practices - Figure 3.
Experience from benchmarking both the practices and theperformance of process plants worldwide demonstrates thata direct correlation between the two has always been demon-strated.3,5
Figure 4. Cost and cycle time from dispensing to finishing.Figure 5. Effects of time compression.
The good news in the pharmaceutical industry is that itdoes have good manufacturing practices. These are welldocumented, with well-trained employees in a good environ-ment. However, the requirements of GMP are often used asan argument against change because of a perception that it iscostly and time consuming, particularly the need for revali-dation. A similar argument applies for safety in the chemicalindustry. Again, in some plants, the argument has been usedthat they cannot change the manufacturing performancebecause it would have an impact on safety. Time and againthis has been proved not to be the case, and by consciouslymanaging the change process, within the environments ofgood manufacturing practice and safety, the performance hasbeen improved quite dramatically.
In looking at what the leaders are doing, it is possible toidentify a number of factors that come together that drive theprocess. These are summarized below:
What practices are the leaders improving?
1. Focus on manufacturing/supply chain2. Targeting process to be “Really Right First Time”3. Continuously aiming to increase OEE4. Automation and on-line quality control5. Reducing non value-added activities6. Measurement and display for productivity7. Reducing stock levels and never replacing8. Focus on continuously improving quality
Really Right First TimeReally Right First Time focuses on quality. Quality is ameasure of the percentage of the products that are perfectfirst time. You will hear in some industries the drive for six-sigma. The well-known examples are GE and Motorola thoughincreasingly the approach (sometimes combined with leanmanufacturing in an approach known as lean sigma) isbecoming much more common in the pharmaceutical indus-try. Table B summarizes the features of sigma in terms ofdefects.5 Notice that it is a logarithmic scale and that if a
Figure 6. Elements of Overall Equipment Effectiveness (OEE).
company is operating in the 2-3 sigma range then its defectsare in the order of 200,000 per million. Put that way it sounds,and is, very large. The means of delivering improved qualityare measurement, process understanding and control, re-moval of root causes, and a period of continuous improve-ment.
Reduce Cycle TimeThe cycle time is the time from starting manufacture todelivery (often to the warehouse). The whole process is one ofcompression as illustrated in Figures 4 and 5.6
The very act of measuring the cycle time highlights somevery “quick” wins which can make a significant reduction intime without costing a great deal of money. Often thelaboratory itself is one of the major delays in the whole of thecycle time and is one of the drivers in many industries for themove to Process Analytic Technology (PAT), and on lineanalysis.
In the example below, it is seen that focusing on valueadding activities, i.e., time when the product is actually beingworked on rather than waiting, e.g., for documentation, QCresults, can result in significant time compression and lowermanufacturing costs. Figure 4 describes a process with a cycletime of 35 days and an actual processing time of three days.It is not uncommon for actual processing time, i.e., excludingdelays, QC testing, transport times, documentation, to be lessthan 10% of the total.8 Figure 5 describes the case where cycletime reduction techniques (on-line analysis, flow path analy-sis)9 have been used to increase the time that the product isbeing processed from 10% of the total cycle time to 50%. Thetotal cycle time in this case is now six days.
Overall Equipment Effectiveness (OEE)The third area is the OEE. Again, Figure 6 illustrates wellhow productive time is easily lost in the manufacturingprocess and it is not unusual for only 10% of the availabletime being used for added value activities. Of the 168 hoursavailable in a 24/7 operation, there may be planned down-time, unpredicted loss of production time, production delaysand poor planning, low speed running, scrap and rework allcontributing to low OEE - Figure 6.
The route to improving the OEE is through loss accountingwhere one first measures the losses in time as they occur.Displaying these in order of priority and attacking them oneat a time results in significant benefits. The impacts can bespectacular and it is not uncommon to improve the OEE by10% per year, year-on-year.
Case HistoryAn international biologicals supplier needed to double vialfilling production capacity using existing equipment withoutsignificant capital investment. In addition to poor line utili-zation, product wastage was high as a result of long cycletimes. The root causes of inefficiency were not well under-stood and were the cause of much argument.
An OEE improvement program was established and pro-duction losses were monitored with a loss accounting system
to track all machine losses (availability, speed, quality). Realmachine data was collected and analyzed to provide a non-subjective picture of causes of downtime. Making this dataavailable to the production operators increased their produc-tivity improvement awareness.
A reliability improvement team was established and metregularly to analyze the OEE data and plan and prioritizeimprovement actions. Real data meant that this team wasable to focus efforts on areas that would have the greatestimpact. The improvement team applied continuous improve-ment techniques: Root Cause Analysis, Cause and Effectdiagrams (CEDAC), and Single Minute Exchange of Die(SMED) to these areas to identify and eliminate the rootcauses of the production problems.
Within two years average production speed had more thandoubled and mean time between line breakdowns had in-creased by a factor of ten.
The Way ForwardThe important message about delivering good manufactur-ing performance is that all the tools and techniques are wellknown, available, and successfully operating in many otherindustries and in general the pharmaceutical industry will beable to “pinch with pride” the ideas, tools, and techniques,and successfully implement them in their process. Some ofthe available technologies and tools are summarized below:
• empowered operations staff skilled in continuous im-provement methodologies (including six sigma, Total Pro-ductive Maintenance [TPM], lean manufacturing, Reli-ability Centred Maintenance [RCM])
• loss accounting and loss management
• improved process understanding
• continuous or one-pot processing
• design for manufacture - stronger links between manufac-turing and development to ensure that newly introducedprocesses are robust and capable
• Process Analytic Technology (PAT) - raw material andproduct characterization for processing predictability andin-process control
• activity based costing to identify and focus on value addingactivities
• predictive, model-based process control
• agile automation of each processing step
• information technology - Integrated information collec-tion, management, and interpretation, and its use incontrolling and managing the process
• enterprise Manufacturing Execution System with Elec-tronic Batch Record (EBR) capability
It is beyond the scope of this article to describe these tools indetail. The author welcomes any correspondence from read-ers who desire a more in-depth review of available technolo-gies. It is sufficient to note that the technology is in place;continuous improvement, cycle time reduction, automation,on-line analysis and control are established techniques inother industries. The regulatory environment is supportive ofimproving competitiveness in the pharmaceutical industry.So why, we may ask, does pharmaceutical production under-perform against almost all established manufacturing per-formance benchmarks?
It is a question of necessity and desire. There is no doubtthat if one has to look for examples of good manufacturingperformance, the best place to look is in manufacturers whohave a very demanding customer. The automotive industryhas been improving for 30 years and still manages to improveproductivity by 3% per year, year on year.7 For the pharmaceu-tical industry, those demanding customers are here today, inthe shape of public and private healthcare providers, empow-ered consumers and retailers, and shareholders grown used tohigh returns on their healthcare stocks. Manufacturing willplay its part in meeting those customer demands, deliveringflexibly and cost-effectively in support of business goals.
Benchmarking against other industries in order to learnand improve will be a key element of this transition. It doesnot follow that pharmaceutical manufacturing has to beworld class, but the opportunity to move from the positiontoday toward world class is enormous and the impacts will besignificant. Some pharmaceutical manufacturers are alreadywell underway on this journey – can you afford not to jointhem?
ConclusionsGood Manufacturing Practice is the necessary, but not suffi-cient, condition for good manufacturing performance. Deliv-ering good manufacturing performance is a journey, whichstarts with measurement, and recognition that the opportu-nity exists. It is a process of continuous improvement; thetools and techniques exist already and can be adopted withspeed. The ensuing transition from the destructive to thevirtuous manufacturing cycle will have a dramatic effect onthe success individual pharmaceutical companies.
References1. Big Trouble for Big Pharma, The Economist, Dec 4 2003.
2. Protecting and Advancing America’s Health: Respondingto New Challenges and Opportunities, Food and DrugAdministration, August 2003, www.fda.org.
3. Ahmed, Prof. M. and Prof. R.S. Benson, “Benchmarking inthe Process Industries,” published by The I Chem E inJune 1999, ISBN 0 85295 411.
4. McCabe, J., “Benchmarking for Performance Improve-ment,” presented at ISPE Continuing Education Seminar,Zurich 2002.
5. Competitiveness of the UK Process Industries, April 2001;www.picme.org.
6. Dean, D. Doug and Frances Bruttin, “Productivity and theEconomics of Regulatory Compliance in PharmaceuticalProduction,” PwC Consulting, Pharmaceutical SectorTeam, Basel, Switzerland.
7. Storm Clouds over Detroit, The Economist, Nov. 14, 2002.
8. Raju, G.K., “The Need for Continuous Quality Verifica-tion,” presented at ISPE Continuing Education Seminar,Zurich 2002.
9. Gerecke, G., and T. Knight, “Improving Performance andReducing Cycle Time Using Flow Path Management: ACase Study,” Pharmaceutical Engineering, Nov/Dec 2001,Vol. 21, No.6.
About the AuthorsProfessor Roger Benson, FREng is ABBProgram Manager for Chemicals, Analytics,and Advanced Solutions for the Petrochemi-cals and Chemicals (ATPC). He has been theDTI nominated Judge of the UK Best FactoryAward for the last six years. A chemicalengineer by degree, he is Visiting Professorat Imperial College, London, and the Univer-
sities of Newcastle and Teesside. He wishes to acknowledgethe contribution of all his colleagues in ABB. He can becontacted by tel: +44 (0) 1642 372379 or by email:[email protected]. com.
ABB, Belasis Hall Technology Park, PO Box 99, Billingham,Teeside, TS23 4YS, United Kingdom.
Jim McCabe is the ABB Engineering Ser-vices Pharmaceutical Sector Manager.McCabe has a chemical engineering degreefrom the University of Cambridge and aMasters in Business Administration fromWarwick University. He can be contacted bytel: +44 (0) 1925 741018 or by email:[email protected].
ABB, Daresbury Park, Daresbury, Warrington, CheshireWA4 4BT, United Kingdom.
USA/EC/ISO Regulatory Considerationsfor Designing Aseptic ProcessingFacilitiesby Manuel A. del Valle, PE
This articlepresentscleanroomclassificationsas defined inISO 14644 andFED STD 209Eand theirapplication fordesigningasepticprocessingpharmaceutical/biotechnologyfacilities asdefined in bothUSA cGMPs andEuropeanCommunityGMPs.
There is a large diversity of medicalproducts and devices manufactured inpharmaceutical/biopharmaceutical fa-cilities. Some of these include topicals
(applied on skin or eyes), oral (taken by mouth,swallowed), and parenterals (intravenous orintramuscular drugs). The means by which theproduct is administered greatly affects how itis manufactured and sterilized (if required).When a medicine is swallowed, the humanbody has some natural defense mechanisms tohelp reduce or eliminate living organisms thatmay be harmful. Example of these defensemechanisms are the saliva in the mouth andacid in the stomach. Parenteral drugs by-passsome of these defense mechanisms and there-fore, require sterilization. The purpose of ster-ilization is to reduce the number of live organ-isms in a parenteral product to an acceptable(aseptic) level rather than to a sterile (no liveorganism present at all). Sterility is practicallyimpossible to obtain for parenteral productsbecause they are typically affected by heat orradiation. Parenteral drugs therefore need tobe manufactured under stricter and cleanerconditions. This article discusses where someof these regulations and guidelines are foundin the USA, the European Community (EC),
and the international standards for the designof aseptic processing facilities such asbiopharmaceutical facilities. Biopharmaceu-tical facilities are those in which biologicalmolecules that are destined to become diagnos-tics and therapeutics (among other products)are prepared and modified. Updates and draftsto these guides also will be discussed.
Clean Space ClassificationsClean space classifications are defined in termsof maximum number of particles per unit vol-ume of air. Typical particle sizes used are 0.1micron through 5 microns. A micron is equiva-lent to 1 millionth of a meter or 1/25000 of aninch.
In the USA, definitions for cleanroom classesused can be found in Federal Standard #209.1
The latest revision of this standard was Fed-Std-209E dated 11 September 1992. Table A(based on Fed. Std. 209E Table 1) “AirborneParticulate Cleanliness Classes” lists in bothmetric and English units, the class numberstypically used in Pharmaceutical/Biophar-maceutical Facilities. These are classes 100through 100,000 (class M3.5 through M6.5 inmetric units). USA Pharmaceutical/Biophar-maceutical industry utilizes Class 100 through100,000 clean areas based on 0.5 micron or
larger size particles per cubic foot and for “in-operation”conditions. For example, for a space to meet a classification ofClass 100, it can contain no more than 100 particles per cubicfoot (3,500 particles per m3).
On 29 November 2001, the GSA-FSS issued a notice ofcancellation of Fed. Std. 209E and stated it had been super-seded by the International Organization for Standardization(ISO) standard “Cleanrooms, and Associated Controlled En-vironments,” ISO 14644-1 “Classification of Air Cleanliness,”and ISO 14644-2 “Specifications for Testing and Monitoringto Prove continuous compliance with ISO 14644-1.”2
ISO 14644 consists of eight parts - Table B. Of these, parts1, 2, and 4 are official published standards, the others are intheir development and/or review stages. The procedure thathas been followed in the European Community is that an ISORegulation becomes a Standard six months after being offi-cially issued.
ISO 14644-1 “Classification of Air Borne Cleanliness:”defines clean spaces in terms of class numbers from 1 through9 based on acceptable number of particles per cubic meter.Table C (based on ISO 14644-1, Table 1) tabulates classestypically used in Pharmaceutical/Biopharmaceutical facili-ties. To define a class, three items must be specified: Classnumber (1 through. 9), occupancy state (as-built, at rest, or inoperation), and particle size (0.1 through 5 micron) includingcount (number of particles).
Definitions of occupancy states:
As-Built: installation is complete with all services connectedand functioning, but with no production equipment, materi-als, or personnel present.
At-Rest: installation is complete with production equipmentinstalled and in operational condition, but not in use and withno personnel present. The ventilation (HVAC) system is inoperation to maintain cleanliness and pressurization.
In-Operation: installation is functioning in the specifiedmanner with the specified number of personnel present andthe specified production equipment in operation.
It should be noted that the phrase “operating, but with nooperating personnel present” should normally be taken tomean that ventilation systems are operating and other equip-ment is present in an operational condition, but not in use.3
An example of an ISO cleanroom classification may be:ISO Class 5; operational state; 0.5 micron particles (3,520particles/m3).
USA Regulations/Guidelines Related toPharmaceutical/Biopharmaceutical Aseptic
Processing FacilitiesIn the USA, all medical products and devices fall under theFood, Drug, and Cosmetic Act. Section 501 of the Act statesthat a drug, device, diagnostic, or bulk pharmaceutical chemi-cal is considered to be adulterated if it is not manufactured inaccordance with current Good Manufacturing Practices(cGMPs). Minimum standards of the cGMPs are found inparts 210, 211, 606 to 680, and 820 of Chapter 1, Title 21 ofthe Code of Federal Regulations (CFR). Sections that greatlyaffect HVAC are found in part 211, subpart C-Buildings andFacilities, Sections 211.42 (Design and Construction Fea-tures), and 211.46 (Ventilation, Air Filtration, Air Heatingand Cooling).
Examining these sections emphasizes the point that theseregulations were written in vague terms to allow for engi-neering to develop creative ways of meeting the GMPs.Examples of the vagueness include terms such as: suitable, tofacilitate, to prevent, as appropriate, adequate. One item thatis specifically mentioned is the use of High Efficiency Particu-late Air (HEPA) filters. Section 211.42, (10) - Aseptic Process-ing, states under sub-section (iii) “An air supply filteredthrough high-efficiency particulate air filters under positivepressure, regardless of whether flow is laminar or nonlaminar.”The author has been asked if HEPAs are required forcleanrooms classified as Class 100,000 in the USA.
Terminal Sterilization vs. Aseptic ProcessingTo ensure product sterility, the US FDA requires that steriledrug products be terminally sterilized. The FDA “Guidelineon Sterile Drug Products Produced by Aseptic Processing”4
defines terminal sterilization and aseptic processing. Termi-nal sterilization involves filling and closing product contain-ers under conditions of a high quality environment; the
ISO 14644 Title StatusPart No.
14644-1 Classification of Air Cleanliness Published
14644-2 Specification for Testing & Monitoring to Prove PublishedContinued Compliance with ISO 14644-1
14644-3 Metrology & Test Methods
14644-4 Design, Construction and Start-Up Published
14644-5 Cleanroom Operations
14644-6 Terms, Definitions & Units
14644-7 Separative Enclosures (Clean Air Hoods, GloveBoxes, Isolators and Mini-Environments)
14644-8 Molecular Contamination
Table B. List of ISO 14644 parts.
ISO Classification Maximum number of particles/CU. Mtr.Number (N) Equal to or larger for sizes shown below
0.5 5 Micron
ISO Class 5 3,520 29
ISO Class 6 35,200 293
ISO Class 7 352,000 2,930
ISO Class 8 3,520,000 29,300
ISO Class 9 35,200,000 293,000
Table C. Excerpts from ISO 14644-1, Table I Airborne ParticulateCleanliness Classes for Cleanroom and Clean Zones.
product, container, and closure are usually of high microbialquality, but not sterile. The product in its final container isthen subjected to a sterilization process (usually by means ofheat or radiation). Unfortunately, most biopharmaceuticalproducts cannot be terminally sterilized without detriment tothe product. The FDA allows an alternative means of steril-ization: aseptic processing.
In aseptic processing, the product, container, and closureare subjected to the sterilization process separately and thenbrought together in the fill line. Some examples of steriliza-tion methods involve using dry heat for glass containers,moist steam for rubber closures, and 0.2 micron filtration forliquid dosage forms.
Guideline on Sterile Drug Products Producedby Aseptic Processing
Complying with the clean space classifications defined in ISO14644-1 helps minimize the number of foreign particles thatend up in medicines during preparation and filling opera-tions. Unfortunately, these classifications do not address liveorganisms that can reproduce themselves forming a ColonyForming Unit (CFU), which may be harmful to a person ordeadly in an extreme situation.
A USA FDA guideline that defines cleanroom require-ments and CFU limits in the pharmaceutical industry for asterile drug product is the FDA “Guideline on Sterile DrugProducts Produced by Aseptic Processing” mentioned above.Section III - “Buildings and Facilities” of that guide refers tosections 211.42 and 211.46 of the GMPs (see above) anddefines two product exposure areas that are particularlyimportant to drug product quality: critical areas and con-trolled areas.
Critical Areas are those in which the sterilized dosageforms, containers, and closures are exposed to the environ-ment. Requirements for these areas include:
• Class 100 (maximum of 100 particles size 0.5 micron andlarger per cubic foot), when measured not more than onefoot away from the worksite, and upstream of the airflowduring filling/closing operations.
• Air supplied at the point of use as HEPA filtered laminarflow air (currently referred to as “unidirectional” flow).
• An airflow rate of 90 feet per minute (FPM) ± 20%. Thisvelocity is typically measured 6 to 12 inches below ceilingterminal HEPAs. Since the purpose of this velocity is tosweep away particulate matter from the filling/closingarea, this author recommends measuring air velocity atthe same location where particle counts are to be mea-sured, that is, no more than a foot away from the work site,and upstream of the airflow during filling/closing opera-tions.
• Maximum of one Colony-Forming Unit (CFU) per 10 cubicfeet of air.
• The fill room static air pressure with all doors closedshould be as least 0.05 inches water gage (12.5 pascals)positive to adjacent less cleanrooms.
Controlled Areas are defined as those in which unsterilizedproducts, in-process materials, and containers/closures areprepared. Requirements for these areas include:
• Class 100,000 (maximum of 100,000 per particles 0.5micron and larger per cubic foot) in the vicinity of exposedparticles during periods of activity.
• Minimum airflow rate of 20 air changes per hour (ac/hr).
• Maximum of 25 CFU per 10 cubic feet.
• Room static air pressure with all doors closed should be atleast 0.05 inches WG positive to adjacent less cleanrooms.When doors are open, outward airflow should be sufficientto minimize ingress of contamination.
Other room classifications such as Class 10,000 are typicallyused at some controlled areas, especially to meet EuropeanCommunity and International Standards. One question thatis frequently asked is what airflow rate should be used forClass 10,000 areas. An airflow rate this author has used formany years with satisfactory results is 60 ac/hr. Some design-ers have obtained satisfactory results (proven by validationon specific cases) with lower airflow rates. The airflow rateshould be sufficient to sweep particulate matter away fromexposed product. Therefore, one would use a larger flow ratefor rooms where the product is in powder form and exposed tothe ambient for a long period of time than for a liquid productwhich is enclosed in vessels and in piping.
Another topic that is often raised as far as air distributionin clean areas is the location of the HEPA filters and thereturn/exhaust grilles/registers. A register is a grille with abalancing damper attached. For cleanrooms Class 10,000and cleaner, the FDA expects ceiling terminal HEPAs andlow wall returns in order to reduce drafts and to removeparticles at floor level where they are mostly collected orgenerated. For Class 100,000 areas the FDA still prefersceiling terminal HEPAs and low wall returns, but acceptHEPAs in the Air Handling Units (AHUs) as well as ceilingreturns.
This author prefers to use ceiling terminal HEPAs and lowwall returns in most Class 100,000 rooms for the followingreasons:
• It is a cleaner operation since the HEPA is at the point ofuse (the ceiling of the room). When HEPAs are located inthe AHU or in the supply duct main, any dust in theductwork (oxidation dust from galvanized ductwork, ordust entering duct when duct access doors are opened)ends up in the room.
Table E. Excerpts from EC GMP Guide, Annex I - Table on Limitsfor Microbial Contamination.
At rest In operation
Grade Maximum permitted number of particles m/3 equal to or above
0.5µm 5µm 0.5µm 5µm
A 3,500 0 3,500 0
B 3,500 0 350,000 2,000
C 350,000 2,000 3,500,000 20,000
D 3,500,000 20,000 Not defined Not defined
Table D. Excerpts from EC GMP Guide, Annex 1 - Table onAirborne Particulate Classification Grades.
• It is easier to upgrade a Class 100,000 room to a cleanerClass (Class 10,000) since the supply air duct is alreadysized for the gauge (thickness) required by the HEPAshigher static pressure loss, and low wall returns arealready in place. Only additional terminal HEPAs and lowwall returns are needed. The existing air distribution canbe re-used. A Class 10,000 room requires about threetimes the airflow rate of a Class 100,000 room (about 10.5cfm/sq. ft. vs 3.5 cfm/sq.ft. for a space with a 9 foot ceiling).
• HEPAs remain cleaner (after in place testing and certifi-cation) with terminal HEPAs than HEPAs in AHUs sinceeach terminal HEPA may be challenged (with DOP sampleor other aerosol) individually instead of as a group inAHUs. This means that the terminal HEPAs do not haveto filter as much DOP since they are exposed to it for ashorter period of time.
An application where ceiling return/exhaust may be recom-mended is in some areas of washrooms in order to removewater vapor where it is generated. Another application whereHEPAs in AHUs and ceiling returns may be practical forClass 100,000 spaces may be perimeter corridors which oftensurround clean areas, when these corridors require Class100,000 classification.
European Community Regulations andGuidelines Related to Pharmaceutical/Biopharmaceutical Aseptic Processing
FacilitiesIn the European Community, cleanroom classification also isbased on ISO 14644-1 and ISO 14644-2.
The EC GMP Guideline that defines cleanroom classes
and CFU limits for aseptic processing is the “Guide to GoodManufacturing Practice for Medicinal Products,” Annex 1 –Manufacture of Sterile Medicinal Products. This guide isfound in the Medicines Control Agency (British Equivalent toUSA FDA) “Rules and Guidance for Pharmaceutical Manu-facturers and Distributors” 2002. This publication also isknown as “the Orange Guide” because of the color its cover.5
This guideline defines four cleanroom classifications (GradeA, B, C, D) for two occupancy states: “at rest” and “inoperation.” It tabulates particle counts for its classificationsin both occupancy states for two particle sizes: 0.5 micron and5 micron - Table D.
Limits of microbiological counts are shown in a secondtable (Table E) for cleanroom Grades A through D, in opera-tion. Four methods of collecting microbiological counts areshown on that table.
The guideline states that components after washing shouldbe handled in at least a Grade D environment. It also statesthat preparation of solutions, which are to be sterile filteredduring the process, should be done in a Grade C environment.Finally, it mentions that “Transfer of partially closed con-tainers, as used in freeze drying, should, prior to the comple-tion of stoppering, be done either in a Grade A environmentwith Grade B background or in sealed transfer trays in aGrade B environment.”
As far as air filtration and airflow rates, the guideline saysthat Grade A zones shall be provided with laminar airflow atan air speed of 0.45 m/s (90fpm) ± 20% at the workingposition. For Grades B, C, and D, all it requires is that HEPAfilters be used, and that the air change rate be related to thesize of the room and the equipment and personnel present inthe room.
In terms of air pressurization between rooms, the guiderequires an air pressure differential of 10-15 pascals (0.04 to0.05 IN wg) between adjacent rooms of different grades.
In regard to changing rooms, the guide states they shouldbe designed as airlocks and that “the final stage of thechanging room should in the at rest state be the same gradeas the area into which it leads.”
Comparison Table - ISO/USA/ECTable F compares USA/EC/ISO guidelines. It uses, as acommon base for comparison, a particle size of 0.5 micron.Note that the left hand section of the table called “AirborneParticulate Cleanliness Classes” only defines cleanroomclasses in terms of allowable number of particles per cubicvolume of air.
The right hand section of the table called “Sterile DrugsEnvironmental Requirements” shows recommendations byUSA and EC GMPs for applying the cleanliness classes andlimiting the number of live organisms.
It should be remembered that USA regulations and guide-lines are concerned with particle counts in size 0.5 micronduring “in operation” conditions while to satisfy EuropeanCommunity regulations both “in-operation” and “at-rest”conditions must be met for two particle size counts, typically0.5 and 5 micron.
In reference to Barrier Isolation, the USA cGMPs, expectthe isolator to be located in a Class 100,000 room environ-ment while the European community expects it to be in aGrade D room environment.
Update to USA Aseptic Processing GuideThe US FDA is scheduled to publish a draft of an update to its1987 Aseptic Processing Guide. The draft title is “SterileDrug Products Produced by Aseptic Processing Draft 2002.”The following are some excerpts from that draft which reviseareas previously covered or add new ones:
1. Addition: “...sterile drugs should be manufactured byaseptic processing only when terminal sterilization is notfeasible.”
2. Addition: an Air Classification Table (Table 1) coveringClasses 100 through 100,000 areas based on size 0.5micron particles per cubic foot for both: particles ingeneral and microbiological counts. The table is based on“in-operation” conditions.
3. Revised Controlled (Class 100,000) rooms nomenclatureand calls it now “Supporting Clean Areas.” It states thatthese supporting clean areas should be Class 10,000 inthe area immediately adjacent to the aseptic processingline. It also states that manufacturers can classify thisarea as Class 1,000 or maintain the entire aseptic fillingroom at Class 100. It mentions that for a Class 10,000
room at least 20 air changes per hour is typically accept-able, but adds no recommendations for other less cleanareas such as Class 10,000.
4. Addition: Poly-Alpha-Oleofin (PAO) as an alternativeaerol for DOP for integrity testing of HEPA filters andstates that filter scanning should be conducted at aposition about one to two inches from the face of the filter.
5. Addition: airflow velocities are measured six inches fromthe filter face or at a defined distance proximal to the worksurface for each HEPA filter.
6. Addition: “lyophilization processes include transfer ofaseptically fixed products in partially-sealedcontainers…assure that the area between the filling lineand the lyophilizer, and the transport and loading proce-dures, provide Class 100 protection.”
7. Addition: airlocks should be installed between the asepticprocessing area entrance and the adjoining uncontrolledarea and those other interfaces such as personnel entriesalso are appropriate locations for airlocks.
8. Addition: drains are not considered appropriate for roomsin classified area of the aseptic processing facility.
9. Addition: microbiological environmental monitoringshould include both alert and action limits.
Environmental Requirements for Sterile Medicinal Products
International Standard ISO USA Federal Standard 209E USA FDA Guideline on Sterile Drug European Commission 1997 GMP Guide,14644-1 Classification of Cleanroom and Workstation Products Produced by Aseptic Processing, Annex 1 Manufacture of Sterile Medicinal Products
Air Cleanliness (1999); ISO Requirements, Controlled June 198714644-2 Specs. For Testing Environment, 1992 (Note 2)
& Monitoring (2000)
State (Note 1) In Operation In Operation At Rest In Operation
Descriptive Max. Number Descriptive Max. Number Descriptive Max. Number Max. Number Descriptive Max. Number Max. Number Max. Numberparticles 0.5 particles 0.5 particles 0.5 of viable particles 0.5 particles 0.5 of viablemicron and micron and micron and microorganisms micron and micron and microorganisms
larger per m3 larger per ft3 larger per ft3 (CFU) per ft3 larger per m3 larger per m3 (CFU) per m3
(per m3) (per m3) (per ft3) (per ft3) (per ft3)
ISO Class 5 3,520 Class 100 Critical Areas 100 0.1 Grade A 3,500 3,500 Less than one100 Class 100 (3,546) (3.5) (99) (99) (0.03)
ISO Class 6 35,200 Class 1,0001,000
ISO Class 7 352,000 Class 10,000 Grade B 3,500 350,000 10 10,000 (99) (9,912) (028)
ISO Class 8 3,520,000 Class 100,000 Controlled 100,000 2.5 Grade C 350,000 3,500,000 100100,000 Areas
Class 100,000 (3,546,100) (88.7) (9,912) (99,129) 2.8
ISO Class 9 35,200,000 Grade D 3,500,000 Not Defined 200(99,129) 5.7
Notes:1. Define state: as built, at rest, or in operation.2. On 29 Nov. 2001 Fed. Std. 209E was cancelled and superseded by ISO 14644-1 and ISO 14644-2.
Table F. Environmental Requirements for Sterile Medicinal Products.
1 Bulk Pharmaceutical Chemicals Published Jun 1996
2 Oral Solid Dosage Forms Published Feb 1998
3 Sterile Manufacturing Facilities Published Jan 1999
4 Water and Steam Systems Published Jan 2001
5 Commissioning and Qualification Published Mar 2001
6 Biopharmaceutical Manufacturing Facilities Published Jun 2004
Packaging, Labeling and Warehousing In draft stage Operations
Maintenance In draft stage
Laboratories In draft stage
Oral Liquids and Aerosols Proposed for future
Table G. List of ISPE Baseline® Pharmaceutical Engineering Guidesfor New and Renovated Facilities.
10. Addition: Appendix 1: “Aseptic Processing Isolators” andstates that they should not be located in an unclassifiedroom and recommends location inside Class 100,000 or10,000 rooms. It states that the interior of the isolatorshould, at minimum, meet Class 100 standards and thatits interior air pressure be between 0.07" to 0.2" watergage above its surrounding environment.
11. Addition: Appendix 2: “Blow-Fill-Seal-Technology” andstates that they should be located in a Class 10,000environment, but that using isolation technology canjustify an alternate classification. Also mentions that airin the critical zone should meet Class 100 particulate andmicrobiological standards.
12. Addition: Appendix 3: “processing prior to filling/sealingoperations” and states that procedures that expose theproduct to the environment, such as aseptic connectionsshould be performed under unidirectional airflow in aClass 100 environment, in a Class 10,000, or better room.It furthers states that microbiological and particulatemonitoring should be performed during operations.
Updated to EC Aseptic Processing GuideThe previous EC Guide to Good Manufacturing Practicedated 1997 was found in Chapter 4 - EU Guidance onManufacture of “The Orange Guide,” the British MedicineControl Agency publication titled “Rules and Guidance forPharmaceutical Manufacturers and Distributors.” Its Guideto Aseptic Processing was found at the end of Chapter 4,under Annex 1 - Manufacture of Sterile Medicinal Products.
This most recent edition of “The Orange Guide” is dated2002. Almost nothing changed in Annex 1 between these twoissues. The Guide still defines clean areas in terms for GradesA through. D and the classification tables still list both at-restand in-operation conditions for two particle sizes: 0.5 and 5micron.
The following are excerpts from the 2002 issue of Annex 1that shows additions or omissions.
1. Omission: under the “General” section, item #5 the 1997issue included microbiological monitoring. The 2002 issuedeleted this requirement. In this author’s opinion, micro-bial monitoring is a “must” in validating an aseptic pro-cess.
2. Addition: the 2002 issue adds under the “Processing”section, Item #42 that “Process simulation tests should beperformed as initial validation with three consecutivesatisfactory simulation tests per shift...” It further addsthat “normally process simulation tests should be re-peated twice a year per shift and process.”
3. Omission: in both the 1997 and 2002 issues, Annex 1defines the “at-rest” condition occupancy state as “thecondition where the installation is complete andoperating…but with no operating personnel present.”This statement gives the misconception that the “pro-cess” equipment is in operation. The last paragraphs ofAnnex 1, 1997 issue included “Notes and References”that clarified that the phrase “operating, but with nooperating personnel present” should normally be takento mean that ventilation systems are operating and otherequipment is present and in an operational condition,but not in use.” In other words, the air conditioningsystem is working to maintain cleanliness and pressur-ization, but the rest of the equipment in the room is not.This very important clarification was omitted from the2002 issue. This author sent an e-mail to the editor of“The Orange Guide” who answered, “…it was felt that itwas not appropriate to include advice from the UK Medi-cines Inspectorate in an official EU Guide Annex. How-ever, we note your comments and we shall consider re-installing a modified version in the next edition.” Let’shope they do because next to personnel, the processequipment is the major contributor to particles in acleanroom and the “at-rest” particle count limits may notbe met if the process equipment is allowed to be inoperation during the “at-rest” condition.
ISPE Baseline® PharmaceuticalEngineering Guides6
As mentioned previously, GMPs are written in a vaguelanguage. This leads some designers of pharmaceutical/biopharmaceutical facilities to overdesign and others tounderdesign. An international association that is helping inthis respect is the International Society for PharmaceuticalEngineering (ISPE) with its “Baseline® PharmaceuticalEngineering Guides.” This is a series of 10 guides that arebeing developed in the USA in cooperation with FDA toprovide minimum design criteria for designing systems thatwould meet FDA requirements. Table G lists the proposedguides and indicates which ones have already been pub-lished.
ConclusionThis article pointed out some of the regulations, guidelines,and standards that must be followed to design an asepticprocessing facility, and also tried to point out the interrela-tionship among them. An important item to remember is thatthe old Fed. Std. 209E and the current ISO 14644-1 thatreplaces it, only defined cleanrooms in terms of particles perunit volume. Where to use these cleanrooms and the maxi-mum allowed CFUs for each class are only found in the ECand USA guidelines for aseptic processing facilities.
One last comment this author wants to make is that it isabout time that both the 2002 EC Guide and the 2002 USAAseptic Processing draft come together and use ISO 14644-1(that both agencies have approved) as the common base toclassify cleanrooms. To avoid confusion, the “Grades” used bythe EC and the “Classes” used by the USA should be deletedand all classification based on ISO-14644-1.
References1. Federal Standard 209E - Airborne Particulate Cleanli-
ness Classes in Cleanrooms and Clean Zones, 11 Septem-ber 1992. Revised 1992 by the Institute of EnvironmentalSciences (and Technology), 940 E. Northwest Highway,Prospect, IL. 60056, Tel: 1-708/255-1561, Fax: 1-708/255-1699.
2. ISO 14644 - Clean Room and Associated Controlled Envi-ronments. International Organization for Standardiza-tion, Case Postale 56, CH-1211 Geneve 20, Switzerland,Internet [email protected]
3. 1997 EC Guide to Good Manufacturing Practice, Annex 1- Manufacture of Sterile Medicinal Products. This Euro-pean Community GMP guide is found in the publicationknown as “The Orange Guide,” published by the British“Medicines Control Agency” or MCA titled, “Rules andGuidance for Pharmaceutical Manufacturers and Dis-tributors 2002,” published by TSO (The Stationary Office),www.tso.co.uk/bookshop, Tel: 44-8706005522.
4. Guideline on Sterile Drug Products Produced by AsepticProcessing - June, 1987 by the Center for Drugs andBiologics, Food and Drug Administration (FDA), Rockville,MD.
5. Commission of the European Communities. The rulesgoverning medicinal products in the EC. Vol. IV. GoodManufacturing Practice for Medicinal Products. Luxem-bourg: Office for Official Publications of the EC, 1992.ISBN92-826-3180-X.
6. ISPE Baseline® Pharmaceutical Engineering Guides, ISPE,3109 W. Dr. Martin Luther King Jr. Blvd., Suite 250,Tampa, FL 33607, Tel: 1-813/960-2105, Fax: 1-813/264-2816, Web site: www.ispe.org.
About the AuthorManuel A. del Valle is Director, HVACDesign at the Greenville, South Carolina,USA office of Fluor, a worldwide design,build, construct, and maintenance company.The bulk of his work has been in HVACdesign for pharmaceutical and biopharma-ceutical facilities. In 1965, he obtained hisBSME from the University of Puerto Rico. He
is a registered professional engineer in Puerto Rico and sixstates in the USA. In 1971, he began designing HVACsystems for pharmaceutical facilities while managing theHVAC design department of Daniel Construction Co. Int’l. inPuerto Rico. In 1987, he began designing HVAC systems forbiopharmaceutical facilities while working for Fluor in theirGreenville, South Carolina offices. His design experienceincludes preparation of conceptual, preliminary, and con-struction documents of HVAC systems, as well as field super-vision and troubleshooting. He has published a number ofarticles and lectured at various universities and seminars ofnational and international associations on HVAC design forPharmaceutical/Biopharmaceutical facilities. He can be con-tacted by tel: 1/864-281-6607 or by email: [email protected].
Fluor, 100 Fluor Daniel Dr., DC C104D, Greenville, SC29607.
Validation Outsourcing Best Practices:What Does this Mean for the Client-Contractor Relationship?by Katie Henchir and Cynthia Ingols
This articledescribes theimportance ofValidationMaster Planningas it relates tothe success ofboth projectmanagementand contractorcoordination.
Introduction
What is “Validation” to you and yourorganization? Does hearing theterm “Validation” make you cringe?If you are a validation engineer, do
you find that people seemingly avoid you? Whyhas validation become the “necessary evil” ofthe pharmaceutical industry? Let’s look to theFDA.
In 1976, the FDA revised the Good Manu-facturing Practices (GMPs), placing signifi-cant emphasis on quality controls to be imple-mented in manufacturing, packaging, stor-ing, and distributing of products, and then inthe validation of this quality. After three yearsof public hearings, the FDA declared the GMPsa substantive regulation and received the rightto prosecute an organization for failing tocomply. To further reinforce their commit-ment to quality and the shift toward valida-tion, the FDA published in 1987, “GeneralPrinciples of Process Validation.” Now theterm validation is widely referred to as estab-
lishing documented evidence that a system orequipment consistently produces a result meet-ing a predetermined specification. How hasvalidation, an important and yet apparentlystraightforward process, come to be seen as a“necessary evil?”
Concurrent with the FDA’s drive towardvalidation standards was private industry’smove to outsourcing. In the 1980s, manage-ment gurus urged executives to focus on theircore competencies and to outsource non-essen-tial functions and services. Validation processesbecame a prime target for outsourcing since theneed for validation work is often cyclical. Inaddition, as more and more companiesoutsourced, firms and individuals who special-ized in outsourcing, such as validation engi-neer specialists and/or contracting firms, grew.Today, outsourcing plays a significant role invalidation efforts.
Outsourcing validation processes paradoxi-cally both simplifies and complicatesmanagement’s tasks. On the one hand,
outsourcing makes valida-tion easier for managementsince outside specialistscome in for a specified timeand for a specific task, do it,and leave. On the otherhand, these outsiders needto learn quickly anorganization’s “ropes,” who’swho, and the specific needsfor a project. People - bothcompany insiders and out-side specialists - are the keyfactors in developing ad-equate or high quality vali-dation work. Management
of relationships, then, is the critical people component inmaking projects run smoothly.
To ascertain the best practices in managing the peopleaspect of outsourcing the validation process, the authorsinterviewed validation specialists. The findings from theinterviews evolved into the following best practices in thedevelopment of a Validation Master Plan (VMP) and inimplementation of it, using outsourced validation special-ists.
Preparing Insiders: Validation MasterPlanning and its Purposes
The purpose of a Validation Master Plan (VMP) is to createa central document to guide a validation effort. Although aVMP can be a powerful management tool, the document isrecommended, but not required by the FDA in its US GoodManufacturing Practices. The European Union’s (EU) Guideto Good Manufacturing Practices notes that “the VMP shouldbe a summary document, which is brief, concise, and clear(note in references).” (See sidebars for recommended catego-ries and data in EU’s Guide and for standard sections inVMPs developed by US companies).
But the human side of developing a VMP is just asimportant as the categories and data placed into such adocument. What are the people elements that are critical tohigh-quality validation work?
Requires LeadershipSelecting a strong interdisciplinary group, pulling them to-gether, and writing a high quality VMP requires leadership.Since the potential for group discord is ever-present, a leaderwho connects members and engages them in strategic think-ing is needed. This person should be appointed by a top-levelexecutive with the mandate to make a “team hum” and a“VMP live.”
Needs Committed Multi-Disciplinary TeamAn equally important task in VMP development is identify-ing who should be involved. Savvy representatives from eachdepartment that will be touched by the validation process
should sit around the table. Generally, representatives frommanufacturing, quality control, regulatory affairs, qualityassurance, and engineering should be part of the conversa-tion. Leaving out any department is likely to weaken thethinking behind a VMP and acceptance of the document.
Once the right people are sitting around the table, thereare three important tasks associated with the development ofthe VMP and its subsequent implementation.
Promotes Strategic ThinkingA VMP can push managers to consider a company’s approachfor “winning” at validation. If the right people are part of theconversation and if conversations are robust, then the VMPcan be a document that promotes strategic thinking. There is,of course, tension between an efficient process where onepulls from files of a previously-used VMP and inserts sectionsinto the next VMP and the more time- consuming approach ofre-thinking and re-writing a VMP.
In developing documents such as VMPs, there is thehuman inclination to make the process routine and bureau-cratic, draining it of strategic intent. The challenge is to makethe process engaging and vigorous enough to draw out people’sbest ideas. Asking people about the implementation of thelast VMP “What went well?” and “What should we improve?”are important questions to answer.
Paints a Clear PictureThere is inherent tension between giving the multi-disciplin-ary group enough time to talk and writing a draft of the VMP.After an initial conversation, circulate a draft of the VMP tothe group and request feedback. Getting an agreed-uponVMP delivers a single message to the multi-disciplinarygroup and company. The VMP, with its concise milestonesand project goals, signals how the validation will be imple-mented. For example, a detailed VMP can include testingstrategy for equipment providing clear guidance during theprotocol generation phase.
Ensures ComplianceBecause many functional areas review the document, thisdevelopmental approach of the VMP will push the group toevaluate how each component complies or does not complywith regulations.
With the underlying force of achieving compliance, thereare five key elements in developing a useful VMP - Figure 1.
PhilosophyMany times we resort to “that’s our philosophy” when theemployed validation practices are questioned. But really,what is a validation philosophy? On the project level, thephilosophy governs details of a validation program; a philoso-phy dictates everyday practices by delivering a sound meth-odology for achieving compliance. When addressing the phi-losophy question, the intent is to develop a rationale for whythings are done the way they are. For a risk-taking company,the philosophy may combine the risk-based approach withthe goal of minimizing redundant testing – this philosophy
“All Validation activities should be planned. The key elements of a validationprogram should be clearly defined and documented in a Validation MasterPlan (VMP). The VMP should be a summary document, which is brief,concise, and clear. The VMP should contain data on at least the following:
(a) validation policy(b) organizational structure of validation activities(c) summary of facilities, systems, equipment, and processes to be
validated(d) documentation format; the format to be used for protocols and reports(e) planning and scheduling(f) change control(g) references to existing document.
In case of large projects, it may be necessary to create separate ValidationMaster Plans.”
Final Version of Annex 15 to the EU Guide to Good ManufacturingPractice, Validation Master Plans.
may then result in the use of commissioning documents tosupport qualification efforts based on the level of risk.
Internal PracticesWhen a validation project is initiated, there can be severalexternal players, such as equipment vendors, a design firm,third-party representative, and contractors. Internal prac-tices become very important to the success of the project andgenerating the VMP is an opportune time to evaluate howand who should be involved. Evaluating internal practicesinclude identifying key players, strategies for managingcontractors, assigning clear roles and responsibilities, anddeveloping (or reviewing) supporting SOPs or policies thatare consistent with the philosophy.
Test Strategies and Test RationalesThe VMP typically includes a list of equipment followed by abrief system description; the document can function as a toolwhen ramping up a validation team. Clearly identifyingequipment-specific test strategies can be very helpful on theproject by reducing the time spent generating protocols. It isalso important to capture your testing rationale in a docu-ment – why not the VMP. The VMP is a central place to houseinformation and can be readily available during an inspec-tion or even a team meeting. The rationale for a testingstrategy must support the philosophy.
Current industry tools to assess validation needs andfacilitate development of testing strategies and testing ra-tionales are the Impact Assessment and the GAP Analysis.
Internal Project ControlsWhen the VMP is prepared prior to the start of a project, theproject team can assess the appropriate internal projectcontrols. The project team can determine the frequency ofmeetings, time constraints, milestones, timelines, budgets,and a variety of performance measures as key indicators toensure the project is on track.
Decision-Making ToolOnce the document has been prepared and approved, it thenserves as a decision-making tool. All the pertinent projectinformation has been captured in the central and accessibledocument, presenting the same message to the entire team.The team players can use this document to make compliancedecisions or to include additional testing as deemed neces-sary. For the project team, it is a reminder of time-lines, theimportance of the effort, and lastly, a tool that governs theactions as the project unfolds.
Evaluating the Impact of aValidation Master Plan
The focus of our discussion has been on the development of aVMP and how the VMP can be used to support validationefforts. The document as we have highlighted plays a signifi-cant role in planning and preparing for the project. Tocontinue our discussion, we have evaluated current practicesfor managing a large-scale validation project with the focus
on the client-contractor best practices and concepts thatshould be addressed in the VMP.
To aid in developing strategies for managing validationcontractors and how to successfully develop and implement aVMP, common threads were identified from the industryvalidation engineers’ interviews.
The findings from the interviews have been placed into thefollowing best practice categories for managing a large-scalevalidation effort and implementing a successful VMP:
Preparing for ContractorsThe question is: how does the operating company need toprepare for contractors? This stage involves four levels ofactivities: 1. diagnosing your need for a contractor; 2. gather-ing documentation to facilitate and minimize the time spentsearching for information and getting the contractor up tospeed; 3. appointing an inside manager as point person forthe contractor; and communicating to key organizationalplayers about the contractors, their work, and 4. theorganization’s expectations.
Laying a Foundation for Quality WorkThe essential task in this stage is to build the relationshipwith the contractor and to make her a member of the team.Once a contractor is on site, the point-person needs to train,share information, and introduce the contractor to key play-ers and establish clear lines of communications. With thisfoundation, the contractor minimizes the time searching forinformation and maximizes the time spent meeting yourgoals.
Implementing for EfficiencyContractors can deliver a scientifically sound qualificationpackage given the appropriate support and tools. However,the vision and tools must come from the operating company.The most powerful tools producing a vision are clear, continu-ous communication and guidance, and proactive project man-agement. The operating company must be aware that thecontractors will continue to need guidance, support and time;the key to success is for the operating company to ensurecontinuous and consistent communications and direction.
Once the decision has been made that validation supportis needed for a specific reason, the client must request aproposal from a preferred contracting firm that outlines theirneeds and staffing requirements. This is the first and fore-most important tool that is essential for establishing a soundproject. The proposal creates a common understanding ofroles, responsibilities, deadlines, and budget constraints.Without this document, the project has the potential forroadblocks, missed deadlines, and an increase in spending.And without this tool, an operating company cannot success-fully implement the suggested best practices for validationoutsourcing - Figure 2.
How to Use the VMP to Establish BestPractices for Managing Contractors
With the proposal in place, the project team needs to evaluate
what, when, and how to prepare for the contractors. Thisexercise can be performed during the VMP development andin tandem with the proposal development or award of bid toensure the most efficient use of the contractors. The VMP willdocument a list of internal and external references that thecontractors will need to fully understand the validationpractices, policies, and project overview. Use this documenta-tion to get the contractor started in the most efficient manner:
“Usually, my biggest complaint is that the operatingcompany doesn’t think about what a contractor needs toget the job done... I spend time digging up informationwhen they don’t have the materials ready for me to workwhen I arrive.”
This documentation typically includes corporate and depart-mental procedures, project plans, design specifications, andprotocol templates. The benefit of compiling the informationfor review is to minimize the contractor’s time spent trackinginformation down and maximizing the time for your project.
“...it can take me five days to learn where materials arelocated. It slows down the project and is very frustrat-ing.”
As the project team identifies timelines for the VMP, consid-eration should be given to schedule minimal time for contrac-tors to become knowledgeable about the owner’s documentsand facilities. By compiling the documentation prior to thecontractor’s arrival, significant time and energy will be savedand utilized more efficiently as the project unfolds. To furtherproactively manage the learning curve, appoint a ProjectLead or point person in the Responsibilities section of theVMP. A single person within the organization familiar withthe culture, practices, and end goal of the project can addressconcerns and begin to lead the validation contractor towardthe common goal.
“Everyone has a learning curve. When I am first on-site,I have to learn my way around and who is who. Peoplehave to get to know my role. Otherwise, I try to schedulea meeting and people respond by saying “who’s this, whydo I have to go meet with him?”
Finally, organizational expectations should be communi-cated with key players and how their role may impact theproject. By simply sharing information internally, the overallproject awareness heightens and the key players can begin toprepare as they see fit.
Laying a Foundation for Quality WorkDelivering the compiled documentation and providing ad-equate training should occur on the first day contractorsarrive. The documentation review and training will allowtime for the contractor to sense the company culture andbecome accustomed to the new environment. This exercise is
not only creating efficiencies in contractor utilization, but italso satisfies the cGMPs:
21 CFR §211.25 Personnel Qualifications(a)“Each person engaged in the manufacture, process-ing, packing, or holding of drug product shall haveeducation, training, and experience, or any combina-tion thereof, to enable that person to perform theassigned functions...”
The initial role of the project lead is to support and facilitatethe development of the contractor. The best practice is tosimply have the project lead at the disposal of the contractorsto address questions, provide guidance, and ensure adequatetraining to support the job function of the contractor has beensatisfied.
“There was one contact person. That was my personalbest experience just because I got thrown into such a messand he was there to help me out.”
There are two best practices for meeting key players thatcould be spelled out in a VMP. The first is to have the projectlead organize a kick-off meeting with all key players in theroom at the same time. The advantage of this approach is thateveryone hears the same message. The second approach is toindividually introduce the contractor to key players. Thelatter approach often is less stressful for contractors sinceone-on-one meetings allow greater in-depth knowledge of oneanother. Once the introductions are made, it is up to thecontractor to partner with the individuals to ensure theproject stays on track.
Finally and most importantly, clear communication is thekey ingredient to a successful validation effort. There are twocomponents to communication. The first is active listeningand the second is articulation.
Active listening is much more than jotting down notes orcatching a word here and there. The word “active” is theessential component of active listening. It means to be en-gaged in the conversation, to observe nonverbal signals, toname emotions, and to get to the heart of a discussion. Thesecond element of communication – articulation – providesan opportunity to clearly convey a request, need, or informa-tion. Articulation is characterized as clear expression oflanguage that is meaningful and accurate. Articulating apoint requires the ability to relate to the audience, adapt tothe audience’s style, and state the position.
Active listening and clear articulation are important skillsfor a point person and contractor. In contrast, a ValidationEngineer talked about a poorly administered validationproject:
“Communication was awful. Key people had a wealth ofinformation, but they would never share it with the restof the team. They would only share little pieces and theyprovided no explanations for their reasoning.”
Figure 2. Validation contractor management overview for the operating company.
Coupling active listening with articulation by the pointperson and contractor paints a clear picture of what ishappening. Such communication allows the project lead todetermine the roadblocks, implement a plan of action fortesting deviations, be informed of accurate updates, andestablish strong relationships.
Implementing for EfficiencyIt is essential for the operating company to take the lead andestablish the foundation for the contractor as well as theproject. The efforts thus far should have created commonunderstandings between the contractor and key players,setting a positive tone and sense of partnership for the
Figure 3. Validation client management overview for the contractor.
validation efforts. This foundation empowers the contractorto optimize her skills and implies continued active involve-ment by your company’s project lead and key players.
Maintaining common goals and sense of urgency amongthe validation team throughout a project is imperative. At thesame time, reality often intrudes into the life of a project:schedules shift, equipment breaks, unions threaten or go onstrike, or other dramatic events occur. The VMP should be aliving document and change as reality changes. But thequestion is: how are deadlines met under these new anddifficult circumstances? How do you continue to share thesame sense of urgency with the Validation team in a differentcontext?
Time and time again, the answer is through relationshipbuilding. The client-contractor relationship is what drivesthe project. A savvy Validation Engineer explained:
“Any relationship in this business is my priority. In thelong run and in the short run, the relationships impact
the success of the project.”
Making the relationships and communication the priority isthe most effective way to resolve project issues. Specificallythe lead person should establish a regular process and sched-ule for communicating and updating contractors. For ex-ample, one point person held regular Monday morning meet-ings to update contractors on progress of the project and totalk about any problems in the proceeding week. If the pointperson does not assume these tasks, then contractors canlead the communication efforts. The savvy Validation Engi-neer continued:
“I am a firm believer in “footstones,” a quick down-and-dirty e-mail about what I accomplished during the week.Especially in some projects, it is difficult to get fiveminutes of face time with the lead person. However, thatdoes not stop me from communicating with him!”
If communication is strong between client and contractor,then small problems are identified early and fixed. In such asituation larger problems are often avoided.
Finally, remember that the client-contractor relationshipis primarily between the project lead and contractor(s). Astrong relationship allows the client to acknowledge thevalidation priorities, work through problems efficiently, andexercise strong project management skills. A long-time vet-eran of validation projects said:
“….what it came down to was having that one contactperson…that’s what made the project work efficiently.”
What are best practices for the contractor?Since the client-contractor relationship is a two-way street,there are two sets of responsibilities. Some aspects of theresponsibilities are the same, while others are different.
Clients may need to remember what motivates contrac-tors. Part of the excitement of contracting is traveling andmeeting new people. More importantly, however, contractingallows a Validation Engineer to broaden his knowledge byvisiting companies around the world. The exposure to manysites increases the contracting Validation Engineer’s experi-ence with equipment, systems, validation philosophies, pro-tocol formats, and approaches to validation project manage-ment. In many ways, this exposure educates the contractorabout best practices.
Since contractors often have broad experience in valida-tion engineering, what specifically do they need to do on eachand every project?
Preparing for the Client SiteMore often than not, a contractor is flying from one site to thenext with minimal downtime. The contractor’s preparationtime often begins upon site arrival - Figure 3.
Developing a Foundation for SuccessA contractor needs to take the initiative in four fundamentalways. First, the contractor needs to read and understand theclients’ documents. Second, meet the point person and the keyplayers for the project. If the company has not appointed alead person, then explain the importance of the role. Ifcompany insiders are evasive about acknowledging key play-ers, then ask several insiders the same question. With smartdetective work, contractors can identify the key players.Third, ask questions based upon the documents and informa-tion that the contractor has been given – in other words, don’task questions that were answered in the written materials.And fourth, establish communication channels, schedules,and what should be conveyed in the messages.
Implementing EfficiencyFor a contractor, the key elements to project success includedemonstrating commitment to quality, leveraging thecontractor’s existing skill set, and strengthening relation-ships - Figure 3.
Preparing for the Client SitePreparation essentially begins when the contractor arriveson-site. It becomes the contractor’s responsibility to quicklyadapt to the operating company’s style and validation goals.The first identified best practice is to meet with the Valida-tion Manager to discuss your role and their expectations. Thisinitial dialogue creates a common ground for the client-contractor relationship and enables the contractor to sensethe management style for the Validation Group.
To gain a clear idea of your role, it is more important to askgood questions. As a contractor, there are stacks of docu-ments to read through, understand, and act upon thosedocuments, whether it is preparing a protocol or gowning fora cleanroom. In all cases, the contractor must learn to askgood questions that lead to direct and clear answers to moveforward on the project. A successful Validation Engineernoted:
“I ask good questions to get the job done.”
Developing a Foundation for SuccessFor the contractor, the first step in the foundation phasefocuses on reviewing documents, undergoing training, andgetting up to speed with the operating company’s policies andproject. Contractors should be careful with this documentand policy review since all companies will be different. Thesecond foundation for success for the contractor is meetingwith the lead person and key players and learning to under-stand and manage their styles and meet their expectations.The third step that contractors must take is to ask smartquestions about the documents and the players in the orga-nization. This is not a moment to gossip, but rather to notehow the company is structured, people’s roles, and how peoplelike to interact. For example, are meetings or e-mail mes-sages the preferred means of communicating?
The fourth step for the contractor is to identify the depart-ments and to name the people upon whom the project de-pends. Often several departments, such as manufacturing,quality assurance or microbiology, perform essential tasksfor the Validation project. As a result; learning how tomanage relationships across the departments is a significantpriority during the foundation phase.
Implementing EfficientlyOnce assigned to a Validation project, the client expects thecontractor to take the lead and drive the project. To do this,it is imperative that the contractor demonstrates commit-ment and dedication to quality work and to the overallproject. One practice that demonstrates commitment is to bevisible in the company, to show support for the key players,and to offer to help with different tasks. A successful Valida-tion Engineer explained his approach:
“I was real hands-on. They saw me every single day; itinstilled in them that I did care about the project andthat I would help in whatever way that I could.”
Contractors also are expected to be or become the expert of theequipment or system. The client anticipates the contractorwill leverage their ability to learn and past experiences todeliver on time and within budget.
“You need to be the expert, especially if no one is there toguide you.”
Exceptional Validation Engineers know that throughout theproject it is their responsibility – as much as the insider leadperson – to communicate continuously both the good and badnews. Smart, constant, and consistent communication helpsto build trust and to strengthen the relationship.
What does it all mean?Validation Master Planning is not only the latest craze; it hasevolved into an instrument enabling operating companies toproactively plan for large-scale validation efforts. The docu-ment forces the project team to ask and answer difficultquestions and has become a practicable management re-source.
Validation Outsourcing, Validation Project Plans, whatdoes it really mean? For operating companies, validationoutsourcing coupled with a VMP is a tool to achieve andensure compliance. For contractors, validation is a way of life.For both, it is establishing common goals, clear communica-tions, and solid relationships.
References1. Code of Federal Regulations, Parts 210 and 211 cGMP
Manufacturing, Processing, Packing, or Holding of Drugsand Finished Pharmaceuticals.
2. European Commission, Final Version of Annex 15 to theEU Guide to Good Manufacturing Practices.
3. Process Validation Requirements for Drug Products andActive Pharmaceutical Ingredients Subject to Pre-MarketApproval, Compliance Policy Guide Sec. 638.100 (7125.38),issued 08/30/1993, revised, 03/12/2004.
4. Guideline on General Principles of Process Validation,May 1987.
5. Guidance for Industry: Q7A Good Manufacturing PracticeGuidance for Active Pharmaceuticals, August 2001.
About the AuthorsKatie Henchir is a Senior Validation Engi-neer, specializing in validation project man-agement and equipment validation. Henchirhas successfully completed validation projectsthroughout New England and is a MBA can-didate at the Simmons School of Manage-ment. She can be contacted by e-mail: [email protected].
Dr. Cynthia Ingols is an Associate Profes-sor of Management at the Simmons School ofManagement, Boston, Massachusetts. Oneof her special areas of study is the art andscience of outsourcing relationships acrossdisciplines and industries. She can be con-tacted by tel: 1-617/521-3837 or by email:cynthia.ingols@ simmons.edu.
This articledescribes how aproperly plannedand executedcommissioningstrategy caneliminatedownstreamproblems andaccomplishmuch of thedata required forqualificationsand plantdelivery.
Figure 1. Typical phasesof a project.
Introduction
Quality, risk management, and time-to-market are probably the most im-portant aspects of a biopharmaceuticalproject. These elements seriously im-
pact the viability of a drug. Missing a launchdate for a product or losing a race to marketmay result in serious loss of revenue and/ormarket share.
Quality of a drug is a prerequisite for suc-cess and it must be built in at every stage
during development, design, construction,manufacturing, and distribution. Quality mustbe established at the outset and the appropri-ate level of quality must be determined for allphases of a project.
As in all industries, anticipation, analysis,and management of risks are a constant chal-lenge requiring appropriate proven methodol-ogy.
The above elements are critical to the suc-cess of a project.
BackgroundThe biopharmaceutical industry is one of themost regulated industries, due to the nature ofthe products and the regulations that governtheir usability. A drug is heavily controlledfrom the point of molecule discovery to thepoint where it reaches the patient either via aprescription distributed through a pharmacy/chemist or in hospital. This has madebiopharmaceutical companies cautious and con-servative with regard to the scope awarded to acontractor in an Engineering, Procurement,and Construction (EPC) project. Most indus-tries would feel comfortable awarding a projectto a contractor and having them conduct con-struction, mechanical testing, commissioning,and performance trials ready for handover andproduction - Figure 1. In the biopharmaceuticalindustry, handover is normally performed at“mechanical completion.” This approach hasput pressure on cost, time-to-market (as inte-gration becomes more difficult), and also hasplaced more pressure on clients to participateextensively throughout the whole project.
Many engineering firms have developed “in-tegrated approaches” to EPC, but the key isachieving a reduced time-to-market and a bet-ter quality product, while managing client risksappropriately and cost effectively. The successof this is still being debated.
Construction companies have a good trackrecord in risk management by nature of their
work. Are such firms more suited to integrate installation,commissioning, qualification, and validation? And throughthis integration, can they reduce time-to-market, provide aquality facility, and manage the client’s risks effectively?
For the last 15 years, the industry, led by many engineer-ing firms, has marketed the concept of integrating engineer-ing, procurement, construction, and validation for new phar-maceutical/biotechnology facilities. Certainly, the idea is acommendable one; however, in reality, it has no significantimpact on time-to-market or cost. In many cases, the costescalated and the schedule was extended due to the failure ofintegration and the lack of quality documentation by theconstructor.
In the last 15 years, engineering design has come a longway in terms of Good Practice (GP) compliance throughproperly documented and executed GP audits. Advanceshave been made to the point where the work product of any
major engineering firm specializing in this business can bedeemed to be GP compliant.
The Mystique of ValidationCommissioning and validation have become a costly andtime-consuming exercise. For large, new capital expansionprojects, an owner’s cost for validation, inclusive of bothinternal and external services, includes spent labor andmaterials, and is on the same order of magnitude as typicalcosts for engineering or construction management services.While actual validation costs will vary depending upon anowner’s approach and the nature and location of the project,the range of costs are shown in Table A.
In the majority of cases, much attention has been paid toqualification/validation at the expense of commissioning.
The effectiveness of commissioning as a proven method toexpedite plant delivery has been overshadowed in the phar-
Figure 2. Typical phases of a pharmaceutical project.
maceutical industry by the emphasis on qualification/valida-tion. Often the problems encountered in qualification are dueto incomplete commissioning.
A properly planned and executed commissioning can elimi-nate many downstream problems and accomplish much ofthe data required for qualifications and plant delivery.
Construction QualificationIn light of the “Risk-Based Approach to Validation” and theincreasing pressures on cost and time-to-market, a newmethodology is needed to ensure the true and successfulintegration of construction, commissioning, and qualifica-tion.
Many API producers in Europe were not familiar withqualification/validation some 15 years ago; however, theyknew that in a regulated industry they needed to ensurequality and competitiveness so they relied on GP and riskanalysis as part of a methodology, namely Design Qualifica-tion (DQ).
In Europe, bulk producers regarded DQ as “qualification”for a long time before installation and operational qualifica-tions were enforced. Typically, Design Qualification encom-passed:
• Design and GP/Audits• Risk Assessment and Criticality Analysis• User Requirements Specifications (URSs)• Traceability of Changes
This methodology worked well from a design perspective, butwas not extended effectively to the field - Figure 2. Thus,providing design compliance without much impact on costand time-to-market. Extrapolate this methodology to thefield and you have Construction Qualification (CQ).
The CQ methodology is aimed at reducing cost and time-to-market through a number of critical steps as follows:
• Risk Assessment and Criticality Analysis• Construction Audits at Approved For Design (AFD), Ap-
proved For Construction (AFC), and during field activities(based on Risk/Criticality Analysis)
• Turnover Package Organization• GP construction forms• Control and traceability of field charges
See Figure 3 (CQ Approach).
“CQ is a prerequisite to successful integration withCommissioning.”
The activities stated above are key to expediting a project toconclusion and delivery. They impact commissioning, asmany of the final construction activities (for mechanicalcompletion) are entwined with pre-commissioning/commis-sioning activities.
Mechanical completion is the phase between installationand commissioning, in which components of the plant/facility
are proved to be mechanically fit for their duty. It can beconsidered as a specialized part of the pre-commissioningactivity in which each component is prepared for processcommissioning. Since installation may be continuing in someareas of the plant while others are being tested and commis-sioned, site safety must be given detailed consideration. Forexample, component suppliers and sub-contractors must becarefully controlled during this phase since areas can changeclassification during the course of construction and commis-sioning.
Generally, pre-commissioning refers to preparing the fa-cility/plant for the introduction of process materials, and itsmain purpose is to eliminate any problems which might ariseat later and more critical stages of facility/plant operations.
The sequence of mechanical completion is governed by theoverall program, but usually starts with electrical power andutilities. The objective of mechanical completion is to provethat an installed plant component is suitable for commission-ing.
CommissioningProperly planned commissioning begins during the pre-con-struction phase of a project. During this time, the parametersfor commissioning and qualification turnover documents areidentified. Also, Factory Acceptance Test (FAT) plans andSite Acceptance Test (SAT) plans are developed for pre-purchased equipment and systems.
The goal is to have the commissioning and closeout docu-mentation requirements identified in outline form prior tothe start of construction. Specific requirements for long leadequipment and modules require definition and will be fullydeveloped for incorporation into the bid documents.
Overall, it is the intent to utilize the project’s commission-ing process to enhance and reduce the time taken for qualifi-cation, hence reducing time-to-market. Properly documentedcommissioning can be leveraged into qualification by sys-tems and completing the process in phases, allowing for earlyproduction and manufacturing.
Commissioning is defined as a well planned, documented,and managed approach to the start-up and turnover of sys-tems and equipment to the end-user that results in opera-tional, safe, and functional systems, which meets establishedoperational requirements and end-user quality expectations.
Engineering Construction Validation1
Management
Bulk Chemical API 10 - 14% 5 - 11% 5 -7%
Bulk Bio API 14 - 18% 5 - 11% 10 - 15%
Secondary 7 - 10% 4.5 - 8% 5 -9%Pharmaceuticals(Solid Dosage,Liquids, andOintments)1 Includes owner spent material and plant labor costs up through qualification
costs.
Table A. Typical Engineering, CM, and Validation Costs (% ofTotal Installed Cost “TIC”).
Commissioning can be accomplished through differentphases and methodologies. There are a number of provenmethods to achieve commissioning in the biopharmaceuticalindustry. For purposes of illustration, it will be broken downinto six phases to reflect the various tasks that will beexecuted through the project. These phases and tasks aresummarized below and described in detail in the followingsections.
1. Design Phase• Kick-off of Commissioning Activities• Focus Design Review (System Impact Assessment)• Documentation Requirements• Commissioning Protocol Writing (toward the end of
3. Construction Phase• Component Impact Assessment• Commissioning Protocol Writing and Approval• Construction Quality Control Activities• Owner Quality Assurance Activities
4. Start-Up Phase• Construction Quality Control Activities• Trade Contractor Pre-Commissioning Checks• Owner Quality Assurance Activities
5. Inspection, Testing, and Documentation Phase• Construction Quality Control Activities• Owner Quality Assurance Activities• Installation Commissioning/Verification “IC”• Initial Calibration• Operational Commissioning/Verification “OC”• Training
6. Handover to End-User Phase• Closeout Reports/Deviation Resolution
Design PhaseThis is the phase of the project when the scope of commission-ing is defined, the commissioning team is assembled, respon-sibilities are assigned, information is obtained, and protocolsdrafting is planned.
Kick-off of Commissioning ActivitiesThe project manager assigns a commissioning team for theproject. The project has a basis of design, a control levelschedule and control estimates, and a preliminary equip-ment list available to assist the commissioning team indefining the commissioning activities. The commissioningteam breaks the project into a series of systems, whichformulates the basis for the commissioning plan.
The first draft of the commissioning plan is produced asoutlined by the team. The commissioning plan is distributedfor review and comment. Subsequent team meetings are heldto review and resolve comments.
Focus Design Review (System ImpactAssessment)The team conducts a System Impact Assessment using thesystem list. The systems list covers the entire scope of theproject, broken up into manageable segments. Typically,these are by equipment package, distribution or piping sys-tems, and architectural items.
The team assesses each system with regard to its effect onproduct quality. In effect, the systems will be categorized intoone of three categories.
• Direct Impact on Product Quality• Indirect Impact on Product Quality• No Impact on Product Quality
The Direct Impact Systems require further qualificationafter commissioning and the Indirect Impact and No ImpactSystems will not require further qualification after commis-sioning. However, this is dependent on company policies, forexample, some companies will further qualify some IndirectImpact Systems depending on the criticality of the interfacewith a Direct Impact System. These categories and theassessment criteria are further defined by the commissioningteam and are usually documented in the System ImpactAssessment Report.
Documentation RequirementsDuring the design phase, the User Requirement Specifica-tion (URS) and Design Specifications for Good Manufactur-ing Practice (GMP) critical systems and equipment should bereviewed with regard to the vendor/contractor documenta-tion required to support commissioning and qualification aswell as operations and maintenance. Where appropriate,documentation numbering, layout, formats, etc., should bespecified. In most instances, the equipment/system vendor isbest placed to provide the documentation required to supportthe commissioning and qualification effort. Therefore, thismust be stated during the design phase of the project so that
the documentation becomes one of the key deliverables forthe vendor/contractor.
Commissioning Protocol WritingThe team will decide how to generate the commissioningprotocols and who will execute them. Two approaches exist.
1. The equipment manufacturer (vendor) provides a SiteAcceptance Test (SAT), which is incorporated into thecommissioning protocol. The vendor also executes theSAT.
2. The commissioning team writes the commissioning proto-col for engineered systems, such as utility distributionsystems. Subject matter experts are consulted as required.The commissioning team also executes the protocol.
Procurement PhaseThe procurement of subcontractors, vendors, and equipmentdesign and fabrications systems could potentially have “addedvalue” to the overall schedule and cost of a project. Withoutproper integration of design, prefabrication, and construc-tion, the maximum benefit may not be obtained. In addition,without a rigorous implementation strategy, not only willinefficiencies result that erode the schedule and cost benefits,but the end product may be viewed as a compromise and fallshort of expectations.
A successful approach must influence the project from theearly stages of preliminary engineering. This early involve-ment will yield dividends for every phase of the project.
1. Objective• Maximize the use of ‘Equipment Modules” to provide the
optimum combined schedule and cost benefit value whileincreasing the overall quality and improving the project’sschedule.
2.How Implemented• Assemble a team of individuals who possess a unique
combination of pharmaceutical/biotech design and con-struction experience.
• A team with the design, construction, and integration ofskidded process equipment.
• A team with know-how in project turnover requirementsfor Good Practice facilities.
• A team with experience in the start-up and commissioningof pharmaceutical/biotechnology facilities.
• Empower the team to be part of the up-front engineering
3.Engineering and Design Recommendations• Develop module boundaries for the project.• Lead in the development of an “Equipment Module De-
sign, Fabrication, and Installation Standard” (EMDFIS).• Review and critique layout and general arrangement
studies in regard to module implementation.• Develop engineering and design boundaries between pro-
cess engineer/design firm and equipment module manu-
Figure 4a. Proposed organizational chart.Figure 4b. Alternative organizational chart (C&Q =Commissioning and Qualification).
facturer to optimize design schedule and cost.• Prepare a construction strategy that identifies field versus
shop work and interface requirements as well as stan-dards for controls, piping, electrical, etc.
4.Procurement Strategies• Conduct pre-qualification visits to potential suppliers of
equipment modules.• Evaluate best methods of design and construction that
may influence the EMDFIS.• Evaluate integration options of module boundaries.• Develop options for the field and module scope boundaries.• Review overall shop capabilities in regard to projected
workload, shop capacity, quality program, maximum as-sembly size, technical capability, turnover documents, etc.
• Develop a procurement strategy to maximize buying powerand scope distribution across vendor availability.
• Evaluate major equipment, instrument, and controls pro-curement with drop ships to equipment module vendorsversus turnkey approach.
Construction PhaseSubcontractors, vendors, operations, maintenance, and engi-neering develop support documentation that will be reviewedduring commissioning.
Component Impact AssessmentTo evaluate the impact of system’s components on product
quality, the team meets to review the details of each system.Similar criteria that were used for the System Impact Assess-ment are used to judge a component’s effect on productquality. Although all systems undergo a component impactassessment, emphasis is placed on the direct impact systems.The spirit and principle of risk analysis and managementplays a vital role here, and this information also is recordedin the Component Impact Assessment Report.
Commissioning Protocol Writing and ApprovalIt is the commissioning team’s responsibility to ensure thatthe proper information is getting to the individuals who arewriting the protocols with the operations and maintenancerepresentatives on the team serving as the focal points of thisinformation flow.
The data for these protocols is gathered from end-users,the construction manager, third parties, or subject matterexperts as required.
The commissioning team reviews and approves all proto-col submittals and then signs the protocols prior to executionaccording to the document approval matrix.
Note: If commissioning is to be leveraged into qualifica-tion then the involvement of the clients’ Quality Assur-ance (QA) organization is a pre-requisite. The level ofinvolvement is critical as this impacts the approvaltimes and the overall schedule.
Construction Quality Control ActivitiesAt this stage of the project, construction groups and equip-ment vendors review documentation and drawings for designcompleteness and adherence to building codes and practices.As construction progresses, the quality control activitiesbecome more physically orientated to ensure installationcomplies with approved design. Deviations are tracked in theproject worklist/punchlist.
Owner Quality Assurance ActivitiesSimilar to the construction control process, but with ownerparticipation along with the construction groups and engi-neers who review documentation and drawings for designcompleteness and adherence with regulatory requirements,operational requirements, and best practices. As construc-tion progresses, the quality control activities will becomemore physically orientated to ensure installation complieswith approved design. Deviations are tracked in the projectworklist/punchlist.
Start-Up PhaseVendors and system representatives power-up the systemsand perform necessary procedures to make the systems fullyoperational. This phase culminates in the handover of sys-tems to the commissioning team.
Construction Quality Control ActivitiesThe construction manager and the commissioning team con-duct periodic reviews of the construction progress and the
quality of the installation (walk downs). Deficiencies aretracked in the project worklist/punchlist, which containscommissioning, qualification, and general items still requir-ing completion.
Trade Contractor Pre-Commissioning ChecksPrior to system or equipment start-up, the trade contractor isresponsible for performing valve/equipment line-ups to en-sure that all equipment is in the proper operating conditionand no equipment or system damage will occur.
Continuity checks also are performed and documented inaccordance with the specifications.
Turnover of a system from construction to commissioningis based on the acceptance of a system by the commissioningteam. The following items are typically required to defineconstruction process as being complete:
• Construction manager, contractors, vendors or systemrepresentatives, engineering, and operations sign off onthe construction turnover package.
• All installation documents required to support commis-sioning are complete and available.
• Required utility services are available in adequate supplyto properly operate the system.
• All controls signals from external sources are available orcan be reliably simulated.
• Equipment/system start-ups requiring lockout/tagout forequipment or personal protection are performed usingowner procedures.
• After the system has been checked, the construction man-ager assembles the completed forms and provides them tothe commissioning leader for review. The constructionmanager provides copies of the completed forms to thecommissioning leader and keeps the originals for inclu-sion in the turnover package.
Owner Quality Assurance ActivitiesSystem walkdowns, which begin when installation of a sys-tem is approximately 90 percent complete, are coordinatedwith the owner representative. The construction managerinforms the owner representative prior to system/equipmentstart-up, and coordinates times when the equipment could beavailable for certain activities, should the owner representa-tive need or wish to access the systems/equipment at anytime.
Coordination with the owner representative is critical toensure that start-up of the equipment does not affect areasoutside of the scope of specified project.
After a successful start-up and commissioning has beencompleted, plant personnel including facilities engineering,operations, and safety are notified that the system is readyfor them to prepare, execute, and issue plant specific readi-ness reviews or an Operational Readiness Report (ORR)indicating it is safe to be turned over to operations for regularuse.
Start-UpThe equipment vendors and/or contractors review their owninternal installation complete checklist to make certain theequipment/system has been properly installed and is ready tobe safely activated.
The equipment vendors/engineers will activate the equip-ment, and perform all necessary activities required to makethe equipment/system fully functional. This includes check-ing liquid/lubrication levels, checking motor rotations, tun-ing loops, debugging installation problems, confirming in-stallation against as shipped drawings, setting system spe-cific parameters, and making the equipment/system readyfor testing.
Calibrations also are performed during this phase, whichvendors require to finish their start-up procedures. Full loopchecks are performed (field device through software to con-sole or vice versa) and documented.
Inspection, Testing, andDocumentation Phase
The commissioning team executes the commissioning proto-cols in this phase. The executed protocols, system closeout,and handover reports are then reviewed by the team. Execu-tion of the commissioning protocols confirms that the instal-lation was performed according to the approved design. Theacceptance criteria are defined in the approved design docu-ments.
Installation Commissioning/VerificationThe commissioning team reviews the available documentscomparing them to the requirements outlined in the Engi-neering Turn Over Package (ETOP). The commissioningprotocol execution ensures that the required documents arecomplete and available.
The installation is checked against the approved draw-ings. This process includes such activities as P&ID verifica-tion, general arrangement drawing verification, and name-plate verification.
Initial CalibrationProper documentation of the calibration is referenced back toa traceable standard depending on the country, e.g., USA:NIST. The initial calibration is performed as part of thevendor or system start-up activities. The documented evi-dence is reviewed at this stage.
Operational Commissioning/Functional TestingThe commissioning team system representatives execute thecommissioning protocols to ensure proper operation of themachine as defined in the approved project documents. Thecommissioning team signs off the executed protocols accept-ing the results of the execution.
TrainingThe commissioning team ensures that operator and mainte-nance training has been addressed to the end-user’s satisfac-
tion. The commissioning protocol ensures that training docu-mentation has been provided to the end-user and that train-ing has been scheduled.
Handover to End-User PhaseCommissioning is complete; the system end-user formallyaccepts the systems. Plant maintenance is responsible for thepreventative maintenance of systems, some of which undergofurther qualification.
For true integration to take place and in order for commis-sioning to be leveraged into qualification, it is stronglyadvised to have the same team members perform qualifica-tion for the systems they commissioned.
Closeout Reports/Deviation ResolutionCloseout reports address open issues, identify correctiveactions required, the responsible party, and dates for comple-tion. Deviations, documented on the project worklist/punchlist,are reviewed to ensure that all remaining open issues aretransferred to the closeout report.
Project Organization and ExecutionThere are many different ways to organize a commissioningteam, but the most time and cost-effective is when commis-sioning and qualification activities are integrated.
It is not the intent of this article to offer details withrespect to project organization and execution; however, the
following structures are proposed as examples - Figures 4aand 4b.
Commissioning in Support of QualificationEarlier in the article, reference was made to the fact thatoveremphasis on qualification/validation has overshadowedcommissioning, resulting in problems during validation dueto incomplete commissioning. Some of these problems can bedetrimental to cost and time-to-market since fixing themrequires a high level of backtracking and mending of instal-lation and documentation.
Commissioning performed in new construction and exist-ing facilities helps to ensure that systems are installed,functionally tested, and capable of being operated and main-tained to perform in conformity with the design intent andthe owner’s needs. This ensures that a new facility begins itslife cycle at optimal productivity. Commissioning also canresult in restoring an existing facility to optimal operation.Furthermore, when commissioning is repeated periodicallythroughout the life of a facility, it improves the likelihood thatthe facility will maintain a higher level of performance.
Placing more emphasis on Documented Commissioning(DC) may have cost and schedule consequences. In general,qualification costs can be at least twice as much as that ofcommissioning. Reversing the emphasis will make the cost ofdocumented commissioning higher. However, the cost ofqualification could come down significantly as documented
Figure 5. Leveraging commissioning into qualification.
commissioning can be the lion’s share of the effort requiredfor qualification. More significantly, by adopting documentedcommissioning as the basis of qualification, clients couldsignificantly reduce the risk of non-compliance and seriousproblems affecting the delivery of a qualified facility, hencereducing time-to-market.
In order for DC to work effectively as the basis for qualifi-cation, early active participation of the client’s quality unit iskey to ensuring that various commissioning activities areeventually accepted for inclusion in support of the installa-tion and operational qualification. These protocols, alongwith any performance qualification protocols that are re-quired, form the basis of the qualification/validation effort.This is in accordance with the framework set forth in theISPE Baseline® Pharmaceutical Engineering Guides for Newand Renovated Facilities, Volume 5, Commissioning andQualification. It is worth referencing the ISPE definition ofcommissioning as a well-planned, documented, and managedengineering approach to the start-up and turnover of facili-ties, systems, and equipment to the end-user that results ina safe and functional environment that meets establisheddesign requirements and stakeholder expectations.
DC must be treated as a unique and discrete activity inaccordance with the above definition to be used as the basisof qualification/validation. In many cases, where attemptswere made to mix commissioning and qualification togetherserious delays and shortfalls occurred - Figure 5.
The FutureThe new order for our industry is, as always, driven bypressures on cost and constant changes to meet market de-mands. The new industry drivers are risks (analysis andmanagement), cost, and time-to-market. If you agree that
those stated above are the real drivers in our industry, wouldit make sense to expect C/CM contractors to deliver a qualifiedfacility rather than a mechanically complete facility. Clearly,the goal is to be cost-effective, fast, as well as comprehensive.Redundancies in testing may be eliminated through the imple-mentation of a smart and efficient approach to installation andoperational qualification of systems and equipment. Thislogic, or “Qualification Rationale” as it is called by the ISPEBaseline® Guide, can be achieved through the integration andimplementation of Construction Qualification (CQ) and Docu-mented Commissioning (DC).
Finally, the next decade may see Documented Commis-sioning replacing Qualification and/or Qualification becom-ing the QA function for Documented Commissioning.
About the AuthorWael Allan is a Senior Vice President of theSkanska Pharmaceutical Group, leading itscommissioning, validation, and regulatorycompliance services globally, and serving asManaging Director of the PharmaceuticalGroup’s U.K. entity. Allan has extensive ex-perience managing a wide range of commis-sioning and validation businesses in the phar-
maceutical and biotechnology realm. He is responsible fordeveloping the group’s global strategy as well as focusing onoperations, business development and growth. Allan holds aMSc. in biochemical engineering and B.Eng. in chemicalengineering, graduating with honors, from the UniversityCollege of Swansea in the United Kingdom. He can becontacted by tel: 1-973/390-9219 or by email: [email protected].
Skanska USA Building Inc., 1633 Littleton Rd., Parsippany,NJ 07054.
Investigation of the CorrosionCharacteristic of AL-6XN FusionWelded to Inconel Alloy 22by Arnie Grant and John Jermain
This articledescribes theresults of theelectrochemicalcorrosion testingof the aswelded,mechanicallypolished, andelectropolishedAL-6XN fusionwelded toInconel 22. Alsotested was asingleelectropolishedweld segment,which wassubmerged in aferric chloridesolution inaccordance withthe ASTM G-48procedure tofurther elucidatecorrosionresistance andsites of attack.
Table A. Specificationlimits for AL-6XN andInconel 22.
Acorrosion study was undertaken insupport of Fluid Line Technology de-velopment efforts to investigate thecorrosion characteristics of AL-6XN
fusion welded (no filler) to Inconel 22. Thepurpose of the study was to compare and rankthe corrosion characteristics of the dissimilarmetallic weld in the 1. as welded condition, 2.mechanically polished (180 grit) after welding,and 3. electropolished after welding. Of inter-est in this work, in addition to measuring thedifference in pitting potential, was to identifythe sites or specific susceptibility on the weldcoupons at which corrosion would initiate, e.g.,Heat Affected Zone (HAZ) and the type ofcorrosion, e.g., intergranular. A singleelectropolished weld segment was tested perASTM G-48, Pitting Corrosion by Use of FerricChloride to further elucidate corrosion resis-tance and sites of attack. All samples weretested as-received with no additional cleaningor passivation.
The specification compositional limits forthe two alloys are listed in Table A. Inconelalloy 22, a nickel-chromium-molybdenum-tung-sten alloy, and AL-6XN, a “super-austenitic”alloy, are utilized in corrosive environments forexcellent resistance to general corrosion, pit-ting, crevice corrosion, and intergranular at-tack. Due to their exceptional corrosion resis-tance, these two alloys have been used in alarge variety of industrial applications, andmost recently in the pharmaceutical/biotechindustry.
Alloy - AL-6XNAL-6XN was initially intended to be used in aseawater environment, but extensive testinghas demonstrated it to be resistant to a varietyof corrosive elements. Its excellent chloridepitting resistance is attributable to its 6.50%molybdenum content, while its significant re-sistance to chloride stress corrosion cracking isa result of its nickel content of about 25.00%.The addition of nitrogen enhances its pitting
resistance as well as mechanicalstrength. Nitrogen also serves tosignificantly reduce the formationof potentially harmful secondaryphases during the manufacture oflarge cross-section products. The Al-legheny Ludlum Corporation, whichdeveloped the AL-6XN alloy, hastested it against other stainless steelalloys and concluded that it is themost corrosion resistant iron-baseaustenitic stainless alloy presentlyavailable.1
Alloy - Inconel 22Inconel alloy 22 is a nickel-base al-
loy made up of 21.5% chromium, 13.6% molybdenum, and 3%tungsten. The material is an adapted version of Inconel alloy622, which offers superior resistance to pitting and crevicecorrosion in acid chloride solutions and resistance to generalcorrosion in mixed and reducing acids. The alloy is used tomanufacture a wide variety of chemical process equipmentsuch as: Flue gas scrubbers, chlorination systems, acid pro-duction and pickling systems, outlet ducting and stack linersfor power plants, sulfur dioxide scrubbers, pulp and paperbleach plants and for weld overlay of less corrosion resistantmetals.
Test MethodologyMaterial - Weld Test CouponsThe weld coupons were made from AL-6XN TUBE, 1.5" OD x0.071, Heat Number 023 BFW, and Inconel 22 Tube, 1.5" OD,Heat Number 024 BIG. Chemical analysis yielded the data inTable B, which are within their respective specificationtolerances. Each EPIT test coupon cylinder was 2" long x 1.5"OD with the weld at the center. No data was provided on thewelding parameters.
Electrochemical Corrosion Test - Cal-ChemPassivation Monitor (PAMO Meter)The pitting potentials (EPIT) of the three AL-6XN/Inconel 22weld coupons, 1. as-welded, 2. mechanically polished, and 3.electropolished, were measured using the electrochemicalPassivation Monitor (PAMO) shown in Figure 1 with the testcell configuration. The electrolyte solution is 1.0 M KCl and
Figure 1. PAMO meter and test cell.
the test temperature is 60°C, the counting electrode was316L, and reference electrode was Calomel. This field por-table monitor was developed by Cal-Chem in conjunctionwith the Materials Science Department at USC to providedata on pitting potential equivalent to ASTM G-61. Thisprocedure tests the resistance of the sample coupon cylinderto electrochemically induced pitting. The pitting potential forthese highly corrosion resistant alloys is measured after twohours at 60°C with an applied current to the test cylinder viathe potentiometer. For comparison, the pitting potential forthe much less corrosion resistant 316L alloy is measured in0.1 M KCl at 15 minutes at ambient temperature. This EPIT
value may then be conveniently used to compare the relativecorrosion resistance before and after passivation for the samealloy as well as comparison of corrosion susceptibility for thethree surface finishes above. The larger the value of the EPIT,the better is the resistance to pitting corrosion. The “Break-through Time” or time to onset of corrosion also was used asan indication of relative corrosion resistance.
ASTM G-48, Pitting Corrosion of Ferric ChlorideSolutionA single electropolished weld segment was supplied for the G-48 immersion test (Method A) in which the test coupon issubmerged in a 6% solution of ferric chloride at a prescribedtemperature and time to determine resistance to pittingcorrosion of stainless steels. The recommended elevatedtemperature is 50°C and time is 72 hours. The sample isremoved periodically, cleaned, examined, and weighed, andthe observations are recorded to give an appraisal of corro-sion susceptibility.
Photographic DocumentationPhotographs were taken of the samples to document theappearance of the internal surfaces before and after thecorrosion tests. Because it was noted that a very significantdiscoloration (oxidation) appeared on the post EPIT test speci-mens, it was decided to cut the cylinders longitudinally andto “derouge” one half of the cylinder to see if the discoloration(oxide) was removed. These discolored and derouged photo-graphs also are presented. After photographing the post testcylinders, they were bisected to document the areas of corro-sion attack.
Experimental ResultsPhotographic Documentation from theElectrochemical Pitting Potential TestingFigures 2, 3, and 4 present the photos of the as-welded,mechanically polished, and electropolished cylinders, (a)before the electrochemical pitting corrosion test looking downthe cylinder, (b) after the electrochemical pitting corrosiontest, and (c) before and after derouging of oxide discoloration.
On the as-welded sample (Figure 2) after EPIT testing, verydark blue/purple discoloration occurred immediately adja-cent to the weld bead on the Inconel 22 side at the down slopearea. To a lesser degree, yellow, blue, green, and purplediscoloration was very obvious on the entire Inconel 22
surface. It is presumed these discolorations are indicative ofoxidation of the various metals on the surface and variationsin oxide thickness. There was no evidence of pitting at 10Xmagnification. The AL-6XN side of the cylinder did notdisplay the bright profusion of oxide coloration. Instead,there were distinct areas (much more localized) of rust spots.These are taken as a precursor or nucleation areas of pitting.In addition, hazy areas appeared on the AL-6XN side imme-diately adjacent to the weld bead and in the HAZ, whichappeared to be intergranular corrosion. Verification of thelatter assumption would require further metallographic test-ing. After derouging (not passivation), the Inconel 22 discol-oration was removed, but the surface appeared mottled grayfollowing the contours of the earlier discoloration. A distinctstraw color remained on the AL-6XN HAZ, but the dispersedrust areas were removed.
The mechanically polished sample (Figure 3) displayed alesser degree of yellow, blue, and purple discoloration on theInconel 22 side and no apparent rust spots on the AL-6XNside after the electrochemical corrosion test. Intergranularcorrosion (hazing) did not appear on the AL-6XN side as it didon the as-welded cylinder. After derouging, the Inconel 22portion was a dull gray with a mottled appearance.
The electropolished sample (Figure 4) displayed the over-all yellow, green, blue, and purple discoloration on the Inconel22 side and a relatively slight uniform yellowing on the AL-6XN side. After derouging, the Inconel 22 was very slightlymottled and the slight yellowing was removed from the AL-6XN leaving a bright metallic shine.
As can be seen in the photographs, the surface of theInconel 22 portions of the as-welded and electropolishedcylinder appear to be pebbly (orange peel) in appearance andthe weld bead protrusion is very apparent. Mechanical pol-ishing removed the pebbly surface and the weld bead protru-sions - Figures 3 and 4. The as-welded coupon also displayedsignificant discoloration in the weldment area not present in
the other two cylinders. Extensive surface discoloration oc-curred on the Inconel 22 portion of all samples after thepitting potential test; the as-welded being the worst.
Results from the Electrochemical PittingPotential TestingThe pitting potential data and breakthrough times for thethree samples are presented in Table C and shown graphi-cally in Figures 5 through 9. With both pitting potential andbreakthrough times as the criteria, the electropolished samplehad the highest resistance to corrosion and the as-welded had
Figure 2. As welded (mill finish ID and OD).A. Before pitting corrosion test.B. After pitting corrosion test.C. Surface comparison of as welded samples (left = after pitting
corrosion test, right = after derouge). AL-6XN is shown abovethe weld and INCONEL 22 is shown below the weld.
Figure 3. Mechanical polished ID - 180 grit (no electropolish) - ODis mill finish.A. Before pitting corrosion test.B. After pitting corrosion test.C. Surface comparison of mechanical polished (left = after pitting
corrosion test, right = after derouge). AL-6XN is shown belowthe weld and INCONEL 22 is shown above the weld.
Figure 4. Electropolish ID and OD (mill finish).A. Before pitting corrosion test.B. After pitting corrosion test.C. Surface comparison of electropolished (left = after pitting
corrosion test, right = after derouge). AL-6XN is shown abovethe weld and INCONEL 22 is shown below the weld.
Table C. Breakthrough time and EPIT for the three surfaces.
Figure 5. Pitting potential (mV) curve for as welded cylinder.
Figure 6. Pitting potential (mV) curve for mechanical polishedcylinder.
Figure 7. Pitting potential (mV) curve for electropolished cylinder.
the lowest resistance.In the electrochemical pitting test, equilibration to the
open circuit potential is achieved at temperature before thepolarizing current is applied. Then the voltage is recorded asa function of time for a period of two hours at which time theEPIT is established. The millivolt vs. time plots for the as-welded, mechanically polished and electropolished weldedcylinders are presented in Figures 5, 6, and 7, respectively.The breakthrough time is taken as the time point at which arapid drop in voltage occurs signifying that corrosion hasbegun. This data is shown in bar graph form in Figure 8. Thepitting potential values of EPIT are taken uniformly at 120minutes elapsed time for comparison. The EPIT values areshown in bar graph form in Figure 9.
These results are tabulated in Table C, comparing break-through time and EPIT for the three surface finishes of the AL-6XN/Inconel 22 fusion weld. Both parameters show the as-welded sample at the lowest value, the mechanically polishedsample at a midpoint value, and the electropolished at thehighest, most corrosion resistant value. Taking the EPIT valueas the more important test parameter in the evaluation,mechanically polishing the ID of the weld improved corrosionresistance by 62% while electropolishing improved corrosionresistance by 93%.
Results from the Ferric Chloride Corrosion Test(ASTM G-48)Table D presents the ASTM G-48 weight loss data for theelectropolished segment at various times. Because no weightloss was observed after 60 hours at 50°C, the test tempera-ture was increased to 60°C for an additional period of 12hours. Photographs of the segment at test initiation (0 hours),and 72 hours (60 hours at 50°C + 12 hours at 60°C) are shownin Figures 10a and 10b. After 12 hours at 60°C, the samplelost 0.72% of its weight in the form of a single large pit (Figure10b) away from the weld bead and HAZ on the AL-6XN sidein the parent metal. No pitting corrosion could be seen withmagnification up to 10X in the Inconel 22, weld bead or theHAZ of either metal.
From this visual data, it would appear that the electro-chemical pitting potential test measures the nucleation stagewhile the G-48 test induced the rapid propagation stage ofpitting.
DiscussionInconel Alloy 22, a Ni-Cr-Mo alloy, demonstrates excellentcorrosion resistance under a wide variety of corrosive envi-ronments including pitting by chloride and oxidation by ferricchloride. The AL-6XN, a superaustenitic alloy with highlevels of nickel (25%) and molybdenum (6.5%) also demon-strates excellent corrosion resistance, especially under chlo-ride pitting conditions.
All three of the Al-6XN/Inconel 22 cylindrical fusion weldcoupons showed outstanding resistance to the two acceler-ated laboratory chloride corrosion tests that were employedto compare the as-welded condition to the post weld mechani-
cally polished and electropolished cylinders.As noted above to illustrate the exceptional degree of
corrosion resistance of these weld coupons compared to 316Lstainless steel, the pitting potential for 316L is measured in0.1 M KCl at ambient temperature after 15 minutes ofapplied potential. No corrosion could be induced on these testspecimens under these test conditions. After a series ofexperiments, it was determined that the test parametersrequired were 1.0 M KCl electrolyte at 60°C for a period of twohours to obtain meaningful pitting corrosion data.
In the as-welded condition, the Inconel 22 side of the weldappears to be susceptible to oxidation discoloration immedi-ately adjacent to the weld and in the parent metal but to asuperficial depth. Corrosion susceptibility appears to be on
the AL-6XN side of the weldment where possible intergranu-lar attack occurs in the HAZ and pitting occurs in the parentmetal.
By mechanically polishing the ID of the weld area, signifi-cant improvement in both breakthrough time and EPIT isseen. The Inconel 22 discolored to a lesser degree and thesignificant attack appeared to be on the AL-6XN portion. Byfar, the best corrosion protection is obtained by electropolishingthe ID after welding. However, oxidative discoloration is stillseen on the Inconel 22, but the significant corrosive attackstill occurs on the AL-6XN. It should be emphasized that thisis a very limited study performed on a single sample of eachsurface condition. Also, the pitting potential and G-48 testparameters were under development as part of this study. We
Id Number Weight at 0 Hours Weight at 24 Hours Weight at 48 Hours Weight at 72 Hours* Weight Loss % Weight Lossat 50°C at 50°C after 72 Hours** after 72 Hours**
should like to expand this study to obtain statistical valida-tion of our conclusions.
Furthermore, since we have shown that CHELANT passi-vation of AL-6XN removes significant levels of iron from thealloy surface, improving Cr/Fe Ratio by 100%, it would bemost prudent to explore the corrosion improvement obtainedby passivation of the as-received welds.1
References1. Grant, A., and Jermain, J., “The Effect of Passivation on
AL-6XN Alloy Compared to the Traditional 304L and316L Stainless Steel Alloys,” Pharmaceutical Engineer-ing, March/April 2003.
2. Grant, A., Henon, B., PhD, and Mansfeld, F., PhD, “Effectof Purge Gas Purity and Chelant Passivation on theCorrosion Resistance of Orbitally Welded 316L StainlessSteel Tubing,” Pharmaceutical Engineering, March/April1997.
3. Grant, A., “Passivation of Welds on Stainless Steel Tub-ing,” Presented at the ASME Bioprocess EngineeringSymposium, San Francisco, California, October 1995.
5. Stainless Steel Selection Guide - Central States Indus-trial, Central States Industrial Equipment and ServicesInc., Springfield, MO., 2001.
6. Suzuki, O., Newberg, D., and Inoue, M., “Discoloration andits Prevention by surface Treatment in High-Purity WaterSystems,” Pharmaceutical Technology, April 1998.
AcknowledgmentsAlloy tubing was provided by Theo Wolfe at Fluid LineTechnology.
DedicationTheo Wolfe at Fluid Line Technology initiated this study andthis article is dedicated to his memory.
About the AuthorsArnie Grant is Director of Research at Cal-Chem Corp., an international service organi-zation providing precision cleaning and pas-sivation to the pharmaceutical industry. Hereceived a BS in chemistry from FairleighDickinson University and pursued post-graduate courses at UCLA and CaliforniaState University. He has more than 35 years
of experience in analytical chemistry methods development,corrosion prevention and control, parts materials and processevaluation, and contamination control and prevention. Hewas past chairman of the ICRPG Infrared Committee and iscurrently active in ISPE and ASME presenting papers andconducting seminars in passivation and corrosion. He can becontacted by tel: 1/800-444-6784 or by email: [email protected]
John Jermain is a research chemist at Cal-Chem Corp., an international service organi-zation providing precision cleaning and passi-vation to the pharmaceutical industry. Hereceived a BS in chemistry from the CaliforniaPolytechnic University at Pomona and planson pursuing post-graduate courses in advancedphysical chemistry and forensic science.
Cal-Chem Corp., 2102 Merced Ave., So. El Monte, CA91733.
Increasing Alkaloid Production fromCatharanthus roseus Suspensionsthrough Methyl Jasmonate Elicitationby Jennifer L. Gaines
This articledescribes theeffects ofmethyljasmonateelicitation on thealkaloidproduction ofCatharanthusroseus cellsuspensions.
This article won theundergraduate levelaward at the ISPEBoston AreaChapter's StudentPoster Competitionin the spring of2003, and went onto compete in theInternationalStudent PosterCompetition at theISPE AnnualMeeting inNovember 2003.
Figure 1. Establishing aplant cell culture.1
Illustration of the stepsinvolved in establishing acell suspension culture.
An exceedingly large population relieson pharmaceuticals derived fromplants. The Catharanthus roseus plantproduces two anti-cancer compounds,
vincristine and vinblastine, as well as the anti-hypertensive and sedative compounds,ajmalicine and serpentine. Through plant cellculture, these pharmaceuticals can be mademore available to those in need. Plant cellculture offers controlled conditions for rapidcell reproduction without depleting natural
resources. Methyl jasmonate elicitation wasutilized to increase the production of the alka-loids in the cell suspension cultures. Two foldand five fold increases in serpentine andajmalicine production were obtained whenmethyl jasmonate was introduced to cell sus-pensions on day 6 of growth at a concentrationof 10µM and 100µM. The implementation ofagar immobilization did not increase alkaloidproduction in conjunction with methyljasmonate elicitation.
Figure 2. Biosynthetic Pathway of Catharanthus roseus Alkaloids.2
Outline of the series of reactions that occur within the C.roseuscell.
IntroductionIt is estimated that 75% of the world’s population is depen-dant upon plant-derived pharmaceuticals. Some very com-mon plant-derived pharmaceuticals include quinine which isan anti-malarial from the Cinchona ledgeriana, codeine fromthe Papaver somniferum, and paclitaxel, an anti-cancer treat-ment from the Taxus brevifolia. Affordability and availabilityof pharmaceuticals are the most prevalent issues facing theworld today. The need for methods of increasing the produc-tion of plant-derived pharmaceuticals cost-effectively andwith environmental consideration is becoming more impor-tant. Through plant cell culture, pharmaceuticals can beproduced on a large-scale in turn increasing the accessibilityof the product.
Of particular interest are the pharmaceutically valuablealkaloids from the Catharanthus roseus (Madagascar peri-winkle). These include ajmalicine (anti-hypertensive), ser-pentine (sedative), vincristine and vinblastine (anti-cancer).The compounds range in price from $30,950 per kg ($14,068per lb) to $36 million per kg ($16.4 million per lb). An increasein production of these pharmaceuticals through plant cellculture will result in greater accessibility and affordability ofthe product.
BackgroundFigure 1 illustrates the steps involved in developing a plantcell culture suspension. Establishing a cell culture suspen-sion begins with a sterile seed or cutting from the plant. Thesterile seed or cutting is placed on a solid media known asagar, usually in a Petri-dish. The agar contains vitamins,sugars, salts, and hormones necessary for growth. The pieceof plant (either from the cutting or plant growth from theseed) is allowed to incubate on the agar until a callus hasformed. The callus is an undifferentiated and aggregated
mass of cells similar to a stem cell in the human body in thatit is not assigned to a specific role as a plant cell. The callusis transferred into liquid medium that contains the samevitamins, sugars, salts, and hormones. Cells are sloughed offof the aggregate and a more homogeneous cell suspension isformed. The cells must be continually transferred into freshmedia in order to keep the plant cells in a live and reproduc-tive state.
There are numerous benefits to utilizing plant cell culture.These benefits include the ability to control the growthconditions of the culture, such as nutritional and supplemen-tal components in the media, light, temperature, pH, andoxygen (or dissolved gasses). Preservation of natural re-sources also can be experienced because plant cell cultureallows for the rapid reproduction of plant cells without theneed for depletion of natural crops or disturbance of sur-rounding wildlife for product extraction. A particularly im-portant benefit is the ability to manipulate and improve theproduction of desired compounds within the plant cell throughexperimentation with cell culture.
In order to manipulate and improve the production of adesired compound, there must be knowledge of where or howthe compound is produced within the naturally occurringplant. Figure 2 illustrates the series of complex reactions thatoccur within the Catharanthus roseus, known as its biosyn-thetic pathway. Ajmalicine, serpentine, vincristine and vin-blastine are known as secondary metabolites and are foundtoward the bottom of the illustrated series of reactions.Secondary metabolites are compounds within the plant thatparticipate in reactions within the metabolic pathway, but donot contribute to the growth of the plant. With knowledge ofthese reactions, experimentation is conducted to manipulatethe production of the pharmaceutical compounds.
Increasing the production of the pharmaceutically valu-able compounds was attempted through methyl jasmonateelicitation. Methyl jasmonate is a compound that has beenknown to stimulate reactions within the metabolic pathwayof the Catharanthus roseus. Given the precursory compoundsnecessary for reaction are present, methyl jasmonate willhelp that the reaction to completion. The following experi-ments were performed with the intent of increasing thealkaloid production of the Catharanthus roseus by the treat-ment of cell suspensions with methyl jasmonate.
Timing of Methyl Jasmonate AdditionThe first of two experiments involved the timing and dosageof methyl jasmonate addition. The purpose of this experimentwas to determine the optimal time and concentration ofmethyl jasmonate to introduce to the cell suspension culture.Effects of methyl jasmonate elicitation may be differentdepending on the concentration of methyl jasmonate and thestate of the cell upon addition. The Catharanthus roseus has
“Through plant cell culture, pharmaceuticals can be producedon a large-scale in turn increasing the accessibility of the product.”
Figure 4. Effect of MeJa Timing and Dosage on Serpentine Concentration. Shows the cumulative serpentine production over the span ofthe experiment for methyl jasmonate addition on day 0, 3, 6, and 9 (day 12 and 15 not shown).
a 15 day growth cycle. From the day that cells are introducedinto fresh media (subculture, day zero) to day three of growth,the cells are in what is known as a lag phase. During the lagphase, cell growth is slow because the cells are acclimating to
the fresh media and begining the uptake of nutrients from themedia. From day three to day nine is an exponential phase ofgrowth where the cells have acclimated to their environmentand are rapidly growing and reproducing. Day nine to day 15
Figure 3. Effect of MeJa Timing and Dosage on Ajmalicine Concentration. Shows the cumulative ajmalicine production over the span of theexperiment for methyl jasmonate addition on day 0, 3, 6, and 9 (day 12 and 15 not shown).
Table A. Effect of Agar Immobilization on Alkaloid Production.Shows the cumulative ajmalicine and serpentine production overthe span of the experiment for cell suspensions, heated cellsuspensions, and agar immobilized suspensions.
Figure 5. Highest Ajmalicine and Serpentine Productions. Focuseson the optimal time and dosage of methyl jasmonate addition.
is a deceleration phase where the cells have used most of theavailable nutrients and are producing some toxic by-prod-ucts. Growth is slowed and transfer into fresh media isnecessary for the survival of the cells. Addition of methyljasmonate at different concentrations and during the differ-ent periods of growth resulted in a relatively comprehensiveoptimization study.
MethodCatharanthus roseus cell line A11 was obtained from thelaboratory of Dr. Carolyn Lee-Parsons at Northeastern Uni-versity. Methyl jasmonate addition was studied at concentra-tions of 0, 10, 100 and 100µM within the cell suspension. Eachconcentration of methyl jasmonate was added on either day0, 3, 6, 9, 12, or 15 of growth. 1g (0.0022lb) of XAD-7 resins(Sigma) were introduced into the cell suspension three daysafter the methyl jasmonate addition and after which ex-changed for fresh resins every three days through the termi-nation of the experiment. The alkaloids produced from thecell suspension were adsorbed onto the resin. The alkaloidswere then removed from the resin with a series of methanol
washes and analyzed for content by high pressure liquidchromatography.
ResultsResults are shown in Figures 3 and 4. The charts indicatecumulative production of the alkaloids (ajmalicine or serpen-tine) over the span of the experiment (21 days). For clarifica-tion, the purple triangles indicate the day of methyl jasmonateaddition and the green circles indicate the day of resinaddition. In terms of both ajmalicine and serpentine, anincrease in production is observed when methyl jasmonate isintroduced on day three as opposed to day zero. A greaterincrease in production is observed with methyl jasmonateaddition on day six as opposed to day three. A decrease inproduction is observed when methyl jasmonate is introducedon day nine. A further decrease in production is observedwhen methyl jasmonate is added on day 12 and also day 15(both not shown). Addition of methyl jasmonate on day six ofcell growth appears to lead to the greatest production of thealkaloids. As seen in Figures 3 and 4, concentrations of 10µMand 100µM of methyl jasmonate elicited the most optimalproduction. Figure 5 focuses on the day six addition of methyljasmonate. There were replicates of each methyl jasmonateaddition on each day in order to determine standard devia-tion and error within the experiment. Figure 5 includes errorbars to illustrate this. The error bars overlap and indicatethat the effects from adding 10µM and 100µM on day six maybe equal for both ajmalicine and serpentine.
DiscussionIt is possible that the greatest alkaloid production wasachieved with methyl jasmonate addition on day six of growthbecause at that point the cells had become accustomed totheir environment, as well as in a healthy state of growth withplenty of nutrients and negligible by-product accumulation.The effects of 10µM or 100µM of methyl jasmonate additionshow a five fold increase in terms of ajmalicine productionand a two fold increase in terms of serpentine production overa 0µM concentration. Although the methyl jasmonate elicita-tion was successful in increasing the production of the alka-loids, multiple tactics may be utilized to further increaseproductivity.
Agar Cell ImmobilizationThe second of two experiments studies the use of agar immo-bilization in conjunction with methyl jasmonate elicitation.Cell immobilization occurs when cells are trapped or encasedin a material such as mesh, gel, or a polymer. The growth isrestricted while at the same time allowing mass transfer ofmedia and dissolved gasses. Agar immobilization involvesthe same agar that was used as a solid growth media as theimmobilizing agent. Benefits to studying cell immobilizationinclude a decrease in shear stress that the cell may experi-ence in suspension because of a protective coating given bythe immobilization material. In some instances, cells havebeen known to excrete the desired product into the media.
This allows for cell reuse because the cell does not need to bedamaged in order to obtain the desired product that is simplywithin the media. Immobilization also may increase produc-tion in certain cell types. The purpose of the second experi-ment was to determine how restricting growth through agarimmobilization in conjunction with methyl jasmonate elicita-tion affects alkaloid production.
MethodThe A11 cell line was used to study agar immobilization. Inorder to determine the effects of restricted growth withmethyl jasmonate elicitation, agar immobilized suspensionswere compared to cell suspensions and heated cell suspen-sions. A heated cell suspension was included because the agaris heated upon mixing with the cells. Heating effects werestudied in order to ensure that the resultant alkaloid produc-tion from the agar immobilization was due to restrictedgrowth and not heat. In order to immobilize a cell suspensionin agar, the plant cells are mixed with warm agar at 50°C andpoured in a flask to cool. The agar which contains thenecessary nutrients and hormones quickly hardens aroundthe cells rendering them immobilized. The immobilized cellssimply resemble a thin film of agar with cells trapped inside.The thin film is then placed in fresh liquid media where itincubates for the duration of the experiment. Methyl jasmonatewas added to all suspensions on day zero at a concentrationof 100µM. 1g (0.0022lb) of XAD-7 Resins were added threedays after the methyl jasmonate addition and exchanged forfresh resins every three days until the termination of theexperiment. The alkaloids were adsorbed onto the resins andthen recovered with a series of methanol washings similar tothe prior experiment. The alkaloid content was analyzed byHPLC.
ResultsResults are shown in Table A. The table indicates cumulativeproduction of the alkaloid (ajmalicine or serpentine) over thespan of the experiment. The experiment was performed inreplicate in order to determine standard deviation and error.From Table A, it is clear that there is not a great differencebetween the alkaloid productions of the cell suspension ascompared to the heated cell suspension. The alkaloid produc-tion resultant from agar immobilization is notably lower thanboth the cell suspension and the heated cell suspension.Because the heated cell suspension was included in theexperiment, it is determined that the decrease in alkaloidproduction can be attributed to restricted growth and notheat effects.
DiscussionNumerous reasons could be given as to why agar immobiliza-tion caused a decrease of alkaloid production. It is possiblethat the restricted growth was too much of a stress to the cellitself or in conjunction with the methyl jasmonate elicitation.It also is possible that agar may not be the optimal immobi-lizing material for this particular cell.
ConclusionMethyl jasmonate elicitation has shown a definite effect onalkaloid production in the Catharanthus roseus. As the firstof two experiments shows, optimal timing and dosage arenecessary in order to obtain the largest alkaloid production.The optimal time and dosage of methyl jasmonate addition tothe Catharanthus roseus cell suspension is day six of growthat either a concentration of 10µM or 100µM. The maximumajmalicine production was 10.4 ± 0.6 mg/L (8.67x10-5 ± 5x10-
6 lb/gal) and the maximum serpentine production was 1.15 ±0.06 mg/L (9.58x10-6 ± 5x10-7 lb/gal). These concentrationswere obtained with methyl jasmonate elicitation during aperiod of rapid and steady cell growth.
Conversely, while growth is restricted, the cell suspensiondecreased alkaloid production. Methyl jasmonate elicitationjoined with agar immobilization did not successfully increasealkaloid production. Catharanthus roseus cell suspensionsand heated cell suspensions obtained ajmalicine productionsof 4.1 ± 0.65 mg/L (3.42x10-5 ± 5.42x10-6 lb/gal) and 4.4 ± 0.97mg/L (3.67x10-5 ± 8.08x10-6 lb/gal) respectively and serpen-tine productions of 0.72 ± 0.12 mg/L (6x10-6 ± 1x10-6 lb/gal)and 0.77 ± 0.18 mg/L (6.42x10-6 ± 1.5x10-6 lb/gal) respectively.Lower concentrations were obtained with the agar immobi-lized suspensions as ajmalicine production was 1.5 ± 0.65 mg/L (1.25x10-5 ± 5.42x10-6 lb/gal) and serpentine production was0.32 ± 0.16 mg/L (2.67x10-6 ± 1.3x10-6 lb/gal).
RecommendationsFurther experimentation is recommended in order to narrowthe findings of the two described above. Timing and dosage ofmethyl jasmonate elicitation can be further optimized bystudying a range of methyl jasmonate concentrations be-tween 10µM and 100µM with day six addition. If desired, itmay be possible to pinpoint the optimal time of methyljasmonate addition to the hour within the sixth day of growth.
Further exploration into cell immobilization could bebeneficial with the study of a different material as theimmobilizing agent. Cell immobilization could simply be anundesirable tactic to increase alkaloid production, or agarcould be an undesirable immobilizing material. It also ispossible that methyl jasmonate and agar immobilizationtogether do not create an optimal environment for the cellsuspension. Additional study into these issues could result inknowledge as to how alkaloid production in the Catharanthusroseus can be further increased.
References1. Kargi, F., and Shuler, M.L., Bioprocess Engineering,
2. Memelink, J., and van der Fits, L., “ORCA3, a Jasmonate-Responsive Transscriptional Regulator of Plant Primaryand Secondary Metabolism,” SCIENCE, Vol. 289, 2000,pp. 295-297.
AcknowledgmentsExperimental work was performed in the laboratory of Dr.Carolyn Lee-Parsons at Northeastern University. Furtheracknowledgments include Kevin Cash, Seda Ertuk, and AmberRoyce for their gracious support as colleagues. The authorwould like to thank ISPE and AIChE for the opportunity topresent this research to its members and for their support ofundergraduate research. Northeastern University under-graduate research fund, NSF, and Louis Stokes Alliance forMinority Participation (LSAMP) all offered their support andhelped make these studies possible.
About the AuthorJennifer Gaines graduated in May 2004with a BS in chemical engineering from North-eastern University. She will continue at-tending Northeastern University to obtain amaster’s degree in chemical engineering.Gaines has had the opportunity to presenther research at a national level to both ISPEand AIChE in the past year. She can be
