Qualification andValidation for Supercritical FluidChromatography

Qualification and Validation for Supercritical Fluid C

hromatography

© Agilent Technologies Inc., 2011Printed in Germany, November 1, 2011Publication Number 5990-9148EN

www.agilent.com/chem/

A Primer

The Mea sure of Confidence

Qualification andValidation forSupercritical FluidChromatography

Dr. Ludwig Huber

I

Preface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . III

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1

1.1 Scope and Content of the Primer . . . . . . . . . . . . . . . . . . . . . . . . . 2

1.2 SFC Overview – History, Current Status and Future . . . . . . . . . . 3

1.3 Importance of Compliance along the Drug Life Cycle . . . . . . . . . 4

1.4 Changing Focus of Analytical Laboratories along the Drug Life . . . 6

1.5 Resources . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7

2. Regulations and Quality Standards . . . . . . . . . . . . . . . . . . . . . . . . 9

2.1 Good Laboratory Practice Regulations . . . . . . . . . . . . . . . . . . . . 11

2.2 Current Good Manufacturing Practice Regulations . . . . . . . . . . 12

2.3 FDA Guidance Documents . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13

2.4 International Conference for Harmonization. . . . . . . . . . . . . . . . 14

2.5 Pharmaceutical Inspection Convention Scheme (PIC/S) . . . . . . 15

2.6 Unites States Pharmacopeia (USP) . . . . . . . . . . . . . . . . . . . . . . 16

2.7 FDA’s 21 CFR Part 11 and the EU GMP Annex 11 – Regulations for Electronic Records and Signatures . . . . . . . . . . 17

3. Requirements for Laboratories. . . . . . . . . . . . . . . . . . . . . . . . . . . 19

3.1 Compliance Overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21

3.2 Quality Assurance and Compliance across the Pharmaceutical Laboratory . . . . . . . . . . . . . . . . . . . . . . . . . . 22

3.3 Compliance across all Workflow Steps . . . . . . . . . . . . . . . . . . . 23

3.4 Compliance for individual Workflow Steps. . . . . . . . . . . . . . . . . 24

4. Qualification of SFC Hardware and Validation of Systems. . . 25

4.1 Analytical Instrument Qualification According to USP <1058> . . 26

4.2 Qualification Planning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28

4.3 Design Qualification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 31

4.31 The Importance of Requirement Specifications. . . . . . . . . 32

4.32 Vendor Assessment. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35

Contents

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4.4 Installation Qualification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 37

4.5 Operational Qualification. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39

4.6 Tests for Operational Qualification . . . . . . . . . . . . . . . . . . . . . . . 40

4.7 Performance Qualification. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 42

4.8 Specific Considerations for Software and Computer Systems. . 44

4.9 (Preventive) Maintenance and Repair . . . . . . . . . . . . . . . . . . . . 45

4.10 Change control. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 47

4.11 Validation Reports . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 48

5. Validation of Analytical Procedures . . . . . . . . . . . . . . . . . . . . . . 49

5.1 ICH Validation Parameters for Target Applications. . . . . . . . . . . 50

5.2 Strategies for Integrated Method Development and Validation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 53

5.3 Verification of Compendial Procedures. . . . . . . . . . . . . . . . . . . . 58

5.4 Transfer of Analytical Procedures . . . . . . . . . . . . . . . . . . . . . . . . 59

6. Managing Electronic Records for FDA Part 11 and EU Annex 11 Compliance . . . . . . . . . . . . . . . . 61

6.1 Risk Assessment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 63

6.2 System Validation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 63

6.3 Data Accuracy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 63

6.4 Limited Access to authorized Users and Authority Checks . . . . 64

6.5 Raw Data and Copies of Records. . . . . . . . . . . . . . . . . . . . . . . . 65

6.6 Protection of Records . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 65

6.7 Computer Generated Time-Stamped Audit Trails . . . . . . . . . . . . 66

6.8 Electronic Signatures. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 67

6.9 Periodic Evaluation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 68

References. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 69

Glossary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73

III

Qualification of analytical hardware and validation of analytical methodsand systems is required by many national and international regulations,quality standards and company policies. If executed correctly, it also helpsto improve instrument uptime and to avoid out-of-specification situations(OOS) in laboratories. This primer “Qualification and Validation forSupercritical Fluid Chromatography” guides analysts, laboratory managers, quality assurance managers and validation professionalsthrough the entire process from SFC instrument qualification to methodand system validation.

The concept, examples, templates and recommended procedures are based on my more than 20 years’ multinational experience and incorporates information from validation and qualification practicesapplied at Agilent Technologies and Labcompliance. Readers of this primer will learn how to speed up their validation and qualificationprocess, thereby avoiding troublesome reworking and gaining confidencefor audits and inspections.

Typically regulations and quality standards are around for a long timewithout significant changes. Guidelines developed by regulatory andindustry task forces are published more frequently. Interpretations, inspection and enforcement practices experience most frequent changes. What is state-of-the art today may not be appropriate tomorrow. Therefore, a timely update of all information is important and only possible using on-line information tools, such as the Internet. To take this fact into account, I recommend the websites listed on the following page which offer regular updates related to compliance in laboratories:

Preface

IV

www.fda.govRegulations and guidelines for the biopharmaceutical industry

www.ema.europa.euEuropean Medicines Agency website

www.ich.org International Conference for Harmonisation of Technical Requirements forRegistration of Pharmaceuticals for Human Use (ICH)

www.picscheme.org Pharmaceutical Inspection Co-operation Scheme website

www.usp.orgUnited States Pharmacopeia website

www.who.orgWorld Health Organization website

www.labcompliance.com A website with tutorials, many references and regular updates related toall quality and compliance issues in laboratories

Dr. Ludwig HuberChief Advisor for global FDA complianceLabcomplianceludwig_huber@labcompliance.com

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Chapter 1

Introduction

Introduction The application of Supercritical Fluid Chromatography (SFC) in the pharmaceutical industry is expanding from research to development andmanufacturing and from primarily chiral separations to other applications. The reason is that new instruments have been developed that provide theperformance, reliability and robustness as required by various regulationsand guidelines such as Good Laboratory Practices, Good Clinical Practicesand Good Manufacturing Practices. Beyond performance these regula-tions have other requirements for analytical instruments and methods, for example, instruments should be qualified and methods should be validated. This primer will give an overview about the requirements of different regulations and guidelines as they apply to SFC.

After a short introduction about history and the current outlook on SFC,the primer will explain applicable regulations during a drug lifecycle fromresearch and development to manufacturing. The second chapter willprovide more detailed information about regulatory documents alongwith requirements for laboratories. Chapter 3 will guide readers throughcompliance requirements for sample and data flow, from sampling toarchiving of test reports and other documents. The main focus of the primerwill be on qualification of SFC hardware, validation of SFC methods andvalidation of complete systems as outlined in chapters 4 and 5. Chapter 6will provide information on what is necessary to manage electronic recordsfor compliance with various FDA and international regulations.

The primer will not only help readers to understand the instrument quali-fication and method validation processes, but also offers templates andexamples to easily implement various tasks. Because of the nature andsize of this primer, all the details of instrument qualification and methodand system validation cannot be given. For more details, please refer toreference text books and primers (1-5). Exact procedures and test para-meters very much depend on the type of instrument and applications.Details on recommendations and services can be obtained from instru-ment vendors. Although the primer has recommendations for validationof standard commercial computerized SFC systems without the need formajor customization, it does not give details on validation activities duringsoftware development but refers to further literature (6-7).

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1.1 Scope and Content ofthe Primer

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1Initially the application of Supercritical Fluid Chromatography (SFC) was limited to quantitative and qualitative analysis of chiral moleculesprimarily in pharmaceutical research and early development. Even thoughthe technique allows analyzing other compounds, these applications have been excluded. The reason is that compared to other chromato-graphic techniques, such as high performance liquid chromatography andgas chromatography, the performance was limited. For example, HPLC can be used to easily meet ICH requirements for quantitation of drugimpurities down to 0.05% related to the drug substance; this was notpossible with SFC because of higher background noise of the SFC UVdetectors. In addition, SFC instruments have not been robust enough that they could be used for reliable routine analysis in late stages of drugdevelopment and for quality control of drug substances and drug products.

Recently new generation SFC instruments have been introduced by majorinstrument suppliers, for example, by Agilent Technologies. These instru-ments offer superior performance and functionality. For example, Dunkleand co-workers could demonstrate that low level impurities down to0.05% can be quantitated with sufficient precision (8). In addition therobustness and automation capabilities have been improved. Also thecompatibility with detectors has been expanded from traditionally vari-able UV/Vis and flame ionization detectors to on-line mass spectrometers.

New generation SFCs offer performance, automation capabilities anddetector versatility comparable to HPLC and GC. They also offer all of theadvantages that are identified with the SFC technology. Benefits of SFCvs. the workhorse analytical tool in pharmaceutical analysis HPLC are:

Less toxic waste is generated. With increasing environmental aware-ness to minimize toxic waste production, SFC is increasingly acceptedas the “green alternative”.

Safer operation because mobiles phases, such as CO2, are not combustible.

3-5 times faster analysis because of lower viscosity of SFC mobilephases. Under SFC operating conditions, the mobile phases exhibitgaseous as well as liquid-like properties. The major advantages of thisalternative in relation to HPLC are improved diffusion characteristics,mass transfer and low viscosity, which result in high separation efficiency and fast separation capability.

1.2 SFC Overview –History, CurrentStatus and Future

1Higher sample throughput through faster column equilibrium.

Better separation through higher number of theoretical plates of SFC columns.

For many applications better selectivity, e.g., through variation of pressure as an additional parameter.

Lower operation cost due to lower cost of mobile phases.

Easier scale-up to semi preparative and preparative applications.

Even though the long-term application range of SFC will be smaller compared to HPLC, especially for biomolecules, in many cases analystsnow have the luxury to pick and choose from more chromatographic techniques. Applications of SFC are no longer limited to samples thatcannot be analyzed by other techniques. Due to improved performancecharacteristics, such as precision, linear dynamic range, robustness and reliability traditional applications such as chiral analysis will move to drugdevelopment and quality control.

SFC’s move out of drug discovery to drug development, clinical trials andquality control in manufacturing, means compliance is becoming increas-ingly important.

Basic research activities, such as disease discovery and drug discovery,are not regulated. This changes as soon as a compound has been identified as a target drug substance.

The drug discovery, development and marketing authorization process isa long process that typically takes more than 10 years. The process canbe divided into phases that are shown in figure 1. The process beginswith basic research and discovery activities, the results of which are usedto define efficacy targets for the potential drug.

Once a target compound has been identified as a drug candidate it goesthrough preclinical studies, which are controlled by good laboratory

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1.3 Importance of Complianceaccompanying the Drug LifeCycle

5

practice regulations. Clinical trials are governed by good clinical practiceregulations and the manufacturing process by current good manufacturingpractice regulations. Quality Control laboratories are also regulated byGMP as well as the manufacturing process of drug substance (APIs). At the end of the preclinical studies the industry submits an InvestigationalNew Drug (IND) Application and at the end of the clinical trials a NewDrug Application (NDA) or New Biological License Application (BLA). The applications are reviewed by the FDA to determine if the drug canmove to the next phase.

Once the drug has been registered and is available on the market, healthagencies regularly control compliance with GMP regulations by testingproducts on the market and inspecting manufacturing establishments. Incase of non-compliance, agencies take enforcement actions, for example,sending the company a warning letter and stopping shipment of productsfor companies in the US or an import alert for non-US based companies.

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Clinical Trials Phase I, II, III

DrugDiscovery

BasicResearch

Manufacturingincl. APIsQC Laboratories

GLPNot Regulated GCP

GLP = Good Laboratory PracticesGCP = Good Clinical PracticesGMP = Good Manufacturing Practices

Submission &Review

IND BLA/NDAPostMarketingSurveillance

Lead toDrug Target

GMP

IND = Investigational New Drug ApplicationBLA = Biologic License ApplicationNDA = New Drug Application

PreclinicalDevelopment

21 CFR 11 Electronic Records & Signatures

Safety, Quality, Efficacy

Submission &Review

Figure 1Regulations for drug development and manufacturing.

Throughout development, the principles of GxPs three pillars – Safety,Quality and Efficacy – should be applied in an incremental approach:Safety to assure the maximum achievable protection against adverseevents in relation to the benefit obtained by the drug, quality to assurehigh technical product excellence and efficacy to demonstrate the product effectiveness.

The focus and objectives of analytical laboratories change between drugdiscovery, preclinical studies, clinical studies and manufacturing. Drugdiscovery is characterized by requirements for high throughput, flexibility,qualitative assessment and compound identification. Linearity and precisionof analytical methods is of secondary importance. Demonstration of dataintegrity is important to protect potential patents, but not from an FDAcompliance viewpoint.

During the development phases from preclinical to clinical phases I to III,accurate and precise results become more important to ensure reliableresults.

Manufacturing control laboratories have the highest requirements for system robustness and reliability. Using the risk-based approach, they fallinto the high risk category. Results are used to release the product intothe market. Wrong test results can mean that products with incorrectspecifications are shipped, for example with too high impurities. Apartfrom compliance, also the business impact can be quite high. For example,out-of-specification results caused by single laboratory tests can cost a company dollars in the five to six digit range. Inadequate follow-up can cause problems with regulatory agencies with consequences such as shipment stop and product recall.

For SFC, like other analysis techniques, working in regulated environmentsmeans:

Instruments should be qualified and well maintained.

Methods should be validated.

Systems should be tested for suitability.

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1

1.4 Compliance for Individual Workflow Steps

7

Analytical results should be traceable to raw data.

The security and integrity of data should be ensured from the time ofdata acquisition or data entry into the SFC computerized system for aslong as data are retained.

The scope of this primer is to give an overview on validation and qualification in SFC. However, there are many resources available where readers can get more details, i.e., regulatory agencies, joint industry/agency task forces and independent authors. Regulatory and other official documents will be discussed in the next chapter.

There are many publications available from independent authors that are published as traditional journal papers, online articles and traditional text books. Readers are referred to well-known search engines to look-up online articles. This chapter gives an overview ofsome resources published by independent authors or organizations.

Taylor gave a comprehensive review of SFC with its history and current trends for analytical and preparative chiral and achiral separations. (9)

Wang and colleagues have published a paper that compares SFCapplications and analytical activities in drug discovery and regulateddrug development. (10)

Agilent Technologies has published several primers about Compliancefor Biopharmaceutical Laboratories (1), Analytical InstrumentQualification and System Validation (2) and Validation of AnalyticalMethods (3). They are useful to obtain a good understanding aboutcompliance and validation in laboratories.

Huber has authored a validation reference book for the analytical laboratory (5). It covers all validation aspects of an analytical laboratoryincluding equipment, analytical methods, reference compounds andqualification of personnel.

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1.5 Resources

The Good Automated Manufacturing Practices Forum (GAMP) hasdeveloped guidelines for computer validation. The most recent versionhas been released in 2008 (6). These guides have been specificallydeveloped for computer systems in general, and because of theirimportance have also been used for laboratory system validation.

Sandra and colleagues gave an overview with examples of SFC applications in pharmaceutical development. (11)

Dunkle and co-workers presented SFC as an alternative method to HPLC for the determination of achiral low level impurities in pharmaceutical products. (8)

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Chapter 2

Regulations andQuality Standards

Regulations andQuality Standards

10

2The pharmaceutical industry is one of the most regulated industries. Drugdevelopment and manufacturing are controlled by government agenciesthrough a set of laws, regulations and guidance documents in all industrialcountries and in an increasing number of developing countries. The fore-most underlying regulations are the so-called GxP regulations consisting of good laboratory practices (GLP), good clinical practices (GCP) and goodmanufacturing practices (GMP). In addition there are special regulationsfor product labeling, the use of computers in a regulated environment andfor marketing authorization.

The main purpose of regulations is to ensure quality, safety and efficacy of drugs. Marketing authorization of new drugs is determined by agenciesevaluating study data and deciding if the benefit of the drug is higher thanthe risk through insufficient drug safety. Regulations for the pharmaceuticalindustry in general follow modern quality system principles with high focuson data accuracy, reliability and integrity.

Qualification of instruments and validation of analytical methods and systems is a requirement of the FDA and equivalent international agencies.No or inadequate qualification can result in regulatory actions, such asshipment stops of drugs and APIs. The rationale behind this assumption is that analytical test results obtained with no or inadequately qualifiedinstruments and validated methods can be wrong. Because of the importance of compliance this chapter is dedicated to regulations and regulatory guidance.

Regulations are quite static and typically don’t change for several years.More dynamic than regulations are inspection and enforcement practices.Information can be found in the FDA’s inspection documents, such aswarning letters, establishment inspection reports (EIR) and 483 forminspectional observations. Most critical are the FDA’s warning letters. They are sent to companies in case of serious regulatory violations.Companies are expected to respond within 15 business days. If there is no response or if the response is inadequate, the FDA will take furtheractions which may cause delay of new product approvals, import alertsand denials, or product recalls. Since March 2003 warning letters arereviewed by higher-level FDA officials and reflect FDA’s current thinking.

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Warning letters are published on the FDA website:www.fda.gov/ICECI/EnforcementActions/WarningLetters The only problem is that there are thousands of them and they mostly relate to marketing and labeling drugs so it is difficult to find the ones that are of interest for laboratories. Interesting are sites that only publish warning letters related to GxP issues. For example,www.fdawarningletter.com has many quotes related to qualificationof instruments, validation of methods and validation of analytical systems.

Good Laboratory Practice (GLP) regulations deal with the organization,processes and conditions under which preclinical laboratory studies areplanned, performed, monitored, recorded and reported. GLP data areintended to promote the quality and validity of study data. GLP regulationswere first proposed by the U.S. FDA in November 1976, and final regulations were coded as Part 58 of Chapter 21 of the Code of FederalRegulations in 1979 (13). The Organization for Economic Cooperationand Development (OECD) published the principles of Good LaboratoryPractice in the Testing of Chemicals in 1982 (14), which has been sinceupdated (15) and incorporated by OECD member countries. In the meantime most industrial countries and some developing countries have their own GLPs.

All GLP regulations include chapters on equipment design, calibration and maintenance, for example, U.S. GLP regulations, Sections 58.61 and 58.63 (13).

Automatic, mechanical, or electronic equipment used in the generation,measurement, or assessment of data shall be of appropriate designand adequate capacity to function according to the protocol and shall be suitably located for operation, inspection, cleaning, and maintenance.

Equipment used for generation, measurement, or assessment of datashall be adequately tested, calibrated, and/or standardized.

Written standard operating procedures shall set forth in sufficient detailthe methods, materials, and schedules to be used in routine inspection,

2

2.1 Good Laboratory PracticeRegulations

2cleaning, maintenance, testing, calibration, and/or standardization ofequipment and shall specify remedial action to be taken in the event of failure or malfunction of equipment.

Written records shall be maintained of all inspection operations.

The GLP principles of the OECD include similar but shorter sections onequipment (15):

The apparatus used for the generation of data and for controlling environmental factors relevant to the study should be suitably locatedand of appropriate design and adequate capacity.

Apparatus and materials used in a study should be periodically inspected, cleaned, maintained, and calibrated according to Standard Operating Procedures.

Records of procedures should be maintained.

Good Manufacturing Practices (GMPs) regulate manufacturing and itsassociated quality control. GMP regulations have been developed toensure that medicinal (pharmaceutical) products are consistently producedand controlled according to the quality standards appropriate to theirintended use. In the United States, the regulations are called CurrentGood Manufacturing Practices (cGMP) to account for the fact that FDA regularly updates its view on cGMP implementation in guidance documents and the industry is advised to follow the current interpretation. CGMPs are defined in Title 21 of the U.S. Code of Federal Regulations,21 CFR 210-Current Good Manufacturing Practice for Drugs, Generaland 21 CFR 211-Current Good Manufacturing Practice for FinishedPharmaceuticals (16). Drugs marketed in the United States must firstreceive FDA approval and must be manufactured in accordance with the U.S. cGMP regulations. Because of the importance of the U.S. pharmaceutical market, FDA regulations have set an international regulation benchmark for pharmaceutical manufacturing.

In Europe, local GMP regulations exist in many countries. These arebased on the EU directive: Good Manufacturing Practice for Medicinal

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2.2 Current Good ManufacturingPractice Regulations

2

13

Products in the European Community (17). This EU GMP is necessary topermit free trade in medicinal products between the member countries.Regulations in the EU allow the marketing of a new drug in the membercountries with the acquisition of just a single marketing approval. The intention of the EU GMP is to establish a minimum manufacturing standard for all member countries.

Like GLPs all GMP regulations include chapters on equipment design, calibration and maintenance, for example, U.S. CGMP regulation forpharmaceutical drugs, Sections 211-140 b and 211-68 (16).

Laboratory controls shall include the calibration of instruments, apparatus,gauges, and recording devices at suitable intervals in accordance withan established written program containing specific directions, schedules,limits for accuracy and precision, and provisions for remedial action in the event accuracy and/or precision limits are not met. Instruments,apparatus, gauges, and recording devices not meeting establishedspecifications shall not be used.

Automatic, mechanical, or electronic equipment or other types ofequipment, including computers, or related systems that will perform a function satisfactorily, may be used in the manufacture, processing,packing, and holding of a drug product. If such equipment is so used, it shall be routinely calibrated, inspected, or checked according to awritten program designed to assure proper performance.

Written records of those calibration checks and inspections shall bemaintained.

Typically regulations are not detailed enough for FDA inspectors and reviewers. In this case the FDA develops inspection guides. They are available on the internet and also available for the industry(www.fda.gov, search for FDA Guidance). In addition the FDA hasdeveloped a large number of industry guides on selected topics. They provide assistance to the regulated industry by clarifying requirementsimposed by Congress or issued in regulations by the FDA and by explaininghow industry may comply with these statutory and regulatory requirements.

2.3 Current Good ManufacturingPractice Regulations

2They also provide specific review and enforcement approaches to helpensure that the FDA's investigators implement the agency's mandate inan effective, fair and consistent manner. Although laws and regulationsare mandatory for the industry, guidance documents are not. Industry candecide to use alternatives to comply with regulations.

Important FDA guidances related to the topic of this primer are:

Analytical Procedures and Methods Validation (draft) (18)

Bioanalytical Method Validation (19)

Part 11, Electronic Records and Signatures, Scope and Applications (29)

The International Conference on Harmonization (ICH) of TechnicalRequirements for Registration of Pharmaceuticals for Human Use bringstogether the regulatory authorities of Europe, Japan and the UnitedStates and experts from the pharmaceutical industry in the three regionsto discuss scientific and technical aspects of product registration.

The purpose is to make recommendations on ways to achieve greaterharmonization in the interpretation and application of technical guidelinesand requirements for product registration in order to reduce or obviatethe need to duplicate the testing carried out during the research anddevelopment of new medicines.

ICH publishes guidelines that are either signed into regulations of membercountries, for example, in Europe or recommended as guidelines bynational authorities, e.g., by the US FDA.

The most important guideline related to validation of analytical methodsis ICH Q2 (R1) (20). It defines terminology and methodology for validationof analytical methods and procedures. It is considered the global standardfor method validation in pharmaceutical laboratories. Chapter 5 of thisprimer mainly refers to ICH Q2 (R1).

The most important ICH document related to equipment qualification and validation of computerized SFC systems is the GMP Guide for Active Pharmaceutical Ingredients, Q7 (21). Contrary to other official

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2.4 International Conference for Harmonization

15

documents, Q7 has very specific requirements for equipment and computer systems in chapters 5.3 and 5.4:

Equipment calibrations should be performed using standards traceableto certified standards, if existing.

Records of these calibrations should be maintained.

The current calibration status of critical equipment should be knownand verifiable.

Instruments that do not meet calibration criteria should not be used.

Deviations from approved standards of calibration on critical instrumentsshould be investigated to determine if these could have had an impacton the quality of the intermediate(s) or API(s) manufactured using thisequipment since the last successful calibration.

GMP related computerized systems should be validated. The depth andscope of validation depends on the diversity, complexity and criticalityof the computerized application.

Appropriate installation qualification and operational qualificationshould demonstrate the suitability of computer hardware and softwareto perform assigned tasks.

Another important ICH guidance related to SFC validation and qualificationis ICH Q9: Quality Risk Management (33). More and more decisions in pharmaceutical development and manufacturing are based on riskassessment. For example, FDA (29) and EMA (31) recommend basing theextent of validation of computerized systems on justified and documentedrisk assessment. ICHQ9 is the global reference document for risk assessmentand management in the pharmaceutical industry.

PIC/S' mission is ”to lead the international development, implementationand maintenance of harmonized Good Manufacturing Practice (GMP)standards and quality systems of inspectorates in the field of medicinalproducts”.

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2.5 Pharmaceutical InspectionConvention Scheme (PIC/S)

2.6 Unites States Pharmacopeia(USP)

This is to be achieved by developing and promoting harmonized GMPstandards and guidance documents; training competent authorities, in particular inspectors; assessing (and reassessing) inspectorates; and facilitating the co-operation and networking for competent authorities and international organizations. As of June 2011 there are 39 participatingauthorities in PIC/S including all EU member countries and several countries have applied for PIC/S membership.

The most relevant PIC/S document related to this primer is the GoodPractice Guide: Using Computers in GxP Environments (22). The guidancedocument is intended to provide a logical explanation of the basicrequirements for the implementation, validation and operation of computerized systems. Recommendations are documented in chapters4.6 and 4.8:

Independent from user acceptance testing (OQ) versus the functionalspecification, the regulated user also has responsibility for the (PQ)performance qualification of the system.

The validation documentation should cover all the steps of the lifecyclewith appropriate methods for measurement and reporting, (e.g. assessmentreports and details of quality and test measures), as required.

Regulated users should be able to justify and defend their standards,protocols, acceptance criteria, procedures and records in the light oftheir own documented risk and complexity assessment.

USP develops methodology for specific application and general chapterson different analytical aspects for FDA regulated industry. According tosection 501 of the Federal Food Drug and Cosmetic act USP methodologiesconstitute legal standards. For marketing authorization, manufacturersmust meet USP standards for the drug substance, for excipients and forthe drug product, if available. USP has developed several general chaptersrelated to equipment qualification and method validation.

Chapter <621> on “Chromatography” defines the terms and proceduresused in chromatography and provides general information on different

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2.7 FDA’s 21 CFR Part 11 and the EU GMP Annex 11 –Regulations for ElectronicRecords and Signatures

chromatographic techniques. It also defines parameters for system suitability testing (23).

Chapter <1058> on “Analytical Instrument Qualification” (24) providesa framework for the qualification of analytical instruments. It covers thecomplete process from writing specifications through installation, initialand on-going testing and maintenance.

Chapter <1224> on “Transfer of Analytical Procedures” is available in draft form. It describes four different options and elements for controlled method transfer (25).

Chapter <1225> on “Validation of Compendial Methods” definesparameters and tests for validation of compendial methods. The recommendations are also useful for laboratories developing and validating their own methods (26).

Chapter <1226> on “Verification of Compendial Methods” (27) hasbeen written for laboratories implementing compendial and standardmethods. The recommendations are also useful for laboratories implementing validated methods from another laboratory.

The USP also develops and provides standards and certified referencematerial that can be used as quality control samples in routine analysisand for validating accuracy of analytical methods.

In 1997 the United States Food and Drug Administration (FDA) issued aregulation that provides criteria for acceptance by the FDA of electronicrecords, electronic signatures and handwritten signatures (28). The regulation, entitled Rule 21 CFR Part 11, states that electronic recordscan be equivalent to paper records and handwritten signatures. The ruleapplies to all industry segments regulated by the FDA that includes GoodLaboratory Practice (GLP), Good Clinical Practice (GCP) and current GoodManufacturing Practice (cGMP). Similar requirements for Europe havebeen published in 1993 and updated in 2011 in the Annex 11 to the EUGMP directives (31).

2

The primary requirements for both documents are:

Computer systems should be validated to ensure accuracy, reliabilityand consistent intended performance.

Systems should have built in user-independent computer generatedtime-stamped audit trails. The audit trail function should be validatedand activated.

System and data security, data integrity and confidentiality should beensured through limited system and data access for authorized users.

Records should be protected to enable their accurate and readyretrieval throughout the required records retention period.

When electronic signatures are used instead of handwritten signatures,they should have the same impact as handwritten signatures andshould be permanently linked to the electronic record.

Persons developing, maintaining or using electronic systems shouldhave the education, training and experience to perform their assignedtasks.

Changes to the system should follow documented change control procedures.

Chapter 7 of this primer will be dedicated to electronic records and signatures.

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Chapter 3

Requirements forLaboratories

Requirements forLaboratories

Reliable and accurate data measured in pharmaceutical laboratories areimportant to ensure that only safe and efficient drugs are authorized formarketing and released for product shipment. Therefore pharmaceuticaldevelopment and QC laboratories have to follow GxP regulations todemonstrate quality of data.

This chapter describes the GxP requirements for pharmaceutical laboratories.It does not differentiate between development and quality control laboratories.When reading though the chapter, scientists and professional analystsmay consider many of the requirements to be common sense and thereshould be no need to write them down. However, in a regulated world it is not enough to understand what should be done and it is even notenough to implement the requirements. Most important is documentingwhat has been implemented. Inspectors simply consider everything notdocumented as not being done.

The requirements listed in this chapter in principle apply to all phases ofdevelopment and manufacturing, although a gradual increase in require-ments is implemented for phases from preclinical studies to finished drugQC laboratories. For example, in clinical Phase I it may be sufficient tocreate a document that describes why an analytical method is suitablefor its intended use. On the other hand in Phase III the statements alwaysmust be supported by experiments. In GMP environments all requirementslisted in this chapter should be fulfilled, although this may not always benecessary for earlier phases.

Requirements for laboratories can be divided into two categories:

1. General quality system requirements apply to all regulated activities within a company, for example, control of documents, internal audits and qualification of personnel. They are typically called quality system requirements.

2. Laboratory specific requirements apply to specific situations in a laboratory, for example, validation of analytical methods, sampling,and review and approval of test reports.

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The overall impact of regulations on a pharmaceutical laboratory can bebest illustrated by looking at the whole sample/data workflow as shownin figure 2. The upper segment shows general quality assurance requirementsthat are applicable to regulated laboratories. The lower part of the figureshows a typical laboratory workflow of samples and test data, togetherwith key requirements. The middle section shows requirements that areapplicable to the entire sample or data workflow.

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Sampling Samplehandling

Testing Test reportsRecordmanagement

Sampling plan &samplingdocumentation

Sample identification &protection of sample integrity

Monitoring the quality of test results, handling OOS

Test conditions,test results, review & approvals

Ensure record integrity & security

Compliance across all workflow steps

• Validation of analyticalmethods & procedures

• Controlled environmental conditions

Compliance across the laboratory

Non-conflicting organizational structure, document control, complaint handling, corrective & preventive actions, supplier & subcontractor management, internal audits, qualification of personnel

Compliance across the sample and data workflow

• Equipment calibrationtesting & maintenance

Figure 2Requirements for Pharmaceutical Laboratories.

3.1 Compliance Overview

3Pharmaceutical laboratories are expected to follow quality assurancepractices that are commonly accepted in regulated industries.They include:

Documentation control GxPs require that regulated documents are controlled from creation andapproval to distribution, archiving and disposal. Typical documentationincludes: policies, quality plans, master plans, standard operating procedures, records, such as analytical test records and trainingrecords.

Organizational Structure and ResponsibilitiesOrganizational structures should be regulated so that departments withconflicting interests do not adversely influence quality and complianceof data. For example, finance and the QA department should operateindependently from laboratory activities. Tasks and responsibilitiesshould be defined for each job.

Qualification of PersonnelPersonnel should be qualified for the assigned task. Qualification can come from education, experience in the job and from training.The effectiveness of trainings should be verified and documented.

Facilities and EnvironmentsThe laboratory should have a procedure to ensure that its facilities andenvironmental conditions do not adversely affect or invalidate samplehandling, instrumentation, instrument calibration and qualification andanalytical testing.

Internal AuditsInternal audits are a key element of any quality system. Their objectiveis to evaluate activities and existing documentation, checking whetherthese meet predetermined internal and/or external standards and/orregulations or customer requirements.

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3.2 Quality Assurance andCompliance across thePharmaceutical Laboratory

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Some required activities are applicable for all workflow steps. They arelisted in the middle section of figure 2. They include:

Validation of analytical methods and proceduresGxPs require analytical methods and procedures to be validated todemonstrate suitability for their intended use. The ultimate objective of the method validation process is to provide evidence that themethod does what it is intended to do, accurately, reliably and reproducibly. Typical method characteristics to be validated are: precision of amounts, reproducibility, specificity, linearity, accuracy,robustness, limit of quantitation and limit of detection. Chapter 5 of this primer will go into more details.

Equipment calibration and qualificationAll equipment that have an impact on regulated activities should bequalified and/or calibrated. The objective of equipment calibration andqualification is to provide evidence that the equipment is suitable for its intended use. Equipment to be calibrated or qualified includes hard-ware, software such as Excel spreadsheets and complete computerizedsystems consisting of hardware and software.

Equipment maintenanceEquipment should be well maintained to ensure proper ongoing performance. Procedures should be in place for regular preventivemaintenance of hardware to detect and fix problems before they canhave a negative impact on analytical data. Chapter 4 of this primer will be dedicated to equipment.

Controlled environmental conditionsEnvironmental conditions such as temperature and humidity should becontrolled and monitored to ensure that they do not adversely affectthe performance of equipment and material. Environmental conditionspecifications are typically provided by suppliers of equipment andmaterial.

3.3 Compliance across allWorkflow Steps

3All workflow steps as shown in the lower section of figure 2 have specificrequirements. They include:

Sampling Sampling of substances, materials or products for subsequenttesting should follow a well-documented procedure. Inspectors want tosee a sampling plan with a description of the sampling system, how sampling is performed and by whom. Sampling data should be recorded,specifically sampling procedure used, location, the identification of the person who took the sample, equipment used for sampling and environmental conditions, if relevant.

Handling of test items Laboratories should ensure proper identificationand protection of samples from the time the sample is taken until its disposal. Receipt, protection, storage, processing, retention and disposalshould be described in a procedure. The procedure should include provisions for protection against deterioration, loss or damage duringtransportation, handling and storage.

Testing Procedures for testing should ensure that only validated methodsare used, that the equipment is qualified and that sufficient system suitability test runs are conducted. Specifications and acceptance criteria should be defined for the sample to be tested. Procedures andparameters for testing should be documented.

Handling out-of-specification test results GMPs require that aninvestigation is conducted whenever a test result is observed that fallsoutside the previously specified acceptance criteria. This includes labora-tory testing during the manufacture of APIs, raw material and testing offinished products to the extent that cGMP regulations apply.

Data validation and reporting of results Test results should be signedby the analyst, reviewed and approved by a second person. A reviewercan be, for example, the analyst’s supervisor or a member of QA staff.

Record management All records associated with testing should bearchived. These records include certificates of analysis (CoA), instrumentand method parameters, supporting information such as chromatogramsand spectra, and equipment qualification records. The archiving period isdefined by individual regulations and can range from 6 to over 15 years.Controls should be in place to ensure security, integrity and availability ofthe records during the entire archiving period.

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3.4 Compliance for IndividualWorkflow Steps

Chapter 4

Qualification of SFC Hardware andValidation of Systems

4.1 Analytical InstrumentQualification According toUSP <1058>

Qualification of SFC Hardware andValidation of Systems

GxPs require that each analytical instrument used in the generation, measurement, and evaluation of analytical data is suitable for its intendeduse. This means, instruments should be well designed and qualified toensure compliance with pre-determined specifications.

Equipment qualification and validation of computerized systems cover the entire life of a product. It starts when someone has a need for a specific product and ends when the equipment is retired. Computer systemvalidation ends when all records on the computer system have beenmigrated and validated for accuracy and completeness to a new system.Because of the length of time and complexity, the process has beenbroken down into shorter phases, so called lifecycle phases. Several lifecycle models have been described for qualification and validation.

This primer will utilize USP terminology and the 4Q life cycle model steps as recommended by chapter <1058> “Analytical InstrumentQualification” (23). Since its release in 2008, this chapter became the global standard for analytical instrument qualification for the pharmaceutical industry. It is the preferred qualification procedure for commercial SFC instruments and with some modifications it also can be used for validation of configurable computerized SFC systems.

This chapter will give an overview on the USP process for qualification,validation and maintenance of SFC hardware and systems. More detailed information on the process is available in the Agilent Technologies: Analytical Instrument Qualification and System Validation primer (5).

USP <1058> adopts the 4Q lifecycle using these four phases: designqualification (DQ), installation qualification (IQ), operational qualification(OQ), and performance qualification (PQ). The process is illustrated in figure 3. In the DQ phase user requirements are compared with the vendor’s specifications. In addition, users conduct an assessment of thevendor. In the IQ phase the user’s environment is checked if it meets the vendor’s environmental specifications. The instrument is installed

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according to the vendor’s recommendations and correct installation isverified and documented. OQ checks if the instrument conforms to thefunctional specifications, as defined in the DQ phase. PQ verifies that thecomplete system works for selected applications. Preventive maintenanceactivities and controlled changes also are part of this phase.

All activities are defined in a validation or qualification plan and resultsare documented in a summary report. Configurations are defined in therequirement specifications document for configurable computerized SFCsystems. They are implemented during the installation and tested as partof OQ.

Figure 4 illustrates which questions should be answered during the fourphases along with tasks and who usually performs the tasks. It also hasinformation on the test objectives, the type of sample and the type of column used for testing.

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Design Qualification

Installation Qualification

Operational Qualification

Performance Qualification

•

•

Compare user requirements with supplier specifications

• Verify environment

• Verify arrival as purchased

• Check proper installation of hardware and software

• Test of operational functions

• Performance testing

• Test of security functions

• Test for specified application

• Preventive maintenance

• On-going performance tests

Supplier assessment

Qua

lific

atio

n Pl

an

Qua

lific

atio

n R

epor

t

Figure 3Analytical instrument qualification according to the 4Q model.

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Qualification activities should be laid out in a master plan. The masterplan documents a company’s approach to specific activities, for example,how to qualify analytical instruments, how to assess vendors or what totest for in commercial computer systems. A master plan serves two purposes: when implemented right, it ensures consistent and efficientimplementation of equipment qualifications, and it answers an inspector’squestion of a company’s approach to instrument qualification and systemvalidation. FDA regulations and guidelines do not specifically require avalidation master plan. However, inspectors want to know what the company’sapproach towards validation is. The qualification master plan is an idealtool to communicate this approach both internally and to inspectors.

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4.2 Qualification Planning

DQ IQ OQ PQ

Question to Is the Is the lab Does the Is thebe answered instrument’s suitable for the instrument instrument design suitable instrument? work in the performance for the intended Is the the user’s suitable for application? instrument environment the intended correctly according to application? installed? specifications? Tasks Compare user Prepare lab Qualify Conduct requirements environment. equipment system with instrument Install and operational suitability specifications verify correct specifications tests installation in the user’s environment

Who User(s) assisted User(s) and User(s) or User(s)performs by vendor(s) vendor(s) vendor(s)the tasks

Test sample N/A Generic Generic Application specific

Column N/A None or Generic Application generic specific

Figure 44Q phase test diagram.

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In case there are any questions as to why things have been done or notdone, the master plan should provide the answers.

Within an organization a validation master plan can be developed for:

The entire company at a corporate level

Multiple or single sites

Departments

System categories

The master plan is a framework for individual project plans and should bewritten at the highest level possible. This ensures consistent implementationacross an organization.

Equipment and computer validation master plans should include:

1. Introduction including the plan’s scope, e.g., sites, systems, processes

2. Responsibilities, for example, user departments, QA, IT

3. Related documents, for example, risk management master plan

4. Products/processes to be validated and/or qualified

5. Qualification/validation approach

6. Risk assessment

7. Steps for equipment qualification and computer system validation with examples on type and extent of testing

8. Vendor assessment

9. Handling existing systems

10. Change Control procedures and templates

11. Instrument obsolescence and removal

12. Training plans (system operation, GMP)

13. Glossary

14. Attachments with templates and examples

4For each individual project a validation project plan should be developed.This plan is derived from the validation master plan.

The project plan outlines what is to be done in order to get a specific system into compliance. For inspectors it is a first indication of the controla laboratory has over a specific instrument or system and it also gives afirst impression of the qualification quality.

For simple equipment qualification a template in table form can be usedto outline planned activities. An example template is shown in figure 5.The left column can be the same for all instruments in the same category,which makes the whole qualification process very efficient.

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Scope This plan outlines qualification activities for SFC equipment hardware.

Product An SFC system consisting of a CO2 pump, modifier pump, description back pressure regulator, automated liquid sampler, column heater, and UV/Vis diode array detector. Instrument control and data acquisition is performed through a chromatographic data system.

Validation The 4Q lifecycle model is used. Individual modules arestrategy tested as a part of the SFC system. This holistic test approach should be used, whenever reasonable.

Responsibilities The end-user department has the ultimate responsibility that the system is qualified. The department leader will decide on a case-by-case basis who will be responsible for individual qualification activities. This person is known as the system owner.

Supplier ISO 9001 or equivalent is sufficient for supplierassessment assessment.

Risk assessment Risk assessment will be performed by the system owner according to ICH Q9. Risk assessment includes defining a risk category for the SFC system, major risk factors and mitigation steps for high risk factors.

Testing Performance functions as specified in the requirement strategies specification list will be tested in the end-user department. Exceptions: functions that are not impacted by the user’s environment.

Figure 5 (continued)Elements and content examples for the SFC hardware qualification project plan.

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Activities in the DQ phase should ensure that the design of the instrumentis suitable for the user’s applications and that the instrument is developed,manufactured, tested and supported by vendors with a certified quality system. Design qualification is a shared responsibility between the vendorand the user of an instrument.

The user writes requirement specifications for the instrument. This includesall functions the instrument should have and the performance specificationsthe equipment should meet as required for the intended application. Next the user compares his/her specifications with the vendor’s specificationsheet. If the vendor’s specifications are equal or better than what isrequired, the design is qualified for the intended use. Also included in the

DQ Includes: 1. A comparison of requirement specification, vendor specifications and confirmation that the vendor’s product design meets the user’s specification. 2. A statement that the vendor designs, develops and manufactures the analytical equipment in a controlled quality environment. 3. A vendor statement that support requirements will be met.

IQ IQ is performed by the vendor and reviewed and signed by the system owner.

OQ OQ is performed before initial use. Requalification is performed annually and after significant system changes. The system owner decides who will perform OQ: the end user department or the vendor.

PQ PQ tests will be performed by the end user on an ongoing basis: daily or whenever the system is used.

Traceability A traceability matrix links product requirement specifications matrix to OQ tests. It is created by the system owner.

Procedures Procedures are required for: DQ, IQ, OQ, PQ, change control, maintenance, operation, training and problem reporting.

Approval Qualification documents will be developed under supervision of the system owner, approved by the user department’s supervisor and QA.

Documentation Qualification documents will be created, reviewed, approvedcontrol and archived following the laboratory’s SOP for document control.

Figure 5 Elements and content examples for the SFC hardware qualification project plan.

4.3 Design Qualification

DQ phase is a formal vendor assessment. This can be made based onexperience with the vendor, through a mail audit or through a direct audit.

Specific vendor activities:

Design, develop and manufacture instruments in a quality control environment

Provide functional and operational product specifications

Provide information on how software and instruments are validated during development and supported during the entire life of the products

Allow user audits, if required, and share approaches for developmentand testing

Specific user activities:

Describe the analysis problem and selection of the technique

Describe the intended use of the equipment

Describe the intended environment (including computer environment)

Select and document the functional and performance specifications(technical, environmental, safety)

Verify that the vendor’s functional and operational product specificationsconform to the user requirements

Select and assess the vendor

Document the rationale for selecting the specific SFC equipment and the vendor

4.31 The Importance of Requirement SpecificationsDQ should ensure that instruments have all the necessary functions andperformance criteria that will enable them to be successfully implementedfor the intended application and to meet business requirements. Errors in

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DQ can have a tremendous technical and business impact, and therefore a sufficient amount of time and resources should be invested in the DQphase. For example, setting wrong operational specifications for an SFCsystem can substantially increase the workload for OQ testing, and selectinga vendor with insufficient support capability can decrease instrument up-timewith a negative business impact.

Figure 6 shows a template for documenting user requirements and vendorspecifications of an SFC system. The user defines his/her requirement specifications and compares them with the vendor’s specifications. Theexact user requirement specifications depend on the intended use of thesystem. To set the functional and performance specifications, the vendor’sspecification sheets can be used as guidelines. However, it is not recom-mended to simply copy the vendor’s specifications, because compliance to the functional and performance specifications must be verified later inthe process during operational qualification and also when re-qualifying the instrument at a later time. Specifying too many functions and settingthe values too stringently will significantly increase the workload for OQ.For example, if a company has a need for an isocratic SFC system, but plansto purchase a gradient system for future use, only an isocratic systemshould be formally specified for regulatory purposes. This means, as long as the instrument is not used for gradient runs no gradient test needs to be conducted. Later on, when the system is used for gradient analysis, the specifications should be changed through a change control procedure.

The specifications should be set so that there is a high likelihood that theinstrument conforms to them, not only during initial OQ but also duringrequalification, for example, a year later. Otherwise users may be expectedto initiate an investigation to determine if the non-qualified instrument couldhave had a negative impact on the quality of the product. For example, thesepossibilities are expressed in ICH Q7A (21): “Deviations from approvedstandards of calibration on critical instruments should be investigated to determine if these could have had an impact on the quality of theintermediate(s) or API(s) manufactured using this equipment since the last successful calibration”.

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Functions and performance User Vendor Passed requirements specification failed

Autosampler

Capacity

Injection volume range

Injection volume precision

Sample carry over

CO2 Pump

Pressure range

Flow rate range

CO2 Pump flow rate precision

CO2 Pump flow rate accuracy

Modifier Pump

Modifier pump flow rate precision

Modifier pump flow rate accuracy

Step gradient composition accuracy at 10, 50 and 90%

Back pressure regulator range

Back pressure regulator precision

Column Heater

Column heater temperature range

Column heater temperature precision

Column heater temperature accuracy

Column capacity

Column length

Diode Array Detector

Baseline noise (peak to peak at 254 nm)

Baseline drift

Linearity

Wavelength range

Wavelength accuracy

Software

Figure 6Template for user requirements and vendor specifications of an SFC hardware.

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4.32 Vendor AssessmentVendors of analytical instruments should be qualified through a formalprocess. The objective is to ensure that vendors provide the quality of products and can give adequate support. For basic equipment, such as pH-meters or a balance, this can be a single page statement describingwhy the vendor XY has been selected. Certification for a recognized quality system is sufficient for simple instruments. The formal assessmentstatement should be supported by the quality system’s certificate. Figure 7shows a template with examples to document vendor assessment criteriafor analytical instruments.

For more complex systems especially for critical computer systems, such aschromatographic data systems, a more detailed assessment is recommended.Depending on the complexity and criticality of the system this can be a mailaudit, 3rd party audit and a direct audit through the user firm.

The purpose of the vendor assessment is to ensure that products aredesigned, developed and manufactured in a documented quality environment.The assessment should also verify that the vendor provides the right servicesand can maintain the instrument through phone and on-site support.

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AssessmentItem

Establishmentin the marketplace

Expertise withFDA regulatedindustries

Experiencewith the vendor

Requirement

Vendor must be a leadingsupplier of SFC systems.

Vendor must berecognized as a leadingsupplier of products andcompliance services forFDA-regulated industry.

Our company must havegood experience with thevendor.

Findings

Vendor is a leading supplierof SFC systems.Vendor is also a leadingsupplier of equipment forpharmaceutical qualitycontrol laboratories.

Vendor has several timesbeen ranked as the #1supplier of products andcompliance services for FDA regulated industry.

Our laboratory already has 2 SFC systems with differentconfigurations and 10 HPLCsystems from the vendor.Experience with the vendorregarding instrument andsupport quality is very good.

Figure 7 (continued)Documenting vendor assessment.

4.4 Installation Qualification During the IQ phase the delivery of equipment is compared with the purchase order for completeness and the vendor’s installation instructionsare executed. This should also include checking if the laboratory conformsto the vendor’s environmental specifications, for example, humidity androom temperature. Finally, the IQ protocols are completed inserting thevendor’s name, model number, serial number and other relevant productinformation.

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4Quality system

Training

Installationqualification

Operationalqualification

Productsupport

Upgradesupport

Defectreporting

Vendor must have ISO9001:2008 certification.

Vendor must provideonsite operator training.

Vendor must install theproduct and performinstallation qualification.

Vendor must provideoperational qualificationservices.

Vendor must providephone and on-sitesupport in case of defectsand other problems.

Vendor must have auser-accessible websitewith information onavailability of newfirmware upgrades.

Vendor must notify usersabout known criticalhardware, firmware andsoftware defects.

Vendor is ISO 9001:2008certified.

Vendor provides onsiteoperator training.

Vendor installs the productand performs installationqualification.

Vendor provides operationalqualification services for SFCsystems.

Vendor provides phone andon-site support in case ofdefects and other problems.

Vendor has a useraccessiblewebsite with information on availability of new firmware upgrades.

Vendor has a procedure tonotify users about knowncritical hardware, firmwareand software defects.

Figure 7Documenting vendor assessment.

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Tasks performed for IQ:

Prepare the laboratory facility according to the vendor’s environmentalspecifications.

Control and record environmental conditions, if critical. For example,temperature and humidity.

Compare equipment received with the purchase order (including, accessories and spare parts).

Check equipment for any damage.

Verify that the instrument conforms to physical and construction requirements, as specified by the user.

Check documentation for completeness (operating manuals, maintenance instructions, standard operating procedures for testing,safety and validation certificates).

Install hardware (instrument, fittings and tubing for fluid connections,columns, power cables, data flow and instrument control cables).

Switch on the instruments and ensure that all modules power up andperform an electronic self-test.

List equipment manuals and SOPs.

Record firmware revision.

Prepare an installation report.

Enter instrument data into an inventory data base.

Prepare, review and sign formal IQ documentation.

Figure 8 shows a template with selected examples that can be used todocument completeness of delivery.

4

All instruments should be documented in the IQ protocol and entered into a database. The IQ documents should be updated whenever there is achange made to any entry in the IQ documents. Examples of changes are a firmware revision and the location of the instrument within a buildingor site.

Installation should verify that the instrument hardware and software areproperly installed. It does not verify that the instrument conforms to thefunctional and performance specifications. This is done later in the OQphase. For individual modules, testing is limited to perform and documentthe instrument’s self-diagnostics when it is switched on.

For SFC systems comprised of multiple modules, correct connectionbetween the modules should be verified. This can be easily achieved byrunning a well characterized test sample and comparing the output with a reference plot. An example is shown in figure 9.

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Purchase order item Vendor Complete PN Yes No

CO2 pump

Modifier pump

Backpressure regulator

Autosampler

UV/Vis diode array detector

Column heater

10 µL flow cell

Figure 8Examples for documenting completeness of delivery for IQ.

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Operational qualification (OQ) should demonstrate that the SFC equipmentwill function according to its operational specifications in the selected environment as defined by the user. OQ tests can be performed by a vendor representative or by the user. In any case, ultimate responsibility lies with the user and a user representative should sign the OQ document.Test engineers should be formally trained. When the tests are conducted by vendor representatives, user firms should get a copy of their trainingcertificate. When the test is conducted by user representatives, documentedevidence that the test engineer has been trained on the principles andrequirements for equipment qualification and on specific SFC OQ testingshould be available.

4Actions Expected result Pass/fail

1) Set instrument conditions according to the installation manual for analyzing the installation verification sample

2) Inject the installation verification sample

3) Verify if actual results are as expected

A chromatogram similar to the one in the installation manual is obtained.

1. Chromatogram must include four peaks.

2. Peaks 1 and 2 are higher than 3 and 4

3. Retention time of 4th peak should be between 1.7 and 2.1 minutes

1)

2)

3)

Figure 9Testing an SFC system for correct installation.

4.5 Operational Qualification

Tools used for testing such as digital flow meter, thermo meters and standards used for wavelength calibration should be formally calibratedand/or traceable back to national standards. Calibration certificates are initially provided by the vendor of the tools and may have to be recalibratedevery year. Inspectors expect OQ tests to be quantitative. This means thatthe test protocol should include expected results and actual results.

The instrument’s OQ is repeated at regular intervals. SFC systems classifiedas high risk equipment should be re-qualified every six to 12 months. Less frequent requalification should be justified based on documented riskassessment. In general, the time intervals should be selected so that theprobability is high that all parameters are still within the operational specifi-cations. Otherwise, analytical results obtained between the last and actualOQ with that particular instrument are questionable. Here the importanceof proper selection of the procedures and acceptance limits becomes veryapparent.

As a part of OQ, SFC equipment hardware should be tested for all specifi-cations after it is installed in the user’s laboratory even if it has been testedat the vendor’s site. Equipment hardware, such as UV/Vis detector gratings,can be impacted by mechanical vibration as a result of shipping. The purposeof OQ testing is to verify the specifications as defined by the user for allindividual modules or system components in the user’s environment. Therecommendation is to use the holistic approach for testing, which means to perform the test using a complete SFC system as shown in figure 10.

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CO2 pumpModifier pump

Automated sampler

Column heater

Computer control

Detector Back pressureregulator

Figure 10Block diagram of an SFC system.

4.6 Tests for OperationalQualification

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Typical tests include precision of injection volume, detector linearity, precisionand accuracy of flow rates and temperature of the column compartment.

Examples for SFC hardware test items are shown in figure 11. The templatecan be used to document test items, and expected and actual results.

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Test

Precision of injection volume: Inject a standardsolution 6 times and calculate the precision ofpeak areas.

Carry over: Inject a high standard concentrationfollowed by a blank solvent injection.

Flow rate accuracy: Measure the flow rate at 1 and 5 mL/min with a digital flow meter over six oneminute fractions. Calculate the average and compare with the setpoint.(Important: The intial words "Flow rate accuracy" should be bold, nor new paragraph before "Calculate"

Flow rate precision: Calculate the standarddeviation of the six readings from the flowaccuracy experiment.

Colum heater temperature accuracy: Measure the temperature inside the column heater at twodifferent locations and compare temperatures withthe setpoint.

Column heater temperature precision: Repeat the column heater accuracy experiment six times and calculate the relative standard deviation of thetemperature readings.

Baseline noise: Measure the peak-to-peakbaseline noise over six one-minute fractionsaccording to ASTM 1657 and calculate theaverage.

Baseline drift: Measure the slope of the baselinenoise according to ASTM 1657 over 10 minutes.

Injection linearity: Inject 6 standard samples withincreasing concentrations. Calculate thecorrelation coefficient of peak area versusconcentration.

Wavelength accuracy: Scan caffeine from 200 to300 nm. Compare the actual WL at 205, 245 and273 nm with the setpoints.

Expected Actual Passed/result result failed

Figure 11Template and examples for SFC hardware testing.

4.7 Performance Qualification Performance qualification (PQ) should demonstrate that an instrument consistently performs according to the specifications as defined by the user,and is appropriate for the intended use.

Here, emphasis is placed on the word ‘consistently’. Important for consistentinstrument performance are regular preventive maintenance checks, makingchanges to a system in a controlled manner and regular testing. The PQtest frequency is much higher than for OQ. Another difference is that PQshould always be performed under conditions that are similar to routinesample analysis. For an SFC system this means using the same column, the same analysis conditions and the same or similar test compounds.

PQ should be performed on a daily basis or whenever the instrument isused. The test frequency depends on the criticality of the tests, on therobustness of the instrument and on anything which may contribute to the reliability of analysis results. For an SFC instrument, this may be theanalytical column or a detector’s UV lamp.

In addition to tests during routine analysis, PQ tests should be conducted:

After developing and validating a new method

In the receiving laboratory when analytical methods are transferredbetween laboratories

When compendial methods are introduced into a laboratory

Whenever a method is changed

Whenever SFC hardware changes

After instrument maintenance

After instrument repair

After software upgrades and other software and computer systemchanges

Before a new column is routinely used

As part of a failure investigation in an OOS situation

In practice, PQ testing can mean system suitability testing or the analysis ofquality control samples. This is supported by USP chapter <1058>: “Somesystem suitability tests (SST) or quality control checks that are performed

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concurrently with the test samples also imply that the instrument is per-forming suitably”. For system suitability testing, critical system performancecharacteristics are measured and compared with documented preset limits.For example, a well characterized standard can be injected 5 or 6 timesand the standard deviation of amounts is then compared with a predefinedacceptance criteria. If the limit of detection and/or quantitation is critical,the lamp’s intensity profile or the baseline noise should be tested.

For chromatographic equipment the following SST tests are recommendedin USP chapter <621>:

Precision of the amounts

Resolution between two peaks

Peak tailing factor

Theoretical plates (N) for column performance

Capacity factor (k’)

USP <621> makes it clear that these tests should be performed frequently:“System suitability must be demonstrated throughout the run by injection of an appropriate control preparation at appropriate intervals.” The results of SSTshould be evaluated before the sample run. If SST does not pass, the systemmust not be used for any sample analysis: “No sample analysis is acceptableunless the requirements of system suitability have been met. Sample analysesobtained while the system fails requirements are unacceptable”.

4

mAU300

250

200

150

100

50

00 2 4 6 min

Figure 12Example of a SFC system suitability test run.

The type and frequency of system suitability tests should be defined in anSOP. Conformance to the SOP should be checked by QA in regular internalaudits. For routine analysis a possible scenario for test frequency is:

Whenever samples are analyzed

For a series of 1 to 9 consecutive sample analyses: before starting the series

For a series of 10 to 20 consecutive sample analyses: before starting theseries and after finishing the last sample analysis

For a series of >20 consecutive sample analyses: before starting theseries, after finishing the last sample analyses and after each tenth sample analyses

Computers with associated software are used to control the SFC hardware,to acquire SFC signals and for data evaluation, printing and storage.Frequently complete computerized systems are purchased from a singlevendor to ensure smooth system set-up and operation and to get full system support from a single vendor. Sometimes computer hardware ispurchased from a different vendor because of economic reasons. The final

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Test # Date Time T21 T22 T23 T24 T25 Comment

Baseline Tailing Peak Precis. Precis. noise factor resol. #1 #2

mm/ a=am Accept Accept Accept Accept Accept Passed/ day p=pm limit limit limit limit limit failed <1x10-4 <1.3 >2.0 <1% <1%

Test # Date Time T21 T22 T23 T24 T25 Comment

001 08/04 06.20 p 4.5x10-5 1.1 2.3 0.61 0.50 passed

002

003

004

Figure 13Template with examples for an ongoing system suitability test protocol.

4.8 Specific Considerationsfor Software andComputer Systems

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decision should be based on risk assessment. In any case, complete computerized systems should be validated. The approach is similar to theone for equipment. The major differences are:

More focus should be put on qualification of vendors. Vendors shouldprovide documented evidence that development followed a documentedprocess and that the software has been validated as part of this process.

Whereas for hardware equipment qualification all user specifications areverified in the user’s environment, this is not required for software. It issufficient to verify a selection of key software functions and to perform a complete system test. Examples of functions that should be tested aresecurity access and electronic audit trail.

Many times users customize computer systems, for example, throughreport generators or when setting network configurations. Users shouldinclude these configurations in the requirement specifications document.Configurations are implemented during installation and tested as part of OQ.

More information on validation and examples for software and laboratorycomputer system validation are included in references 2 and 6.

SFC instruments should be well maintained to ensure proper ongoing performance. Procedures should be in place for regular preventive maintenance of hardware to detect and fix problems before they can havea negative impact on analytical data. These procedures should describe:

The maintenance to be done.

When it is to be done.

What should be re-qualified after maintenance is done. For example, a PQ test should always be performed after instrument maintenance.

How to document maintenance activities.

Instruments should be labeled with the dates of the last and next scheduledmaintenance.

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4.9 (Preventive) Maintenanceand Repair

Planned maintenance activities should follow a documented instrumentmaintenance plan. Some vendors offer maintenance contracts with servicesfor preventive maintenance at scheduled time intervals. A set of diagnosticprocedures is performed and critical parts are replaced to ensure ongoingreliable system uptime.

Unplanned activities that are necessary in addition to the planned activitiesshould be formally requested by the user of the instrument or by the person who is responsible for the instrument. The reason for the requestedmaintenance should be entered as well as priority. All maintenance activitiesshould be documented in the instrument’s logbook.

Defective SFC systems must not be used. Procedures should be availableon how to handle most common problems. Procedures should also includeinformation if and what type of requalification is required after repair.Uncommon problems should be handled through a special procedure thatguides users of instruments through the repair and reinstallation process,for example, if a CO2 pump becomes defect. In this case the impact of thefailure on previously generated data should be evaluated.

Figure 14 shows a flow chart on how defective instruments should be handled.The following steps are recommended:

The problem should be reported to the laboratory supervisor, or to theperson responsible for the instrument, who will decide on further action.

The instrument should be clearly labeled as “Out of Service”.

After repair, correct functioning must be verified. The type and extent of testing depends on the failure and possible impact on the system.Depending on the failure, this may require partial or full requalification or only system suitability testing.

The impact of the defect on previous test results should be examined.

Suitable actions should be taken in case the defect instrument couldhave had an impact on the validity of data, for example, samples mayhave to be re-analyzed.

An entry on the defect, repair and performance verification should bemade in the instrument’s logbook.

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Analytical instruments and systems go through many changes during theirlifetime. New hardware modules may be added to enhance functionality,for example, an automated sampling system replaces a manual one forunattended operation. Vendors may change the firmware to a new revisionto remove software errors or application software may be upgraded to becompatible with a new operating systems. Or a complete system is movedto a newly designed laboratory.

Any changes to instrument hardware, firmware and software should followwritten procedures and should be documented. Requests for changesshould be submitted by users and authorized by the user’s supervisor ordepartment manager and by QA. Before any change request is approvedbusiness benefits should be compared with risks a change may bring. USPchapter <1058> states: “Implementing changes may not always benefitusers. Users should therefore adopt changes they deem useful or necessaryand should also assess the effects of changes to determine what, if any,requalification is required”.

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Repair Perform OQ and system suitability tests

Check impact on validity of data

Enter repair into logbook

Continue with analysis

New OQ sticker

Suitable actionsLabel as defective

Report to system owner

Identify defective instrument

Figure 14Flow chart for handling repair of SFC instruments.

4.10 Change control

USP also recommends following the same 4Q model for changes as for initial qualifications. This requires:

Specifications should be updated, for example in case a new automatedsampling system replaces a manual one.

IQ documents should be updated, if a new firmware revision is installed.Installation documents should also be updated when a system is movedto a new laboratory.

OQ documents with new test cases and test protocols should be added if the software is upgraded with new functionality.

PQ tests need to be performed to verify correct functioning of the completesystem, for example, when updating firmware of any SFC module.

Before any change is approved and implemented, a thorough evaluationshould be made if OQ tests should be repeated. Depending on what thechange is an instrument may need no, partial or full testing of a system.

At the end of validation a summary report should be developed. This shouldbe a mirror of the validation project plan. It should be organized so that it contains all the elements and follows the outline of the validation plan.This makes it easy to check if all plan items have been completed successfully.Deviations should be documented if there are any, together with correctiveactions and/or work around solutions. The report should include a statementthat the instrument or system is qualified or validated. When the statementhas been signed by laboratory and QA management, the product can bereleased for operation.

Typically the validation plan and the report are the first documents inspectorswant to see when they inspect a validation project. If everything is wellorganized and documented it may well be that after looking at both documents inspectors get such a good impression about the validationwork that they will focus on other inspection areas.

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4.11 Validation Reports

Chapter 5

Validation ofAnalyticalProcedures

5.1 ICH Validation Parametersfor Target Applications

Validation ofAnalyticalProcedures

GxPs require analytical methods and procedures to be validated todemonstrate suitability for their intended use. The ultimate objective ofthe validation process is to provide evidence that the procedure doeswhat it is intended to do, accurately, reliably and reproducibly.

Regulatory agencies and other official organizations have developed several documents for validation of analytical methods and procedures.For example, the FDA has published a draft Guidance on “AnalyticalProcedures and Methods Validation” (18) and a guidance on“Bioanalytical Method Validation”. USP has a chapter on “Validation of Compendial Methods” (26), one for “Verification of CompendialMethods” (27) and a recent draft for “Transfer of Analytical Procedures” (25).The reference document for validation of analytical methods is the ICH Q2(R1) guide “Validation of Analytical Procedures: Definitions andMethodology”. (20)

This section will give a brief overview on method validation according to ICH Q2. More detailed information is available in the Agilent primer“Validation of Analytical Methods” (3). For example, it includes a definition and detailed test parameters for all validation characteristics as required by ICH Q2.

The ICH Q2 describes validation parameters and gives recommendationsfor validation of SFC methods. Validation parameters are listed in figure 15.Robustness is usually included in the lists, but ICH recommends conductingthis test during method development. FDA and other agencies expectthat related robustness tests are included in the method validation package.

In concept, it is not always necessary to validate all analytical parametersas listed in figure 15. For example, if the method is to be used for qualitative trace level analysis, there is no need to test and validate themethod’s limit of quantitation, or the linearity, over the full dynamic rangeof the equipment. The extent of validation also depends on the lifecyclephase of the drug. While agencies expect full validation in clinical PhaseIII and for drug manufacturing control, most time-consuming tests, suchas intermediate precision, reproducibility and ruggedness, most likely arenot necessary in preclinical studies and for Phase I clinical studies.

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However, a statement is expected as to why the manufacturer believesthat the method is suitable for its intended use.

According to ICH Q2 the selection of validation parameters and acceptancecriteria should be based on regulatory requirements and should be justifiedand documented.

ICH defines different types of analytical procedures to be validated:

Identification test

Quantitation tests for impurities content

Limit test for the control of impurities

Quantitative tests of the active ingredient or other main components of the drug

Accuracy, any type of precision and limits of detection and quantitationare not required if the analytical task is identification. For assays themajor component or active ingredient to be measured is normally presentat high concentrations; therefore, validation of limits of detection andquantitation is not necessary. In quantitative impurity tests, all parametersshould be validated except limit of detection. Limit tests only require validation of specificity and limit of detection.

5

Analytical task Impurity testing

Parameter Identification Quantitative Limit tests Assay

Accuracy No Yes No Yes

Precision Repeatability No Yes No Yes

Intermediate precision No Yes No Yes

Reproducibility No Yes No No

Specificity Yes Yes Yes Yes

Limit of detection No No Yes No

Limit of quantitation No Yes No No

Linearity No Yes No Yes

Range No Yes No Yes

Figure 15ICH Validation characteristics for target applications. (For a detailed descriptionof terminology and methodology see reference 3).

5ICH Q2 also gives recommendations for test procedures. Typically, foreach test parameter several options are provided. Examples are shown in figure 16.

Figure 17 shows the specificity of SFC for a racemic mixture of R-1,1’-bi-2-naphthol and S-1,1’-bi-2-naphthol. The separation was performed at 2 ml/min on a Chiracel OD-H column. In the upper chromatogram the racemic mixture was separated, in the middle andlower panel a so called pure formulation was analyzed. LOQ studiescould demonstrate that the LOQ was below 0.05% at acceptable precision.

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Parameter Tests

Accuracy Minimum at 3 concentrations, 3 replicates

Precision

Repeatability Minimum of 9 determinations over the specified range

Intermediate Over 3 days, 2 operators, 2 instruments

Reproducibility Only required if routine testing is done in different laboratories

Specificity Prove with specific techniques: SFC, DAD, MS. or by using

different SFC columns

Limit of detection Visual, Signal to Noise ≥3

Limit of quantitation Visual, Signal to Noise ≥10, Standard deviation of response

Linearity Min. 5 concentrations covering 70 to 130 % of the expected

concentration range: visual examination, correlation coefficient

Range 70 to 130 % of test concentration, from linearity, precision

and accuracy tests

Figure 16Selected tests as recommended by ICH Q2.

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The validity of a specific method should be demonstrated in laboratoryexperiments using samples or standards that are similar to unknown samplesanalyzed routinely. The preparation and execution should follow a validationprotocol, preferably written in a step-by-step instruction format. Just likeequipment qualification and computer system validation, method validationalso is not a one-off event. It starts when somebody wants to implementa new method in a laboratory and ends when the method is no longerused. Because of the length of time and complexity, the process is brokendown into phases.

Method development and validation are not independent from eachother. For example ICH Q2 suggests robustness testing to be done duringmethod development. Figure 18 shows the progression of integratedmethod development and validation.

5.2 Strategies for IntegratedMethod Development andValidation

mAU

150100

500

1 2 3 4 5 6 7 minmAU

300400

200100

0

300400

200100

0

1 2 3 4 5 6 7 minmAU

1 2 3 4 5 6 7 min

A) R, S - 3

B) R - 3

C) S - 3

S

R

S

S

R

Racemic mixture

Impurity of 'S' in 'R'

Impurity of 'R' in 'S'

Figure 17Specificity of SFC for a racemic mixture R-1,1’-bi-2-naphthol and S-1,1’-bi-2-naphthol. Mobile phase: CO2/Modifier (MeOH with 0.1 % TFA and 0.1 % DEA)75/25), Column: ChiralCel OD-H, 4.6 ID x 250 mm, 5 µm.

Preparation

Development

Validation

Routine operation

•

•

Select preliminary method, scope & specificationsAssure performance of equipmentAssure that operators are qualified

•

••

•

Select and optimize method & parametersRobustness testingDefine operational limits and SST Preliminary validation experiments

•

•

•

Define and document finalacceptance criteriaDocument final scopePerform validation tests

•

•

•

Controlled transfer Regular reviewControlled changes & revalidation

•Reg

ulat

edN

ot R

egul

ated

Figure 18Integrated SFC method development and validation.

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The entire project includes four phases:

Preparation phase

Development phase

Validation phase and

Routine operation

In the preparation phase a project plan is created and the analysis tech-nique is selected. In addition, the preliminary scope, method parametersand acceptance criteria are defined. The performance of the selectedinstruments and material should be checked and operators should betrained on SFC instrument operation and analysis procedure.

In the development phase instruments are selected and method parametersare optimized. This phase also includes extended robustness testing.Good knowledge of a method’s robustness allows defining criteria forrevalidation. It is also recommended to perform preliminary method validation experiments. Contrary to the formal validation studies, thedevelopment phase is not regulated and these studies will help to ensure successful validation studies without out-of-specification results.

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The development phase is also an appropriate time to determine thetype, frequency and acceptance criteria for system suitability tests.

At the beginning of the validation phase a method validation plan is createdwhich includes the final scope, final acceptance criteria and owners,responsibilities and deliverables. The plan also includes the compoundswith concentration range, the sample matrix, the specific equipment thatshould be used and the location where the method should be used forsample analysis. The equipment is formally qualified, all materials shouldbe qualified and operators must be trained and the efficiency of the trainingshould be documented. Validation experiments are conducted accordingto the plan and the results are compared with acceptance criteria. Finallya validation report is written, which summarizes the validation results andconclusions. Figure 19 shows components and content recommendationsof a method validation plan.

Items

Plan approvals(Lab supervisor, QA representative and QCU director or any otherperson responsible forbatch release)

Glossary

Method description

Validation team

Content examples

• Verify plausibility of validation parameters and acceptance criteria• Verify conformity with internal procedures, e.g., with SOP: Validation of Analytical Methods• Make sure that the plan is signed before the experiments start

• List all abbreviations used in the plan• Use the abbreviations from your company’s general glossary to avoid any confusion

• Purpose of the SFC method, e.g., Quantitative analysis of impurities in drug (be as specific as possible)• Intended environment• Regulatory environment

• Team members: analysts, lab supervisor, QA department member, QCU director or delegate• Responsibilities• List intended analysts by name• List substitutes

Figure 19 (continued)Components and content recommendations of a method validation plan.

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5Equipment

Software andcomputer systems

Reference material

Reagents andsolutions

Requireddocumentation

Validation parameters,experiments, andacceptance criteria

Schedule anddeliverables

• List SFC equipment with manufacturer and model number Make sure that the same equipment is also available or can be provided for routine analysis• List all accessories required to run the method• Make a statement that the equipment is qualified according to USP <1058> or equivalent

• List intended software for instrument control, data acquisition, data evaluation, reporting and archiving• Make a statement that the software is validated according to your company’s validation master plan (this also applies to MS Excel® spreadsheets used for statistical evaluation)

• List all reference material, such as standards for method calibration and certified reference material for accuracy studies• List all the suppliers of the material• Make sure that the vendor and material is qualified

• List all reagents, solutions and other chemicals and consumables that are required to perform the experiments• List all material suppliers

• List all documents that are required to execute the experiments and to evaluate the data.

Describe methodology, detailed step-by-step description of the experiment, description of data evaluation and presentation and acceptance criteria for:• Accuracy• Limit of quantitation• Repeatability• Intermediate precision• Selectivity• Linearity• Range• Robustness

• Include tasks, responsible persons and deliverables

Figure 19Components and content recommendations of a method validation plan.

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Figure 20 shows a template with selected examples of a summary reportof validation studies. The method is used for the release of drugs in theform of tablets for shipment. The scope of the method is assay testing ofthe drug substance. All validation experiments have been carried out asdescribed in the validation plan. Validation experiments included accuracyat 70, 100 and 130 % of the label claims, short term and intermediateprecision, specificity, linearity, range and robustness. Reproducibility testswere not necessary because the routine method is used in the same labwhere it has been developed. Because all parameters meet acceptancecriteria, the method is released as validated.

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Validationparameter

Accuracy

Method precision

Intermediateprecision

Specificity

Linearity

Range

Robustness

Measure

Recovery – 70 %Recovery – 100 %Recovery – 130 %of label claim

RSD

RSD

Peak resolutionFactor R

CorrelationCoefficientVisual inspection of plot

Correlation CoefficientVisual inspection of plotRecovery at 3 conc.

Column temp. ±2 ºC% Org, mobile phaseStability of DSA1

Acceptancecriteria

97 – 103 %97 – 103 %97 – 103 %

≤ 1.5 %

≤ 2.0 %

R for all peaks >1.5

≥ 0.9900

Linear response plot

≥ 0.990

Linear response plot97 – 103 %

R for all peaks ≥ 1.5R for all peaks ≥ 1.5≤ 2 % loss in 96 hours

Results

98.3 %97.4 %98.6 %

0.4 %

0.8 %

All peaks >2.0

0.9900

Shows linearity

0.995

Shows linearity99.6%

All peaks ≥ 2.0All peaks ≥ 2.00 % loss

Figure 20Method validation summary results.

5.3 Verification ofCompendial Procedures

Laboratories working in regulated or quality standard environments arerecommended to use official methods as those developed by organizationssuch as the American Society for Testing and Materials (ASTM), ISO or the USP. For example, the US Food, Drug & Cosmetic Act requires FDA regulated industries to use compendial procedures or demonstrateequivalency. These methods are validated and many analysts believe that a method can be used without any further validation, verification or testing done in the laboratory. This is a wrong assumption. The USFDA cGMP regulation states in 21 CFR 211:194 a(2); “If the methodemployed is in the current revision of the United States Pharmacopoeia, or in other recognized standard references, or is detailed in an approvednew drug application and the referenced method is not modified, a state-ment indicating the method and reference will suffice. The suitability ofall testing methods used shall be verified under actual condition of use.”

This makes it clear that official methods don’t need to be validated aslong as they are not changed, but the laboratory should demonstrate that it is capable of successfully running the method. The question is how to do this: should some or all validation experiments be repeated, or are successful system suitability tests or the analysis of quality controlsamples enough?

Help came from the USP through its chapter <1226>: Verification ofcompendial methods (27). The given recommendations are not only useful to implement compendial methods but can also be useful for anystandard method.

Key recommendations:

Demonstrate the performance of the laboratory and system throughsystem suitability tests.

Assess the criticality and complexity of the method.

Select most critical performance characteristics of the method.

Depending on the criticality and complexity of the method, repeat one to three most critical validation experiments.

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Just as the validation of methods developed in-house, the evaluation and verification of standard methods should also follow a documentedprocess for example, a validation plan or an SOP. Results should be documented in the validation protocol. Both documents will be a majorsource of the validation report.

When validated methods are transferred between laboratories, thereceiving laboratory should demonstrate that it can successfully performthe method. Typical instances when method transfer occurs are from theResearch and Development (R&D) laboratory to the Quality Control (QC)laboratory, Site A to Site B when a product line is moved, from a sponsorcompany to a contract laboratory and from Company X to Company Ywhen a product is purchased by another company.

Currently there is no official document available that can be used as aguidance document on how the performance of the receiving laboratorycan be demonstrated. Most promising is the draft USP chapter <1224>entitled “Transfer of Analytical Procedures” (25).

Key recommendations:

Train analysts in the receiving laboratory on methodology and theapplication.

The transferring laboratory defines one or more well characterizedsamples, documents method parameters and acceptance criteria, for example, for accuracy of the method. The sample should cover the complete concentration range as specified when the method wasoriginally validated.

The samples are analyzed in the receiving laboratory and the resultsare compared with the acceptance criteria.

Depending on the criticality of the analysis and on the complexity of the method, one or two validation tests should be repeated, for example, limit of quantitation for quantitative impurity analysis.

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5.4 Transfer of AnalyticalProcedures

The method transfer process follows a documented process e.g., a transfer plan or an SOP. The extent of testing depends on specific situations, for example, on the method, samples to be analyzed, howmany different instruments and how many analysts will run routine tests.

Variables to be considered are:

Number of samples, lots, batches: 1-3

Concentrations: 1-3

Number of repetitive analysis / sample: 4-6

Number of analysts: 1-2

Number of days: 2-4

Equipment from different manufacturers: 1-all

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Chapter 6

Managing ElectronicRecords for FDA Part 11and EU Annex 11Compliance

Managing ElectronicRecords for FDA Part 11and EU Annex 11Compliance

Most SFC instruments are connected to computer systems with specificapplication software that controls instrument parameters, acquires signaland spectral data, convert the original digital data into meaningfulresults, and finally print results and store and archive instrument andmethod parameters, original digital data and processed data for therequired retention period. The United States and the European Unionhave regulations for managing computers and electronic records and signatures. The US regulation is the FDA’s 21 CFR Part 11 (28) supportedby the industry guidance document “Scope and Applications” (29). The equivalent regulations in Europe are Chapter 4 of the EU GMPs (30)which deals with documentation and Annex 11 to the EU GMPs (31)with requirements for managing computer systems. 21 CFR Part 11applies to all computer systems used in an FDA-regulated environment,the cited EU documents apply to environments regulated by EuropeanGMPs. The most specific document for GLP is the OECD ConsensusDocument #10 “Using computerized systems in GLP environments” (32).

The objective of all these documents is to make sure that electronicrecords and signatures are as trustworthy and as reliable as paperrecords and handwritten signatures and that the use of computer systemsdoes not adversely impact the product quality and quality assurancewhen compared to manual systems.

Since several years EMA and FDA investigators have focused on computersystems and electronic records during inspections. The main reason is thatthe FDA and EMA inspectors found several manipulations of electronicrecords. For example Rivera (12) reported at the 2007 international GMPconference in Athens that chromatograms were cut and pasted and sampleweights were changed to bring out of specification results in specifications.Since then more than 30 warning letters have been published with violations related to computer systems and data.

Because of the importance of this topic, a chapter of this primer is dedicated to computer system and electronic record management. It will cover all important requirements as defined in references that are relevant to computerized SFC used in pharmaceutical developmentand QC laboratories.

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Risk management should be applied according to ICH Q9 throughout thelifecycle of the computerised system taking into account patient safety,data integrity and product quality. As part of a risk management system,decisions on the extent of validation and data integrity controls should be based on a justified and documented risk assessment of the computerised system.

Recommendations are:

Determine the risk level of the SFC system: high, medium, or low.Criteria are: impact of the system on data integrity, (medicinal) productquality and patient safety.

Apply type and extent of compliance activities according to the defined category, for example, for the extent of validation and the frequency of revalidation, whether electronic audit trail should beimplemented or not and the frequency of back-up.

Computer systems used to generate, maintain, and archive electronicrecords should be validated to ensure accuracy and reliability of therecords.

Recommendations are:

Follow the lifecycle model for validation as described in chapter 4 of this primer.

Apply the concept of risk based validation.

Check Annex 11 chapter 4 (31) for the type of documentation thatshould be generated and available during inspections.

Systems exchanging data electronically with other systems shouldinclude appropriate built-in checks for the correct and secure entry andprocessing of data, in order to minimize the risks. For critical data enteredmanually, there should be an additional check on the accuracy of the data.This check may be done by a second operator or by validated electronic means.

66.1 Risk Assessment

6.2 System Validation

6.3 Data Accuracy

Recommendations are:

Validate the accuracy of data transfer between computer systems. This can be done during operational or performance qualification.

Use software functionality, if available, to check the plausibility andaccuracy of manual data entries.

If there is no software functionality to validate manual data entries, for high risk data verify the accuracy through a second person.

Procedures and technical controls should be in place to limit access tosystems and data to authorized individuals. Suitable methods of preventingunauthorized entry to the system include the use of keys, pass cards, personal codes with passwords, biometrics, and restricted access to computer equipment and data storage areas.

The system should conduct authority checks to ensure that only authorizedindividuals can use the system, electronically sign a record, alter a record,or perform the operation.

Systems should be designed to record the unique identity of operatorsentering, changing, confirming or deleting data including date and time.

Recommendations are:

Develop policies for generation, distribution, use, and maintenance ofpasswords.

Develop procedures for limited access to the system to individuals, forexample, through user ID and password. Each employee should havehis/her unique user ID and should select his/her own password.

Develop procedures for limited access to the system operation anddata.

Make sure that the installed software can be used to implement yourcompany’s password policies and procedures.

Configure the system to implement your company’s password policy.

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6.4 Limited Access to authorized Users andAuthority Checks

65

Configure the system to implement your company’s procedure for limited access to systems and data.

Verify correct implementation of password policy and procedures forlimited access to the system and data.

Establish a list of authorized users.

Many documents exist in hybrid forms, i.e. some elements as electronicand others as paper based. Regulated users should define which dataare to be used as raw data. At least, all data on which quality decisionsare based should be defined as raw data. When copies of records aremade, the copies should be accurate and complete or should provide the content and meaning of the original record.

Recommendations are:

For each application define what raw data are, for example, originalelectronic records, intermediate processed data or computer print-outs.

For chromatographic systems, including SFC systems, define originalelectronic records as raw data.

Keep electronic raw data for review and copying by the agency afterprinting SFC results.

Contact the agency if there are any questions regarding the ability ofthe agency to perform this review and copying of electronic records.

Records should be protected to enable their accurate and ready retrievalthroughout the required records retention period. Data should be securedby both physical and electronic means against damage. Stored datashould be checked for accessibility, readability and accuracy. Access todata should be ensured throughout the retention period. Regular backupsof all relevant data should be done. Integrity and accuracy of backup dataand the ability to restore the data should be checked during validation andmonitored periodically.

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6.5 Raw Data and Copies ofRecords

6.6 Protection of Records

Recommendations are:

Follow your company’s procedure for retention of electronic records.

Check the required retention period for your records according torequirements as specified in relevant GxP regulations.

Migrate the electronic records when the SFC systems are upgraded or replaced by new ones. Ask the vendor for validated file conversionroutines.

Regularly check availability and integrity of the electronic records during the entire archiving period.

Make a back-up of electronic records.

Validate back-up and restore procedure.

Secure, computer-generated, time-stamped audit trails should be used toindependently record the date and time of operator entries and actionsthat create, modify, or delete critical electronic records. Record changesshould not obscure previously recorded information. The audit trail documentation should be retained for a period at least as long as thatrequired for the subject electronic records and should be available foragency review and copying. The audit trail documentation should be regularly reviewed. For records supporting batch release it should bepossible to generate printouts indicating if any of the data has beenchanged since the original entry.

Recommendations are:

Include electronic audit trail in the user requirement specification forcomputerized SFC systems.

Specify as requirements for audit trail: what was changed, who madethe change, when the change was made by date and time and as anoption the reason for the change.

Make sure that audit trail documentation is available in human readable form.

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6.7 Computer GeneratedTime-Stamped AuditTrails

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Make sure that changed records do not obscure original records.

Review electronic audit trail documentation based on risk of the records.

Verify correct functioning of the electronic audit trail.

Retain the audit trail documentation for as long as the required retention period for subject records.

Use software that can recognize changed records based on print-outs.If such software is not available, implement a manual procedure.

Part 11 and Annex 11 allow signing records electronically. Part 11 hasmore specific requirements for the execution of electronic signatures.Information associated with the electronic signature should include:

The printed name of the signer

The date and time when the signature was executed

The meaning of the signature, for example, review, approval, responsibility, ownership

Handwritten and electronic signature should be permanently linked totheir retrospective electronic records

Recommendations:

Decide and document the decision whether to sign records electronicallyor through handwriting.

If electronic signatures are to be used include the required softwarefunctions in the system’s user requirement specifications document.

If electronic signatures are to be used send a letter to the FDA thatyour company will we using electronic signatures (only applies to US FDA regulated industries).

Train personnel on the meaning and accountability of electronic signatures.

Configure the SFC system for use with electronic signatures.

Validate correct functioning of electronic signatures.

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6.8 Electronic Signatures

6.9 Periodic Evaluation Computerized systems should be periodically evaluated to confirm thatthey remain in a valid state.

Recommendations:

Develop and implement procedures to periodically review computerizedSFC systems.

Set the time frequency to twice / year.

Include in the evaluation the current range or functionality, deviationrecords, incidents, problems, upgrade history, reliability, security and validation reports especially results of regular performance qualifications.

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References

70

R1. Agilent Technologies, Compliance for Biopharmaceutical Laboratories,

Primer, Publication Number: 5990-7001EN, 2011

2. Agilent Technologies, Analytical Instrument qualification and SystemValidation, Primer, Publication Number: 5990-3288EN, 2009

3. Agilent Technologies, Validation of Analytical Methods, Primer,Publication Number: 5990-5140EN, 2010

4. C.C.Chan, H. Lam, Y.C.Lee, X.M. Zhang, Analytical Method Validationand Instrument Performance Verification., Wiley Interscience, HobokenUSA, 2004

5. L. Huber, Validation and Qualification in Analytical Laboratories,Interpharm, Informa Healthcare, New York, USA, 1998, Second revision 2007

6. GAMP, Good Automated Manufacturing Practice, A Risk-basedApproach for Compliant GxP Computerized Systems, Version 5: 2008

7. GAMP, Good Practice Guide for Validation of Laboratory Systems, 2005

8. M.N.Dunkle, A. dos Santos, F. David and P. Sandra, Sensitive determination of impurities in achiral pharmaceutical by supercriticalFluid Chromatography Using the 1260 Infinity analytical SFC SystemAgilent Application Note, Publication Number: 5990-6413EN, 2010

9. L.T. Taylor, Supercritical Fluid Chromatography for the 21st Century, of Supercritical fluids 47, 566-573, 2009

10. Z. Wang, O. Liu and B. Donovan, The Path Forward for SupercriticalFluid Chromatography in the Drug Development Environment – anIndustrial Perspective, APR, 94-99, October 2009

11. P.Sandra, A.Pereira, F. David, M. Dunkle and C. Brunelli, Green chromatography (Part 2): The Role of GC and SFC, ChromatographyOnline, September 2010

References

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12. E. Rivera-Martinez, U.S. Food and Drug Administration, Data Integrity and Fraud – Another Looming Crisis, presented at the31st International GMP Conference, Athens (GA), 2007

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Glossary CDS Chromatographic Data System

cGMP Current Good Manufacturing Practice

CFR (US) Code of Federal Regulations

DQ Design Qualification

EMA European Medicines Agency

EU European Union

FDA Food and Drug Administration

GAMP Good Automated Manufacturing Practice

GCP Good Clinical Practice

GLP Good Laboratory Practice

GMP Good Manufacturing Practice

HPLC High Performance Liquid Chromatography

ICH International Conference for Harmonization

IQ Installation Qualification

ISO International Organization for Standardization

OECD Organization for Economic Co-operation and Development

OOS Out of specification

OQ Operational Qualification

PIC/S Pharmaceutical Inspection Convention / Cooperation Scheme.

PQ Performance Qualification

QA Quality assurance

QC Quality control

SFC Supercritical Fluid Chromatography

SOP Standard operating procedure

SST System suitability testing

USP United States Pharmacopeia

G

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Qualification andValidation for Supercritical FluidChromatography

Qualification and Validation for Supercritical Fluid C

hromatography

© Agilent Technologies Inc., 2011Printed in Germany, November 1, 2011Publication Number 5990-9148EN

www.agilent.com/chem/

A Primer

The Mea sure of Confidence