Analytical Instrument Qualification (AIQ)

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©2010 Waters Corporation

Paul St. JeanProduct Marketing ManagerGlobal Compliance ProgramsWaters CorporationMilford, MA 01757 USA

Analytical Instrument Qualification (AIQ)Analytical Instrument Qualification (AIQ)

IVT Method Validation ConferenceJuly 28-30, 2010Sir Francis Drake HotelSan Francisco, CA

©2010 Waters Corporation 2

Agenda

Regulatory Expectations for Qualification— Terms— Warning Letters

USP <1058> AIQ Guidance Chapter— How it came about

Developing a roadmap for the AIQ Process— DQ IQ OQ PQ and more

Instrument Qualification Case Study Application— Developing an understanding of what’s involved in qualification— Examining various approaches— Can we be more efficient?

Interactive Exercise— Discuss real challenges in your lab— Group Q and A

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II

Regulatory Expectations for QualificationRegulatory Expectations for Qualification


Data Integrity Objective

Data that supports the design, development and production of government regulated products must be accurate and defensible.

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Calibration Requirements

CFR211.68 cGMP Equipment— Automatic, Mechanical and Electronic Equipment.

— “…equipment so shall be routinely calibrated, inspected or checked according to a written program designed to assure proper performance. Written records of these calibration checks and inspections shall be maintained.”


Calibration Requirements

CFR 211.160 General Requirements— Laboratory controls shall include…

— “The calibration of instruments, apparatus gauges and recording devices at suitable intervals in accordance with an established written program containing specific directions, schedules and limits for accuracy and precision…”

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Validation versus Qualification

VALIDATION:− Refers to the total life cycle of a product from development through

use and maintenance.

− Customers (Owners) are responsible for Validating Their Processes (personnel, equipment, methods, SOPs) to ensure compliance to CGMP/GLP regulations.

QUALIFICATION: (Inspection,functional testing and documentation review)

− Is a part of the validation process which verifies module and system functional performance prior to being placed on-line and thereafter according to a standard operating procedure.

− Calibration verification is included as part of qualification.

− Vendors can assist customers in meeting their validation requirements but cannot validate customers’ processes.



According to an FDA inspector…..

QUALIFICATION:Answers the question…..

Was the instrument built right?

VALIDATION:Answers the question…..

Was the right instrument built?

• Vendors are in the best position to verify proper instrument operation

• Only users can determine if a system is suitable for their analysis

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According to an USP 1058 …..

Instruments are QUALIFIED

Processes are VALIDATATED

• It is easiest to qualify instruments using standardized tests

• Procedures and methods are validated for each specific analysis


FDA Guideline

Provide Documented Evidence of HPLC System…— Accuracy, Linearity and Precision

o from: • “Validation of Computerized Liquid Chromatographic

Systems” Furman, W.B., Layloff, T.P. & Tetzalff, R.F. August, 1992

o This may be old, but it is still valid and still gets occasional mention in journals

Regulations tell us what to do, but not how to do ito CFR 211.68o CFR 211.160

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Qualification Time Line

Equipment used should be of Appropriate Design, Adequate Capacity and Properly Maintained.

Functional Testing/VerificationFunctional Testing/Verification

CalibrationCalibration

PQPQ

StructurallyStructurallyValidatedValidatedProductsProducts

IQIQ OQOQ PQPQ

InstallationInstallation OperationalOperational PerformancePerformance

QualificationQualification

BeforeBeforePurchasePurchase

Owner'sOwner'sSiteSite

DQDQ

Owner'sOwner'sSiteSite

Vendor'sVendor'sSiteSite

MaintenanceMaintenance

(Initial Calibration)

System Suitability During UseSystem Suitability During UseBefore UseBefore Use After UseAfter Use


Method Validation

As you know, Method Validation is another area that regulated users must be concerned with but we should not confuse method validation with system qualification

Methods should be validated on qualified systems…• Qualification shows that the module or system performs as the

vendor intended• Qualification is not intended as a complete re-validation of the

design but rather a way to provide documented evidence of proper system performance in terms of accuracy, linearity (accuracy across a range) and precision

• Qualification does not demonstrate suitability for intended use• Method validation shows suitability of a system to perform a

specific analysis

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ICH

International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use (European Union, Japan and USA)

Purpose:— Harmonization of interpretation / application of technical guidelines

and requirements for product registration

— Reduce or obviate need for duplicate testing during the research and development of new medicines

Objective:— Economical use of human, animal and material resources

— Eliminate unnecessary development delays

Sections of interest related to this talk: — Web site is WWW.ICH.Org

— Q7 Good Manufacturing Practice for Active Pharmaceutical Ingredients

— Q9 Quality Risk Management


483 Warning Letters Issued

Personnel Issues (21 CFR 211.25)— Analysts not qualified

— Problems traced to inadequate training

— No training records

— No list of approved computer system users (21 CFR 211.68)

Analytical Method Issues (21 CFR 211.160)— Analytical methods not validated

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483 Warning Letters Issued

Data Audit Trail and Software Issues— No computer system validation report (211.68)— Software not revision controlled (211.68)— No audit trails (21 CFR Part 11)— Lack of audit trails (electronic) proving retention of all

raw data files (211.194 & 21 CFR Part 11)

Equipment Issues— No equipment PM records (211.67)— Un-qualified computer systems (211.68)— Failure to calibrate…no written program (211.68)— Instruments not qualified/calibrated (211.160)— Calibration not conducted at suitable intervals (211.160)


IIII

USP <1058>USP <1058>

Analytical Instrument Qualification (AIQ)Analytical Instrument Qualification (AIQ)

Guidance DocumentGuidance Document

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INTRODUCTION


Workshop on AIQ

AAPS Workshop co-sponsored with FIP and ISPE “Scientific Approach to Analytical Instrument Validation”,

o Held in Arlington, VA, March 3-5, 2003

Proceedings are published and entered in FDA Docket— Bansal SK, Layloff T, Bush ED, Hamilton M, Hankinson EA, Landy JS, Lowes S, Nasr MM, St. Jean PA,

Shah VP. Qualification of Analytical Instruments for Use in the Pharmaceutical Industry: A Scientific Approach.AAPS PharmSciTech. 2004; 5(1): article 22.

Proceedings form Basis of new USP Guidance —<1058> Analytical Instrument Qualification

o Pharmacopeial Forum Vol. 31(4) July-Aug 2005

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Terminology

Ambiguous terms: Validation or Qualification

Agreed at the workshop that — Processes are validated— Instruments are qualified

AIQ (Analytical Instrument Qualification)— “AIQ Is documented evidence that an instrument performs suitably

for its intended purpose and that it is properly maintained and calibrated”

— Does not include people (training), processes performed on instruments (analytical methods)

— Helps to justify the continued use of instrument, but it alone does not ensure quality of the data


Components of Data Quality

Analytical Instrument QualificationAnalytical Instrument Qualification

Analytical Methods ValidationAnalytical Methods Validation

System Suitability TestsSystem Suitability Tests

Quality Control Checks


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Component of Data Quality: Analytical Method Validation

Documented evidence that an analytical method does what it purports to do and addresses the required attributes of the method

Use of a validated method should instill confidence that the method can generate data of acceptable quality


Component of Data Quality: System Suitability Tests

Equipment, electronics, software, analytical operations and samples to be analyzed constitute an integral system

SST verifies the system’s performance according to the analyst’s expectation and criteria set forth in the method

H/W, S/W and analysis checks to provide assurance of system integrity before, during and following analysis of unknown samples

System Precision

Separation Parameters

System suitability testing alone is not sufficient, Qualification is still required.

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Component of Data Quality: Quality Control Checks

Most analyses are performed on calibrated or standardized instruments

Single or multi-point calibration performed

Additional quality control check samples used in some analyses (e.g. bioanalyses) provide an in-process assurance of the test’s suitable performance


Extent of SST or QC checks

Chemical analyses subject to GMP regulations, requiring tighter precision and accuracy may require more SST

Bioanalyses, largely subject to GLP regulations, requires low level and broad range analysis, is performed with more QC checks during sample analysis

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Components of Data Quality

Analytical Instrument QualificationAnalytical Instrument Qualification

Analytical Methods ValidationAnalytical Methods Validation

System Suitability TestsSystem Suitability Tests




IIIIII

Developing a Roadmap for the AIQ ProcessDeveloping a Roadmap for the AIQ Process

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PROCEDURES


Performing AIQEssential Parameters

Several options were discussed, but DQ/IQ/OQ/PQ model was preferred

Most widely understood parameters

Need to provide AIQ specific definitions and scope for these parameters

Recommended set of parameters for AIQ:

o Design Qualification (DQ)o Installation Qualification (IQ)o Operational Qualification (OQ)

o Performance Qualification (PQ)

o Change Control

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Design Qualification

Needed when designing new instrument

Most suitably Performed by developer/manufacturer

Not necessary for user to repeat DQ

Users should ensure:— The instrument is fit for their intended use

— Manufacturer has adopted a quality system for development, manufacturing and testing

— Manufacturer has adequate support for installation, service and training

Procedure— Depends on the intended use, complexity of instrument, acceptability of

instrument in industry and prior experience

— Fulfilled by vendor documentation/audits, peer reviews and testing


Installation Qualification

IQ is a documented collection of activities needed to install the instrument in the environment where it will operate— System Description

— Delivery: Did we receive what was ordered?

— Facility/Environment/Utilities

— Network and Data Storage

— Assembly and Installation

— Installation Verification

IQ can be performed for new, pre-owned or existing unqualified instruments

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Operational Qualification

OQ Parameters: Test operation of instrument as per specifications in the user’s environment— Test critical parameters to assure required performance

— Parameters based on manufacturer’s recommendation and on user’s intended use

— Secure data storage, backup and archive process

Performance of OQ— Can perform holistic or modular— Modular when applicable— Modular approach will allow mix and match of modules in a system — Use non-method specific testing— Repeat relevant OQ tests when instrument undergoes major repairs

or modifications


Performance Qualification

PQ tests are performed on a periodic basis to ensure that the instrument remains in qualified state after IQ/OQ— Run tests to check and verify satisfactory performance of the

instrument

— Preventive maintenance and repairs

— Maintain an SOP for operation/calibration/maintenance

Performance of PQ— Perform at specified intervals (Same tests to generate history of

instrument performance)— Can perform holistic (preferred) or modular— Specifications for tests can be broader than for OQ

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Timing and Applicability

DQ IQ OQ PQ

Prior to purchaseof a new type ofinstrument

At installation ofeach instrument(new, old or existing unqualified)

After installation or major repair of each instrument

Periodically at specified intervals for each instrument


Qualification Activities

DQ IQ OQ PQAssurance of vendor’s DQ

System description Fixed parameters Preventive maintenance and repairs

Assurance of adequate support availability from manufacturer

Instrument Delivery SOPs – operation,calibration and maintenance

Instrument’s fitness for use in laboratory

Utilities/facility/environment

Network and data storage Secure data storage, backup and archive

Assembly and installation

Installation verification Instrument Function Tests

Performance Checks

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Software Validation

Software for analytical work classified into the following categories

— Firmware (low-level software for integrated chips in instruments)o Considered a component of the instrument itself o Qualified at the user site simultaneously with qualification of

instrument

— Instrument control, data acquisition and processing softwareo Hardware and software are inextricably intertwined and both are

necessary for the operation of the instrument o Manufacturers perform the validation and provide users a summaryo User perform holistic qualification involving entire instrument and

software system through AIQ process


Software Validation (Contd.)

—Stand-alone software (e.g. LIMS)o An authoritative guide for validating software is available:“General Principles of Software Validation”, Final Guidance for

Industry and FDA Staff, Jan. 11, 2002o Software developer specifies the development model appropriate

for the softwareo Validation takes place in a series of activities planned and

executed through various stages of development cycleo User site testing is performed as an essential part of the

software development cycleo User testing, though essential, is only a part of the full validation

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Change Control

Changes are inevitable as new features are added and corrections are madeFollow DQ/IQ/OQ/PQ classification process— DQ: Review the change; adopt only useful or necessary changes— IQ: Install the changes

— OQ/PQ:

o Revise tests and specifications as necessitated by the change

o Change SOP as necessary

o Perform changed OQ or PQ tests


AIQ Documentation

Static Documents: — Obtained during DQ, IQ and OQ phases

— For multiple instruments place common documents in one binder or section and instrument specific documents in separate binders or sections

Dynamic Documents:— Obtained during OQ and PQ phase during instrument

maintenance or performance check

— Provide a running record of instrument performance

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Instrument Categories for AIQ

— A single set of principles to qualify the wide variety of instruments is scientifically inappropriate

— Three categories of instruments are suggested

— Users are most qualified to establish the level of qualification needed

— Exact category of an instrument should be determined by the user for their specific instrument or application

— Modern Laboratories typically have a suite of instruments from simple to complex automated


Instrument Categories for AIQ :Group A

Conformance to user requirements determined by visual observations – No independent qualification required

Example instruments:

o Magnetic Stirrers

o Nitrogen evaporators

o Ovens

o Vortex mixers

o Spatula

o Glass pipettes

o Mortar and pestle

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Instrument Categories for AIQ :Group B

Conformance to user requirements is generally unambiguous and established according to instrument’s SOPs. Causes of failure are discernible by simple observations.

Example Instruments:

o Balances

o pH meters

o Variable pipettes

o Refrigerator/Freezers

o Thermometers

o Viscometers

o IR spectrometers

o Titrators

o Melting point apparatus


Instrument Categories for AIQ :Group C

Conformance is complex, highly method specific and the conformity bounds are determined by their application. A full qualification process applies.

Example Instruments:

o HPLC

o Mass spectrometers

o GC

o Diode-array detectors

o Atomic absorption spectrometers

o Electron Microscope

o Micro-plate readers

o Thermal gravimetric analyzers

o X-ray fluorescence spectrometers

o Elemental analyzers

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Roles and Responsibilities


Users

Users include the analysts and management— Are most familiar with their analysis

— Ultimately responsible for AIQ

— Can contract out parts of AIQ, but are still responsible

— Must keep instrument qualified through PQ

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Quality Assurance (QA)

—QA role remains as in any other regulated study

—Ensure that AIQ meets regulatory requirements

—Have users attest to the scientific validity of AIQ


Manufacturer

— Perform DQ when designing instrument

— Validate the software and hardware

— Provide summary of their validation work

— Provide results of final test on hardware

— Should provide critical functional test scripts for qualification of instrument/software at the user site

— Make available any software/hardware bugs found after release

— Support installation, training and service

— Be open to audit by users

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AIQ Conclusions

Use the term “Qualification” instead of “validation” for instruments

AIQ provides a “level of confidence” to users that the instrument is suitable for its intended use

AIQ is only one component in delivering reliable and quality data

Use DQ/IQ/OQ/PQ model with specific definitions and scope for each qualification stage

White paper is available on FDA docket, and AAPS Pharm SciTech (AAPS website).

AAPS PharmSciTech. 2004; 5(1): article 22.


Affect on Your Laboratory

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What does AIQ mean to You?

AIQ is formalizing a long used process….but:

It is an opportunity to revisit what you are doing— Evaluate using a Scientific Approach

— What value added are you getting from your tests?

— Is there a way to simplify what is being done?

Are you doing enough?— Are you re-qualifying on a regular scheduled basis?

— Are you covering the high risk areas?

Are you doing too much?— Does it take you two years to implement a new Data System?

— Do you take measurements on hardware merely because you can?

— Are you re-qualifying more frequently than needed?

— Are you testing things that need not be tested as part of AIQ?

Risk analysis is a good way to decide

©2010 Waters Corporation 50Questions?

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IVIV

Qualification Case StudiesQualification Case Studies


Agenda for Module IV

Determining what is important to test

Some questions asked by equipment users

How various instruments work and the effect on testing

Various approaches in qualifying LC modules and systems

Automating and streamlining the qualification process

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Considerations

As we proceed lets — Consider the importance of the parameters we test

o What value added does the test supply

— Evaluate to what level they need to be tested

o Time to test versus benefit or risk mitigation

— Consider the various approaches we might use to test these

— Gain an understanding how each test and each module under test functions

— Remember that there is no one right way to qualify that an instrument or a system is functioning as intended


Considerations (2)

Remember, in the end, the user must be prepared to answer any questions an auditor may have

Understanding how the equipment functions and how it has been qualified leaves you better prepared to support the process in an audit situation

Qualification testing indicates that a module or system performs as intended

Qualification testing does not indicate suitability for a particular application

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What about Part 11 Requirements?

Electronic Records and Signatures

Recent Changes

Risk Based Approach

Predicate Rules Apply— Laboratory Data

— Equivalency to paper based Systems

Can qualification results be made part 11 compliant?— If the results are treated as chromatography data, they can


Determining what is important to test (continued)

On LC, we are generally concerned with — Accuracy

— Linearity

— Precision

There are multiple ways to get this information— There is no one right way

— The FDA does not dictate the method to use

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Manual SOPs for Qualification

Traditional/Manual SOPs (Qualification Workbooks) are designed to test key product specificationsGenerally, on Liquid Chromatographs, we verify Accuracy, Linearity and Precision where possible (per FDA guideline)

Workbook tests are derived from factory tests for individual modules adapted for use in the fieldThe workbook tests also allow for typical testing error— Errors in class a volumetric flasks— Errors in tests performed by NIST on the test solutions— Errors in meters and temperature probes— Test errors are additive with the product error

Workbooks provide a “high degree of assurance” that product specifications are metThey test the module as a module…not as it will be used in the system so system error is not be taken into accountThe main disadvantages of this form of testing are the — time required— systems not tested similar to the way they will be used


Exploring Ways to Qualify LC Systems

• A new streamlined approach is needed

• A scientific look is taken at the modular tests• Redesigning existing modular tests can simplify the process

• Using the power of the Chromatography Data Software (CDS), the system itself can be used to verify calibration and provide qualification data• Calculations can be performed internally avoiding the need to

deal with external spreadsheets• Automatic reporting and calculations reduces the risk of

human error and interference• Results can be kept on-line and be part 11 compliant

• The concept of Advanced Qualification Technology (AQT) and the Waters Systems Qualification Tool (SystemsQT) is created

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Design Criteria

A design requirement of the SystemsQT was that it could perform all the tests normally performed using a traditional qualification workbook

The system was to be tested the way it will be used

Workbooks were initially developed using tests off the Waters manufacturing floor

SystemsQT was developed with a clean sheet of paper

All procedures were reviewed and new methodologies developed that combined tests wherever possible

New test solutions were incorporated with the aim of minimizing the various chemical kits used


Development

Original factory tests and specifications were developed to test a specific module and only that module

Interference from other modules, solvents, columns and other factors were minimized or eliminated when testing at the modular level

The SystemsQT is designed to test the chromatograph as a system the way it will be used, not as individual modules

New pass/fail criteria had to be developed based on all the factors that could affect the outcome

A number of qualified, characterized systems were used to build tolerance windows around procedures

Pass/fail criteria were empirically derived from qualified systems

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UV Detector Qualification

User question:

I have a UV detector (Variable λ or Photo Diode Array)

I want it qualified

How are all the wavelengths tested for accuracy?

Do we need to test all the wavelengths?

Why or why not?

What happens if I change a lamp?

What if an inspector asks how I know λ is right?— Don’t know?

— A better answer

Did you document it?


UV Detector Qualification

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Wavelength Accuracy (1)

Example Specification

Wavelength accuracy is one of the most important test we run and inspectors frequently ask about this

It is NOT necessary to test every wavelength in a detector’s range to “provide a high degree of assurance” that an absorbance detector is working properly in terms of wavelength accuracy— Not enough suitable standards are available— It would take lots of time

Replicate gratings are highly reproducible

Design of the wavelength algorithms are validated as part of the instrument design qualification (DQ)



For an example, if we are qualifying a Waters Tunable UV detector, the unit has on-board diagnostics that can be used to verify proper wavelength accuracy at every power-up— The user is alerted if a wavelength is found out of specification— The cause could be due to something in the cell

The detector uses wavelengths from both the lamp and from an internal filter that is inserted into the light path. A number of choices are available: — Zero Order diffraction grating energy spike (tunable detectors)

o reflects all wavelengths (used in some older detectors)

— 257 nm erbium nitrate filter absorbance maxima— 379 nm erbium nitrate filter absorbance maxima— 486 nm deuterium energy line (used as anchor for 500nm top of range)

— 487nm erbium nitrate filter (generally not used because of 486nm deuterium line)

— 521 nm erbium nitrate filter absorbance maxima— 656 nm deuterium energy line (anchor wavelength peak)

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The OQ is used as a means of qualifying not only the detector but also the internal diagnostics

Caffeine can be used for qualification purposes and has peaks at 205nm and 273nm for 1nm Slit Band Width

Testing 205nm with Caffeine is about as far as we can get from the 656nm anchor wavelength provided by the lamp and used in the start up calibration— If a wavelength 452nm away from the calibration wavelength

is accurate… those closer should also be accurate

Using the diagnostics and the caffeine solution, we have verified 6 different wavelengths, — 205, 257, 273, 379, 521 and 656nm

Using the power up diagnostics between qualifications allows us to check 257, 379, 521 and 656nm— It is good practice to power down and back up weekly



Absorbance detector test procedures are based on ASTM E 1657-94 *— *Standard Practice for Testing Variable-Wavelength

Photometric Detectors Used in Liquid Chromatography

Qualification of wavelength accuracy in this manner is common practice been audited successfully on many occasions

Knowing about the onboard diagnostics in this case and documenting their use allows you to explain to an inspector how you know the wavelengths are accurate after a lamp change between qualifications

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The test solutions used were tested by NIST with an accuracy of + or- 0.3nm— Wavelength is fixed by the physics of the test solution— Lambda max can change with detector slit bandwidth

depending upon peak shape of each peak in the solution — Concentration of the accuracy test solutions is not critical

The product specification of 1nm is added to the NIST 0.3nm error and rounded up to a pass/fail criteria of +/-2nm because errors are additive and because the detector “bins” the results to the nearest nanometer — The pass/fail criteria in this case becomes +/- 2nm



One way of automating this test is by having the Chromatography Data System make successive injections of the caffeine solution, incrementing the wavelength on each injection and determining the lambda max for the two caffeine peaks— Since there are two detector channels, both 205 and 273nm

can be tested at once— Alternating between 2 wavelengths also verifies λ precision

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0.00

0.30

0.60

0.90

1.20

1.50

1.80

190 210 230 250 270 290 310

Wavelength (nm)

Abs

orba

nce

204.7 nm

272.0 nm

λ Max’ For Caffeine


Absorbance Linearity (1)

Tested by injecting different concentrations of a solution

Caffeine is also a suitable solution for this test— Peaks are symmetrical

Different cell path lengths require different concentration test solutions to test the linear range of the detector— Beer’s law

— The shorter the path length, the more concentrated the solution must be to absorb the same amount of light

A passing detector is generally defined as one where the drop from a theoretical straight line is ≤ 5% at the top end of it’s specified linear range (per ASTM 1657)

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Absorbance Linearity (2)

A line can be drawn from the different injection areas and the straightness of that line or the linearity can be determined by the R2 of the curve

Acceptable pass/fail values are determined by the linearity specification of the detector and the test solutions used

It is impractical to test linearity at every wavelength— Linearity will very at every wavelength due to lamp energy

— Linearity will vary with every mobile phase, diluent and sample

— Qualification provides assurance that the instrument is functioning as intended

— Method validation should be used to test specific samples

o Suitability for intended use


Detector Linearity and Sensitivity

R2 = 0.9993

0.00

0.20

0.40

0.60

0.80

1.00

1.20

1.40

1.60

1.80

0 20 40 60 80 100

Amount Injected

Pea

k H

eigh

t (AU

)

0.000

0.002

0.004

0.006

0.008

0.010

0.012

0.014

0.016

0.018

0.020

Pea

k AU

/Am

ount

% RSD = 3.96

Linearity is R2 of curve

Sensitivity is %RSD Of Peak Heights / Amounts

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Injection Accuracy, Linearity and Precision

Injection Accuracy Linearity and Precision are affected by many factors including:— Mobile phase used

— Degassed state of mobile phase

— Sample diluent used

— Viscosity of the sample

— System dead volume

— Sample loop volume

— Syringe size

— Syringe draw rate

— Number of motor steps per microliter (varies with syringe size)

— Vial septa used

— Injection mode used


Injection Accuracy

While precision is most important, injection accuracy allows for easier movement of an analysis from one system to the next

One way to test is by weighing the aspirated amount of the sample and translating that to volume— Using water for the sample makes this easier at room temps

— Enough weight must be removed from a vial such that the weighing error is not a significant part of the measurement

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Injection Accuracy

An example injection accuracy specification:

The accuracy specification is at 50uL with specific solvents

— Why was the specification written this way?— Six injections are made to remove enough weight (300uL)— The injector is deemed to meet specification if it passes the test— Apparent performance at different injection volumes, with

different solvents, sample loops, syringes, mobile phases columns and samples will vary

— It is not practical to test at all volumeso Method Validation and System Suitability should be used to

demonstrate suitability for intended use


Injection Accuracy

Another way to test, one that can be automated, is by performing actual injections at different volumes to determine the linearity and then using the x intercept as a means to determine the accuracy error

While this may be less metrologic, it demonstrates the injector in the same way that it will be used and provides a high degree of assurance that the injector is operating as intended

The qualification test assures that the syringe drives, needle, valves and injection algorithms are all functioning as intended— Method Validation and System Suitability should be used to

demonstrate suitability for the specific intended use

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Injector Linearity and Accuracy

y = 25000x + 1250

0

200000

400000

600000

800000

1000000

1200000

-1 9 19 29 39 49

volume injected (uL)

area

(uV*

sec)

X-intercept = - 0.05 uL

Linearity = R2 of curve

Accuracy = X Intercept


Injection Linearity

Linearity o Linearity, for this example, is specified from 1 to 100uLo Linearity above 100uL is not specifiedo Injections above 150uL will provide best results with the

2500uL syringe so they can be made in one syringe drawo Making injections at 5uL using a large size syringe and large

volume sample loop is not the best practice because a large loop means more air in the system that will stretch before the actual sample is pulled in

o But, a 250uL sample loop is not large enough for accurate 200uL injections. At least a 330uL loop needed for 400uL of total sample loop volume

o One system configuration is not the best for all injection volume sizes (just like columns!)

o Think about how you will be using your system before choosing the configuration

o Use method validation and system suitability to demonstrate suitability for intended use

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Injection Linearity

Injection linearity testing in the OQ is used as a means of determining if the injector is functioning as expected across the typically used analytical range

The algorithms that determine motor drive functionality and hence linearity are validated as part of the design of the product

It is always best to perform linearity tests with specific compounds, mobile phases, wavelengths, sensitivities, sample loops, syringes and columns to be used— Method Validation and System Suitability

— Demonstrates suitability for intended use


Injection Linearity

A user Question:— An HPLC injector has been qualified using at 5, 10, 20 and 50uL

— Is it still linear at 200uL?

The answer is “it depends” what you mean— By design, the motor will step the proper amount of steps and the

syringe will move the specified amount so in that sense, it will be linear but:

— Physics will get in the way to make the actual results less linear

o Any injection volume much over one half the sample loop volume will start to disperse out of the loop

o A 200uL injection will require a larger sample loop and that will allow more air in the system. That air stretches when aspirating an injection. This will particularly affect small injection volumes

o A 200uL injection with a standard syringe will require multiple syringe draws and valve movements…this will affect accuracy and linearity

o Mobile phase and degassing used will affect accuracy and linearity

o The column used will affect accuracy and linearity

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Column Effects

A rule of thumb for columns

Do not inject more than 5% of the column void volume

To inject 200uL into a column, the column should have a 4mL void volume— Otherwise the sample diluent becomes the mobile phase

Injecting any more than that alters the chemistry and will make the analysis non-linear

A 4.6 x 150mm column has a void volume of only ≈2.7 mLs

Using a 4mL void volume column will mean waiting about an hour for the peaks to come out or a much higher flow rate

If you are using a system in a non-standard configuration, you can qualify using the standard column and loop, then perform PQ and/or System Suitability with the column and loop to be used for the specific analysis


UPLC Flow Rate Linearity

y = 0.8333x + 0.0417

0.0000.200

0.4000.600

0.8001.000

1.2001.400

1.6001.800

-0.1 0.2 0.5 0.8 1.1 1.4 1.7 2

flow rate (mL/min)

1/vo

id ti

me

X-intercept = 0.050 mL/min.Linearity =R2 of curve

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HPLC Flow Linearity and Accuracy


Flow Rate Accuracy

HPLC specification example: A column is in place when using the SystemsQTBecause of this, flow rates are limited to the capabilities of the columnFor HPLC, flow rates are tested at 0.5, 0.75, 1 and 1.5, the most common range used by users with analytical columnsFlow rates across the entire range are validated as part of the design qualification (DQ) for the productIf the HPLC passes at the flow rates tested, there is a “high degree of assurance” that it will work within specification at all flow rates (i.e. very low risk of failure)For the most part, flow error is fixed by the check valve size and designThe fixed error of a properly functioning check valve shows up as a higher percentage of total flow at lower flow rates

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System Precision

%RSD of6 ReplicateInjections


Gradient Proportioning and Compositional Accuracy

If you use two high pressure pumps (or a binary pump) to create gradients and they use a structurally validated gradient algorithm, so long as the flow rates on both pumps test accurately at multiple flow rates (flow rates are linear), gradients will be accurate and precise.

If you use low pressure mixing on a system that has a structurally validated gradient algorithm, generally any gradient inaccuracy is caused by improper operation or sticking of the Gradient Proportioning Valves (GPVs) so focusing on testing these is one way to assure proper gradient performance— If the valves remain unused for a long time (usually the C or D

valves), they can become sluggish or sticky— Exercising them before use or qualification in such cases is

advised

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Gradient Performance

SD of RetentionTime from each of 7 Peaks overSix Injections

Note: In this UPLC case, there are two high pressure pumps.

SD must be < or = 1.0 sec


Compositional Accuracy

Note: In this case, we are testing one pump with low pressure mixing capability of 4 solvents

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Demonstrating “Fit for Intended Use”

A PQ based on specific user and method requirements

Performed by the user AFTER the vendor supplied IQ,OQ and vendor PQ (system level OQ).

Demonstrates that the instrument system can operate reliably under routine, minimum, and maximum operating conditions. Fit for intended use.

Recommended combination of user requirements:— Method specific system suitability

o Precision of quantity of analite injected

o Peak resolution

o Peak tailing

o Column efficiency

— Method specific concentration standards

— Calibration check


FDA Requirements

Vendor Supplied IQ, OQ and vendor PQ (system level test)— Works according to manufacturer’s specifications

PQ based on User Requirementso Demonstrates Fit for Intended Use

Ongoing system calibration (qualification) and documented scheduled maintenance

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Advantages of CDS Managed Qualification

Accurate Qualification Testing and Analysis— Less opportunity for human error

— Measures peak areas, peak heights and retention times accurately and consistently

— Custom field calculations and regression analysis

— Testing consistent from system to system

— Reduces time that system is off-line by about half

— Qualifies software and systems in analytical configuration


Advantages of CDS Managed Qualification (cont)

Secure and Auditable Qualification Data— 21 CFR Part 11 Compliant Qualification Data

— On-Line Qualification Documentation for Easy Inspection

— Easy tracking and trending of qualification results

— Audit trails and change control part of the data system

— No need for external spreadsheets or third party software

— Secure data environment

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Summary

There are many ways to qualify LC systemsAdvanced technology can save time in your laboratorySystemsQT is the Next Generation in automated Instrument QualificationIt enhances qualification testing and data management in an Empower Chromatography Data System environmentIt offers a whole system test approach for a more accurate indication of overall system-level complianceOperates on a simple wizard built into Empower using Run SamplesBuilt into Empower – no separate softwareto install and uninstallProjects to qualify new modules/systemscan be easily addedDocument review is substantially reduced


Questions?