Thermo Scientific UltiMate User manual

M1393R0

UltiMate

™

Capillary- and Nano HPLC Systems

Operational Qualification

and

Performance Qualification

Operating Instructions

P/N 163960

www.lcpackings.com

Page 2 of 60 M1393R0 OQ/PQ Operating Instructions

The material included in this manual is provided to assist authorized personnel in

performing operation qualification (OQ) and performance qualification (PQ) on the LC

Packings UltiMate Capillary and Nano HPLC system. It is assumed that the individual using

this manual has sufficient training in the use of analytical instrumentation and is aware of

the potential hazards including (but not limited to) electrical hazards, chemical solvent

hazards, exposure to UV radiation and the exposure to pressurized solvents.

All information in this manual is subject to change without notice and does not represent a

commitment on the part of LC Packings, BV.

No part of this manual may be reproduced or transmitted in any form or by any means

without the written permission of LC Packings, BV.

The following are Registered Trademarks of LC Packings - A Dionex Company: Probot,

µCarrier, UltiMate, UltiChrom, FAMOS, Thermos, Switchos , Acurate, UZ-View,

UV-Booster, FluoBoost, Pepmap, µ-Dumper, µ-Fluidics, Fusica, Nano Series, Pico Series,

µ-Guard, µ-Precolumn, Nano-Precolumn.

Dionex™ is a Trademark of Dionex Corporation.

CHROMELEON®is a Registered Trademark of the Dionex Corporation.

Analyst® is a Registered Trademark of Applied Biosystems/MDS Sciex.

HyStar™is a Trademark of Bruker-Daltonics Group.

MassLynx™is a Trademark of Waters® Corporation.

Xcalibur® is a Registered Trademark of ThermoFinnigan.

Windows® is a Registered Trademark of Microsoft Corporation.

PEEK™ polymer is a Trademark of Victrex plc.

Other product names or company names mentioned herein may be the trademarks of their

respective owners.

Printed in the Netherlands.

OQ/PQ Operating Instructions M1393R0 Page 3 of 60

Warnings

The Danger sign, Warning sign and the Caution sign shown below are included in various

locations in this manual or in the manuals provided with the instruments which are to be

tested. These signs provide the following information:

DANGER Danger: The information in a danger statement relates to a procedure, practice condition or

action that if not done correctly or adhered to could lead to personal injury or loss of life.

WARNING Warning: The information in a warning statement relates to a procedure, practice,

condition or action that if not done correctly or adhered to could lead to severe injury

and/or damage or destruction to parts or all of the equipment.

CAUTION Caution: The information in a caution statement relates to a condition that could lead to

damage to equipment and/or lead to invalid analytical results.

Note: The information in a note statement relates to important information that should be

read and understood before continuing.

Safety Precautions

Note: The following precautions should be followed to minimize the possibility of personal

injury and/or damage to property.

Note: Make certain that you are familiar with the contents of this manual before working

on the system.

Page 4 of 60 M1393R0 OQ/PQ Operating Instructions

The operator should follow all safety precautions, warnings, etc provided with the

instruments, in addition, please note the items presented below:

1. All components of the system should be plugged into a common power line that is

directly connected to a true ground.

2. Repair or replace faulty power cords and all communication cables.

3. If a leak occurs, turn off power to the instrument and remedy the situation

immediately.

4. If the mobile phase includes volatile or flammable solvents, avoid open flames and

sparks.

5. Many organic solvents and buffers are toxic. Make certain that you know the

toxicological properties of all mobile phases that you are using.

6. The toxicological properties of many samples may not be well known. If you have

any doubt about a sample, treat it as if it contained a potentially harmful substance.

7. Wear protective eye goggles when handling mobile phases or operating the

instrument. An eye wash facility and a sink should be close to the unit. If any mobile

phase splash on the eyes or skin, wash the affected area and seek medical attention.

8. Dispose of all waste mobile phase in an environmentally safe manner that is

consistent with all local regulations. Do not allow flammable and/or toxic solvents to

accumulate. Follow a regulated, approved waste disposal program. Never dispose

flammable and/or toxic solvents through the municipal sewage system

9. Wear protective eye goggles when handling fused silica tubing (i.e. installation,

cutting etc.)

10. If a buffer is used as a part of the mobile phase, flush the system with several

volumes of a methanol/water (50/50) solution before it is shut down. This will

prevent salt buildup inside the unit.

11. Do not use the instrument in ways other than those indicated in the instructions

given in this manual.

WARNING Warning: The OQ/PQ kit (P/N163932) contains a chemical or chemicals known to the State

of California to cause cancer and/or birth defects or other reproductive harm. For additional

information, consult the product Material Safety Data Sheet (MSDS).

OQ/PQ Operating Instructions M1393R0 Page 5 of 60

Table of Contents

Warnings ................................................................................................. 3

Safety Precautions .................................................................................... 3

CHAPTER 1 Introduction .........................................................................7

1.1 The Purpose of OQ/PQ.................................................................... 7

1.2 Defining the Limits ......................................................................... 7

1.3 General Notes and Recommendations ............................................... 8

1.4 How this Manual is structured ......................................................... 8

1.5 For Additional Information ............................................................... 8

CHAPTER 2 Requirements for a Successful OQ/PQ .....................................9

2.1 Overview ...................................................................................... 9

2.2 Checks and Acceptance Limits....................................................... 10

CHAPTER 3 Process ............................................................................. 13

3.1 Required Materials ........................................................................ 13

3.2 Preparations ................................................................................ 15

3.2.1 CHROMELEON® Setup ................................................................ 15

3.2.2 System Setup ............................................................................ 18

3.3 Performing the Checks .................................................................. 21

3.4 Evaluating the Tests ..................................................................... 22

3.4.1 Repeating Tests ......................................................................... 22

CHAPTER 4 Test Procedures.................................................................. 23

4.1 Overview .................................................................................... 23

4.2 Lamp Intensity of the UV Detector ................................................. 24

4.2.1 Performing the Lamp Intensity Test .............................................. 24

4.3 Wavelength Check ....................................................................... 25

4.3.1 Performing the Test.................................................................... 25

4.4 Flow Cell Check ........................................................................... 26

4.4.1 Performing the Intensity Test of the Flow Cell ............................... 26

4.5 UltiMate Fluid Path Test ................................................................ 27

4.5.1 Performing the Test.................................................................... 27

4.6 Baseline Noise and Drift Test of the UV Detector and Oven Test........ 28

4.6.1 Drift and Noise........................................................................... 28

4.6.2 Theory of the Noise and Drift Calculation of CHROMELEON ............ 29

4.6.3 Oven accuracy........................................................................... 29

Table of Contents

Page 6 of 60 M1393R0 OQ/PQ Operating Instructions

4.6.4 Performing the UV Detector Drift and Noise Test and the Oven Test. 30

4.7 Linearity of the UV Detector...........................................................31

4.7.1 Performing the UV Detector Linearity Test ..................................... 32

4.8 Reproducibility of the Injection Volume ............................................ 33

4.8.1 Performing the Test .................................................................... 33

4.9 Linearity of the injection ................................................................ 34

4.9.1 Performing the Test .................................................................... 34

4.10 Gradient Accuracy ........................................................................ 35

4.10.1 Performing the Test .................................................................... 37

4.11 Switchos Fluid Path Check ............................................................. 38

4.11.1 Performing the test .....................................................................38

4.12 Switchos Flow Rate and Pressure Stability Test................................ 39

4.12.1 Performing the test .....................................................................39

4.13 Switchos Valve Position Check ....................................................... 40

4.13.1 Performing the Test .................................................................... 40

4.14 Completing and Printing the OQ/PQ Report ......................................41

4.14.1 Result of the Gradient Accuracy Test ............................................ 41

4.14.2 Completing the Cover Page and Printing ........................................ 42

CHAPTER 5 Troubleshooting ................................................................. 43

5.1 Overview ..................................................................................... 43

5.2 Sample and Mobile Phase Considerations......................................... 43

5.3 Probable Causes and Solutions ....................................................... 44

CHAPTER 6 CHROMELEON®Listings ...................................................... 45

6.1 Overview ..................................................................................... 45

6.2 Wavelength Check (NAN Configuration)........................................... 46

6.3 Noise and Drift Test (NAN Configuration).........................................47

6.4 UV Linearity (NAN Configuration).................................................... 49

6.5 Autosampler Reproducibility (NAN Configuration).............................. 51

6.6 Autosampler Injection Linearity (NAN Configuration) .........................53

6.7 Gradient Accuracy Test (NAN Configuration).................................... 55

6.8 Switchos Pump Test (NAN Configuration) ........................................ 57

6.9 Stop Flow .................................................................................... 59

OQ/PQ Operating Instructions M1393R0 Page 7 of 60

CHAPTER 1 Introduction

1.1 The Purpose of OQ/PQ

An increasing number of standards and official regulations require that the end user is able

to provide evidence that the instrumentation used for analytical work is working in a

satisfactory manner. In the same manner, quality management according to ISO 9000

(and similar standards) requires that the user monitor and document the ability of the

equipment to obtain valid data on a periodic basis. This manual provides a detailed series

of procedures to perform these Operational Qualification (OQ) and Performance

Qualification (PQ) protocols on the LC Packings UltiMate system components.

The validation procedures described herein are designed to demonstrate that the

instrument was working in an acceptable manner using the standards provided on the day

of validation. It is likely that the instrument performance will vary over time due to small

changes in the various components in the instrument and the validation protocol should be

performed on a periodic basis. The frequency of validation is dependent on the level of

usage of the system and the degree of tolerance that is acceptable for the system.

1.2 Defining the Limits

According to ‘The development and application of guidance on equipment qualification of

analytical instruments’ of P. Bedson and M. Sargent [Accred. Qual. Assur. (1996) 1: 265 -

274] the following definitions apply:

•Operational Qualification (OQ) – The purpose of the Operational Qualification is to

demonstrate and document that an analytical system functions according to its

specifications when specific environmental conditions are taken into account. In this

specification, the supplier must define exactly the conditions that must be observed

for the measurement.

•Performance Qualification (PQ) – The purpose of the Performance Qualification is to

demonstrate and document that an analytical system is capable of accurately

measuring the concentration of one or more compounds in a standard sample.

To simplify the overall qualification protocol, the same procedures can be used for both OQ

and PQ, but the tolerances used for Performance Qualification are less restrictive than

those used for Operational Qualification.

Introduction

Page 8 of 60 M1393R0 OQ/PQ Operating Instructions

1.3 General Notes and Recommendations

•After the validation of the different modules, a cytochrome C separation should be

performed according to the LC Packings factory qualification protocol (as described in

the user’s manual).

•For Ultimate Dual Gradient system configurations, the gradient accuracy test

(Section 4.10 ) must be performed on both pumps.

•Channel C & D should only be tested on customer request.

•If the customer ordered a system with two configurations (e.g. a NAN configuration

and a CAP upgrade kit), the OQ/PQ should be performed for the system configuration

that the customer will be using on a routine basis.

•If required, test any other configuration according to the factory qualification protocol

(cytochrome C digest separation).

•After the instrument has been validated via the protocols described in this document,

the analyst should perform a validation of the assay with standards of the compounds

of interest.

1.4 How this Manual is structured

This manual describes the Operation and Performance Qualification for the LC Packings

UltiMate Systems in its different configurations (including the FAMOS Microautosampler

and the Switchos Advanced Microcolumn Switching Unit) and provides the following

information:

Chapter 2: Requirements for a Successful OQ/PQ provides an overview of the parameters

to be tested, a short description of the tests and lists the required acceptance limits for OQ

and the recommended limits for PQ.

Chapter 3: Process lists all required materials, standards and solvents that are necessary

to perform the OQ/PQ test procedures. In addition, it describes how to prepare

CHROMELEON and the UltiMate system to perform the tests.

Chapter 4: Test Procedures provides step-by-step instructions about how to perform and

evaluate the various OQ/PQ tests.

Chapter 5: Troubleshooting discusses how the operator can determine the cause of a

difficulty in the performing of the OQ/PQ.

Chapter 6: CHROMELEON

Listings provides the listings of all CHROMELEON programs

used to perform the QQ/PQ test procedures on an UltiMate system in NAN configuration.

1.5 For Additional Information

For more detailed information about the operation, maintenance or troubleshooting of the

instruments of the UltiMate system or how to use the CHROMELEON software package,

please refer to the documentation provided with these products and to the online help of

CHROMELEON (F1 key).

OQ/PQ Operating Instructions M1393R0 Page 9 of 60

CHAPTER 2 Requirements for a Successful

OQ/PQ

2.1 Overview

The Operational Qualification (OQ) and Performance Qualification (PQ) procedures are

system-specific procedures. The procedures provided with this document apply for the

components of the LC Packings UltiMate Capillary HPLC system listed in TABLE 2-1:

TABLE 2-1 List of UltiMate System Components with available OQ/PQ Procedures

Instrument Version Option

UltiMate [Plus] Nano- and

Capillary HPLC System (b)

Standard / Inert - Flow Sensor

- Without UV Detector (a)

- Manual Injection Valve

UltiMate [Plus] Dual Gradient

Nano- and Capillary HPLC

System (a) (b)

Standard / Inert - Flow Sensor(s)

FAMOS Well Plate

Microautosampler

Standard / Inert - Sample Cooling

FAMOS Carousel

Microautosampler

Standard / Inert - Sample Cooling

Switchos II Advanced

Microcolumn Switching Unit

Standard / Inert N/A

Notes: (a) If no UV Detector is installed, an extra detector is required to perform the OQ/PQ procedures.

(b) For test procedures regarding the MIC versions of the UltiMate system, contact LCP.

Note: The OQ/PQ procedure are identical for LC Packings UltiMate and UltiMate Plus

Capillary- and Nano HPLC Systems. When the LC Packings UltiMate system is mentioned,

the reader should assume that the material applies to both systems.

Note: If the system does not include an UltiMate UV Detector, a standalone UltiMate UV

Detector is required to perform the OQ/PQ procedures.

The instruments should be controlled by CHROMELEON® 6.6 SP1 or higher (previous

versions will have compatibility problems with the CHROMELEON report file). All necessary

CHROMELEON programs and sequences are provided on the CD ROM ‘IQOQPQ on

UltiMate™(Plus) Systems’ (P/N 163935). If a different software package (e.g. Xcalibur™,

Analyst™, HyStar™, MassLynx™, UltiChrom™, etc.) is used to control the instruments, all

programs will need to be prepared manually. Some limitations may apply due to different

or limited control capabilities of these software packages.

Requirements for a Successful OQ/PQ

Page 10 of 60 M1393R0 OQ/PQ Operating Instructions

2.2 Checks and Acceptance Limits

TABLE 2-2 lists all OQ/PQ test procedures that are to be performed in the order they must

be performed. In most instances, it is necessary that a test is passed before the next test

in the overall is attempted. As an example, if the ‘Linearity of the UV Detector’ test is

failed (Section 4.7), the result of the ‘Gradient Accuracy’ test will be questionable (Section

4.10).

TABLE 2-2 List of OQ and PQ Test Procedures to be performed

Test Procedure Performed Section

Lamp Intensity of the UV Detector Manually 4.2

Wavelength Check CM Sequence 4.3

Flow Cell Check Manually 4.4

UltiMate Fluid Path Test Manually 4.5

Baseline Noise and Drift Test of the UV Detector CM Sequence 4.6

Linearity of the UV Detector CM Sequence 4.7

Reproducibility of the Injection Volume CM Sequence 4.8

Linearity of the injection CM Sequence 4.9

Gradient Accuracy CM Sequence 4.10

Switchos Fluid Path Check Manually 4.11

Switchos Flow Rate and Pressure Stability Test CM Sequence 4.12

Switchos Valve Position Check Manually 4.13

Note: According to GLP, a test procedure that failed needs to be repeated and all test

procedures following the one that failed must be repeated.

TABLE 2-3 presents an overview of the parameters to be tested and a short description of

the tests. In addition, it presents the required acceptance limits for OQ and the

recommended limits for PQ.

TABLE 2-3 Overview of the OQ and PQ Test Procedures and Limits

Limits (a)Instrument Parameter Description

OQ PQ

Wavelength

accuracy

External holmium filter is required. ± 2 nm ± 2 nm

Lamp Intensity

(Section 4.2)

The dummy flow cell is installed

and the lamp intensity is read

from the detector SIGNALS

screen.

0.4 <SIG <0.9

0.1 <REF <0.9

0.4 <SIG <0.9

0.1 <REF <0.9

Flow cell

transmittance

(Section 4.4)

(b)

The flow cell is installed and filled

with water. The transmittance of

this flow cell is indicated by the

‘SIG’ value and is read from the

detector SIGNALS screen.

> 15%

of reference

intensity

> 15%

of reference

intensity

Baseline drift

(Section 4.6)

< 4000 µAU/hr

Baseline noise

(Section 4.6)

The drift and noise are recorded

for 21 minutes at 254 nm with a

flow cell (filled with mobile phase

A). < 50 µAU < 50 µAU

UltiMate

UV Detector

(Note: If the

system does

not include an

UltiMate UV

Detector, a

standalone

UltiMate UV

Detector is

required to

perform the

OQ/PQ test

procedures).

Oven test

(Section 4.6)

(d)

An external thermometer is used

to measure the oven temperature.

Accuracy

+/- 1°C

Accuracy

+/- 1°C

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