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Ultra High Performance Liquid Chromatography (UHPLC):

Quality Operations Laboratory

Jack Leary, PhD

Global Quality Operations

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Why is the implementation of new technology important for

Pharmaceutical ualit O erations laboratories?

Productivity improvements accelerate speed of labdecisions, which translates to less idle time for

manufacturing equipment

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Why is the implementation of new technology important for

Pharmaceutical ualit O erations laboratories?

Productivity improvements accelerate speed of labdecisions, which translates to less idle time for

manufacturing equipment

For the phamaceutical industry, how is new technology

defined in the anal tical and re ulator arena?

Depends on frame of reference

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Why is the implementation of new technology important for

Pharmaceutical ualit O erations laboratories?

Productivity improvements accelerate speed of labdecisions, which translates to less idle time for

manufacturing equipment

For the phamaceutical industry, how is new technology

defined in the anal tical and re ulator arena?

Depends on frame of reference

One example of new technology is Ultra High Performance

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What is UHPLC?

Ultra High Performance Liquid Chromatography (UHPLC)

uses:

Chromatographic columns with smaller diameter particles

S stem ressures which are reater than t ical Hi h

Performance Liquid Chromatography (HPLC), e.g. > 6000

psi

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A Technique with Many Names:

Ultra High Performance Liquid Chromatography (UHPLC)

Ultra High Pressure Liquid Chromatography (UHPLC)High Speed Liquid Chromatography

Fast Liquid Chromatography

Ultra Performance Liquid Chromatography (UPLC) WatersRapid Resolution LC System (RRLC) Agilent

treme qu romatograp y - asco

Accela High Speed Chromatography System - Thermo

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Effects and Potential Benefits of sub 2 um particle size and

hi her column ressures:

Decreased run timeImproved sensitivity

Enhanced peak resolution

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Development of UHPLC Method or Conversion of Existing

HPLC Method

UHPLC is based upon the same principles of HPLC.However, since the column packing material is of smaller

diameter, modification of the chromatographic conditions

.

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Development of UHPLC Method or Conversion of Existing

HPLC Method

When converting an HPLC method to UHPLC, use of thesame type of column/packing material is recommended, when

possible, with the exception of the particle diameter and the

.

with 0.20 um filter, use a titanium stone, and install

recolumn filters to minimize column blocka e from

particulates.

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UHPLC Flow Rate

e ow rate o met o s genera y range rom .

0.75 mL/min, but are dependent upon the column.

Focus on system pressure is important. Currently, Agilent

RRLCs can only be used up to a maximum of ~9000 psi whileWaters UPLCs can be used up to 15000 psi. Therefore, it is

mpor an o cons er e ype o ns rumen a on a w e

used at all the site(s) running the UHPLC method being

method.

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UHPLC Injection Volume:

Typical injection volumes range from 0.3 0.8 L, but is

dependent upon peak shape and peak intensity.

Although injection volumes for UHPLC are less than for

reason for the improved sensitivity is because of the greaterefficiency and tighter eluting bands.

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Other UHPLC Recommendations:

The dead volume from the column outlet should be kept to a

minimum to reduce band broadening effect

Cell volumes of detectors may need to be reduced for the

reduced or eliminated.

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Sample Gain in EfficienciesHPLC vs UHPLC

Data from Waters Corp.

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Comparison of HPLC and UHPLC Assay& Deg Method for Capsules

Run Time-16min

Deg 2 API 9.01min

Inertsil, ODS-2, 4.6x150mm,5Flow Rate: 0.6mL/min

Inj. Vol: 20L

M.P. 0.05M Potassium PhosphatepH3.5:ACN(45:55)

Deg 13.80min

4.72 min

Run Time-0.4min

1.7 m C18 2.1x50mm

Deg 1 0.15 min

API 0.34min

Flow Rate: 1.5mL/min

Inj. Vol: 0.4LM.P. 0.05M Potassium

Phosphate

pH3.5:ACN(45:55)

Deg 2 0.25min

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HPLC Chromatogram for Cleaning Method (22 min)

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Strategies for Implementing New Technology

Focus on areas of operation where mode of regulatory change

is self-approvable, such as HPLC to UHPLC conversion forCleaning Verification

Allows Control Labs to gain familiarity with instrumentation

Interface the creation of the new technology in tandem withthe ongoing with development and process improvements of

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Pfizer Experience with UHPLC

purchased or have plans to purchase UHPLC systems

Waters & Agilent systems have been more commonly acquired

Local vendor support for introduction of this new technologyhas been productive

Current focus has been on self approvable new technologyextens ons

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Acknowledgements

Jeff Snider

Ed ardo Ortiz

Leon Clark,Scott Afman,

Avish Ramcharan

Prasad KPP