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Me This website is intended for academic support of HPLC/LC-MS users in Israel first of all. The rest of the world is welcome too.
האתר הזה מוקדש קודם כל למשתמשי כרומטוגרפיה נוזלית בישראל- משתמשים מכל רחבי העולם (פירוט למטה) מתכבדים גם
REVIEW (draft of pre-published chapter):
High
Performance Liquid
Chromatography (HPLC) in the
pharmaceutical analysis
1. Types of HPLC: Reversed Phase chromatography
2. HPLC Theory: System Suitability parameters
3. The Role of HPLC in Drug Analysis
Discovery: High throughput screening and bioanalysis
Development: Method development and validation
Manufacturing: Dissolution, content uniformity, assay, drug impurities and stability, in process and cleaning verification.
4. Specialized HPLC Separations.
Ultra-High-Performance Liquid Chromatography.
Preparative HPLC
Chiral Separations
Introduction
Drug manufacturing control requires high
level and
intensive analytical and chemical support of all stages to ensure the
drug's
quality and safety (1). The
pharmacopeia constitutes a collection of recommended procedures for
analysis
and specifications for the determination of pharmaceutical substances,
excipients, and dosage forms that is intended to serve as source
material for
reference or adaptation by anyone wishing to fulfill pharmaceutical
requirements. The most important
analytical technique used during the various steps of drug development
and
manufacturing is the separation technique: High Performance Liquid
Chromatography (HPLC).
The key to a proper HPLC system operation is
knowledge
of the principles of the chromatographic process, as well as
understanding the
reasons behind the choice of the components of the chromatographic
systems such
as column, mobile phase and detectors. A
scheme of an HPLC system is shown in Figure 1. A
high pressure pump is required to force the
mobile phase through the column at typical flow rates of 0.5-2
ml/min.
The sample to be separated is introduced into the mobile phase by
injection
device, manual or automatic, prior to the column. The detector
usually
contains low volume cell through which the mobile phase passes carrying
the
sample components eluting from the column. There
are books describing the practicality of
HPLC operation (2-11). It is
expected of any proper HPLC system that is used in the pharmaceutical
laboratories to produce highly accurate and precise results, due to
health
related issues of improper measurements.
Every HPLC system must be qualified to comply with the strict
demands
from health authorities for high quantitative performance.
Quality standards in
pharmaceutics
require that all instruments should be adequately designed, maintained,
calibrated, and tested. The approach
that has been adopted in the environment of the analytical instrument
has
become known as the "Four Qs": design qualification (DQ),
installation qualification (IQ), operational qualification (OQ), and
performance qualification (PQ). Design qualification is
performed at the
vendor’s site, and it is representative of the way an instrument is
developed
and produced, usually governed by International Organization for
Standardization (ISO) criteria.
The installation
qualification (IQ)
process can be divided into two steps: pre-installation and physical
installation. During pre-installation, all information relevant to
the
proper installation, operation, and maintenance of the instrument is
checked.
Workers confirm the site requirements and the receipt of all of the
parts, pieces,
and manuals necessary to perform the installation of the specific HPLC
unit.
During physical installation, serial numbers are recorded and all
fluidic,
electrical, and communication connections are made for system
components.
Documentation describing how the instrument was installed, who
performed the
installation, and other various details are archived.
Figure 1: A Scheme of an HPLC System
The operational
qualification process
ensures that the separate modules of a system (pump, injector, and
detector)
are operating according to the defined specifications such as accuracy,
linearity, and precision. Specific tests are performed to verify
parameters
such as detector wavelength accuracy, flow rate, or injector precision.
The performance
qualification (PQ) step
verifies system performance as a whole. Performance qualification
testing is
conducted under real operating conditions in the analytical laboratory
that is
going to be using the instrument. In practice, sometimes operational
and
performance qualification blend together, particularly for linearity
and
precision (repeatability) tests, which can be conducted more easily at
the
system level.
The performance
qualification test of
the HPLC system uses a method with a well-characterized analyte
mixture,
column, and mobile phase. It incorporates type of measurements from the
system
suitability section of the general chromatography chapter <621>
in the U.S.
Pharmacopeia (12).
In the end of the process proper documentation is archived.
1. Types of HPLC: Reversed Phase chromatography
2. HPLC Theory: System Suitability parameters
3. The Role of HPLC in Drug Analysis
Discovery: High throughput screening and bioanalysis
Development: Method development and validation
Manufacturing: Dissolution, content uniformity, assay, drug impurities and stability, in process and cleaning verification.
4. Specialized HPLC Separations.
Ultra-High-Performance Liquid Chromatography.
Preparative HPLC
Chiral Separations
Conclusion
HPLC technology has matured to the extent
that almost
any existing organic compound can be analyzed by an existing method
that can be
found in the analytical literature, such as professional journals,
protocol
books such as Pharmacopeia's or AOAC manuals.
The most remarkable change in the
pharmacopoeias in the past 25 years has been the increasing importance
of HPLC
technology in the analysis of all aspects of drug development and
manufacturing.
References
1. Velagaleti, R., Burns, P., and Gill, M. (2003) Drug Information Journal 37, 407-438.
2. Dong, M. W. (2006) Modern HPLC for practicing scientists, John Wiley & Sons, New Jersey.
3. Rochet, J. C. (2006) American Journal of Pharmaceutical Education 70.
4. Katz, E. (1998) Handbook of HPLC, CRC Press.
5. Farb, D., Luttrell, A., and Kirsch, R. (2005) Pharmaceutical Quality Control Lab Guide book, University Of Health Care.
6. Wellings, D. (2006) A Practical Handbook of Preparative HPLC, Elsevier Science.
7. Winslow, P., and Meyer, R. (2004) Compliance Handbook for Pharmaceuticals, Medical Devices, and Biologics.
8. Hage, D. S. (2006) Handbook of Affinity Chromatography, Taylor & Francis Group.
9. Walker, J. M. (2002) The Protein Protocols Handbook, Humana Pr Inc.
10. Ahuja, S., and Dong, M. W. (2005) Handbook of pharmaceutical analysis by HPLC, Elsevier Academic Press.
11. Ahuja, S., and Scypinski, S. (2001) Handbook of Modern Pharmaceutical Analysis, Academic Press.
12. USP-NF (2005) United State Pharmacopeia National Formulary, United State Pharmacopeial Convention, Inc.