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Topics
in Liquid Chromatography:
Collection of Open
Acess materials on the web אוסף של חומר פתוח וחינמי (באנגלית)
באינטרנט
|
Table
of Content תוכן העניינים:
Topics' are marked by color codes הנושאים מסומנים בצבעים שונים:
Search
and/or Development of HPLC Method חיפוש/פיתוח השיטה לעבודה
Where to begin if you
need to perform
HPLC analysis?
First search in the literature for existing
methods, do not re-invent the wheel. Go to Google and search
for
your compound plus the two pharases "mobile phase" and "liquid
chromatography":
Understanding and
selecting the
right HPLC column בחירת הקולונה המתאימה:
Now, you got an application note or a method
from
the literature. Chances are (90%) that you got a reversed
phase
application for your compound. What is next? There is a huge
selection of reversed phase columns, do I need to buy exactly the same
column as the literature shows? Not necessarily, there is a
term
called: equivalent column.
Optimize and validate
the HPLC
method אופטימיזציה וולידציה של השיטה:
After you set up your HPLC method, you need to
test
it on known samples, and you need to validate your method to be sure
that it
will work properly and accurately from now on. The next links
describe
the
process and the reason for optimization and validation of HPLC methods.
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Validation
of HPLC Method for Quantitative Work
ולידציה של השיטה לעבודה כמותית
Instrumentation
Qualification תיקוף מכשירים
Topics related
to the
hardware of HPLC systems
 Committing
to Calibration of Instruments
Wikipedia:
Verification
and
validation
Operational
Qualification of HPLC Instruments:
LCGC North America Vol. 20 No 4 Apr 2002 (Grisanti and Zachowski, Ed:
Krull&Swartz)
The
procurement, qualification and calibration of lab Instruments: An
Overview
(By
Herman Lam, Ph.D.) Laboratory Focus-Gazette Edition, 1999, vol.: 3, no.
3, Page 8
Qualification
of
HPLC
(Scribd)
Equipment
Specifications
for
Liquid
Chromatography -MS/MS (QQQ) with HPLC System
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to Top
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Operational
Instructions and Troubleshooting הוראות תיפעול ואיתור תקלות
Now
that you have a
validated method, you start working daily, now you need to know how to
operate the instrument, chemistry and software smoothly and
trouble-free. For instructions see Operational
Instructions
The following are topics
related to
the proper operation of the HPLC
system, chemistry, fluidics, software, plumbing, etc.
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Preparing
the
Laboratory
for
HPLC
Work
הכנת
המעבדה לעבודה
Running
HPLC analysis
routinely is not a simple task. See instructions
in: Successful
Operation
of
HPLC
System.
The laboratory
must be properly prepared for it. The following links are
some
pointers to manufacturers of HPLC-compatible chemistry and accessories.
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Water
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Buffers
Solvent
and Sample
handling
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Special
Topics and Applications נושאים נבחרים
שונות Miscellaneous
Chiral
Chromatography כרומטוגרפיה כיראלית
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to
Top
Amino
acids Peptides/Proteins חומצות אמיניות, פפטידים וחלבונים
Proein Purification Links קישורים לטיהור חלבונים
Ion
Analysis and Capillary Electrophoresis אנליזה של יונים
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to Top
Gel
Permeation (GPC) or Size
Exclusion
Chromatography (SEC) הפרדת מאקרומולקולות :
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to Top
Clinical
Applications יישומים רפואיים
Bio-chromatography ישומים ביולוגיים
Sample
Preparation Topics נושאים בהכנת דוגמאות
Solid
Phase Extraction
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Introduction
Before beginning to work with
any HPLC
instrument one must
understand the basic principles of liquid chromatography at its various
modes.
The liquid
chromatograph is
very
complex and it consists of sophisticated technology and sometimes
delicate and
total ignorance in the manner it works might sabotage its parts which
are very
expensive. In
addition, there are
chemical processes of distribution between the two phases, the mobile
and
stationary, while the analysis goes on, and there is a need for a
minimum
understanding of their theoretical basis to operate the system
correctly. Also, a
minimum understanding
of the method,
in which the system performs data analysis, is needed so that no false
data or
results will be obtained in the analysis.
LISTS
OF STEPS NEEDED
BEFORE ANY RUN BY HPLC רשימת צעדים מקדימים לפני הרצה :
- Filter
the solvents with membranes with cutoff of 0.22-0.45 mM
- Use
clean and transparent reservoirs through which precipitates and
colloids can be
distinguished.
- Make
sure that the solvents will be easily mixed with the previous solvents
in the
same inlets (A, B, C, D). For
example
methanol or water should not be placed instead of hexane directly, or
any
organic solvent should not be placed directly instead of a buffer
reservoir.
- Degass
the solvents and purge all the tubing that lead to the pump.
- Connect
the column carefully according to the flow direction marked on it (do
not
connect directly to the detector).
- Flow
the appropriate solvents through the column at a low flow-rate (0.1-0.5
ml/min)
or reach the composition gradually using the appropriate gradient.
- Select
the appropriate
wavelength (or
other
type of setting) in the detector and wait for stable baseline
- Prepare
the set of methods in the workstation: Instrument method for the
control on the
system, Processing method for the data processing and the Report method
for the
report of final results.
- When
the system and the methods are ready a blank run should be performed to
test
the system and verify that it is clean from interferences.
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on
this specific page is
my own, some of them are revision of Waters Inc.'s
educational
presentations
)
LITERATURE ספרות מקצועית:
