Chemistry Reference
In-Depth Information
blends of multiple organic solvents, multiple buffers and pHs, and operating four
or more columns at different temperatures. This multiparameter mobile phase and
multicolumn capability provides access to many of the potential variables the analyst
needs to investigate when manipulating selectivity in method development. In addi-
tion, using multiple detectors in combination, there is less dependence on individual
analyte properties such as UV chromophores or ionization.
Scouting or screening systems are also often used to identify the most promis-
ing conditions (solvent, column pH, etc.) for further optimization (Section 3.5) or
development. By considering the physical characteristics and analyte properties,
templates (collections of instrument methods) can be written in the chromatography
data system (CDS) to generate the various mobile phase conditions, equilibrate or
switch columns, perform all the chromatographic runs, and run wash or shutdown
procedures for both the columns and the system. These templates are usually written
once and then used repeatedly as new methods need to be developed, allowing the
analyst to run the system in a semi-automated manner. A typical screening experi-
ment might generate in excess of sixteen chromatograms; for example, using four
different columns, high and low pH, and two different solvents (e.g., methanol and
acetonitrile) as outlined in Figure 3.5. Of course, multiple additional conditions gen-
erate a great deal more chromatographic data, and once generated, all the data must
be scrutinized. However, rather than go through each chromatogram individually,
the analyst can use the custom reporting features of the CDS relational database to
generate summary plots that at a glance reveal which runs gave the greatest number
of peaks, most resolution, etc., as illustrated in Figure 3.6.
pH 3, Acetonitrile
pH 3, Methanol
pH 10, Acetonitrile
pH 10, Methanol
C18
Shield
RP18
C8
Phenyl
FIgure 3.5
Example column scouting method development approach. In this example,
four different columns, two different buffers (high and low pH), and two different organic
solvents (acetonitrile (ACN) and methanol (MeOH)) are methodically scouted to generate a
set of conditions that can be chosen for additional optimization if needed.
