CombiFlash Torrent® User Manual - Isco
CombiFlash Torrent® User Manual - Isco
CombiFlash Torrent® User Manual - Isco
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Section 4 <strong>User</strong> Help Reference<br />
4.4.7 Monitoring the Wavelength Ratio<br />
When using two peak detection options, such as 1 with 2 or<br />
All Wavelength Detection with External, a ratio of the<br />
detection sources can also be displayed, which at times can<br />
provide the best indication of compound purity. Refer to the<br />
following discussion.<br />
If a pure compound is eluting, the absorbance is linearly<br />
related to the concentration of the compound in the solvent. If<br />
the compound absorbs differently at different wavelengths,<br />
the absorbance at each wavelength may be different, but still<br />
linearly related. For example, assume a compound eluting<br />
from the module has an absorbance equal to 2 times the<br />
concentration at 254 nm. This same compound at 220 nm has<br />
an absorbance of 1.5 times the concentration. The ratio of<br />
these signals will be 1.33. Since the relationship of absorbance<br />
to concentration is not variable, the ratio remains steady while<br />
the concentration changes from the beginning to the end of the<br />
peak. During the duration of the peak, the ratio will be 1.33<br />
and this constant value is displayed as a horizontal line.<br />
Now assume a case where there is a second compound eluting,<br />
only slightly shifted in time from the original compound. It is<br />
possible that the detection absorbance trace alone would<br />
indicate a single, valid chromatographic peak. In reality, it is a<br />
combination of two peaks. By monitoring a second wavelength,<br />
it may be possible to reveal the second compound. Because of<br />
the slight shift in time and the different absorbance properties<br />
of the two compounds, the changing ratio during the detected<br />
peak would reveal the impurity. Therefore, one can therefore<br />
assume that if the ratio is not constant for the entire duration<br />
of the peak, the compound eluting may not be pure.<br />
To display the ratio, open the METHOD EDITOR [4.3.2] window<br />
and select the Show Ratio option.<br />
4.4.8 Bypassing the Solid Load Cartridge<br />
Occasionally, impurities precipitate in the solid load cartridge<br />
as compounds are purified. This may cause high back<br />
pressures resulting in reduced flow rates and long run times.<br />
Clicking this button changes the injection valve position,<br />
which removes the solid load cartridge from the solvent path<br />
and relieves the back pressure.<br />
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