Unresolved Peaks

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Unresolved Peaks

Unresolved Peaks

The following data is extracted from three experiments performed by scientists at the R. W. Johnson Pharmaceutical Research Institute.

Experimental Conditions
Sample	Conc: 2.0 mg/ml	Size: 12 ml
Wavelength	LP: 670 nm	UV: 254 nm
Column	Type: Chirobiotic-T	Size: 150 x 4.6 mm
Mobile Phase	A=H₂O / B=MeOH	Flow: 0.75 ml/min
Experiment 1	70% A / 30% B	R_s = 1.4
Experiment 2	80% A / 20% B	R_s = 0.8
Experiment 3	100% A / 0% B	R_s = 0.4

Chromatographic separations of L- and D-phenylalanine with the different mobile phases, listed in the table above, were run to determine the degree of chromatographic resolution as shown here.

Next, comparisons of the responses between a UV detector and the PDR-Chiral Advanced Laser Polarimeter were made at reasonably good chromatographic resolution (R_s = 1.4). The following example indicates that with good resolution the UV detector has a better MMQ than the laser polarimeter.

However, in the following separations (shown below), as the resolution degrades, the detection limit for the polarimeter improves relative to that of the UV detector. The first set of chromatograms are at a resolution of 0.8 while the second set are at 0.4.

In the two tables below we have provided a summary of the correlation coefficients for the Chiral and UV detectors' responses, respectively. From this data, as expected, we see that using peak area will offer better quantitation than simply using peak height. And this is true for both detectors.

Correlation Coefficient with Chiral Detector
Mobile Phase	L Enantiomer		D Enantiomer
Mobile Phase	Pk Ht	Pk Area	Pk Ht	Pk Area
70/30	999	.999	999	.999
80/20	.998	.997	.998	.993
100/0	.978	.989	.966	.976

Correlation Coefficient with UV Detector
Mobile Phase	L Enantiomer		D Enantiomer
Mobile Phase	Pk Ht	Pk Area	Pk Ht	Pk Area
70/30	999	.999	998	.999
80/20	.998	.998	.999	.999
100/0	.979	.999	.994	.999

In summary, the data presented above clearly shows that the PDR-Chiral Advanced Laser Polarimeter, when used as a screening tool, can save valuable time and money by eliminating time-consuming chromatographic runs to achieve baseline resolution during method development.