CORRELATION OF QUANTITATIVE-ANALYSIS PRECISION TO RETENTION TIME PRECISION AND CHROMATOGRAPHIC RESOLUTION FOR RAPID, SHORT COLUMN ANALYSIS

When chromatographic analysis involves the use of two or more chromatograms, e.g., for performing calibration or for assessing composition changes among different samples, optimization based solely on chromatographic resolution, R(s), may not yield the most rapid or precise analyses. A simple model is developed that predicts quantitation precision for analysis of ill-resolved peaks as a function of retention time precision and R(s). This model implies that use of shorter columns can provide rapid and precise quantitation. Short-column analyses have improved retention time precision and S/N ratio, which offset the detrimental quantitation effects of decreased R(s). A quantitation precision study for liquid chromatography was done beginning with well-resolved peaks having R(s) = 1.06. Quantitation precision as percentage relative standard deviation (%RSD) was theoretically calculated and experimentally measured for two diverse experimental paths in which R(s) was diminished by decreasing either the selectivity ratio or the column length. The quantitation method chosen was deconvolution of mixture chromatograms by performing a classical least-squares fit to chromatograms of pure standards. The quantitation precision model agreed with calculated and measured %RSD values to within 3 percentage points. The short-column analyses yielded improved quantitation precision and shorter analysis time at equal R(s) compared to selectivity-limited analyses. For an analyte having a peak height ratio of 1:2.5 and a peak width ratio of 1:1.2 relative to an adjacent overlapping peak, similar quantitation precision as %RSD was obtained at equal selectivity ratio for a 50 cm column (3.1%) and a 7.5 cm column (4.3%), despite the decrease in R(s) from 1.06 to 0.27. The short-column advantages are also applicable to gas and supercritical fluid chromatographies and possibly to capillary electrophoresis,