Peak asymmetry factor

Elution volume, exclusion chromatography Flow rate, column Gas/liquid volume ratio Inner column volume Interstitial (outer) volume Kovats retention indices Matrix volume Net retention volume Obstruction factor Packing uniformity factor Particle diameter Partition coefficient Partition ratio Peak asymmetry factor Peak resolution Plate height Plate number Porosity, column Pressure, column inlet Presure, column outlet Pressure drop... 

Band Asymmetry. The peak asymmetry factor AF is often defined as the ratio of peak half-widths at 10% of peak height, that is, the ratio b/a, as shown in Fig. 11.2. When the asymmetry ratio lies outside the range 0.95-1.15 for a peak of k =2, the effective plate number should be calculated from the expression... 

In practice, the calculation of peak skew for highly tailing peaks is rendered difficult by basehne errors in the calculation of third moments. The peak asymmetry factor, A, = b/a, at 10 percent of peak height (see Fig. 16-32) is thus frequently used. An approximate relationship between peak skew and A, for taihng peaks, based on data in Yau et al. is Peak skew= [0.51 -t- 0.19/(A, — 1)] . Values of A, < 1.25... 

FIG. 16-32 Exponentially modified Gaussian peak with Xq/Gq = 1.5. The graph also shows the definition of the peak asymmetry factor at 10 percent of peak height. 

In the elucidation of retention mechanisms, an advantage of using enantiomers as templates is that nonspecific binding, which affects both enantiomers equally, cancels out. Therefore the separation factor (a) uniquely reflects the contribution to binding from the enantioselectively imprinted sites. As an additional comparison the retention on the imprinted phase is compared with the retention on a nonimprinted reference phase. The efficiency of the separations is routinely characterized by estimating a number of theoretical plates (N), a resolution factor (R ) and a peak asymmetry factor (A ) [19]. These quantities are affected by the quality of the packing and mass transfer limitations, as well as of the amount and distribution of the binding sites. 

Having chosen the test mixture and mobile diase composition, the chromatogram is run, usually at a fairly fast chart speed to reduce errors associated with the measurement of peak widths, etc.. Figure 4.10. The parameters calculated from the chromatogram are the retention volume and capacity factor of each component, the plate count for the unretained peak and at least one of the retained peaks, the peak asymmetry factor for each component, and the separation factor for at least one pair of solutes. The pressure drop for the column at the optimum test flow rate should also be noted. This data is then used to determine two types of performance criteria. These are kinetic parameters, which indicate how well the column is physically packed, and thermodynamic parameters, which indicate whether the column packing material meets the manufacturer s specifications. Examples of such thermodynamic parameters are whether the percentage oi bonded... 

The peak asymmetry factor should be scrutinized first. In this discussion we refer to the peak asymmetry factor measured at 10% of the peak height (see section 1.5). Some column supply companies use the baseline measurement to specify peak asymmetry, leading to larger limiting values than those given here. Peak asymmetry, especially of unretained or weakly retained peaks (k < 3), is typical of poorly packed columns (if instrumental contributions can be excluded). If only the unretalned peak (k < 1) is asymmetric and/or there is a significant difference (> 15%)... 

Peak asymmetry factor AF Ratio of peak half-widths at 10% peak height... 

The greater the value of N, the greater is the column efficiency. The peak asymmetry factor should be between 0.8 and 1.8, and is determined as shown in Figure 1.10.1. 

The point at which a column used for HPLC will fail depends largely upon how the operator uses it. Eventually, however, all HPLC columns will fail. The onset of column failure can be monitored by two common failure parameters the peak asymmetry factor, As, and the peak tailing factor. These parameters are defined according to the figure below ... 

Peak Asymmetry Factor (As, 10%) Peak Tailing Factor (5%)... 

Peak asymmetry factor Column efficiency Capacity factor... 

The peak asymmetry factor (tailing factor) can be calculated by several different methods. By the USP,... 

Method for hand-calculating peak asymmetry (peak asymmetry factor CB/AC)... 

Graph of first-approximation relationship between peak skew and the peak asymmetry factor... 

Peak shape is usually expressed by the peak asymmetry (AJ. In Fig. 3, the peak asymmetry factor for substance B is given by... 

Peak Tailing (Peak Asymmetry Factor > 1.2). This is attributed to the wrong pH value, wrong column, wrong sample solvent (mobile phase is better to be used), void volumes at column inlet (the column may need repacking), as well as to active sites within the column which can be solved with the use of a competing basic or acidic modifier. 

Peak Fronting (Peak Asymmetry Factor < 0.9). This indicates that a small band is eluting before a large band, a wrong pH value of the mobile phase is used, an overloaded column, a void volume at the inlet, or that the sample solvent is incompatible with the mobile phase. 

Keeping records of column backpressure and important chromatographic parameters [number of theoretical plates (AO, peak asymmetry factor (As), retention factor k ), resolution factor (Rs)] helps to monitor the required column performance, while storage in the appropriate organic solvent extends column lifetime. Table 8 presents the preventive actions for column protection. 

Fig. 2 (A) Measurement of peak tailing using USP tailing factor and peak asymmetry factor. (B) Peak resolution. (From Merck...

The simplest form of an HPLC SST involves comparison of the chromatogram with a standard one, allowing comparison of the peak shape and the peak width baseline resolution. Additional parameters that can be experimentally calculated to provide quantitative SST report include the number of theoretical plates, separation factor, resolution, tailing or peak asymmetry factor, accuracy, and precision (RSD of six measurements). Resolution may also be combined with a selectivity test to check the resolution of the analytes from components present in the sample matrix. If matrix components interfere with a method, a matrix blank may be included in the SST. Peak shape and asymmetry, or tailing factor, can... 

---