Step gradient profile

Once the retention models have been established by the procedures described above, no further experiments are conducted. The software automatically generates tens of thousands of linear and multi-step gradient profiles in a very short time. It predicts the retention of compounds for each gradient profile generated, evaluates the predicted separation using an optimization function, and searches for the best gradient profile using a super-fast Monte Carlo optimization method [5]. 

A pH, B flow rate, C, D, and E buffer eomposition of firsf, second and third step of gradient profile... 

FIGURE 6.24 (a) Isocratic, (b) step gradient, and (c) linear gradient used in the CEC separations of (1) anthracene, (2) pyrene, (3) 1,2-benzofluorene, and (4) benzo[a]pyrene. The injection time was 20 s. The concentrations of analytes 1 1 were 2.8, 0.9, 5.8, and 5.0 M, respectively. The dashed lines show the gradient profiles as seen at the detector [148]. Reprinted with permission from the American Chemical Society. 

Step 6. Sketch out the velocity profiles and velocity gradient profiles and see if they are reasonable for the problem at hand. 

Figure 3.13 Representation of programmed step gradient showing (a) the ideal gradient and (b) typical gradient profile with rounding, ripple, and inconsistent steps highlighted.

Preparative liquid chromatography (in opposition to analytical chromatography) introduces specific requirements, which are mostly related to the separation selectivity and the loadability of a solid phase rather than the separation efficiency or peak resolution. While analytical chromatography separations may use complex profiles of elution gradients, in most of the cases, preparative chromatography utilizes linear or step gradients or a combination of both. 

A suitable gradient must be determined empirically (solvents involved, period between steps, shape of gradient profile), possibly following certain rules or by computer aid " ... 

Figure 25.2 Evaluation of the gradient profile (as an example, only one step is shown). Forthe graphical determination it is q necessary to enlarge every step of the chromatogram individually and to print it out. The height of the steps is determined 3 graphically in mV or mAU. The particular proportion of channel B (in %) is calculated from these step heights (Section 25.8)...

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