Stacked injections

Reducing Cycle Time with Stacked Injections (Case of Isocratic Eluents) For chromatographic separations performed using isocratic eluents (i.e., whose composition does not change over time), once the injected amount has been increased to its ophmal value, eluent consumption can then be reduced by stacking... 

Figure 12.7 Use of stacked injections to decrease cycle time.

Figure 12.8 Example of the chromatogram of 19 stacked injections performed in 3.5h..

However, much of the stationary phase and eluent inside the column is not in contact with the product, even in the case of stacked injections. To utilize the unused stationary phase and eluent to perform the separation, the mixture is reintroduced at the inlet of the column via a loop, as depicted in Figure 12.10b. In this way, separated compounds can be collected continuously while both feed mixture and eluent are injected continuously. Thus, the whole process becomes continuous. ... 

The process daily throughput is linked to both the injected amount per run and the time between two successive injections. This time has to be minimized using stacked injections in order to optimize the process productivity and further decrease the eluent consumption. Minimum time between two successive injections corresponds to the time needed for eluting the two enantiomer peaks. Under the selected conditions, this time was equal to 100 s. 

Figure 12.23 shows the superposed chromatograms obtained for 6 stacked injections at preparative scale. 

The reproducibility of the electrokinetic injection is poorer than that of the dynamic pressure injection and strongly depends on the ionic strength. Internal standards are usually added to improve the accuracy and precision [44]. For a stacking injection, a mathematical model has been developed to account for the increase in the migration time with increasing sample injection time a good agreement with the theory has been found [45]. 

The aforementioned injection of a solution plug is termed as plug injection. Another sample introduction mode is stack injection. Stack and plug injections used in DNA separation have been compared (see Figure 4.2). Under similar conditions, the plug injection produced a better resolution, whereas the stack injection produced a higher sensitivity (see Chapter 6, section 6.2 for more on CE separation) [315]. 

Although stack injection has been employed previously [316,582], the benefit of stacking for sample pre-concentration was only studied in detail later [346]. With the sample buffer (0.5 mM) at a 10-fold lower conductivity than the separation buffer (5 mM), simple EK stacking using the gated injection was observed. This was applied to the separation of dansylated amino acids (dansyl-lysine, didansyl-lysine, dansyl-isoleucine, and didansyl-isoleucine) [346]. 

Owing to the EK bias in favor of faster migrating species in stack injection, the signal enhancement ranged from 31 to 8. However, stack injection produced less resolution, i.e., fewer plate numbers (N), as compared to those obtained in pinched injection. In addition, RSD (n = 6) in peak areas for stack, non-stacked, and pinched injection are 2.1%, 1.4%, and 0.75%, respectively [346]. 

Figure 10 Electropherograms obtained with various channel sizes and matrix conditions for HaeIII digests of 0X1 74 phage DNA. (A) Separation channel, 30 pm cross channel, 30 pm sample (plug injected for 5 s), 100 ng/pL separation in the presence of 1 pM TO. (B) Separation channel, 50 pm cross channel, 30 pm sample (plug injected for 1 s), 10 ng/pL separation in the presence of 1 pM TO. (C) Separation channel, 50 pm cross channel, 120 pm sample (stack injected for 1 s), 10 ng/pL separation in the presence of 0.1 pM T06. (D) Separation channel, 70 pm cross channel, 120 pm sample (plug injected for 1 s), 10 ng/pL separation in the presence of 0.1 pM T06. Sensitivities of DNA detection with TO and T06 are comparable at the concentrations used. (Reprinted with permission from Ref. 62.)...

Figure 5.2 Separation of two components under touching band and stacked injections conditions for three consecutive injections.

Palmer, J., Bnrgi, D.S., and Landers, J.P, Electrokinetic stacking injection of neutral analytes under continuous conductivity conditions. Anal. Chem., 74, 632, 2002. 

FIGURE 13.12 Illustration of the mechanism of electrokinetic stacking injection. The sample matrix is prepared devoid of micelles to an equal conductivity of the BGE. 

Gong, M., Wehmeyer, K.R., Limbach, P.A., and Heimeman, W.R., Unlimited-volume electrokinetic stacking injection in sweeping capillary electrophoresis using a cationic surfactant. Anal. Chem., 78, 6035, 2006. 

P. Zhang, G Xu, J. Xiong, Y. Zheng, Q. Yang and F. Wei, Determination of arsenie species by eapdlary electrophoresis with large-volume field-amplified stacking injection. Electrophoresis, 22,3567—3572, 2001. 

FIGURE50.9 Electrokinetic stacking injection on a microchip. Sample 67 nM BODIPY (a neutral fluorescent dye) in 150 mM NaCl. Run bnffer 80 mM sodium cholate, 10% ethanol, 5 mM tetraborate, pH 9. Sample was first injected by applying an electric field of 183 V/cm between the sample reservoir (S) and the outlet (O) for various periods of time as indicated in the figure. Separation was then carried out by applying 366 V/cm between the inlet (I) and outlet (O). An 80-s injection of 134 nM BODIPY in run buffer is included to show nonstacking injection. (From Palmer et al., Anal. Chem., 73, 725, 2001. With permission.)... 

Gong, M. J., Wehmeyer, K. R., Limbach, R A., Arias, R, and Heineman, W. R., On-line sample preconcentration using field-amplified stacking injection in microchip capillary electrophoresis, Analytical Chemistry, 78, 3130-5131, 2006. 

A second point concerning the subject of how much local sources Interfere with precipitation concerns the effect of the emission height. This came up earlier tdien we were talking about Sudbury. I had to do a few sums first to make sure of my ground, but In the case of a tall stack Injecting Into the free atmosphere above the mixed layer, pollutants might remain there for a considerable period. At this latitude radioactive fallout studies Indicate residence times of as much as ten days. But a cloud gets much of Its air from the mixed layer. 

FIGURE 12 Semipreparative SFC chromatogram illustrating stacked injections for impurity isolation. 

Figure 42 shows stacked injections. In this case, as a result of using stacked injections, the single 8 min injection time observed in Figure 40 is reduced to a 4.5 min injection cycle time allowing for the purification of two targets to be completed in only 47 minutes. An optimized SFC method took approximately 1 h to develop. Optimized SFC method conditions were determined to be 10 mL/min on a 2-ethylpyridine 1 cm x 25 cm column using isocratic methanol cosolvent 40/60, methanol/COi conditions. The outlet pressure was set at 140 bar and the column temperature set at 35 C. Ten injections were stacked over 47 minutes. 

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