Gradient elution, introduction

We should therefore use small bore tubing in short lengths for making the injector-column and the detector-column connections, and the injector and detector must be designed so that their internal volume is as small as possible. Dead volume before the introduction of the sample should also be minimised, to facilitate rapid changes of mobile phase composition that may be required during gradient elution. 

The introduction of the commercially available mass detector meant an expansion of the possible conditions for TG separations, including that of gradient elution. The mass detector showed very narrow and symmetrical peaks, which indicated its usefulness for this application. The goal was to find a system that could be used to separate any natural TG mixture regardless of its carbon number distribution. The basic principle of this detector, i.e., the light scattering, originally was presented by Charlesworth (104). 

Introduction of the light-scattering detector has made the development of more efficient methods for the separation of natural complex mixtures of TG possible, including those of gradient elution. The evaporator temperature in the mass detector oven may influence the response of thermolabile compounds or compounds with low boiling points. Consequently, low tempera-... 

These problems have been circumvented by both the introduction of modern micromembrane suppressors and the development of short clean-up columns for eliminating inorganic impurities in the eluent [130]. Therefore, the gradient elution technique in ion chromatography today is as common as in the area of conventional HPLC. 

Phenylisothiocyanate (PITC). The use of PITC (Edman Reagent) to form thiohydantoin (PTH) derivatives of amino acids for protein sequencing is well known. The development of reversed phase systems for the separation of these PTH derivatives has resulted in the introduction of both isocratic and gradient elution systems (Fig. 11.2.11). A microbore HPLC unit for PTH-amino acid analysis is now available along with the recently introduced gas phase sequenator... 

The introduction of gradient elution greatly improved these analyses. The reason IC easily deals with complex matrices lies not only in the stability and resistance to contamination of the stationary phases used, but also in the sensitivity and specificity of the detection methods employed. 

Appendix 3.3 A Brief Introduction to Snyder s Theory of Gradient Elution... 

Figure 10 Chromatography of mouse interferons on agarose-bound Cu(ll)-IDA column. Sample introduction was followed by stepwise elution with 0.02 M sodium phosphate (1.0M NaCI), pH 7.4, and thereafter with 0.1 M sodium acetate (1.0M NaCI), pH 6.0. The chromatogram was obtained by gradient elution with the same acetate solution at decreasing pH. ( ) Glysosylated (O) nonglycosylated. (From Ref. 48.)...

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