Gradient elution high-pressure mixing

Compared to the isocratic elution mode, the equipment necessary for gradient elution is extended by a second solvent reservoir, an additional pump (in the case of a high-pressure-mixing gradient system), a mixing chamber, and a proper-... 

An HPLC method using progressive electrochemical detection of SPA was described by McCabe and Acworth (128). Samples were mixed with hexane, and SPA were extracted with acetonitrile. An HPLC analysis of the extracts was performed, without an evaporation step, on a high-pressure Coul Array system in which analytes were detected on two coulometric array-cell modules, each containing four electrochemical sensors attached in series after the column. Analytes were separated on a Supelcosil LC-18, 5-/tm column using gradient elution and detected at potentials of —50, 0, 70, 250, 375, 500, 675, and 825 mV. To remove oxidative impurities to be coeluted with BHT, a guard cell with applied potential of 900 mV was also placed in the system. 

Figure 21.21. Gradient devices used in liquid chromatography. A Simple mixing device for gradient elution, gradient formed on low-pressure side of pump. Ri is the flow rate of A into the flask. Ra is the flow rate of A + B to column. B Gradient profiles obtained from A. C Schematic for two-pump gradient chromatograph. Gradient formed on high-pressure side of pump.

The main advantages of gradient over isocratic systems are provided in the analysis of unknown or multicomponent samples where a range of solvent strengths may be required both to ensure that aU the components loaded are eluted and also that the retention times of later eluting peaks are minimised. Gradient systems may be subdivided into two classes those where solvents are mixed prior to the pump (low pressure systems) and those where solvents are mixed after the pump (high pressure systems). 

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