Programming the Mobile Phase Composition

In gradient elution, the concentration of the mobile phase is varied during a run. There are two possibilities: the change in concentration is linear with time or it is not linear. In the latter case, the plot describing the program might be convex or concave. Instruments which are capable of mixing liquids for the mobile phase are also capable of carrying out gradient elution automatically.

Each step of the linear program is described by four variables the initial and final concentration (%) of Component A in the mixed mobile phase, the rate of program (%/min) and the length (time) of the program. Three of these four fully describe the program; in practice, the instruments permit the setting of the two concentrations and either the time or the rate of the program for each step. The electronic control system will then automatically set either the rate or the time of the program.

The situation with curved gradient profiles depends on the particular instrument. Some less convenient programmers require step-by-step entry of multiple percentage vs. time plateaus to approach curved gradient profiles while in more sophisticated systems, one can select from a number of programmed profiles.

In the actual instruments, one has to set the initial and final concentrations, the time of the program and select which program profile is desire This is done by certain keys corresponding to the various exponent values.

Naturally, in sophisticated instruments where various programs can be combined, curved programs can also be included together with linear programs and isocratic periods.

The most important consideration in gradient elution is that the whole program - a simple as well as a complex multistage program - can be exactly reproduced upon command since inaccurate gradient repetition can contribute to significant qualitative and quantitative errors.