Box 25-3 Choosing Gradient Conditions

The retention of a compound in isocratic elution depends on solvent composition  

For gradient elution, an average capacity factor k applies to all peaks  

If you have a successful gradient separation and want to transfer it from column 1 to column 2, whose dimensions are different, the scaling relations are  

Suppose that you have optimized a gradient on a 0.46 X 25 cm column and you want to transfer it to a 0.21 X 10 cm column. The quotient V2/V is ( nr2L)2/( nr1L), where r is column radius and L is column length. For these columns, V2/Vj = 0.083. Equation 25-8 tells us to decrease the volume flow rate, the sample mass, and the delay time to 0.083 times the values used for the large column. The gradient time should not be changed. 

When you make these changes, you will discover that k is the same for both columns. If you change a condition that affects k, you should make a compensating change to restore k. For example, Equation 25-6 tells us that if we choose to double tG we should cut the flow rate in half so that the product tcF is constant and k remains constant. 

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