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FAQs

17. What should I do to scale my analytical sample to a semi-prep column? How much can I load?

March 14, 2021

Analytical separations can be predictably scaled up to the semi-preparative level using some fairly simple calculations. When resolution of a given quantity of racemate is the desired objective, it is best to first optimize using an analytical column to yield the maximum possible loading. If you are fortunate to have more than one possible separation method, each method can be tested separately to determine which one gives the best overall loading. For each attempt to achieve maximum loading, start by making injections of a solution of the target racemate, as concentrated as possible, in the mobile phase. Using a detection wavelength selected to keep the peaks on scale, increase the injection volume until the valley between the enantiomers begins to rise.

This should give you an experimental loading weight (WE): WE = Cmax x VAmax
where Cmaxis the maximum concentration of racemate and VAmax is the largest analytical injection volume before overload.
For ballpark estimation purposes, typical WE values for an analytical column (0.46 x 25 cm) are 1 10 mg/injection.

The relative loading capacity (LCR) on the 0.46 x 25 cm analytical column is assumed to be “1.” For various-size semi-preparative and preparative columns, the relative loading capacity and associated flow rates can be determined from the following table.

Column Size
(i.d. x Length)
Weight of Packing (g)Loading Capacity (relative)
LCR
Flow Rate (ml/min.)
0.46 x 25 cm2.5011.0
1 x 25 cm11.855.0
2 x 25 cm47.41919
5 x 50 cm59225050
10 x 50 cm23701000200

How much can I load?

The loading on the preparative column (WPREP) would be calculated with the experimental weight on the analytical column (WE) and the relative loading capacity (LCR).

WPREP = WE x LCR

Thus, if the typical load on an analytical column (0.46 x 25 cm) is 1-10 mg, then a typical load on a 2 x 25 cm semi-preparative column is 19 –190 mg/injection.

What size column do I need? This is a typical preparative question that is inter-related to the question: How many injections (N) are you willing to make? To determine the size column that you need, the following equation can be used:

where LCR is the loading capacity of the column needed to achieve your objectives, WDES is the weight of desired enantiomer you want to separate and WE  is the experimental weight on the analytical column.

EXAMPLE: A researcher needs to isolate 1 gram each of two enantiomers from a racemic mixture (WDES). It has been determined that 48 half-hour runs (N) made in a 24-hour period would be a reasonable number of preparative injections. The maximum analytical loading weight is 2 mg = .002 g (1 mg of each isomer, WE). What relative loading capacity is required if 48 injections are to be made?

Consulting the table, it can be seen that a 2 x 25 cm column with a relative loading capacity of 19 would probably best meet the researcher’s needs.

/wp-content/uploads/2021/03/Daicel-Chiral-Tech_logo-1.png 0 0 Ryan S /wp-content/uploads/2021/03/Daicel-Chiral-Tech_logo-1.png Ryan S2021-03-14 18:33:262024-11-22 22:54:2017. What should I do to scale my analytical sample to a semi-prep column? How much can I load?
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