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Tuesday, May 21, 2013

Importance of Column Packing & Efficiency Calculations


Column Packing is one of the most important step in the chromatographic purification process. A well packed column yields higher resolution and yield.

Column Packing Efficiency can be checked by calculating:
  • HETP - Height Equivalent Theoretical Plates
  • Peak Asymmetry Factor (AF / As)
HETP Calculation:

HETP = ( d2 * L ) / (5.54* t2)

Where: t = retention value expressed as units of time, volume or measured distance on the Chromatogram
d = peak width at half height expressed in same units as “t”
L = column length (in units of cm for HETP or m for N/m)

N/m =  (5.54* t2) / ( d2 * L )

Asymmetry Factor (AF / As) Calculation:


chromatography peak asymmetry



AF = b/a

Where: a is the peak width (left half) at 10% of peak height, b is the peak width (right half) at 10% of peak height, 


Column Packing Guidelines - TOSOH Biosciences 




References
TOSOH Biosciences - Technical Resources
GE Lifescience - Technical Resources
Pall Lifesciences - Technical Resources