Bio-Rad Media Sampler Pack - Manual (Page 10)

V
e
= Peak elution volume or time
W
½h
= Peak width at peak's half height in volume or time
V
e
and W
½h
should always be the same units
Peaks should be symmetrical. The best achievable asymmetry factor is 1. Values of 0.8
to 1.8 are excellent for large columns >30 cm in diameter. If functional performance is
acceptable, A
s
factors more distant from 1 put less demand on the column packing
procedure in production. A change in the shape of the peak is usually the first indication
of deteriorating performance. Peak asymmetry factor calculation:
A
s
= b/a
a = Front section of peak width at 10% of peak height bisected by line denoting V
e
.
b = Latter section of peak width at 10% of peak height bisected by line denoting V
e
.
8
Fig. 2. A simulated chromatography profile from which HETP and As values are calculated.
Bed height = 20 cm
Column volume = 20 L
Flow rate = 100 cm/h
V
e
= 10 L
W
½h
= 1 L
HETP = 0.036 cm
N = 554
a = 1
b = 1.2
A
s
= 1.2
W
½h
width
at half height
10% of peak
a b
V
e
(elution volume)
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