Chromatography Forum

In the recent Gradient Performance Checks Article of LC/GC magazine (10/2004 issue), the Step-Test was calculated using the "theoretical (programmed) value." But, if the data acquisition software output is in absorbance units and the programmed value is in % A and % B, what is the theoretical value? Is the maximum absorbance of 100% B taken into account? Is the acetone level in line B adjusted to obtain close to a 1 AUFS?

I am writing an IQ/OQ that will be used as a model for future HPLC calibrations, and I am incorporating the methods outlined in this article.

I am sure this is just easy math, but it does not make sense to me.

Any help would be appreciated.

In theory, each step will generate the same percentage change in absorbance.

mathematically, the article indicates you want:
|Bp – Be| ≤ 1

where:
Bp = programmed %B
Be = experimental %B

Be = 100 He / H0+H100

He = step height at Bp
H0 = height at 0 %B
H100 = height at 100 %B

following on from what Tom said, perhaps you could look at the consistency of the step height change (%RSD). For method validations or transfer I think this test is important in terms of accuracy, hence the criterion chosen.

HTH = hope that helps

Thanks for your help. I basically figured it out once I sat down with pen and paper and graphed it out.

The only difference is that I interpreted the end result wrong. I was calculating it as 1-(Be/Bp).