HPLC linearity

[noswald]

Hi Everyone,

This is probably basic question for most of you (apologies for that!), but I would like to know how to deal with the trendline from my linearity test going through the y-intercept.

i.e. To determine the response factor for my compound by HPLC, I measure the absorbance of a range of concentrations then construct a graph which gives a straight line. The line has a good R-squared value and I use the gradient as my reponse factor for measuring samples. However, the line passes through the y-axis above the origin. How should I deal with this in my calculations.

thanks,

Nick.

[Noser222]

scroll down this forum to page 2 and you'll find a thread called "Importance of Y-intercept in Method Validation"

http://www.sepsci.com/chromforum/viewtopic.php?t=4906

[tom jupille]

First of all, how non-zero is the intercept? Your data package should calculate a standard error for the y-intercept. If zero is included in that range, then the intercept is statistically indistinguishable from zero (in other words, don't worry about it).

If the intercept is significantly above zero, what you do depends on how you are using the results.

The most stringently regulated case applies to pharmaceuticals; even there, if you are doing something like drug purity or content uniformity, you only have to demonstrate linearity in a range bracketing your actual concentration (in other words, don't worry about it). If you are doing trace analysis, then a significantly non-zero y-intercept is something to worry about.

In no case should you "force" the intercept to zero.