DavidHPLC wrote:
A debate started yesterday at work .
What are your experiences when preparing buffers? Do you weight the exact amount, water volume and pH? Or do you have +/- tolerance for buffer mass and pH and/or use measuring cylinders instead of volumetric flasks ...
I'd like to comment, my 2 cents worth.
Even when we followed a procedure from elsewhere (compendial, supplier, etc.), I often found instructions for preparation of buffers to be vague. Some did not even specify if a certain pH was measured for just the aqueous portion, or for the mix including the organic.
First thing I'd do would be to follow instructions as written to confirm that the procedure actually worked. But I also measured how much phosphoric acid (for example) was added to reach the specified pH in the procedure. Then from then on, I'd make the buffer adding components by weight or volume, and did not use a pH meter, much faster, and in my mind more accurate and reproducible. Of course my pointy-haired boss
figured EVERYTHING I did was wrong, so he didn't like that, wanted us to use pH meter. I tried telling him that other chromatographers also liked the non-pH meter way after working things out, and that I did use pH meter initially, but he'd periodically return to ME making buffers wrong.
Anyway, buffers exist so that small variations do not affect pH significantly. Yes, I wrote stuff so it would state like" "add 3.20 - 3.40 grams of Na2HPO4" so that anything within that range was acceptable, and anything out of that range would be NOT acceptable, and non-cGMP, and would need to be re-made.