Chromatography Forum

Dear members of this forum,

I would like to ask a question about the best method to prepare a mobile phase. Always I adjust the pH in the buffer aqueous solution because the pH scale (0 - 14) is to aqueous solution.
The last week I had to do an hplc analysis where I had to adjust the pH in a mixture of methyl alcohol and phosphate buffer (pH: 5.5), the relation was 600:400. The technique was an official method, I disagree by two reasons, first at this pH the phosphate not work as buffer and second I adjust the pH of phosphate buffer to pH 5.5 and when prepared the mixture 600 ml methyl alcohol and 400 ml of buffer solution.
I adjusted the pH again according to the official tecnique and I would like to know other opinions about if it the preparation of this mobile phase is good or not.
Thanks in advance for your help,

Diego Delmonte

What you are achieving using this method is an apparent pH, not a true pH.

However, if the method you are following works and gives reproducible results, then the mp phase prep is good.
J

Agree with John; it's just very important that the preparation instructions are very specific, and not ambiguous. I have seen quite a few published procedures where things could be interpreted more than one way.

I actually think that the situation is even worse.

when you dissolve a monobasic phosphat salt in water, you get a pH of around 4.5. When you add organic to this (such as your 60% methanol)and measure again, you get a higher pH (prbably just around 5.5 with 60% methanol).

Conclusion: you don't have a buffer, and it is measured in a silly way.