Hi all,
This is a pretty basic question, but it's been a while since University and I'm a bit rusty with buffers (so please bear with me!)
So I'm trying to prepare a range of buffers for LCMS. I want to prepare buffers at pH 4, 5, 6, 7, 8. Buffer concentration is 1M. I'll need to use volatile buffers such as Ammonium acetate and formate buffers. pKa of Formic is 3.75, Acetic is 4.76, so I know these buffers won't be too effective at or above pH6, but I'm hoping the high concentration (1M) will be enough to buffer a urine sample for example.
For 100mL buffer I've worked out the mass of NH4.acetate and volume of Acetic acid I need with HH eqn. When I use an online buffer calculator (Chembuddy) I get different results so I'm not sure what I'm doing wrong.
Columns are:
pH of buffer mass (NH4.acetate).g Volume (Acetic). uL 100mL buffers
4 1.141 4877 1M NH4.Oac buffer (pH4)
5 4.893 2091 1M NH4.Oac buffer (pH5)
6 7.289 311 1M NH4.Oac buffer (pH6)
7 7.664 33 1M NH4.Oac buffer (pH7)
Buffer calculator
pH of buffer mass (NH4.acetate).g Volume (Acetic). uL
4 1.78 4.4mL 1M NH4.Oac buffer (pH4)
5 6.52 879.7uL 1M NH4.Oac buffer (pH5)
6 7.59 90.09uL 1M NH4.Oac buffer (pH6)
7 7.71 0.1476uL 1M NH4.Oac buffer (pH7)
When I plug in the values from the buffer calculator (above) into the HH equation I don't get the target pH. I'm not sure what I'm doing wrong. Any ideas?
Thanks for your help