I have been trying to prepare some buffers for pH assays in the range of 1-11. Starting with pH1, I noticed that mixing the solutions as shown in the table does not give the correct pH. Instead, it is 1.5, whereas the protocol states that at room temperature, pH values should only be off by 0.02 units. I continued through to pH3.6, transitioning with Glycine-HCl and Citric acid/Phosphate buffer. With every pH, I always require much more or much less than the volumes listed in the tables. I solved this to some extent by adding the required volume of solution A and then titrating with solution B. With this, the concentrations of buffer components is altered, but at least the pH is correct. Are these buffers still good to use in assays? The only difference I made is that I prepare 100 ml of the buffer and not 200 ml, therefore everything is halved, except the stock solutions. Last but not least, when I adjust the buffer to 100ml with water, the pH changes(increases), which should not happen since it is only distilled water. Has anyone encountered something similar? Suggestions and solutions are welcome.
Regards,
AD
Hi!
i am just wondering did you make a calibration with a pH meter and with standard buffers?
Regrads
Hi,
I use the exact same recipes you do, and I use citrate/phosphate a lot. I prepare solution A, and titrate with solution B, and there is a difference of less than 0.5 ml to the reported values; my suggestion is to check your reagents: that they are at the correct hydration (otherwise recalculate to get the correct molarity), that they are dry, and of high enough reagent grade. (i.e "for the preparation of buffers")
Now, I find faster (and with no deleterious effects so far seen) to prepare this buffer (at a different molarity than reported) by rather than using the stock solutions recipes, to weigh the total amount of citric acid to use per the pH/molarity desired, dissolve in 80% of the required water, adjust pH with 0.2 M dibasic sodium phosphate and then fill to the correct volume.
Hope this helps
Thank you JMC Rubio, this is very helpful. What about buffers between pH 1 - 2, have you ever prepared the the same way?
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