I am trying to prepare a bicarbonate buffer for TNBS analysis which requires a 0.1 M bicarbonate buffer of pH 8.5. Most of the chemical suppliers have a reagent of 1 M and of different pH. I have been told that I can use carbonate and bicarbonate to make the buffer using the Henderson-hasselbalch equation, but I am not sure how to calculate exactly how much concentration or volume must be added in order to obtain a specific pH buffer of the same. Can anyone point out the exact process to make the same? Thanks.
[I have also attached the protocol steps for reference.]
It will be necessary to clarify whether buffer pH = 8.5 is meant at a temperature of 20 or 37oC. Since the required pH is at the lower end of the buffer range this system, adjusting the pH of 0.1M NaH(CO3)2 solution with Na2CO3 solution will probably be more accurate. Only a very small amount of Na2CO3 will be needed.
viz.: https://www.researchgate.net/post/How_do_you_adjust_pH_of_NaHCO3_buffer
@Dalibor Matýsek, Thankyou for your reply. As per the protocol, the sample needs to be incubated at 37 degrees so the pH must be 8.5 during that time. I will try adjusting the pH with Na2CO3 as mentioned. Thanks again.
This complicates the situation a bit, but it will be manageable. It will be necessary to properly calibrate the pH meter to 37oC, heat the solutions to this temperature in a water bath, stir the solution preferably with a magnetic stirrer and titrate the bicarbonate solution to a stable pH 8.5. The second possibility is to find out in the pH temperature dependence tables what pH value at 21oC corresponds to pH 8.5 at 37oC. Then it would be enough to do it at room temperature. The estimated pH of 8.5 / 37oC corresponds to approx. pH 9 / 20oC.
Titration preparation has one advantage over the calculation. This is an activity, not a calculated concentration.
Yes, that seems manageable Dalibor Matýsek. I will go with the titration process. Thankyou.
For related RG discussions; see: https://www.researchgate.net/post/How_can_stabilise_the_pH_of_01M_sodium_bicarbonate_solution_made_in_the_lab_to_80 https://www.researchgate.net/post/Anyone_know_how_to_prepare_2_M_of_sodium_bicarbonate_pH_100_solution_no_buffer
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