Buffer preparation
What to weigh out for a buffer, with the pKa corrected for temperature.
Better suited to pH 7.4:
The temperature of the meter when you titrate.
Where the experiment actually runs.
Weigh out
Tris·HCl
C4H12ClNO3 · 157.59 g/mol · 20.51 mmol
Tris base
C4H11NO3 · 121.14 g/mol · 4.488 mmol
pKa 8.06 at 25 °C · base fraction 0.18 · reads pH 7.4 at 25 °C
About TrisThe largest temperature coefficient in common use. Adjust the pH at the temperature you will work at, not on the bench. Tris also binds some metal ions and interferes with protein assays that use the Bradford reagent.
pH = pKa(T) + log₁₀([base]/[acid]); pKa(T) = pKa₂₅ + (dpKa/dT)(T − 25)- Hydrogen ion buffers for biological research — Biochemistry, 1966
- The Henderson–Hasselbalch relationship — Journal of Chemical Education, 2001
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No correction for ionic strength is applied. Activity effects shift real pH by roughly 0.1 units at physiological salt, so use this to get close and then check with a meter at your working temperature.
About this tool
Calculates the masses needed for a buffer at a chosen pH, concentration and volume, using a pKa corrected to the temperature you are working at. Tells you how far the pH will drift if you adjust it on the bench and use it in the cold room — which for Tris is more than half a pH unit.
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Science last reviewed .