Buffer & pH
Acid and base amounts, or titrant volume, for a target pH. pKa corrected for temperature.
Result
Dissolve in ~80 % of the volume, adjust pH at the working temperature, then make up to 1 L.
Strong temperature dependence: a pH 8.0 Tris buffer made at 25 °C reads ≈ 7.7 at 37 °C and ≈ 8.6 at 4 °C.
Formula
pH = pKa + log₁₀([A⁻] ÷ [HA]), so with r = 10^(pH − pKa) the base fraction is r ÷ (1 + r) and the acid fraction 1 ÷ (1 + r). Starting from the base form, HCl equal to the acid fraction is added; from the acid form, NaOH equal to the base fraction. pKa(T) = pKa(25 °C) + ΔpKa/°C × (T − 25). Ionic strength and activity effects are ignored, so treat the result as a starting point and finish with a calibrated pH meter at the working temperature.
Questions
How much HCl do I add to make 50 mM Tris pH 7.5?
Dissolve 50 mmol of Tris base per litre, then titrate. At 25 °C the pKa is 8.07, so pH 7.5 needs a 3.7 to 1 acid to base ratio, which is 39.4 mL of 1 M HCl per litre. Finish with a pH meter.
Why does my Tris buffer read a different pH at 37 °C?
Tris has a temperature coefficient of about −0.028 pH units per °C. A buffer set to pH 8.0 at 25 °C reads roughly 7.7 at 37 °C and 8.6 at 4 °C. Set the pH at the temperature you will use, or enter that temperature here.
Should I weigh both forms or titrate one?
Weighing both the acid and base forms is quicker and avoids adding extra ions from HCl or NaOH, but it depends on the tabulated pKa being right for your ionic strength. Titrating one form with a pH meter is more accurate. Both routes are calculated here.