Question

To prepare 100 mL of 25 mM Tris buffer at pH 8.0, calculate the amount of Tris base and Tris·HCl needed to make the buffer. (Hint: find out the pKa of Tris·HCl and use the Henderson-Hasselbalch Equation.) Describe how do you want to make the solution, including how much water to add, how to adjust pH, etc.

Answer 1

Let's denote the Tris-HCl by HA its conjugate base Tris base by A^-.

Total moles of acid and conjugate base = Molarity*Volume = 0.025*0.1 = 0.0025

For Tris-HCl, we have: pK_a = 8.08

Using Henderson Hasselbach equation:

pH = pKa + log(moles of A^-/moles of HA)

Putting values:

8 = 8.08 + log(moles of A^-/moles of HA)

So,

(moles of A^-/moles of HA) = 0.832

Also,

Moles of A^- + moles of HA = 0.0025

Solving these two equations we get:

Moles of A^- = 0.0011

Moles of HA = 0.0014

So,

Mass of Tris base needed = Moles*MW = 0.0011*121.1 = 0.133 g

Mass of Tris HCl needed = Moles*MW = 0.0014*157.6 = 0.22 g

Thus to make this buffer, mix the above quantities of Tris HCl and Tris base in 100 mL of distilled wwater.

Hope this helps !