To prepare 100 mL of 25 mM Tris buffer at pH 8.0, calculate the amount of Tris b

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. 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. 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.

Explanation / Answer

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: pKa = 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 !

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