How to Prepare Buffers

Chemical buffers prevent sudden changes in the solution acidity expresses using a pH scale. Buffer systems can maintain a certain pH nearly constant during the course of biological reactions and are crucial for many physiological processes. All laboratory works in biochemistry or biophysics require a preparation of buffer solutions. As an example, make 200 ml of the buffer solution of 4-morpholineethanesulfonic acid (MES) with the concentration 0.1 molar at pH 6.0.

Things You'll Need

• pH meter
• Magnetic or mechanic stirrer
• Stir bar
• Beaker 250 or 500 ml
• Graduated cylinder 250 or 500 ml
• Distilled water
• Plastic pipette
• Solution of sodium hydroxide (NaOH) 3M
• Solution of Hydrochloric acid (HCl) 3M
• 4-morpholineethanesulfonic acid (MES), 5 g

Instructions

1. Calculate the mass of the compound needed to make the buffer solution using the following formula: mass(compound)=molar mass x molarity (solution) x volume (solution)/1,000 ml. Molar mass of MES is 195.24 g/mole. Hence, mass (MES)= 195.24 g/mole x 0.1 M x 200 ml/1,000 ml= 3.905 g.

2. Weigh the calculate amount of the compound on the scale and place the chemical in the beaker. Pour the distilled water into the beaker. The volume of the water added should be about 10 ml less than the final volume. In our example, you would add 190 ml.

3. Place a stir bar in the beaker with the solution. Put the beaker on the magnetic stirrer and turn it on to start solution stirring.

4. Place the glass electrode of the pH meter in the solution and wait 20 to 30 seconds. Read the pH displayed by the instrument.

5. Compare the pH of the solution with the target value. If the pH is higher than the desired one you need to use hydrochloric acid solution for the adjustment. On contrary, if the pH is lower that it should be, you need to use the solution of sodium hydroxide. For example, if the current pH is 5.1, then you need to use sodium hydroxide to raise it to 6.0.

6. Fill in the plastic pipette with either hydrochloric acid or sodium hydroxide solution as determined in Step 5.

7. Add three to five drops from the plastic pipette to the solution in the beaker. Wait 20 to 30 seconds.

8. Read the value displayed by the pH meter and compare it with the desired pH. You should see a pH change towards the target value.

9. Repeat Steps 7 and 8 until the pH of the solution reaches the designated value.

10. Switch off the magnetic stirrer, take off the stir bar and the pH glass electrode.

11. Pour the solution into the graduated cylinder and add the distilled water to the final volume. In our case, it would be 200 ml.

Resources

• Buffer Center
• pH measurement