Preparation of 3 M Sodium Acetate Solution, pH 5.2 from Sodium Acetate Trihydrate (CH3COOH.3H2O)

Sodium acetate is a sodium salt of acetic acid which dissolves readily in water. It is commercially supplied as both anhydrous (molecular weight: 82.03) and trihydrate (molecular weight: 136.08) forms and both can be used to prepare 3M Sodium acetate solution. Since anhydrous sodium acetate is extremely hygroscopic, sodium acetate trihydrate is a better choice to prepare solution.

Here we show a procedure to prepare 100 ml of 3M Sodium acetate, pH 5.2 from sodium acetate trihydrate. Briefly, to prepare a 100 ml solution of 3M Sodium acetate of pH 5.2, dissolve 40.824 g Sodium acetate trihydrate in 80 ml water, adjust the pH 5.2 using acetic acid, and then make the final volume to 100 ml. Remember that this solution act as a buffer solution (Acetic acid-Sodium acetate buffer)


Reagents and solutions
Sodium acetate trihydrate (CH3COONa.3H2O) (Molecular weight = 136.08)
Deionized / Distilled water

Equipment and disposables
Measuring cylinder
Conical flask / Beaker
Magnetic stirrer

3 M Sodium acetate

Preparation of 100 ml of 3M Sodium acetate solution, pH 5.2 in water from sodium acetate trihydrate  (CH3COONa.3H2O)


Step 1: To prepare 100 ml aqueous solution of 3M Sodium acetate (CH3COONa) solution, weigh out 40.824 g Sodium acetate trihydrate (Molecular weight = 136.08). Transfer to 250 ml beaker/conical flask. Add 80 ml deionized / Milli-Q water. Mix until sodium acetate dissolves completely.

One can use manual shaking using a glass pipette to mix the ingredients. Magnetic stirrer makes the dissolving process easy and convenient.

Do not dissolve in 100 ml of deionized / Milli-Q water. In most cases, solution volume increases when a large amount of solute dissolves in the solvent.

Step 2: Adjust the pH to 5.2 with glacial acetic acid.

Since pH depends on temperature, one should adjust pH at room temperature (25°C).

Step 3: Adjust the volume to 100 ml with deionized / Milli-Q water. Mix it again.

The solution will appear colorless and transparent.

Step 4: Transfer the solution to an autoclavable bottle. Sterilize the solution by autoclaving (20 minutes at 15 lb/ (psi) from 121-124°C on liquid cycle).

Depending on the consumption, one can make small aliquots of the solution.

One can sterilize a solution by passing through 0.2μ filter unit. Filter sterilization removes all suspended particles with size more than 0.2 μ which includes most bacteria and their spores but not mycoplasma. However, it does not inactivate enzyme activities (e.g., DNases). Autoclaving inactivates most enzymes except some (e.g., RNases) and kills most microorganisms including mycoplasma.

Solution can be stored at room temperature.


  • Precipitation of nucleic acid (DNA and RNA)
  • Buffer solution
  • Protein crystallization
Follow the table to prepare Sodium acetate (CH3COONa) solution of different concentrations and volumes from Sodium acetate trihydrate.
Conc. / Volume10 ml50 ml100 ml250 ml500 ml1000 ml
0.1 M0.136 g0.68 g1.36 g3.402 g6.804 g13.608 g
0.5 M0.6804 g3.402 g6.804 g17.01 g34.02 g68.04 g
1 M1.36 g6.804 g13.608 g34.02 g68.04 g136.08 g
3 M4.08 g20.412 g40.824 g102.06 g204.12 g408.24 g

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