Date: 12-JUNE-2023  Last Updated: 22-JANUARY-2026

Step‑by‑Step Instructions to Prepare 16 mM Ammonium Acetate in 90:10 Acetonitrile:Aqueous Phase

This procedure produces a mobile phase containing 16 mM ammonium acetate in a solvent mixture of approximately 90% acetonitrile and 10% aqueous phase (made from buffer stock and DI water).

1. Prepare the Buffer Stock (500 mM Ammonium Acetate)

1. Weigh 3.85 g of ammonium acetate.

2. Transfer it into a 100 mL volumetric flask.

3. Add DI water up to the 100 mL mark.

4. Mix until the salt is fully dissolved.

5. Filter the solution through a 0.45 µm nylon filter.

6. Store the buffer stock at 2–8°C.

Result: This is a 500 mM ammonium acetate stock solution.

2. Prepare the Final 90:10 Mobile Phase (16 mM Ammonium Acetate)

1. Add 32 mL of the 500 mM buffer stock into a 1000 mL (1 L) volumetric flask.

2. Add 68 mL DI water to the same flask.

3. Fill the flask to the final 1000 mL mark with acetonitrile.

4. Mix thoroughly.

5. (Recommended) Filter the final mixture through a 0.45 µm nylon membrane before use.

3. Final Composition 

• The total buffer added contributes to a final concentration of:
  (32 mL ÷ 1000 mL) × 500 mM = 16 mM ammonium acetate

• The solvent composition is:
  ~90% acetonitrile + ~10% aqueous phase (buffer + water)

These instructions help to ensure you get reproducible results with respect to the mobile phase used in our Application Notes.