Buffer calculator
Work out how much of each component to weigh for a buffer at the pH, strength and volume you need.
- Base : acid ratio
- 1.58 : 1
- pKa used
- 7.20 at 25 °C
| Component | Amount | Weigh |
|---|---|---|
| Sodium dihydrogen phosphateAnhydrous, 119.98 g/mol | 38.69 mmol | 4.641 g |
| Disodium hydrogen phosphateAnhydrous, 141.96 g/mol | 61.31 mmol | 8.704 g |
Dissolve both in about 80% of the final volume of water, check the pH with a calibrated meter at the temperature you will use it, adjust with a little acid or base, then make up to 1 L.
pH = pKa + log₁₀([base] ÷ [acid])
The Henderson–Hasselbalch equation
A buffer is a weak acid mixed with its conjugate base, and its pH depends on their ratio: pH = pKa + log₁₀([base] ÷ [acid]). Rearranged, the base-to-acid ratio is 10pH − pKa. The calculator splits the total buffer concentration between the two forms in that ratio, then turns each into a mass to weigh.
Phosphate buffer recipe
Sodium phosphate buffers mix sodium dihydrogen phosphate (the acid form) with disodium hydrogen phosphate (the base form) around their pKa of 7.20. At pH 7.4 the ratio is 100.2 = 1.58, so 1 L of 100 mM buffer needs 38.7 mM of the acid form (4.64 g anhydrous) and 61.3 mM of the base form (8.70 g anhydrous). Potassium phosphate works the same way.
Tris buffer and temperature
Tris has a pKa of 8.07 at 25 °C, but it falls by about 0.028 per °C, more than any other common buffer. A Tris buffer set to pH 8.0 at 25 °C reads about 8.6 at 4 °C. Enter the temperature you will use it at, and the calculator corrects the pKa.
PBS buffer recipe
Phosphate-buffered saline (PBS) is a fixed recipe rather than a single buffer pair. 1× PBS is 137 mM sodium chloride, 2.7 mM potassium chloride, 10 mM disodium hydrogen phosphate and 1.8 mM potassium dihydrogen phosphate, at pH 7.4. Per litre that is 8.01 g NaCl, 0.20 g KCl, 1.42 g Na₂HPO₄ (or 2.68 g of the heptahydrate) and 0.245 g KH₂PO₄. A 10× stock uses ten times each amount.
Choosing a buffer
A buffer holds its pH best within about one unit of its pKa, so pick one whose pKa is close to your target. Check it won't interfere with the experiment: phosphate precipitates with calcium, and Tris has a reactive amine. The calculator does not correct for ionic strength, which lowers phosphate's pKa towards 6.8 in concentrated solutions. Treat every recipe as a starting point and finish by adjusting with a calibrated pH meter.
Many buffer salts are sold as hydrates, so choose the form on the bottle's label, or look it up with the molecular weight calculator. The dilution calculator takes a concentrated stock down to working strength.