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Miscalculating buffer pH by even 0.1 units can invalidate enzymatic assays, crash protein crystallization trials, or produce unreliable HPLC separations. This calculator applies the Henderson-Hasselbalch relationship pH = pK_a + log₁₀([A⁻] ÷ [HA]) across 15 common buffer systems with literature pK_a values sourced from CRC and Good’s buffer references. Temperature correction uses published dpK_a/dT coefficients because a Tris buffer prepared at 25°C shifts by approximately −0.028 pH/°C when moved to 4°C storage.

The tool also computes buffer capacity β, which quantifies resistance to pH change upon acid or base addition. A buffer operates effectively only within pK_a ± 1; outside this window, buffering collapses rapidly. Approximation assumes dilute aqueous solutions at atmospheric pressure. Activity coefficient corrections are not applied; for high ionic strength solutions (>0.1 M), measured pH will deviate from calculated values.

Formulas

The Henderson-Hasselbalch equation for a weak acid buffer system:

pH = pK_a + log₁₀ ( [A⁻][HA] )

Where [A⁻] is the molar concentration of the conjugate base and [HA] is the molar concentration of the weak acid. For basic buffer systems the equivalent form uses pK_b:

pOH = pK_b + log₁₀ ( [BH⁺][B] )

Temperature-corrected pK_a at temperature T:

pK_a(T) = pK_a(25) + dpK_adT × (T − 25)

Buffer capacity β measures the moles of strong acid or base required to change the pH by one unit per liter of buffer:

β = 2.303 × C × K_a × [H⁺](K_a + [H⁺])²

Where C = total buffer concentration ([HA] + [A⁻]), K_a = acid dissociation constant (10^−pK_a), and [H⁺] = hydrogen ion concentration (10^−pH).

Reference Data

Buffer System	pK_a (25°C)	ΔpK_a/ΔT (°C⁻¹)	Useful pH Range	Mol. Weight (g/mol)	Common Use
Phosphoric acid (pK₁)	2.15	0.0044	1.15 - 3.15	98.00	Low-pH chromatography
Glycine (pK₁)	2.35	0.0025	1.35 - 3.35	75.03	SDS-PAGE running buffer
Citric acid (pK₁)	3.13	0.0024	2.10 - 4.10	192.12	Food preservation, citrate buffer
Acetic acid	4.76	0.0002	3.76 - 5.76	60.05	Acetate buffer, histology
MES	6.15	-0.011	5.50 - 6.70	195.24	Plant tissue culture
Phosphate (pK₂)	7.20	-0.0028	5.80 - 8.00	98.00	PBS, biological research
MOPS	7.20	-0.015	6.50 - 7.90	209.26	Electrophoresis, cell culture
HEPES	7.55	-0.014	6.80 - 8.20	238.30	Cell culture, neuroscience
Tris	8.07	-0.028	7.00 - 9.00	121.14	Molecular biology standard
Bicine	8.35	-0.018	7.60 - 9.00	163.17	Electrophoresis
Boric acid	9.24	-0.008	8.25 - 10.25	61.83	TBE buffer, PAGE
CHES	9.50	-0.018	8.60 - 10.00	207.29	High-pH protein studies
Glycine (pK₂)	9.78	-0.025	8.80 - 10.80	75.03	Western blot transfer
CAPS	10.40	-0.032	9.70 - 11.10	221.32	Alkaline PAGE
Phosphate (pK₃)	12.35	-0.026	11.30 - 13.30	98.00	Extreme alkaline buffers

Frequently Asked Questions

Each buffer system has a characteristic temperature coefficient (dpK_a/dT). Tris is particularly sensitive at −0.028 pH/°C. A Tris buffer titrated to pH 7.5 at 25°C will read approximately pH 8.1 at 4°C. Good's buffers (MES, MOPS, HEPES) have smaller coefficients. Always prepare buffers at the temperature of intended use, or apply the correction factor this calculator provides.

The Henderson-Hasselbalch equation remains mathematically valid, but the buffer has negligible practical capacity outside the pK_a ± 1 window. At a ratio of 10:1, adding a small amount of strong acid or base will cause a disproportionately large pH shift. The calculator flags this condition and reports low buffer capacity β values, indicating that you should select a buffer system with a pK_a closer to your target pH.

The Henderson-Hasselbalch equation assumes ideal solution behavior (activity coefficients of 1.0). At ionic strengths above 0.1 M, ion-ion interactions reduce effective concentrations. Phosphate buffers are especially affected because they carry multiple charges. Additionally, pH electrodes have junction potential errors, temperature calibration drift, and response lag in viscous solutions. For precision work, use the calculated pH as a starting guide and fine-tune with a calibrated meter at working temperature.

This calculator treats each ionization step independently. Phosphoric acid has three pK_a values: 2.15, 7.20, and 12.35. Select the appropriate step for your target pH range. At pH 7.0 - 8.0, the pK_a2 = 7.20 equilibrium dominates, and the other two ionizations contribute negligibly. The cross-talk between ionization steps becomes relevant only within about 2 pH units of another pK_a.

Select a buffer whose pK_a is within ±0.5 units of your target pH for maximum capacity. Then consider: (1) does the buffer interact with your system (phosphate precipitates divalent cations like Ca²⁺ and Mg²⁺); (2) temperature sensitivity (avoid Tris if temperature varies); (3) UV absorbance (MES absorbs below 230 nm); (4) metal chelation (HEPES weakly chelates copper). Good's buffers (MES, MOPS, HEPES, PIPES) were designed to minimize these interferences for biological research.

Buffer capacity β quantifies resistance to pH change, measured in mol/(L·pH). A 50 mM buffer at its pK_a has maximum β of approximately 0.029 mol/(L·pH). It matters critically in fermentation (metabolic acids accumulate), cell culture (CO₂ equilibration), and enzyme kinetics (proton-releasing reactions). If your calculated β is below 0.01, increase total buffer concentration or choose a system with pK_a closer to your working pH.