Biological Buffers

Buffer Reference Center

Choosing the Right Buffer

You’ll want to choose a buffer based on your pH requirements as well as the pKa, a measure of acid strength that accounts for pH, concentration, and temperature. Regulatory or purity needs for your exact application should also be considered.

The following tables can help you navigate preparation of many common buffer solutions by pH and pKa. Refer to our biological buffers selection guide for complete information on all our buffers.

 

What are buffer grades?

Buffer grade indicates the quality and impurity levels appropriate for different uses. We provide six grades of buffers indicated for general lab use, final pharmaceutical formulation and manufacturing, and applications in between that may need trace metal testing or materials of a specified purity.  

 

 

Useful pH Ranges of Selected Biological Buffers Chart (25 °C, 0.1M)   

 

Buffers Useful pH Range pKa
(at 20°C)
pKa
(at 25°C)
pKa
(at 37°C)
Reagent Grade  BioPerformance Certified Grade Pharma Grade
MES 5.5–6.7 6.16 6.10 5.97 M8250 M2933 PHG0003
Bis-Tris 5.8–7.2 n/a 6.50 6.36 B9754 B4429 RES1161B-A7, PHG0004
ADA 6.0–7.2 6.65 6.59 6.46 A9883 n/a n/a
ACES 6.1–7.5 6.88 6.78 6.54 A9758 A3594 RES1000A-A7
PIPES 6.1–7.5 6.80 6.76 6.66 P6757 P1851 RES0703P-A7
MOPSO 6.2–7.6 n/a 6.90 6.75 M8389 n/a RES0196M-B7
Bis-Tris Propane 6.3–9.5 n/a 6.8, 9.0 n/a B6755 B4679 PHG0005
BES 6.4–7.8 7.17 7.09 6.90 B9879 B4554 n/a
MOPS 6.5–7.9 7.28 7.20 7.02 M1254 M3183 PHG0007
TES 6.8–8.2 7.50 7.40 7.16 T1375 T5691 n/a
HEPES 6.8–8.2 7.55 7.48 7.31 H3375 H4034 RDD002, RES6003H-B7
DIPSO 7.0–8.2 n/a 7.60 7.35 D9648 n/a n/a
MOBS 6.9–8.3 n/a 7.60 n/a M3295 n/a n/a
TAPSO 7.0–8.2 n/a 7.60 7.39 T9269 T5566 n/a
Tris or Trizma® 7.0–9.0 8.20 8.06 7.72 T1503 T6066 RES3137T-A7, RES3193T-A7
HEPPSO 7.1–8.5 n/a 7.80 6.66 H3137 n/a n/a
POPSO 7.2–8.5 n/a 7.80 7.63 P3405 n/a n/a
TEA 7.3–8.3 n/a 7.80 n/a T1377 n/a n/a
EPPS 7.3–8.7 n/a 8.00 n/a E9502 E0276 n/a
Tricine 7.4–8.8 8.16 8.05 7.80 T0377 T5816 RES3077T-A7
Gly-Gly 7.5–8.9 n/a 8.20 n/a G1002 G3915 G0674
Bicine 7.6–9.0 8.35 8.26 8.04 B3876 n/a RES1151B-B7
HEPBS 7.6–9.0 n/a 8.30 n/a H6903 n/a n/a
TAPS 7.7-9.1 8.49 8.40 8.18 T5130 T5316 n/a
AMPD 7.8–9.7 n/a 8.80 n/a A9754 n/a n/a
TABS 8.2–9.6 n/a 8.90 n/a T1302 n/a n/a
AMPSO 8.3–9.7 n/a 9.00 9.10 A6659 n/a n/a
CHES 8.6–10.0 9.55 9.49 9.36 C2885 n/a RES1299C-B7
CAPSO 8.9–10.3 n/a 9.60 9.43 C2278 n/a n/a
AMP 9.0–10.5 n/a 9.70 n/a A9879 n/a n/a
CAPS 9.7–11.1 10.56 10.40 10.02 C2632 n/a n/a
CABS 10.0–11.4 n/a 10.70 n/a C5580 n/a n/a

 

Tris or Trizma® Buffer Preparation Table – pH vs. Temperature

 

pH at Temperature   g/L for 0.05 M Solution
5 °C 25 °C 37 °C   Trizma® HCL Trizma® Base
7.76 7.20 6.91   7.02 0.67
7.89 7.30 7.02   6.85 0.80
7.97 7.40 7.12   6.61 0.97
8.07 7.50 7.22   6.35 1.18
8.18 7.60 7.30   6.06 1.39
8.26 7.70 7.40   5.72 1.66
8.37 7.80 7.52   5.32 1.97
8.48 7.90 7.62   4.88 2.30
8.58 8.00 7.71   4.44 2.65
8.68 8.10 7.80   4.02 2.97
8.78 8.20 7.91   3.54 3.34
8.88 8.30 8.01   3.07 3.70
8.98 8.40 8.10   2.64 4.03
9.09 8.50 8.22   2.21 4.36
9.18 8.60 8.31   1.83 4.65
9.28 8.70 8.42   1.50 4.90
9.36 8.80 8.51   1.23 5.13
9.47 8.90 8.62   0.96 5.32
9.56 9.00 8.70   0.76 5.47

 

Phosphate Buffer Preparation Table – 0.2M solution  

 

Potassium 
Phosphate Monobasic Anhydrous g/L
Sodium Phosphate Dibasic
 Heptahydrate g/L
23 °C pH   Potassium Phosphate Monobasic Anhydrous g/L Sodium Phosphate
Dibasic Heptahydrate g/L
23 °C pH
22.4 3.49 5.7   10.80 29.51 6.9
22.08 4.29 5.8   9.36 32.73 7.0
21.60 5.37 5.9   7.92 35.95 7.1
21.05 6.60 6.0   6.72 38.63 7.2
20.40 8.05 6.1   5.52 41.31 7.3
19.56 9.93 6.2   4.56 43.46 7.4
18.60 12.07 6.3   3.84 45.07 7.5
17.64 14.22 6.4   3.12 46.68 7.6
16.44 16.90 6.5   2.52 48.55 7.7
15.00 20.12 6.6   2.04 49.09 7.8
13.56 23.34 6.7   1.68 49.89 7.9
12.24 26.29 6.8   1.27 50.81 8.0

Preparation of Citric Acid – Na2HPO4 Buffer Solutions, pH approx. 2.6–7.61

Citric acid monohydrate, C6H8O7 • H2O, M. wt. 210.14; 0.1M-solution contains 21.01 g/l. Na2HPO4, M. wt. 141.98; 0.2M-solution contains 28.40 g/l, or Na2HPO4 • 2H2O, M. wt. 178.05; 0.2M-solution contains 35.61 g/l.

x ml 0.1M-citric acid and y ml 0.2M-Na2HPO4 mixed.

pH x ml 0.1M-citric acid y ml 0.2-Na2HPO4
2.6 89.10 10.90
2.8 84.15 15.85
3.0 79.45 20.55
3.2 75.30 24.70
3.4 71.50 28.50
3.6 67.80 32.20
3.8 64.50 35.50
4.0 61.45 38.55
4.2 58.60 41.40
4.4 55.90 44.10
4.6 53.25 46.75
4.8 50.70 49.30
5.0 48.50 51.50
5.2 46.40 53.60
5.4 44.25 55.75
5.6 42.00 58.00
5.8 39.55 60.45
6.0 36.85 63.15
6.2 33.90 66.10
6.4 30.75 69.25
6.6 27.25 72.75
6.8 22.75 77.25
7.0 17.65 82.35
7.2 13.05 86.95
7.4 9.15 90.85
7.6 6.35 93.65

Preparation of Citric Acid – Sodium Citrate Buffer Solutions, pH 3.0–6.21

Citric acid monohydrate, C6H8O7 • H2O, M. wt. 210.14; 0.1M-solution contains 21.01 g/l. Trisodium citrate dihydrate, C6H5O7Na3 • 2H2O, M. wt. 294.12; 0.1M-solution contains 29.41 g/l.

x ml 0.1M-citric acid and y ml 0.1M-trisodium citrate mixed.

pH x ml 0.1 M-citric acid y ml 0.1M-trisodium citrate
3.0 82.0 18.0
3.2 77.5 22.5
3.4 73.0 27.0
3.6 68.5 31.5
3.8 63.5 36.5
4.0 59.0 41.0
4.2 54.0 46.0
4.4 49.5 50.5
4.6 44.5 55.5
4.8 40.0 60.0
5.0 35.0 65.0
5.2 30.5 69.5
5.4 25.5 74.5
5.6 21.0 79.0
5.8 16.0 84.0
6.0 11.5 88.5
6.2 8.0 92.0

Preparation of Sodium Acetate – Acetic Acid Buffer Solutions, pH 3.7–5.61

Sodium acetate trihydrate, CH3COONa • 3H2O, M. wt. 136.09; 0.2M-solution contains 27.22 g/l.

x ml 0.2M-NaOAc and y ml 0.2M-HOAc mixed.

pH, 18 °C x ml 0.2M-NaOAc y ml 0.2M-HOAc
3.7 10.0 90.0
3.8 12.0 88.0
4.0 18.0 82.0
4.2 26.5 73.5
4.4 37.0 63.0
4.6 49.0 51.0
4.8 59.0 41.0
5.0 70.0 30.0
5.2 79.0 21.0
5.4 86.0 14.0
5.6 91.0 9.0

Preparation of Na2HPO4 – NaH2PO4 Buffer Solutions, pH 5.8–8.0 at 25 °C1

Na2HPO4 • 2H2O, M. wt. 178.05; 0.2M-solution contains 35.61 g/l. Na2HPO4 • 12H2O, M. wt. 358.22; 0.2M -solution contains 71.64 g/l. NaH2PO4 • H2O, M. wt. 138.01; 0.2M-solution contains 27.6 g/l. NaH2PO4 • 2H2O, M. wt. 156.03; 0.2M-solution contains 31.21 g/l.

x ml 0.2M-Na2HPO4, y ml 0.2M-NaH2PO4; diluted to 100 ml with H2O.

pH, 25 °C x ml 0.2M-Na2HPO4 y ml 0.2M-NaH2PO4
5.8 4.0 46.0
6.0 6.15 43.85
6.2 9.25 40.75
6.4 13.25 36.75
6.6 18.75 31.25
6.8 24.5 25.5
7.0 30.5 19.5
7.2 36.0 14.0
7.4 40.5 9.5
7.6 43.5 6.5
7.8 45.75 4.25
8.0 47.35 2.65

Preparation of Imidazole (glyoxaline) – HCl Buffer Solutions, pH 6.2–7.8 at 25 °C1

Imidazole, C3H4N2, M. wt. 68.08.

25 ml 0.2M-imidazole (13.62 g/l), x ml 0.2M-HCl, diluted to 100 ml with H2O.

pH, 25 °C x ml 0.2M-HCl
6.2 21.45
6.4 19.9
6.6 17.75
6.8 15.2
7.0 12.15
7.2 9.3
7.4 6.8
7.6 4.65
7.8 3.0

Preparation of Sodium Carbonate – Sodium Bicarbonate Buffer Solutions, pH 9.2–10.81

Na2CO3 • 10H2O, M. wt. 286.2; 0.1M-solution contains 28.62 g/l. NaHCO3, M. wt. 84.0; 0.1M-solution contains 8.40 g/l.

x ml 0.1M-Na2CO3 and y ml 0.1M-Na2HCO3 mixed.

pH x ml 0.1M-Na2CO3 y ml 0.1M-NaHCO3
20 °C 37 °C
9.2 8.8 10 90
9.4 9.1 20 80
9.5 9.4 30 70
9.8 9.5 40 60
9.9 9.7 50 50
10.1 9.9 60 40
10.3 10.1 70 30
10.5 10.3 80 20
10.8 10.6 90 10

 

Helpful Buffer Preparation Formulas and Equations

Percentage by weight (w/v)

(% buffer desired / 100) × final buffer volume (ml) = g of starting material needed.

Molar Solutions

desired molarity × formula weight × solution final volume (L) = grams needed

Henderson-Hasselbach Equation

Henderson-Hasselbach Equation

Reference

 

  1. Dawson, R. M. C.; Elliot, D. C.; Elliot, W. H.; Jones, K. M. Data for Biochemical Research; 3rd ed., Oxford Science Publ., 1986.

 

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