Table 2. buffers for anion-exchange chromatography – Bio-Rad UNO® Monolith Cation Exchange Columns User Manual
Page 4
while maintaining maximum buffering capacity. In any case, a buffer concentration
of 20 mM is recommended.
As can be seen in Table 2 and 3, the pK
a
and hence the pH of the buffer, changes
with temperature. Therefore the pH of the buffer must be adjusted at the working
temperature.
Table 2. Buffers for Anion-Exchange Chromatography
pH range
Buffer
Mwt
pK
a
@25 °C
Counter-ion
∆
pK
a
/°C
5.0 - 6.0
Piperazine
86.1
5.7
Cl
-
/HCOO-
-0.015
5.5 - 6.0
L-Histidine
155.2
6.15
Cl
-
5.8 - 7.2
Bis-Tris
209.2
6.5
Cl
-
-0.017
6.4 - 7.3
Bis-Tris Propane
282.3
6.8, 9.0
Cl
-
7.3 - 8.3
Triethanolamine
149.2
7.8
Cl
-
/CH
3
COO
-
-0.020
7.6 - 8.6
Tris
121.1
8.1
Cl
-
-0.031
8.4 - 8.8
Diethanolamine
105.1
8.9
Cl
-
-0.025
9.0 - 9.9
Ethanolamine
61.1
9.5
Cl
-
-0.029
9.8 - 10.3
1,3-diamino-propane 74.1
10.5
Cl
-
-0.026
Table 3. Buffers for Cation-Exchange Chromatography
pH range
Buffer
Mwt
pK
a
@25 °C
Counter-ion
∆
pK
a
/°C
3.6 - 4.3
Lactic acid
90.1
3.8
Na
+
4.2 - 5.2
Citric acid
192.1
3.1
Na
+
5.5 - 6.7
MES
195.2
6.1
Na
+
-0.011
6.1 - 7.5
PIPES
302.4
6.8
Na
+
-0.009
6.5 - 7.9
MOPS
209.3
7.2
Na
+
-0.006
6.7 - 7.6
Phosphate
120 (Monobasic)
7.2
Na
+
-0.003
142 (Dibasic)
6.8 - 8.2
TES
229.2
7.4
Na
+
-0.020
6.8 - 8.2
HEPES
238.3
7.5
Na
+
-0.014
7.4 - 8.8
Tricine
179.2
8.1
Na
+
-0.021
Always use buffer components of the highest purity available as UV-absorbing
impurities may cause baseline disturbances and interfere with the detection of pro-
tein peaks.
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