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p. 7
The buffering ion should carry the same charge as the ion exchange group
and should have a pKa within 0.5 pH units of the pH used in the separation.
Buffering ions of opposite charge may take part in the ion exchange process
and cause local disturbances in pH.
Starting pH
Cation exchangers (SP): At least 1 pH unit below the pI of substance to be
bound.
Anion exchangers (Q): At least 1 pH unit above the pI of substance to be
bound.
Table 2. Buffers for cation exchange chromatography.
pH interval Substance Conc. (mM) Counter-ion pKa (25 °C)
1
1.4–2.4 Maleic acid 20 Na
+
1.92
2.6–3.6 Methyl malonic acid 20 Na
+
or Li
+
3.07
2.6–3.6 Citric acid 20 Na
+
3.13
3.3–4.3 Lactic acid 50 Na
+
3.86
3.3–4.3 Formic acid 50 Na
+
or Li
+
3.75
3.7–4.7; 5.1–6.1 Succinic acid 50 Na
+
4.21; 5.64
4.3–5.3 Acetic acid 50 Na
+
or Li
+
4.75
5.2–6.2 Methyl malonic acid 50 Na
+
or Li
+
5.76
5.6–6.6 MES 50 Na
+
or Li
+
6.27
6.7–7.7 Phosphate 50 Na
+
7.20
7.0–8.0 HEPES 50 Na
+
or Li
+
7.56
7.8–8.8 BICINE 50 Na
+
8.33
1
Ref: Handbook of chemistry and physics, 83
rd
edition, CRC, 2002–2003.