27
IEC
Peptide Purifications
Cation exchange chromatography is commonly used for
peptide purification. Table V shows the same particle size
availability of TOYOPEARL and TSKgel PW strong cation
exchange resins. Based on the needs for capacity and
resolution, an appropriate S or SP resin is selected for a
particular peptide application. TSKgel SP-3PW (30) is based
on a 25 nm pore resin. It was developed to provide high
binding capacities for peptides and small proteins. It also
has a different selectivitiy than TSKgel SP-5PW (30).
It is especially suited for insulin purification. Table VI
compares the capacity of TSKgel SP-3PW (30) to TSKgel
SP-5PW (30) and Source 30S resin. The improved resolving
power of TSKgel SP-3PW (30) resin is demonstrated in
Figure 14.
The new TOYOPEARL GigaCap Q-650S resin offers a low
pressure alternative to oligonucleotide purification with
TSKgel resins while preserving the selectivity, resolution
and yields of those higher pressure processes.
TABLE VI
Column: 4.6 mm ID x 7.5 cm L; Eluent: Acidic buffer (pH 3.0) containing neutral salt and 1-propanol
Flow rate: 0.75 mL/min (270 cm/h); Sample: Recombinant insulin (7.2 g/L)
DBC calculated at 10% breakthrough
INSULIN DYNAMIC BINDING CAPACITIES
TSKgel SP-3PW (30)
TSKgel SP-5PW (30)
PS-DVB (30) S Type Resin
Matrix
polymethacrylate
polymethacrylate
polystyrene divinylbenzene
Particle size
30 µm
30 µm
30 µm
Insulin DBC
49 g/L
24 g/L
45 g/L
Ion exchange capacity
0.12 eq/L
0.09 eq/L
0.08 eq/L
Pore size
25 nm
100 nm
NR
0
50
100
-15
35
85
135
185
235
285
335
385
435
0
50
100
150
200
Concentration B-buffer (%)
Concentration B-buffer (%)
Detector Response (mAu)
Detector Response (mAu)
Elution volume (mL)
0
50
100
80
180
280
380
480
580
680
780
0
50
100
150
200
Elution volume (mL)
N- 1
N- 1
shoulder
Purification of Oligonucleotides
Resin: TSKgel SuperQ-5PW (20); Column size: 6.6 mm ID × 18.5 cm (6.3
mL); Mobile phase: A: 20 mmol/L NaOH; B: 20 mmol/L NaOH, 3.0 mol/L
NaCl; Gradient: 50 % B (2 CV) 50-100 % B (15 CV), 100 % B (2 CV); Flow rate:
200 cm/h (1.14 mL/min); Detection: UV @ 254 nm; Sample load: 1.0 mg;
Sample: crude phosphorothioate deoxyoligonucleotide
Resin: TOYOPEARL GigaCap Q-650S; Column size: 6.6 mm ID × 18.5 cm
(6.3 mL); Mobile phase: A: 20 mmol/L NaOH, B: 20 mmol/L NaOH, 3.0 mol/L
NaCl 50 % B (2 CV); Gradient: 50-100 % B (15 CV), 100 % B (2 CV); Flow rate:
200 cm/h (1.14 mL/min); Detection: UV @ 254 nm; Sample load: 1.0 mg;
Sample: crude phosphorothioate deoxyoligonucleotide
Figure 13
Ion exchange
chromatography
