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Note: In CelerityCE capillaries virtually all of the residual silanols on the capillary wall have been eliminated by
replacement with hydride (Si-H) and basic peptides are not adsorbed assuring the intraday and interday repeatability
of migration times. The results are reproducible between different lots of capillaries. The capillary involved in the
study was used for 1128 runs, demonstrating the long-term stability.
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Synthetic Peptides by CE v. HPLC
Method Conditions:
Capillaries:
- CelerityCE™ C18 Capillary, i.d. 50µm, total length 50.0 cm, effective length 25.0cm, lot# 011028.
Column was first subjected to 109 injections of other peptides. Analyzed peptide was injected first time (injection
# 110).
- CelerityCE™ C18 Capillary, i.d. 50µm, total length 50.0 cm, effective length 23.5cm, lot# 011028.
Injection # 205.
Catalog No: 04918-50
- HPLC Column: TSK ODS 120T, 4.6 x 150 mm, 5 µm. Catalog No.: 04925-50
CE Conditions
Injection: 3 seconds @ 50 mbar
Stock Buffer: 38 mM Tris + 1.6 mM phosphoric acid, pH = 2.14
Run Buffer: stock buffer diluted 1:10, pH = 2.14 + 10% ethanol
Voltage: 25 kV
Instrument: Agilent 3D HPCE
HPLC Conditions
Linear gradient of 0-100% B over 60 min. Solvents: A, 0.1% TFA in water; B, 0.09% TFA in 60:40 (v/v)
acetonitrile/water.
Flow rate: 1 mL/min.
Detection: UV @ 214 nm for OTCE and HPLC
Sample: synthetic therapeutic peptide
Sequence: HTNIHQDQHNHFHR, 1 mg/mL of peptide in the running buffer
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Discussion:
A significant factor that often limits the usefulness of CZE is the lack of reproducibility of data from run to run.
CelerityCE™ capillaries eliminate these problems as shown in the data in Figures A and B. Two electro-chromatograms
show results after many injections: Figure (A) was done after the capillary was used for 109 injections of various
peptide samples.
Figure (B) represents the run after the capillary was used for more then 200 analyses. Reproducibility of the number
of peaks, peak shape, and migration times is excellent from run to run.
The RP-HPLC profile of the analyzed peptide sample obtained using a standard HPLC column and a standard linear gradient
exhibited only a broad peak (Figure C).
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