Page 30 - Shimadu Consumables Catalog

Shim-pack Arata Peptide Columns

Excellent Seperation Performance of Peptides

Typically, in order to obtain good peak shape of peptides under reversed phase chromatography, TFA containing mobile phases
are frequently used which the ion pairing effect is relatively strong. However, TFA could cause ion suppression in LC/MS(/MS)
analysis. Excellent peak shape and separation performance for peptides could be achieved on the Shim-pack Arata LC column
even under formic acid (weak ion paring acid) containing mobile phase conditions, which are suitable for LC/MS(/MS) without the
use of typical ion pairing agents.

Increased Assurance of Peptide Analysis ~ Shim-pack Arata Peptide C 18 column ~
In order to ensure lot-to-lot reproducibility in peptide analysis, each lot of Shim-pack Arata Peptide
C18material is tested using a mixture of peptide standards in addition to the standard Shim-pack
Arata C18 lot QCtest. This test is carried out under severe condition using 0.1 % formic acid mobile
phase to help ensure consistent column performance for requirements of customers under regulated
requirements.use of typical ion pairing agents.

Excellent Seperation Performance of Peptides
When analyzing peptides on a typical ODS column with 0.1% formic acid mobile phase, not only poor peak shape but also
long equilibration time required to obtain stable retention times and area is a common problem. Shim-pack Arata Peptide C18
columnscan be rapidly equilibrated even in 0.1% formic acid containing mobile phase, achieving excellent peak shape, stable
retention andarea of peptides at the same time.
H
N
N
O
N OH
O NH H O
O
ON H2 H O H O H O N
HO N N N N N N H
O H O H O
NH OH
H
HN NH2 N
N Angiotensin ᶗ
Compounds Equilibration time (min) 30 60 120 180 240 360 CV
Retention time (min) 5.05 5.03 5.03 5.03 5.04 5.06 0.3
Angiotensin ᶗ Symmetry factor 1.26 1.24 1.25 1.25 1.24 1.25 0.4
Area 2.17×10 5 2.13×10 5 2.26×10 5 2.25×10 5 2.23×10 5 2.24×10 5 1.9

Analytical Conditions
LC system : NexeraX2 MP_SPD20A (Semi-micro Cell) Detection : 214 nm
LC column : Column temp. : 40 °C
Shim-pack Arata Peptide C18 (3.0 × 75 mm, 2.2 μm) Injection volume : 1 μL
Typical ODS (3.0 × 75 mm, Sub 2 μm) Sample : Angiotensin I
Mobile Phase A : 0.1% HCOOH in H2O Vial : Torast-H Bio Vial
Mobile Phase B : 0.1% HCOOH in CH3CN
Flow rate : 0.4 mL/min
*Peptide is usually analyzed using gradient conditions. Isocratic condition was used for this
application in order to show the difference of LC columns more clearly. Acetonitrile concen-
tration was adjusted in order that the retention time of peptide on each column become
similar.
Compounds Equilibration time (min) 30 60 120 180 240 360 CV
Retention time (min) 7.78 7.15 6.53 6.20 5.98 5.82 11.0
Angiotensin ᶗ Symmetry factor 4.89 4.38 4.85 4.87 4.93 4.92 3.9
Area 1.65×10 5 1.58×10 5 1.76×10 5 1.78×10 5 1.83×10 5 1.84×10 5 5.3
Both columns were new columns (shipping solvent: acetonitrile) and equilibrated with mobile phase without any conditioning.
Angiotensin I was analyzed after a certain period of equilibration and retention time, symmetry factor and area of Angiotensin I
were compared. The typical ODS showed poor peak symmetry and unstable retention time even after 360 minutes of
equilibration. In contrast, Shim-pack Arata peptide C18 already showed stable retention after 30 minutes of equilibration and
excellent peak shape was obtained.

Pg A-26