Showa Denko Shodex Asahipak NH2P-50 10E User manual

Operation Manual Shodex Asahipak NH2P-50 10E, NH2P-90 20F, NH2P-130G 7B Version 150305E
Operation Manual
Shodex™ Asahipak™NH2P-50 10E,
NH2P-90 20F, NH2P-130G 7B
Europe, Middle East, Africa, Russia
www.shodex.de
Sales Office:
Showa Denko Europe GmbH
Shodex Business
Konrad-Zuse-Platz 3
81829 Munich
Germany
Technical Support:
[e-mail]
+49 (0)89 / 93 99 62 37
Quotes and Orders:
[e-mail]
+49 (0)89 / 93 99 62 34
All manuals and user guides at all-guidesbox.com
all-guidesbox.com

Operation Manual Shodex Asahipak NH2P-50 10E, NH2P-90 20F, NH2P-130G 7B Version 150305E
2
Operation Manual
Shodex™ Asahipak™ NH2P-50 10E,
NH2P-90 20F, NH2P-130G 7B
(Please read this manual carefully before using the column to ensure performance and life.)
1. Introduction
The Shodex Asahipak NH2P series is packed with a polymeric gel containing chemically bonded
polyamine groups. Under the chromatographic conditions generally employed for the silica-base
amino columns, the NH2P column provides equal resolution and far greater reproducibility. NH2P
column can be used as Hydrophilic Interaction chromatography [HILIC] mode and ion-exchange
chromatography [IEC] mode.
2. Handling Instructions
Caution! Please consult the MSDS for the reagents and solvents used with the columns for health
concerns caused by acute exposure due to leakage from the column or adjoining tubing.
Attention! Use the column within the regular range of flow rate, pressure and temperature. The
column performance may deteriorate when it is handled beyond the permissible range even for a short
time. See the Section 4. Usable Conditions for the permissible ranges.
3. Specifications
Product code
Product name
Size (mm)
Particle size
(μm)
Plate Number
(per column)
ID
Length
F6830001
Asahipak NH2P-50 10E
10.0
250
5
≥10,000
F6710016
Asahipak NH2P-50G 4A
4.6
10
5
(Guard column)
F6830031
Asahipak NH2P-90 20F
20.0
300
9
≥10,000
F6710017
Asahipak NH2P-130G 7B
7.5
50
13
(Guard column)
Packing material: Polyvinyl alcohol
Functional group: Amino (Polyamine)
Column material: Stainless steel (SUS 316)
Screw type: Unified Thread Standard No. 10-32 UNF
Shipping solvent: Acetonitrile
4. Usable Conditions
Product name
Flow rate (mL/min)
Max pressure
(MPa)
pH range
Temperature
range (°C)
Standard
Max
NH2P-50 10E
2.0 - 3.0
5.0
15
2-13
4-50
NH2P-50G 4A
-
-
-
NH2P-90 20F
5.0 - 10.0
12.0
10
NH2P-130G 7B
-
-
-
All manuals and user guides at all-guidesbox.com

Operation Manual Shodex Asahipak NH2P-50 10E, NH2P-90 20F, NH2P-130G 7B Version 150305E
3
Attention! High-temperature operation may result in the generation of an air bubble, necessitating
degassing. Low-temperature operation may require reduced flow rates, because of increased eluent
viscosity.
5. System Clean Up
Clean up the LC system including the injector and the sample loop by switching the valve and flow the
eluent before column installation.
Attention! The previous eluent used for analyses in the system may damage the column, if it is not
compatible with the column.
Attention! When replacing nonpolar solvent to water, replace first with methanol and then replace
with water. When replacing buffer solution to acetonitrile/water, replace first with 100% water and
then replace with eluent. Substances adsorbed in the pump and tubing may not be compatible with
the column.
6. Column Installation
Install and use the Asahipak NH2P column with the flow through the column matching the flow
direction arrow on the column tag. Set the flow rate at half of standard rate and connect the column.
Flow at the low rate until the column temperature increases to the setting temperature, and then
increase the flow rate to the analytical condition.
Attention! The column should always be installed in the manner above, for safe and effective
operation.
7. Eluents
HILIC mode
IEC mode
Operational
solvents
Aqueous solutions of acetonitrile (or ethanol*1) at
any ratio, is used as eluent.*2
Any salts*3soluble in the solution above (such as
tetrapropylammonium acetate or sodium acetate)
are available.
*1With aqueous ethanol solutions, the flow rate
should be lower than normal, due to relatively high
viscosity.
*2 Other organic solvents are not guaranteed as the
eluent.
*3Maximum concentration of the buffer depends on
the ratio of acetonitrile (or ethanol). Please note any
precipitation of salt when usinq buffer!
Buffer solutions such as
phosphate, acetate, and tris.
Aqueous solutions of NaCl,
KCl, or Na2SO4
(pH 2-13)
Eluent
modes
lsocratic, gradient, or stepwise elution
Attention! Filter the eluent with a membrane filter (0.45 μm) to prevent chromatogram noise and
column performance deterioration by small particles or undissolved materials.
All manuals and user guides at all-guidesbox.com

Operation Manual Shodex Asahipak NH2P-50 10E, NH2P-90 20F, NH2P-130G 7B Version 150305E
4
Attention! When the shipping solvent (100% acetonitrile) is replaced with a buffer solution, please flow
100% water to the column before flowing the buffer solution to avoid the precipitation of the buffer.
When replacing buffer solution to acetonitrile solution, also flow 100% water to the column before
flowing acetonitrile.
Attention! Shodex line filter, Asahipak NH2P-LF (F6710100), is a filter for removing undesired
contaminants in the eluant. By installing the filter between the pump and the injector, deterioration
of the column is reduced.
Attention! Column equilibration for using acetonitrile/buffer eluent
Anions in the buffer interact with the amino functional groups of NH2P packing gel. The reproducibility
of the analysis will not be attained until the ionic equilibrium is attained between the anions and the
amino groups. In such case, the equilibration time may be shortened by the operation below. This
equilibrium operation is not needed when acetonitrile/water or buffer eluents are used.
HILIC mode
IEC mode
Eluent
Acetonitrile / buffer
Acetonitrile / water
Buffer
Solution
The same buffer without acetonitrile
Unnecessary
Unnecessary
Quantity
10 to 20 times the column volume
Flow rate
Lower than half of the normal flow rate
8. Sample Preparation
HILIC mode
IEC mode
Sample
Preparation
Solid
Sample
Eluent:
Acetonitrile/
water
Dissolve sample in water or the
eluent and add acetonitrile to
obtain 50% or higher acetonitrile
aqueous solution.
Dilute with the
eluent
Eluent:
Acetonitrile/
buffer
Dissolve sample in the buffer or
the eluent; and add acetonitrile
to obtain 50% or higher
acetonitrile aqueous solution.
Aqueous
Sample
Add acetonitrile to obtain 50% or higher
acetonitrile aqueous solution.
Injection volume
Smallest possible quantity
Optimum separation efficiency is generally obtained with sample matrix similar to the eluent
composition. Use acetonitrile in the sample matrix whenever possible.
Attention! Filter the sample with a membrane filter (0.45 μm) to prevent blockage.
Attention! In case of gradient condition, dilute the sample with the initial eluent.
All manuals and user guides at all-guidesbox.com

Operation Manual Shodex Asahipak NH2P-50 10E, NH2P-90 20F, NH2P-130G 7B Version 150305E
5
9. Column Cleaning
Elution characteristics of a column may change considerably after long, repeated usage, due to the
accumulation of pollution components on the packing material, for example metal ions from the LC
system or the sample. The cleaning procedures outlined below may be used. Clean the guard column
and analytical column separately by flowing the cleaning solution in the opposite direction of the arrow
on the column tag. The applied flow rate should be lower than standard flow rate.
Cleaninq method 1 (Standard cleaninq)
Cleaning solution
NH2P-50 10E
NH2P-50G 4A
NH2P-90 20F
NH2P-130G 7B
Flow rate
1.5 mL/min
1.5 mL/min
5.0 mL/min
5.0 mL/min
1
H2O
15 min
1 min
23 min
1 min
2
0.1 M NaOH aq.
191 min
5 min
275 min
2 min
3
H2O
30 min
2 min
46 min
2 min
4
Eluent
95 min
3 min
138 min
1 min
Cleaninq method 2 (Cleaninq metal ions)
Cleaning solution
NH2P-50 10E
NH2P-50G 4A
NH2P-90 20F
NH2P-130G 7B
Flow rate
1.5 mL/min
1.5 mL/min
5.0 mL/min
5.0 mL/min
1
H2O
15 min
1 min
23 min
1 min
2
0.1 M HClO4aq.
191 min
5 min
275 min
2 min
3
H2O
15 min
1 min
23 min
1 min
4
0.1 M NaOH aq.
191 min
5 min
275 min
2 min
5
H2O
30 min
2 min
46 min
2 min
6
Eluent
95 min
3 min
138 min
1 min
Caution! Do not mix the HClO4solution with the NaOH solution, as NaClO4is produced when mixed -
NaClO4is a combustible and explosive compound. Flow the acidic and alkali waste into separate
bottles, never concentrate or dry the wastes, and process each waste promptly.
Attention! Complete the washing procedure steps continuously. Do not store the column with either
acid or alkaline solution as it will advance the deterioration of the column.
Attention! Flow the alkaline waste into a separate bottle without flowing to the detector. The strong
alkaline solution may cause damage to the cell of the detector.
10. Column Inspection
Column inspection method is described in Certificate of Analysis (CoA).
11. Attention
1) Do not remove the end fittings of the column to prevent performance deterioration and for safety
reasons.
2) Do not make a strong impact on the column: such as hitting or dropping on the floor.
3) Replace the solvent in the LC system with the eluent to be used before connecting the column.
4) Connect the column so that the flow direction corresponds to the arrow mark on the tag.
5) When the column is not used for two weeks or more, replace the in-column solvent with the
shipping solvent, remove it from the LC system, close each end with a stopper, and store it at controlled
room temperature.
6) Contact Shodex website (www.shodex.de) or Shodex partners regarding product and analysis
applications.
All manuals and user guides at all-guidesbox.com

Operation Manual Shodex Asahipak NH2P-50 10E, NH2P-90 20F, NH2P-130G 7B Version 150305E
6
12. Warranty (Version 3)
1) Showa Denko K.K. warrants that the ShodexTM Column, at the time of delivery to the user, will
conform to the specification of the attached Certificate of Analysis, if the Shodex Column is used in
accordance with the attached operating manual. The foregoing warranty is exclusive and is in lieu of
all other warranties with respect to the Shodex Column, whether written, oral, implied, statutory or
otherwise. No warranties by Showa Denko K.K. are implied or otherwise created, including, but not
limited to, the warranty of merchantability and fitness for particular purposes.
2) Any claim of inconformity to the specification must be notified to Showa Denko K.K. within ten (10)
days after delivery to the user. User’s exclusive remedy and Showa Denko K.K.‘s exclusive liability for
such claim are limited to the replacement of the Shodex Column in question. In no event is Showa
Denko K.K. liable for any indirect, incidental or consequential damage arising out of in connection with
the Shodex Column, whether or not such damage is allegedly based on breach of warranty, negligence
or otherwise.
3) No warranty is made in any of the following cases:
3-1) If the Shodex Column is not used in accordance with the operating manual
3-2) If the Shodex Column is remodelled by anyone other than person or firm designated by
Showa Denko K.K.
3-3) If the Shodex Column is disposed of
3-4) If the Shodex Column is resold by the user without giving prior written notice to Showa
Denko K.K.
3-5) If the performance of the Shodex Column is not conform to the specification of the
attached Certificate of Analysis due to any of the reasons below:
a) Computer virus
b) Impurities contained in the sample, reagent, gas air or cooling water provided by
the user
c) Breakdown or malfunction of equipment, apparatus or component used in
combination with the Shodex Column
d) Force majeure such as fire, earthquake, flood, other natural disaster, crime, riot, act
of terrorism, war or radioactive contamination
4) In no event is Showa Denko K.K. liable for
4-1) the results of analyses or preparations using the Shodex Column or any portion of the
same, including, but not limited to, the reliability, accuracy, efficacy and safety of said results,
and
4-2) the occupational hazard in the use of the Shodex Column, whether or not such use is made
in accordance with the attached Conditions for use.
5) The Shodex Column is for laboratory use only. It must not be used for clinical diagnosis. Showa
Denko K.K. is not liable for any use of the Shodex Column except laboratory use.
All manuals and user guides at all-guidesbox.com
all-guidesbox.com

Operation Manual Shodex Asahipak NH2P-50 10E, NH2P-90 20F, NH2P-130G 7B Version 150305E
7
Product Information: Shodex HILIC Columns
Shodex HILIC columns are polymer-based (polyvinyl alcohol particles) for better stability (pH 2-13). This
results in a longer life time of the columns than silica-based columns. They can be used in a
temperature range from 4 to 60°C. The HILIC columns are suitable for RI (isocratic), evaporative light
scattering, corona charged aerosol and LC-MS/MS detectors (gradient mode). The PEEK housing of the
HILICpak series prevents adsorption of ionic compounds. Guard columns are available for each series
and size. Please refer to the individual operation manuals for detailed specifications or ask for technical
support for your application.
Asahipak NH2P: Amino (primary -NH2)
For very polar small organic compounds (like stevia, water-soluble vitamins)
Saccharides, oligosaccharides (up to DP15)
Pharmaceuticals
HILICpak VG: Amino (tertiary -NR2)
For very polar small organic compounds (like melamine)
For reducing sugars (especially aldoses, also lactulose)
Pharmaceuticals, organic acids, amino acids
HILICpak VT: Quaternary Ammonium (-NR3+)
For polar anionic compounds (like glyphosate, phosphorylated sugars)
HILIC + anion exchange mode
HILICpak VC: Carboxyl (-COO-)
For polar cationic compounds (like choline, acetylcholine, amino acids, neurotransmitters)
Pharmaceuticals (like metformin)
HILIC + cation exchange mode
HILICpak VN: Diol (-OH)
For oligosaccharides (up to DP25) (like hydrolysed cellulose or dextran)
For biomolecules (oligo DNA)
Product
code
Product name
Column Size
(mm)
Particle
Size
Pressure
Plate Number
(per column)
Housing
F7630005
Asahipak NH2P-50 4B
4.6 x 50
5 µm
5 MPa
≥1,500
Steel
F7630002
Asahipak NH2P-50 4D
4.6 x 150
5 µm
10 MPa
≥ 5,500
Steel
F7630001
Asahipak NH2P-50 4E
4.6 x 250
5 µm
15 MPa
≥ 7,500
Steel
F7630006
Asahipak NH2P-50 2D
2.0 x 150
5 µm
10 MPa
≥ 3,500
Steel
F7630007
Asahipak NH2P-40 3E
3.0 x 250
4 µm
13 MPa
≥ 8,500
Steel
F7630010
Asahipak NH2P-40 2E
2.0 x 250
4 µm
20 MPa
≥ 7,000
Steel
F7630200
HILICpak VG-50 4D
4.6 x 150
5 µm
10 MPa
≥ 5,500
Steel
F7630100
HILICpak VG-50 4E
4.6 x 250
5 µm
15 MPa
≥ 7,500
Steel
F7630300
HILICpak VG-50 2D
2.0 x 150
5 µm
15 MPa
≥ 3,500
PEEK
F7630400
HILICpak VT-50 2D
2.0 x 150
5 µm
10 MPa
≥ 4,500
PEEK
F7630700
HILICpak VC-50 2D
2.0 x 150
5 µm
10 MPa
≥ 3,500
PEEK
F7630500
HILICpak VN-50 4D
4.6 x 150
5 µm
10 MPa
≥ 10,000
PEEK
F7630600
HILICpak VN-50 2D
2.0 x 150
5 µm
10 MPa
≥ 3,500
PEEK
All manuals and user guides at all-guidesbox.com