Shodex CLNpak EV-2000 User manual

ver.211118E
SHOWA DENKO K.K. (https://www.shodex.com/)
1
Operation Manual
Shodex

CLNpak

EV-2000
（Please read this manual carefully to achieve the best and consistent column performance for a long time）
Important Handling Instructions
Caution! · Please consult the Safety Data Sheet (SDS) of reagents and solvents used with the column
and understand their proper handling methods to prevent potential health hazards or death
from occurring.
· Please wear appropriate personal protective equipment such as lab goggles and gloves
when handling organic solvents and acid and alkaline reagents. Avoid any direct physical
contact to prevent chemical injuries.
Before Using the Column
(1) Please visually inspect the package and outside of the column for any damage.
(2) Please check if product name and serial number written on the column package, column adhesive label, and
enclosed Certificate of Analysis (CoA) are matching and correct.
1. Introduction
Thank you for purchasing the Shodexproduct. ShodexCLNpakseries is a GPC clean-up column
packed with a styrene divinylbenzene copolymer. It is suitable for clean-up of residual pesticide components in
foods.
2. Column Components
3. Column Specifications
Product
Code Product Name Column Size Particle
Size (μm)
Theoretical
Plate Number
(Per Column)
I.D. (mm) Length (mm)
F6090001 CLNpak EV-2000 20.0 300 16 ≥4,000
F6090002 CLNpak EV-G 20.0 100 16 (Guard Column)
Base Material : Spherical porous particles of styrene divinylbenzene copolymer
Column Housing : SUS-316
Screw Type : Internally-threaded type No.10-32 UNF
Shipping Solvent : Ethyl acetate/Cyclohexane = 3/7

ver.211118E
SHOWA DENKO K.K. (https://www.shodex.com/)
2
4. Usable Conditions
Usable solvents are listed below.
(1) The standard eluent is a mixture of ethyl acetate and cyclohexane.
Recommended ethyl acetate:cyclohexane ratio is between 2:8 and 3:7.
(2) Up to 100 % (v/v) acetone, ethyl acetate, cyclohexane, toluene, and dichloromethane are usable.
Attention! · Use the column within above stated flow rate, pressure, and temperature ranges. Using
the column outside the given range may damage the column and lower its performance.
· Column pressure is influenced by the eluent composition, flow rate, and column
temperature. When changing the eluent compositions, adjust the flow rate and column
temperature so that the column pressure remains below the usable maximum pressure.
· The high flow rate setting prevents the eluent that going through the column to reach the
set column oven temperature. It is recommend to equip a “coiled SUS tubing” inside the
column oven to help the situation. It should be placed in-between the injector and the
column. A 0.5 mm I.D. x 10 - 20 m SUS tubing is recommended for making the coil.
5. Eluent Preparation
(1) Degas the eluent fully to prevent the formation of air bubbles.
(2) Presence of small debris or insoluble substances may result in deterioration of the column and/or they appear
as noise on the chromatograms. Filter the eluent with a 0.45-μm disposable filter to prevent the problems.
Attention! · Use HPLC grade organic solvent whenever possible. Solvents left in an opened bottle for
a long time should not be used. The content may have been changed, absorbed moisture,
or has been contaminated.
· Always use freshly prepared solvents. Solvents stored for a long time may have changed
their compositions and may influence elution patterns and/or damage the column.
Note · Use of on-line degasser is recommended.
6. Sample Preparation
(1) If possible, use the eluent for analysis to dissolve or dilute samples. If this is difficult, use a solvent which
has a composition that is as close as possible to the eluent's composition, but which fully dissolves or dilutes
the sample.
(2) Filter the sample solution using disposable 0.45-μm filter to prevent the column from clogging or deteriorating.
(3) Recommended injection volume is less than 5 mL per column.
(4) Samples with large amount of lipids have high viscosities. This delays the elution of lipids and interferes its
separation from pesticides. Another potential problem of highly viscous samples is that a portion of sample
may be retained in the injector and not fully injected. Thus, dilution of such samples with eluent is
recommended. The approximate maximum lipid load per injection is less than 500 mg.
Note · Use of guard column is recommended to protect the analytical column.
Product Name Recommended
Flow Rate (mL/min)
Maximum Pressure (MPa)
(Per Column)
Recommended
Temperature (°C)
CLNpak EV-2000 4 - 5 2.5 40
CLNpak EV-G - -

ver.211118E
SHOWA DENKO K.K. (https://www.shodex.com/)
3
7. Column Usage Procedure
7.1 HPLC System Preparation
Wash entire LC system prior to the column installation, including all flow-lines and sample loop by switching the
valve, and then replace the washing solution with the eluent to be used. If desired new eluent has low
miscibility/solubility to the eluent of previous analysis, first use the eluent that is miscible/soluble to both eluents,
and then replace it with the desired eluent.
e.g. When replacing water to ethyl acetate/cyclohexane, first run methanol and then replace it with the eluent.
Attention! · If the eluent left in the system is not compatible with the column to be used, it may damage
the column.
· A drastic change in the eluent compositions may remove substances adsorbed on the
system and they may enter and deteriorate the column.
7.2 Column Installation
(1) Connect the column to LC system by following the “flow direction arrow” (➡) indicated on the column
adhesive label. If guard column is used, position the guard column in front (before the inlet) of the analytical
column.
(2) Make sure to insert the tubing all the way to the end fitting and secure it with the male nut. It is important that
there is no extra space between the tubing and the column side of the end fitting. Presence of an extra space
will let the sample to spread out and may result in wide peaks.
(3) Set the initial flow rate at less than 2.5 mL/min and start the system. If using the column at an elevated
temperature, keep a low flow rate until the temperature of the column reaches to the set temperature, and
then gradually increase the flow rate to the desired.
(4) Multiple number of columns can be connected in series. When connecting multiple columns with each having
different exclusion limits, set the column with higher exclusion limit at the upper stream position.
Caution! · Verify that there is no solvent leak. It may cause electronic leakage, rust, and/or chemical
injury.
Attention! · Make sure not to let air bubbles enter the column while installing the column. The air
bubbles may damage the column.
· When restarting the system after column installation or after holding the eluent flow, start
the system at less than 2.5 mL/min. A rapid increase in pressure can damage the column.
· If the column was used at an elevated temperature, lower the flow rate to less than
2.5 mL/min at the end of analysis. Then, turn off the column oven, and let the column
temperature return to room temperature before stopping the pump. If the pump was
stopped while the eluent inside the column was still hot, as the eluent temperature
decreases, its volume also decreases. This may result in creating an empty space in the
column and deteriorates the column.
Note · It is recommended to set the pump limiter to avoid exceeding the maximum pressure.
7.3 Solvent Exchange
(1) When replacing the solvent, set the column temperature at 40 °C and start the system at less than
2.5 mL/min. Recommended solvent volume to introduce at each step is 3 to 5 times of the column volume.
(2) Check miscibility/solubility of the desired new solvent and the solvent currently filled in the column.
(3) When replacing the current solvent with a miscible solvent, first introduce 1:1 mixture of the current solvent
and the new solvent, and then replace it with 100 % of the new solvent.

ver.211118E
SHOWA DENKO K.K. (https://www.shodex.com/)
4
(4) When replacing the current solvent with a solvent with low miscibility/solubility to the current solvent, first
use a solvent that is miscible/soluble to both eluents, and then replace it with the new solvent.
Attention! · Frequent solvent replacement deteriorates the column, and thus not recommended.
7.4 Column Cleaning
Problems in peak shapes and elution time changes or elevated column pressure are often caused by insoluble
or adsorbing components present in the eluent and reagents being deposited inside the column. These
problems may be resolved by cleaning the column.
If a guard column is used with the analytical column, first remove the guard column and check the performance
of the analytical column alone. If the problem is solved, most likely the cause is from the guard column. In this
case, clean the guard column.
If the problem is not solved by removing the guard column, clean both guard and analytical columns. Make sure
to clean the guard and the analytical columns separately. When washing the column, let the washing solution
flow from the column outlet go directly into the waste container and not let the solution go through the detector.
<Cleaning Method>
Insoluble components that block the column inlet may be removed by reversing the flow direction, i.e.,
introducing the eluent from the column outlet, with flow rate at less than half of the recommended flow rate.
8. Column Storage
Remove the column from the system keeping the solvent used for the analysis. Securely tighten the end caps
and store the column at a location with stable temperature (a cool and dark space is recommended). Refer to
section 7.3 Solvent Exchange for how to replace the eluent.
Attention! · Never allow inside of the column to dry. It can damage the column.
9. Column Inspection
Inspection method is described in the Certificate of Analysis (CoA).
Theoretical Plate Number (N) and Asymmetry Factor (Fas) were calculated using the below equations.
(1) Theoretical Plate Number (N)
(2) Asymmetry Factor (Fas)

ver.211118E
SHOWA DENKO K.K. (https://www.shodex.com/)
5
10. Additional Warnings
(1) Do not remove end fittings.
(2) Do not make a strong impact on the column. Do not drop or hit the column on a hard surface.
Refer to the Shodex™ website (https://www.shodex.com/) for product details and their applications.
For additional assistance, contact the distributor from whom you purchased the column or contact your regional
Shodexsupport office (https://www.shodex.com/en/support_office/list/).

ver.211118E
SHOWA DENKO K.K. (https://www.shodex.com/)
6
11. Warranty (Ver. 4COLUMN)
(1) Showa Denko K. K. warrants that the Shodexproduct, at the time of delivery to the user, will conform to
the specification of the attached CERTIFICATE OF ANALYSIS, if the Shodex product 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 product, 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 product 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 product, 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 product is not used in accordance with the operating manual
3-2 If the Shodex product is remodeled by anyone other than person or firm designated by Showa Denko K.K.
3-3 If the Shodex product is disposed of
3-4 If the Shodex product is resold by the user without giving prior written notice to Showa Denko K.K.
3-5 If the performance of the Shodex product 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
product
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 analysis or preparations using the Shodex product 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 product, whether or not such use is made in accordance
with the attached Conditions for use.
(5) The Shodex product 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 product except laboratory use.
Following names are trademarks or registered trademarks of SHOWA DENKO K.K.
Shodex, AFpak, Asahipak, AXpak, CLNpak, CXpak, HILICpak, MSpak, ODP, OHpak,
ORpak, RSpak, SUGAR, USPpak.
Products described in the operation manual are not intended for medical use or medical applications.