HPLC Column Care Instructions

Shipping and Storage Solvents

Phase Shipping Solvent Storage Solvent
Reverse Phase – C18, C8, C4, C30, AQ, PH, PFP, DP, Phenyl, Phenyl-hexyl, etc. MeCN:Water, e.g. 65:35 MeCN or MeOH
Normal Phase - CN, Diol, Silica, NH2* n-Hexane:Ethanol, e.g. 95:5 n-Hexane:Ethanol or Heptane:IPA

* MeCN (Inspire NH2)
Polar Reverse Phase - CN, Diol, NH2* n-Hexane:Ethanol, e.g. 95:5 MeCN

* MeCN (Inspire NH2)
HILIC and Mix-Mode Phase - Hilic, MMP-G, MMP-H, MMP-L, MMP-M, MMP-Q, MMP-Y, MMP-Z, etc. MeCN MeCN
Ion exchange Phase - SAX, SCX MeOH MeOH

General Column Care

  • Dropping or otherwise "shocking" columns can disrupt the column bed and cause peak splitting.
  • The lower the operating pressure, the longer the operating life.

HPLC packings are subject to a rigorous array of QC tests in a ISO9001 compliant facility, with special emphasis on reagent purity, raw material traceability, and consistency in raw materials and finished products.

A detailed analysis of all physical properties, chemical purity, chromatographic selectivity, and column packing efficiency is used to confirm that each lot of column is identical to all previous lots.

We know how important column consistency is to your work, so Dikma Technologies goes to great lengths to make certain that every HPLC column we ship to our customers is of the highest quality possible.

When You Receive an HPLC Column

When you receive an HPLC Column from Dikma Technologies, please take the following simple steps:

  • Check the column for signs of physical damage which may have occurred during shipping. Contact Dikma Technologies immediately to report any problems.
  • Make sure that the column you received is the column that you ordered.
  • Take note of the solvent contained in the column during shipping. The solvent used for shipping is the same as that used as the mobile phase on the QC test chromatogram, except for HILIC, Inspire NH2, Platisil CN, SAX, SCX, MMP-G, MMP-H, MMP-L, MMP-M, MMP-Q, MMP-Y and MMP-Z. Before attempting to change solvents, make certain that the eluent you will be introducing into the column is COMPLETELY miscible with the eluent contained in the column, to avoid precipitation of buffer salts or other mobile phase additives.
  • Test the column to verify column efficiency and back pressure using one of the later-eluting components of the QC test sample. Contact Dikma Technologies immediately to report any problems.

Method Considerations

There are several important things about your analytical method that will greatly affect column life and column performance, including sample preparation, solvent selection, and solvent preparation.

Sample Preparation

Non-ideal chemical and physical interaction of samples with the column frits and column packings is a primary source of problems. Most columns fail because the frit becomes clogged or the stationary phase at the head of the column becomes contaminated.

Samples should be filtered prior to injection. Even sample solutions which appear to be particulate-free can contain small solids which can clog the pores of the column inlet frit. Samples should be filtered through a 0.45µm or 0.2µm syringe filter before injection.

In addition to filtering samples, Guard cartridges can be used to trap "problems" before they reach the analytical column. Guard cartridges are essentially tiny HPLC columns that are cheaper to replace than the analytical column. Please contact Dikma Technologies for ordering information for guard cartridges.

Solvent Selection

Columns last longest when they are used with benign eluents. Using eluents of high pH or low pH can dissolve silica or catalyze hydrolysis of the bonded phase. Try to stay within the pH range of the column. If you must use a pH outside this range, column life might be reduced.

Solvent Preparation

Use a 0.45µm or 0.2µm filter for solvents as well as samples, even HPLC grade solvents!

Column Storage

Column storage conditions can have a profound effect on column lifetime and performance-after-storage.

Before extended storage (e.g. greater than 2 days), rinse the column COMPLETELY free of eluents containing buffers, ion-pair reagents, or inorganic solutes, by flushing with 20-50 column volumes of the eluent WITHOUT the dissolved additives.

Then flush the column with 20 column volumes of storage solvent shown above.

In-Situ Column Cleaning

Columns that become fouled over time can sometimes be rejuvenated with an aggressive rinsing sequence, as shown below.

Flush the column with 20-30 column volumes of the indicated solvents in the indicated sequences.

Please confirm the column volumes before initiating the rinsing sequence.

Reverse Phase Mode C18, C8, C4, PH, NH2*, CN, Diol, AQ, C30

10% Methanol-90% H2O → Methanol → Isopropanol → Methanol
NH2 50% Acetonitrile-50% H2O → Acetonitrile → Isopropanol → n-Hexane → Isopropanol → Acetonitrile
Normal Phase Mode Silica, CN, Diol, NH2

n-Hexane → Dichloromethane → Isopropanol → Dichloromethane → n-Hexane
HILIC and Mix-Mode Hilic, MMP-G, MMP-H, MMP-L, MMP-M, MMP-Q, MMP-Y, MMP-Z

40% Acetonitrile-60% H2O → Acetonitrile
Ion Exchange Mode SAX, SCX

H2O → Methanol
Note: Isopropanol has a higher viscosity, and the flow rate should be reduced for flushing.

Maximum Operating Conditions

Maximum Operating Temperature

60°C

Maximum Operating Pressures

Particle Size Maximum Operating Pressure
5 µm, 3 µm 400 bar
2.7 µm 600 bar
1.8 µm 1200 bar

Form 1 — Column Volume Calculation Table

Column Size Column Volume Rinse Volume Range
250 × 4.6 mm 4.15 mL 80–125 mL
150 × 3.0 mm 1.06 mL 20–32 mL
100 × 2.1 mm 0.35 mL 7–11 mL

Form 2 — Recommended pH Range

Endeavorsil

Bonded Phase Particle Size (µm) Carbon Loading (%) Endcapping pH
C18 1.8 20 Yes 1.5–9.0
C8 1.8 13 Yes 1.5–9.0
C18-A 1.8 20 Yes 1.5–9.0
C18-B 1.8 21 Yes 1.5–9.0

Leapsil

Bonded Phase Particle Size (µm) Carbon Loading (%) Endcapping pH
C18 2.7 27 Yes 1.5–10.0

Navigatorsil

Bonded Phase Particle Size (µm) Carbon Loading (%) Endcapping pH
C18 2.7 8 Yes 1.5–9.0
C8 2.7 5 Yes 1.5–9.0

Spursil

Bonded Phase Particle Size (µm) Carbon Loading (%) Endcapping pH
C18 3, 5, 10 25 Yes 1.5–10.0
C18-EP 3, 5, 10 24 Yes 1.5–10.0
C18-Amide 3, 5, 10 25 Yes 1.5–10.0
AQ 3, 5, 10 24 Yes 1.5–9.0

Inspire

Bonded Phase Particle Size (µm) Carbon Loading (%) Endcapping pH
C18 3, 5, 10 27 Yes 1.0–11.0
C8 3, 5, 10 17 Yes 1.0–11.0
Silica 3, 5, 10 – No 2.0–7.5
Diol 3, 5, 10 7.5 No 2.0–7.5
Hilic 3, 5, 10 – No 1.5–7.5
PFP 3, 5 15 Yes 1.5–7.5
DP 3, 5 16.5 Yes 1.5–8.0
Phenyl 3, 5 17.0 Yes 1.5–9.0
Phenyl-hexyl 3, 5 21.0 Yes 1.5–9.0
NH2 5 4.5 No 2.0–7.5

Luster

Bonded Phase Particle Size (µm) Carbon Loading (%) Endcapping pH
C18 5, 10 20 Yes 2.0–9.0
C8 5, 10 12 Yes 2.0–9.0
Diol 5, 10 5 No 2.0–8.0
Silica 5, 10 – No 1.5–7.5

Bio-Bond

Bonded Phase Particle Size (µm) Carbon Loading (%) Endcapping pH
C18 3, 5, 10 8 Yes 2.0–8.0
C8 3, 5, 10 5 Yes 2.0–8.0
C4 3, 5, 10 3 Yes 2.0–8.0

Platisil

Bonded Phase Particle Size (µm) Carbon Loading (%) Endcapping pH
ODS 5 15 Yes 1.0–11.0
CN 3, 5 12 Yes 1.5–7.5
PH 5 14 Yes 1.5–7.5
NH2 3, 5 7 No 2.0–7.5
Silica 5 – No 1.5–7.5

Silversil

Bonded Phase Particle Size (µm) Carbon Loading (%) Endcapping pH
C18 5 20 Yes 1.5–9.0
C8 5 12 Yes 1.5–9.0
Silica 5 – No 1.5–7.5

Diamonsil Plus

Bonded Phase Particle Size (µm) Carbon Loading (%) Endcapping pH
C18 3, 5 14 Yes 1.5–10.0
C18-A 5 15 Yes 1.5–10.0
C18-B 5 16 Yes 1.5–10.0
C8 5 9 Yes 1.5–10.0
C30 3, 5 16 Yes 1.5–8.5
SAX 5 3.5 No 2.0–7.0
SCX 5 4.5 No 2.0–7.5

Aurorasil

Bonded Phase Particle Size (µm) Carbon Loading (%) Endcapping pH
MMP-G 3, 5 17 Yes 1.5–10.0
MMP-H 3, 5 17 Yes 2.0–9.5
MMP-L 3, 5 17 No 2.0–7.5
MMP-M 3, 5 16 No 2.0–7.5
MMP-Q 3, 5 17 No 2.0–7.5
MMP-Y 3, 5 17 Yes 1.5–9.0
MMP-Z 3, 5 20 Yes 1.5–9.5
HPLC Column Care Instructions — Dikma Technologies
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