Waters Protein-Pak PW Ion-Exchange Column - Manual (Page 3)

[ Care and Use ManUal ]
Protein-Pak PW Ion-Exchange Columns 3
f. Column Purge Volumes
III. CaRe aND Use
Liquid chromatography columns have a finite life influenced by their care
and use, number of injections, sample and eluent cleanliness, frequency
of eluent changeover and handling and storage procedures among other
factors. If a change is observed in the:
• Retention of a particular compound
• Resolution between two compounds
• Peak shapeTake immediate steps to determine the reason for the
changes, and until the determination is made, the results of any
separations using the column must not be relied upon. Follow
generally accepted procedures for quality control and methods
development when using these columns.
Important Note: Before running the first analysis on your new column
perform the test sample separation given in the Test Conditions section.
a. Sample Preparation and Filtration
Use HPLC grade water. All buffer solutions should be filtered to remove
microparticulate matter above 0.45 μm. This reduces the problem of
plugged filters and preserves column life. If organic solvents are required,
use HPLC grade solvents.
Adequate sample cleanup (using Sep-Pak cartridges designed for
this purpose) prevents alteration of the column chemistry by strongly
adsorbing or precipitating sample components.
b. Flow Rate Range
Flow rate with Protein-Pak DEAE-5PW and SP-5PW, 7.5 mm i.d.
columns should be maintained at less than 1.2 mI/min. In general, flow
rates of 0.5-1 mI/min are recommended for 7.5 mm i.d. columns.
Maximum flow rate for 21.5 mm i.d. columns is 10 mL/min and the
recommended flow range is 5-10 mL/min.
c. Precautions
Maximum Flow Rate should not exceed: 1.2 ml/min for 7.5 mm i.d. columns
and 10 mL/min for 21.5 mm i.d. columns.
• Protein-Pak ion-exchange columns are compatible with acids or
bases with buffers in the pH range of 2-12.
• If using buffers containing halide ions, flush the entire system with
HPLC grade water following use.
• Try to dedicate columns to specific applications. Constant switching
of sample and solvents will cause a more rapid column contamina-
tion and loss of resolution.
• Filter all aqueous buffers. Avoid using turbid or cloudy buffers. Be
sure that any eluents containing buffers, salts, etc., are compatible
with the wetted surfaces of the column and equipment.
• DO NOT exceed 20% organic content in the mobile phase.
• DO NOT use the following on the type of column they are listed under:
• Protect column from vibration, mechanical shock, and rapid changes
in pressure. Column packings are based on a highly porous and rigid
resin. Any thermal, physical or chemical shock (such as changing
eluents rapidly or at high flow rates) can cause the particles to shift
and may result in a loss of efficiency.
• When using water, distill or treat with a Milli-Q or equivalent sys-
tem. De-ionized water is not acceptable because it contains organic
compounds which alter column selectivity.
• DO NOT expose columns to freezing temperatures.
• To avoid precipitation of sample and assure reproducible results,
dissolve sample in the initial buffer. Highly ionic samples may give
irreproducible or ambiguous results.
• Protect the column from rapid changes in eluent composition.
DO NOT change the flow rate faster than 0.5 ml/min increments.
DEAE- 5PW SP-5PW
Sodium Azide Cationic Detergents
Sodium Dodecylbenzenesulfonate
Anionic Detergents
Column Size (mm x cm) Recommended Volumes (mL)
7.5 x 75 20
21.5 x 150 300-400