28/04/2019 Care of HPLC Columns

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POSTED ON NOVEMBER 20, 2013 BY DR. DEEPAK BHANOT 3 COMMENTS

Care of HPLC Columns

A HPLC column is the most critical component of the HPLC system. Quality of your results will
greatly depend on its performance. Though it is possible to monitor the HPLC column condition
by keeping track on number of theoretical plates you are advised to look for other tell-tale signs
such as :

Degradation of peak shapes

Variations in retention times
Sudden increase in backpressure
Poor resolution between peaks
Appearance of ghost peaks

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It is possible to extend the useful life of your HPLC column by adhering to recommended
guidelines in handling, use and storage
TOPICS
Handling AAS (53)
  Avoid dropping and other mechanical shocks. Shocks often lead to channeling which can
Analytical Chemistry (3)
contribute to peak tailing
Operate the HPLC column within recommended pressure limits. Excessive pressure leads to Calculations (5)

same problems as mechanical shocks Chemical (2)

Use only compatible nuts, ferrules and and ttings. Incompatible ttings can lead to leaks
Chromatography (9)
and damage to column threads
Never over tighten the end ttings Environmental Analysis (2)
Never get tempted to use the column as a stirring rod for mixing of mobile phases and
Featured (2)
bu ers.
Free e-courses (2)

Use Gas Chromatography (55)

Install the HPLC column so that the ow direction matches the direction of the arrow General Topics (140)
Mobile phase should be the degassed prior to use and ltered using a 0.45 µm lter.
Guest Posts (27)
Additionally, use in-line lter in the mobile phase reservoir.
Equilibriate the column with the mobile phase which is compatible with the manufacturer HPLC (60)
storage solvent (iso-propylalcohol). Use at least 5– 10 volumes of equilibriating mobile ICP Techniques (13)
phase
Jobs (1)
Before storing rinse the column to remove any residual bu er or ion- pairing agents. This is
essential to prevent degradation or bu er salt deposition on drying. Lab Bytes (17)
Recommended pH range for silica-based columns is 2.0 – 7.5. Some columns permit range
Lab Management (95)
from pH 1- 11 but for safe operation and long column life you should use it as far as

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28/04/2019 Care of HPLC Columns

possible between 2.0-7.5 pH range Laboratory Insurance (8)

When changing mobile phase the selection of solvents should be gradual in terms of
Laboratory Safety (13)
polarity . Avoid drastic changes or extreme gradients.
Reverse ush can be used if excessive back pressure develops Method Validation (1)

Filter all samples through 0.45 or 0.2 μm lter prior to injection Microbiology (14)
User guard column between the injector and analytical column to prevent contaminants
News (3)
and particulates from reaching the column
Use mobile phase for dissolving the sample or if it that is not possible then another solvent Paper Chromatography (10)
or a mixture of solvents of compatible polarities should be used
pH Measurement (3)
Never exceed the prescribed temperature limits and for consistency of results use a column
oven Preformulation Studies (3)

Quality System (9)

Storage Separation Science (11)
HPLC Columns should be stored under ambient temperature conditions and never in a
Spectroscopy (80)
refrigerator or oven
Storage conditions will depend on time interval between use: Stability studies (8)

Training (21)
Short storage of a few hours to overnight- same solvent as last analysis can be used.
Weighing (3)

Medium interval storage (two days to weekend ) – ush with pure water to prevent microbial or
mould growth.

Long-term storage – use eluent which should not contain more than 50% water.

Use of soap or detergent solutions is not recommended for cleaning columns after use.
Strong acids or alkalis cannot be used as they will result in total irreversible damage.
During storage end plugs should be tightly tted to prevent solvent evaporation leading to
drying of stationary phase

We hope you found the guidelines useful for getting good quality results and increasing useful
life of your column. Please share any experiences that you may have by leaving your comment.

Filed Under: HPLC Tagged With: Column, Hplc Columns, Hplc System

About Dr. Deepak Bhanot

Dr Deepak Bhanot is a seasoned professional having nearly 30 years expertise
beginning from sales and product support of analytical instruments. After
completing his graduation and post graduation from Delhi University and IIT
Delhi he went on to Loughborough University of Technology, UK for doctorate research in
analytical chemistry. His mission is to develop training programs on analytical techniques and
share his experiences with broad spectrum of users ranging from professionals engaged in
analytical development and research as well as young enthusiasts fresh from academics who
wish to embark upon a career in analytical industry.

Comments

farida says:
July 21, 2015 at 4:07 am

what is about conditions storage of columns whish are used in bioanalyse

because this columns must be stored between 4 and 8 °c

thank you
nb : excuse my anglish it is not verry good

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28/04/2019 Care of HPLC Columns
Reply

Dr. Saurabh Arora says:

July 23, 2015 at 11:59 am

Hi Farida, You are absolutely correct! GPC and many columns used in analysis of
large molecules do ned to be stored at lower temperature.

Reply

jahangir khandakar says:

July 6, 2018 at 5:41 pm

Informative

Reply

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