Waters Analytical Columns and Cartridges User Manual

[ Care and Use ManUal ]

analytICal Columns and CartrIdges

I. IntroduCtIon

Waters offers a large selection of analytical columns and cartridges

packed with silica-based packing materials. Follow this instruction

sheet to obtain the best performance, reproducibility, and longevity

from your analytical columns and cartridges. Waters silica-based

materials are packed in:

TM

• Steel and PEEK

analytical columns are durable and highly efficient. PEEK

columns are used with nonmetallic systems.

• Steel Cartridge Columns — The cartridge columns feature reus-

able end fittings that permit you to replace the cartridge quickly

and easily without tools.

• Polyethylene Radial-Pak™ Column Segments — The Radial-Pak

column segments (also referred to as Radial-Pak cartridges)

employ Waters patented technology to improve efficiency

by eliminating the wall effect and voiding common to rigid

columns.

(polyetheretherketone) Columns — The steel

Contents

I. IntroduCtIon

II. start-up test proCedure

a. Measuring Intrument Band Spreading

b. Installing the Cartridge or Column

c. Equilibrating the Column

d. Testing the Column Efficiency and Calculating Column Backpressure

III. narrow-bore Columns

IV. Column maIntenanCe

a. Caring for the Column

b. Cleaning and Regenerating the Column

c. Storing the Column

V. troubleshootIng

Analytical Columns and Cartridges 1

VI. orderIng and addItIonal InformatIon

[ Care and Use ManUal ]

Spacer

0.25 mm (0.009 inch) Tubing

Sentry Guard

Column

C-Clip

Cartridge Column

End Connector

II. start-up test proCedure

Follow the four step start-up procedure for successful operation of

your Waters column or cartridge: A. The pH range for the mobile

phase is pH 2 to 8. However, phosphate buffer at pH 8 in combination

with high temperatures can lead to shorter column life. For high

temperature work, this buffer at neutral pH is preferred.

a. Measuring Instrument Band Spreading

Poor column performance can sometimes be attributed to excessive

instrument band spreading. Test the band spreading of your HPLC

system before you install your column or cartridge. Band spreading

of a properly operating analytical system should be less than

100 μL and <40 μL for 2.1 mm i.d. microbore columns.

Test the instrument band spreading using the procedure below.

1. Remove the column or cartridge from the system. Connect the

inlet and outlet tubing with a low-dead-volume union.

2. Inject the test sample used for a plate count determination (see

Section II. d.) using a fast chart speed or sampling rate. You may

need to adjust the detector sensitivity or dilute the sample to

keep the peak on scale.

Determine the width of the peak at 4.4% of the peak maximum.

Determine how many microliters per minute your eluent is flowing.

Multiply the two terms to yield the band spread of your instrument.

If the band spreading is excessive, check all tubing and fittings. If it

is necessary to repair fittings, refer to the procedure below. If you

have a 2.1 mm i.d. microbore column, you may need to modify your

HPLC system. Refer to the System Modifications Guidelines outlined

in Section III.

Repair a damaged compression screw or a worn ferrule assembly

using the procedure below.

1. Scribe the circumference of the tubing at the desired break point

using a tube cutter or a file with a cutting edge.

2. Grasp the tubing on both sides of the scribe mark with cloth

covered pliers. Gently work the tube back and forth until it sepa-

rates at the scribe mark. Ensure that the tubing end is straight,

open and free of burrs.

3. Slide the compression screw, followed by the ferrule (large end

of the taper first), over the tube. Seat the tubing in the fitting. If

the tubing is not completely seated, the resulting dead volume

can lead to poor chromatographic results.

3. Use the 5-Sigma method shown in Figure 2 to obtain the band

spreading value in μL.

Figure 1: Installing a Cartridge Column with a SentryTM Guard Column

Analytical Columns and Cartridges 2