Waters CORTECS 1.6 um Columns - Manual (Page 8)

8
CORTECS Columns
V. ADDItIonAL InFoRMAtIon
a. tips for Maximizing CoRteCs Column Lifetimes
1. To maximize CORTECS Column lifetime, pay close attention to:
 Water quality (including water purification system)
 Solvent quality
 Mobile-phase preparation, storage and age
 Sample, buffer and mobile-phase solubilities
 Sample quality and preparation.
2. When problems arise, often only one improper practice must
be changed.
3. Always remember to:
 Use in-line filter unit or, preferably, a VanGuard
Pre-column.

Discourage bacterial growth by minimizing the use of
100% aqueous mobile phases where possible.
 Change aqueous mobile phase every 24 – 48 hours
(if 100% aqueous mobile phase use is required).
 Discard old 100% aqueous mobile phases every 24-48
hours to discourage bacterial growth.
 Add 5% -10% organic modifier to mobile phase A and
adjust gradient profile.
 Filter aqueous portions of mobile phase through
0.2 µm filter.
 Maintain your water purification system so that it is in
good working order.
 Only use ultra pure water (18 megohm-cm) water and
highest quality solvents possible. HPLC grade water is not
UPLC grade water.
 Consider sample preparation (e.g., solid-phase extraction,
filtration, etc).
4. Avoid (where possible):
 100% aqueous mobile phases (if possible)
 HPLC-grade bottled water
 “Topping off” your mobile phases
 Old aqueous mobile phases. Remember to rinse bottles
thoroughly and prepare fresh every 24 to 48 hrs
 Using phosphate salt buffer in combination with high
ACN concentrations (e.g., > 70%) due to precipitation.
5. Don’t: assume a “bad” column is the culprit when high
backpressure or split peaks are observed:

Investigate cause of column failure
• Backpressure

Mobile phase(s), bacteria, precipitation and/or samples
 Peak splitting
 Sample quality
 Injection solvent strength.
6. Remember: the diameter of CORTECS Column (2.1 and 3.0 mm
ID) are often lower than that of a conventional HPLC column and
therefore, mobile phases last much longer. To reduce the chances of
mobile-phase contamination or degradation, only prepare what you
need for analysis or store excess bulk quantities in a refrigerated
environment.
7. Mobile-phase related questions to ask:
 Am I using 100% aqueous mobile phases? Am I able to
add a small amount of organic modifier to my mobile
phase A?
 Do I filter my aqueous mobile phases through 0.2 µm
filters?

How old is my mobile phase? Do I label the bottle with
preparation date?
 Do I “top off” or do I prepare fresh mobile phases every
24 – 48 hrs?
 What is the quality of my water? Has the quality recently?
changed? How is my water purification system working?
When was it last serviced?
 Am I working with pH 7 phosphate buffer (which is
VERY susceptible to bacterial growth)?
8. Sample-related questions to ask:
 If I inject neat standards prepared in mobile phase do I
observe these problems?
 If I prepare my standards in water and prepare them
like samples (e.g. SPE, filtration, etc.) do I still observe
these problems?
 Has the quality of my samples changed over time?
b. Getting started with CoRteCs HILIC Columns
1. Due the fact that CORTECS HILIC Columns do not possess a
bonded phase, the pH operating range is 1 to 5, and they can be
operated at temperatures up to 45 °C.
2. As with any LC column, operating at the extremes of pH,
pressures and temperatures will result in decreased column
lifetime.
Column equilibration
1. When the column is first received, flush in 50% acetonitrile/
50% water with 10 mM final buffer concentration for
50 column volumes.
2. Equilibrate with 20 column volumes of initial mobile-phase
conditions before making first injection.
3. If gradient conditions are used, equilibrate with 8-10 column
volumes between injections.
4. Failure to appropriately equilibrate the column could result in
drifting retention times.