Waters XBridge Peptide BEH C18 130A and 300A Columns - Manual (Page 6)

b. Storage
For periods longer than four days at room temperature, store the
column in 100% acetonitrile. Immediately after use with elevated
temperatures and/or at pH extremes, store in 100% acetonitrile
for the best column lifetime. Do not store columns in highly
aqueous (<20% organic) mobile phases, as this may promote
bacterial growth. If the mobile phase contained a buffer salt, flush
the column with 10 column volumes of HPLC grade water (see
Table 1 for common column volumes) and replace with 100%
acetonitrile for storage. Failure to perform this intermediate step
could result in precipitation of the buffer salt in the column or
system when 100% acetonitrile is introduced. Completely seal
column to avoid evaporation and drying out of the bed.
Note: If a column has been run with a mobile phase that contains
formate (e.g., ammonium formate, formic acid, etc.) and is then
flushed with 100% acetonitrile, slightly longer equilibration times
may be necessary when the column is re-installed and run again
with a formate-containing mobile phase.
V. CONNECTING T HE COLUMN TO T HE HPLC
a. Column Connectors and System Tubing Considerations
Tools needed:
■ ■
3/8 inch wrench
■ ■
5/16 inch wrench
Handle the column with care. Do not drop or hit the column on a
hard surface as it may disturb the bed and affect its performance.
1. Correct connection of 1/16 inch outer diameter stainless steel
tubing leading to and from the column is essential for high-
quality chromatographic results.
2. When using standard stainless steel compression screw
fittings, it is important to ensure proper fit of the 1/16 inch
outer diameter stainless steel tubing. When tightening or
loosening the compression screw, place a 5/16 inch wrench on
the compression screw and a 3/8 inch wrench on the hex head
of the column endfitting.
Note: If one of the wrenches is placed on the column tube flat
during this process, the endfitting will be loosened and leak.
3. If a leak occurs between the stainless steel compression screw
fitting and the column endfitting, a new compression screw
fitting, tubing and ferrule must be assembled.
4. An arrow on the column identification label indicates correct
direction of solvent flow.
Correct connection of 1/16 inch outer diameter stainless steel
tubing leading to and from the column is essential for high-quality
chromatographic results. To obtain a void-free connection, the
tubing must touch the bottom of the column endfitting. It is
important to realize that extra column peak broadening due to
voids can destroy an otherwise successful separation. The choice
of appropriate column connectors and system tubing is discussed
in detail below.
Figure 1. Waters and Parker Ferrule Types.
Due to the absence of an industry standard, various column
manufacturers have employed different types of chromatographic
column connectors. The chromatographic separation can be
negatively affected if the style of the column endfittings does not
match the existing tubing ferrule settings. This section explains
the differences between Waters style and Parker style ferrules and
endfittings (Figure 1). Each endfitting style varies in the required
length of the tubing protruding from the ferrule. The XBridge
Column is equipped with Waters style endfittings that require a
0.130 inch ferrule depth. If a non-Waters style column is presently
being used, it is critical that ferrule depth be reset for optimal
performance prior to installing a XBridge column.
In a proper tubing/column connection (Figure 2), the tubing
touches the bottom of the column endfitting, with no void
between them.
Figure 2. Proper Tubing/Column Connection.
The presence of a void in the
flow stream reduces column
performance. This can occur if
a Parker ferrule is connected
to a Waters style endfitting
(Figure 3).
Note: A void appears if tubing with a Parker ferrule is connected to
a Waters style column.
Waters Ferrule Setting
.130”
Parker Ferrule Setting
.090”
6
[ CARE AND USE MANUAL ]
XBridge Peptide BEH C
18
, 130Å and 300Å Columns