Agilent Technologies and C8, User Manual

Page: 8 / 90

Recommended starting gradients
Stationary phase Usage notes
Most reversed-phase
columns
5% methanol or acetonitrile initially and 100%
methanol or acetonitrile as the final solvent.
ZORBAX Eclipse PAH* 30 or 40% acetonitrile initially, to 100%
acetonitrile as final solvent. It may be necessary
to cool column to 15 to 20 °C for improved
resolution.
ZORBAX Bonus-RP Lower concentration of organic mobile phase
modifier is required for compound elution,
compared to traditional long-chain alkyl
stationary phases.
Large molecule
columns (for example,
Poroshell 300 or
ZORBAX 300SB
columns)
Separations of polypeptides, proteins, DNA
fragments, etc. often use trifluoroacetic or
formic acid modifier for pH control and/or as
an ion-pairing additive for desired retention
and selectivity. Organic buffers such as TRIS
are especially useful for intermediate pH
applications.
* Best column lifetime is achieved at <40 °C.