ParticleSize
• Analytical separations generally are on5µmor
smaller particles.
• Semi-preparative andpreparative analyses general-
ly are on7µmor larger particles.
• Smaller particles provide greater efficiency (N)
than larger particles (a 3µmparticle provides
~50% greater efficiency than a 5µmparticle).
• Smaller particles createhigher backpressure.
PoreSize
• Smallmolecules (<2000mw) generally analyzedon
material withpore size
≤
200Å(typical size:
60Å–180Å).
• Largemolecules (>2000mw) generally analyzedon
material withpore size
≥
300Å.
• Surface area inversely related topore size (the
smaller the pores the larger the surface area).
• Stationaryphase loadingdirectly related to surface
area (the larger the surface area the higher the sta-
tionaryphase loading).
ColumnDimensions
ID
• Theoretically, resolution andpressure are inde-
pendent of ID, if themobile phase flow rate is
adjusted tomaintain the same linear velocity.
• Smaller column ID can increase sensitivity if sam-
ple is limited.
• Smaller column ID can reduce / eliminateneed for
flow splitting inLC/MS.
• Column ID<3.2mm requires reducing extra col-
umn volume (tubing, injector, detector flow cell).
Length
• If all else is equal: shorter columns provide faster
analyses and less backpressure; longer columns
provide greater resolution
• Efficiency (N) is directly related to column length,
but is a square root factor in resolution: therefore,
a 4-fold increase in length is needed todouble res-
olution
[R=
1
/
4
(
α
-1 /
α
) (
√
N) (k' / k'+1)]
α
: selectivity k': capacity
ChoosingaStationaryPhase
• Most critical decision in column selection.
• Basedon analyte solubility and chemical
differences among analytes.
• Analyte solubilitydictatesmode of separation:
Reversedphase - analytes soluble inwater and/or
polar organic solvent (e.g., acetonitrile,methanol).
Mobile phasemore polar than stationaryphase.
Normal phase - analytes soluble innonpolar sol-
vent (e.g., hexane).Mobile phase less polar than
stationaryphase.
AnalyteFunctionalGroup
Trident™DirectGuardColumnSystem
Unlike “one size fits all” guard systems, the Trident™Direct system gives you
the power to select the right level of protection for your analysis. The system
offers three levels of protection and guard cartridges in four dimensions,with
a varietyof bondedphases tomatch your analytical column.The economical,
leak-free cartridgedesignprovides anunprecedented combinationof conven-
ience, economy, and reliability.The foundationof theTrident™Direct system
is a reusable direct connect holder that easily attaches to any HPLC column
using CPI- orWaters®-style end fittings.* The system is available in the fol-
lowing configurations tomatch different protection level needs: in-line filter,
in-line filter with holder for 10mm guard cartridge, and in-line filter with
holder for20mmguardcartridge.Theguardcartridges areavailable in2.1and
4.0mm ID and are interchangeablewithin the appropriate length holder. Use
4.0mm ID guardwith 4.6 or 4.0mm ID analytical column, 2.1mm ID guard
with2.1 - 3.2mm ID analytical column.
*The standard PEEK
®
tip inTrident
™
Direct systems is compatiblewith Parker
®
, Upchurch
®
,
Valco
®
, and other CPI-style fittings.To useTrident
™
Direct systemswithWaters
®
-style end fittings,
the tipmust be replacedwith cat.# 25088.
Easy toUse, LowDeadVolume—TheUltimate
CombinationofConvenienceandColumn
Protection
Trident™DirectGuardColumnSystem
Please seeRestek’s
Chromatography
ProductsCatalog for
moreHPLCcolumns
andaccessories.
Steps inScoutingaMethod
1.Choose stationaryphase basedon chemistry
of analytes.
2.Use a 150 x 4.6mm, 5µm column; determine
appropriatemobile phase strengthneeded
(2<k<5) byusing a gradient test run.
3.If resolution is close to requirements, optimize
columndimensions and conditions. If large
gain in resolution is needed changemobile
phase compositionor stationaryphase.
5µmParticles
3µmParticles
ID (mm)
FlowRate (mL/min.)
FlowRate (mL/min.)
4.6
1.00
1.5
3.2
0.50
0.73
2.1
0.20
0.31
1.0
0.05
0.07
OptimumFlowRate
ReversedPhase (RP)
Normal Phase (NP)
ChoosingaMode
R
P - reversedphase
SEC - sizeexclusion chromatography
GFC - gel filtration chromatography
RP-IP - reversedphase-ionpair ionexchange
NP - normal phase
GPC - gel permeation chromatography
IEX - ionexchange
Description
qty.
cat.#
High-pressure filter
ea.
25082
10mm guard cartridge holder without filter
ea.
25083
10mm guard cartridge holder with filter
ea.
25084
20mm guard cartridge holder without filter
ea.
25085
20mm guard cartridge holder with filter
ea.
25086
Connection tip forWaters
®
-style end fittings
ea.
25088
PEEK
®
tip standard fittings
ea.
25087
Replacement cap frits: 4mm, 2.0µm
5-pk.
25022
Replacement cap frits: 4mm, 0.5µm
5-pk.
25023
Replacement cap frits: 2mm, 2.0µm
5-pk.
25057
Ultra100ÅPores
Allure™60ÅPores
SampleCharacteristics
Sample soluble in:
Ionic Strength
Analytes primarily differ by:
RecommendedMode:
water and/or polar organics
nonionic
hydrophobicity/hydrophilicity
RP
weakly ionic
hydrophobicity/hydrophilicity
RPwith ion suppression
nonionic/weakly ionic
size (>2000MW)
SEC (GFC)
strongly ionic
hydrophobicity/hydrophilicity
RP-IP, IEX
nonpolar organics
nonionic
hydrophobicity/hydrophilicity
NP
weakly ionic
hydrophobicity/hydrophilicity
NPwith ion suppression
nonionic/weakly ionic
size (>2000MW)
SEC (GPC)
Neutral
Base
Acid
Acids, Bases, &Neutrals
Mixed
Ultra IBD
UltraAqueousC18
Allure™AqueousC18
Allure™OrganicAcids
Allure™Basix
Allure™PFPPropyl
Ultra IBD
UltraPFP
Pinnacle™DBCyano
UltraCyano
UltraAmino
Pinnacle
II
™Amino
Pinnacle™DBC18
UltraC18
Pinnacle
II
™C18
Allure™C18
UltraC8
Pinnacle
II
™C8
Pinnacle
II
™Phenyl
Ultra IBD
Ultra AqueousC18
Allure™AqueousC18
Allure™OrganicAcids
Ultra IBD
Allure™Basix
Hydrophobic
Hydrophilic
HPLCColumnSelectionGuide
INNOVATIVE
PRODUCTS
TurningVisions intoReality
™
800-356-1688 • 814-353-1300
20
YEARS
Presorted Standard
US Postage
PAID
Restek
RestekCorporation
110Benner Circle
Bellefonte, PA16823-8812
Lit. Cat.#59454B
© 2005 Restek Corporation.
20
YEARS
RestekTrademarks: Allure, Pinnacle, Pinnacle II, Viva,Trident,TurningVisions intoReality, Restek logo.
OtherTrademarks: Aquasil, Betamax, BioBasic, Fluophase, Hypersil, Prism, HyPurity Advance (Thermo ElectronCorp.); Delta-Pak, Spherisorb, Symmetry,Waters (Waters
Corp.); Develosil, Jupiter, Luna, Maxsil, Ultracarb (Phenomenex, Inc.); Discovery, Supelcosil (Sigma-AldrichCo.); Fluosep-RP (ES Industries); Inertsil (GL Sciences, Inc.);
Kromasil (EkaChemicals); LiChrospher (Merck KgaA); Parker (Parker InstrumentationDivision); PEEK (Victrex plc); Platinum (AlltechAssociates, Inc.); Upchurch
(Upchurch Scientific); Valco (Valco Instruments Co., Inc.); Zorbax (AgilentTechnologies, Inc.).
