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  • LC and GC Supplies in Stock
  • Autosampler Chromatography Vials & Accessories
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    • HPLC Vials & Closures
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      • Screw Thread Vials
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      • Max Recovery Vials
    • GC Vials & Closures
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      • Crimp Top Vials
      • Headspace GC Vials
    • 12 x 32 mm LC and GC Autosampler Vials, Closures & Inserts
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      • 8-425 Standard Screw Thread Vials
      • 9 mm Wide Mouth Screw Thread Vials
      • Pre-Loaded Inserts in 11 mm Autosampler Vials
      • 11 mm Wide Mouth Snap Vials
      • 11 mm Standard Crimp Vials
      • 11 mm Wide Mouth Glass Vials
      • PFAS Testing
    • 15 x 45 mm WISP and Shimadzu Vials, Closures & Inserts
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      • Standard Screw Thread Vials
      • Snap Seal Vials
    • Headspace Chromatography Vials and Closures
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      • Crimp Vials
      • Screw Thread Vials
      • PolyMag Screw Caps
    • Miscellaneous Chromatography Vials
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      • Silanized Vials
      • Storage Vials, Caps & Seals
      • V-Vial Reaction Vials
      • Vial Racks
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      • EPA/VOA Vials
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    • Low Adsorption Vials
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      • Special Surface Properties
      • Screw Thread RSA Vials
      • Snap Top RSA Vials
    • Agilent Vials
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      • 12 x 32 mm, Wide Mouth Screw Thread
      • Wash Vials
      • 12 x 32 mm, Wide Mouth Crimp
      • Headspace Vials
  • 96-Well Plates
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    • 96 Deep Well Plates
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      • Glass Inserts in Plates
      • Chrom Tech MS Polypropylene Plates
      • 96 Well ELISA Plates
      • PCR Plates
      • Pulled Point Inserts in Plates
      • Reservoir Trough Polypropylene Plates
      • Greiner Bio-One Plates
      • Friction Seals, Pierceable Cap Mats
    • Sealing Films & Foils
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      • Heat Sealing Film and Foil Rolls
      • 96-Well Sealing Film & Tape
      • Porvair Heat Seals & Foils
    • 96-Well Microplate Instrumentation
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      • Plate Heat Sealers
      • Positive Pressure Processor
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  • Sample Preparation
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    • Laboratory Filtration Supplies
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      • Centrifuge Filter Tubes
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      • Filter Vials
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        • Agilent Econofilters
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    • Solid Phase Extraction
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      • Agilent Bond Elut Plexa Polymeric SPE
      • Agilent Bond Elut Reversed Phase (Non-Polar) Silica SPE
      • Agilent Bond Elut Ion Exchange Silica SPE
      • Agilent Bond Elut Ion Mixed Mode Silica SPE
      • Agilent Bond Elut Inorganic SPE
      • Agilent Specialty SPE
      • Bond Elut Lipid Extraction SPE
    • Supported Liquid Extraction
    • Instrumentation
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      • Evaporators
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        • Bar Vap
        • Cerex Concentrator 48
        • Chrom Tech Low Cost
        • Porvair
      • Positive Pressure Processor
      • Vacuum Manifolds
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        • BeadBlaster 24
        • BeadBug 6
        • BeadBlaster 96
      • Dried Blood Spot Card Punch
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        • BioMixer
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  • HPLC Columns
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    • Agilent for Small Molecule
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      • InfinityLab Poroshell 120
      • ZORBAX Eclipse Plus
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      • ZORBAX 80Å Extend-C18
      • ZORBAX Bonus-RP Columns
      • ZORBAX Eclipse PAH
      • Zorbax HILIC Plus
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      • ZORBAX Rx
      • Pursuit XRs
      • Polaris
    • Agilent BioHPLC
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      • Poroshell 300
      • AdvanceBio RP-mAb
      • AdvanceBio Glycan Mapping
      • InfinityLab Poroshell 120 AdvanceBio Peptide Mapping
      • PLRP-S
      • Zorbax 300A Stablebond
      • AdvanceBio SEC
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      • Bio SEC-3
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    • Agilent for GPC/SEC
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      • Aqueous GPC/SEC Columns
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        • PL aquagel-OH SEC columns
        • PolarGel GPC Columns
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        • Aqueous Polymer Standards
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        • PLgel MIXED Columns
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        • PLgel Olexis Columns
        • PLgel Individual Pore Size & Preparative Columns
        • HFIPgel Columns
        • PL Rapide Columns
        • PlusPore Columns
        • EnviroPrep Columns
        • Polystyrene Standards
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    • Concise Separations
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      • Carbohydrate Analysis
      • Organic Acid Analysis
      • Protein Analysis
      • Amino Acid Analysis
      • Ion Analysis
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      • Reversed-Phase
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      • Ion Exclusion
    • Imtakt
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      • Presto FF-C18
      • Dacapo DX-C18
      • Cadenza CD-C18
      • Intrada Amino Acids Separation Column
      • Scherzo C18 Mutli-Mode Column
    • Jordi
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      • Jordi Resolve Aqueous GPC
      • Jordi Resolve Organic GPC
    • Regis
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      • Regis Pirkle Chiral HPLC Columns
      • Immobilized Artificial Chromatography
    • Restek
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      • UltraShield UHPLC Precolumn Filter
      • Raptor LC Columns
      • Ultra HPLC
      • Force
      • Roc
    • Millipore Sigma
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      • Ascentis Express
    • Thermo Scientific
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      • Accucore
      • Hypersil Gold
    • Vydac
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      • HPLC/MS Columns
      • 201TP and 202TP
      • Columns for Peptides & Polypeptides
    • Zirchrom
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      • ProTain In-Line Protein Removal System
      • Zirconia Based HPLC Columns
  • GC Columns
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    • Agilent J&W GC Columns
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      • DB-WAX Ultra Inert
      • DB-1ms Low-Bleed
      • DB-1ms Ultra Inert
      • HP-1ms Low-Bleed
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    • Agilent J&W Polysiloxane
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      • DB-1
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    • Agilent J&W Specialty Columns
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        • DB-1ht
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        • CP-Sil PONA CB
        • HP-PONA
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        • Select Low Sulfur
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      • Select Biodiesel
      • Amines
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        • CP-Volamine
        • CP-Sil 8 CB Amines
      • Pesticides
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        • VF-5 Pesticides
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        • DB-1701P
        • DB-608
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        • Select PAH
        • DB-EUPAH
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        • DB-UI 8270D
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        • DB-624 UI
        • DB-624
        • CP-Select 624 CB
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        • HP-VOC
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    • Restek High Performance GC Columns
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      • Rxi-1301Sil MS
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    • Restek General Performance
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      • Rtx-1301
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      • Rtx-35 Amine
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  • HPLC Accessories
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    • HPLC Instrumentation
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      • Chrom Tech Pumps
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        • Single Piston Isocratic Pumps
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          • Low Profile M1 Class
          • MXT MODEL
          • LST MODEL
        • Dual Piston Isocratic Pump
        • Constant Pressure Isocratic Pump
        • High Flow Dual Piston Pumps
        • Super Critical Fluid Pumps
        • Flash Pumps
        • Pulse Dampener
      • Vacuum Degasser
      • Column Heaters
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        • H-705 Column Oven
        • H-105 Column Heater
        • Column Hot Sleeve
      • Detectors
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        • UV/VIS
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          • Fixed Wavelength
          • Variable Wavelength
          • Multi Wavelength & Diode Array
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            • FLASH 10MWD
            • FLASH 14 DAD
        • Refractive Index
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          • RI-S2020
      • Fraction Collectors
      • DataApex Clarity Software
      • Post Column Derivatization
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        • Application Kits, Reagents & Columns
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          • Amino Acid Analysis Kits
          • Pesticide & Herbicide Analysis Kits
        • Pickering PINNACLE PCX System
        • Pickering VECTOR PCX System
        • Sensivate System
    • HPLC Accessories - Empty Columns
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      • Glass Columns
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        • EZ Column
        • EZ Solvent Plus Columns
        • Replacement Parts, EZ & Solvent Plus
      • SS Columns
    • Valco Fittings & Accessories
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      • Internal Unions
      • Internal Reducing Unions
      • Internal Reducers
      • Stainless Steel Fittings
      • Syringe Adapter
      • Tees
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      • National Pipe Thread Adapters
      • Valco Sample Loop for C6W
    • HPLC Fittings
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      • Flangeless Fittings
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        • Flangeless
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      • National Pipe Thread Adapters
      • Large Bore Fittings
      • Fingertight Fittings
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        • Rheodyne RheFlex
        • Agilent A-Line Quick Connect Fitting
        • Upchurch One-Piece
        • Upchurch Two-Piece
        • SealTight
        • LiteTouch
      • Stainless Steel Fittings
      • UHPLC Fittings
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        • Agilent InfinityLab Quick Connect Fitting
        • Agilent A-Line Quick Turn Fitting
        • Idex VHP Fittings
        • Optimize EXP2 Fittings
        • PK VHP Fittings
      • Agilent InfinityLab Quick Connect Fittings
      • MicroTight Fittings & Sleeves
      • Tubing Sleeves
      • Plugs & Caps
      • Tools
    • HPLC Connectors/Adapters
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      • Unions, Tees & Crosses
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        • Union, High Pressure
        • Union, Low Pressure
        • Tees & Crosses, High Pressure
        • Tees & Crosses, Low Pressure
        • Manifolds
      • MicroTight Connectors
      • MicroTee & Cross
      • NanoPort Assemblies
      • Mixing Tees
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        • ASI Static Mixers
        • Idex Static and Mixers
      • Threaded Adapters
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        • English Threads
        • Metric Threads
      • Luer Adapters & Luer Fittings
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        • LuerTight Fittings
        • Quick Connect Adapters
      • Barbed Adapters & Connectors
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        • Barbed Connectors
        • Luer to Barb Adapters
        • Swivel Barb Adapter
        • Thread to Barbed Adapter
      • Peristaltic Tubing & Conical Adapters
    • Polymeric Liquid Chromatography Tubing
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      • PEEK Tubing
      • FEP Fluoropolymer Tubing
      • ETFE Tubing
      • PFA Tubing
      • Tubing Cutters
    • High Pressure HPLC Tubing
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      • VICI Precut Stainless Steel Tubing
      • 316 Stainless Steel Tubing
      • Tubing Cutters
      • Fused Silica Tubing
      • Hastelloy C Custom Length Tubing
      • Agilent Miscellaneous Capillaries
      • PEEKsil Tubing
    • Chromatography Solvent Containment
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      • Solvent Waste Container Kit
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        • System kit
        • Replacement Parts
        • Accessories
      • Solvent Waste Manifolds
      • Popular Manifold Fittings and Tubing
      • Vaplock Solvent Waste Manifold Caps for GL45, GL38 and 38-430
      • Vaplock Solvent Waste Manifold Caps for Carboy Containers
      • Mobile Phase/Storage Bottles
      • Glass Filtration Assembly
      • Bottle Caps
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        • IDEX Bottle Caps
        • Vaplock Bottle Caps & Evaporation Control Valves
        • Omnifit Q-Series Bottle Caps
      • Solvent Filters and Inlet Filters
    • Valves
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      • Injection Valves
      • Sample Loops
      • Switching Valves
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      • RheBuild Kits, Rapid Replacement Pods & MXX Replacement Fittings
      • Rotor Seals & Stators
      • Valve Accessories
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        • Syringes for Manual HPLC Valves
      • Hamilton Manual Valves
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      • Back Pressure Regulators (BPRs)
      • Pressure Relief Valves
    • Inline & Precolumn Filters
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      • Agilent Quick Connect Inline Filters
      • Inline Solvent Filters
      • Standard Precolumn Filters
      • Replacement Frits
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        • PEEK frits
        • Stainless Steel frits
      • EXP® Pre-Column Filter Holder
      • Biocompatible Precolumn Filters
      • Precolumn MicroFilters
      • Frit-In-A-Ferrule™
    • Guard Columns
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      • EXP Guard Column Cartridges
      • IDEX Guard Columns
      • Capillary Sample Trap Columns
      • Agilent AdvanceBio Desalting-RP Cartridges
    • LC Instrument Replacement Parts
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      • Agilent Parts for Agilent HPLCs
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        • Loop Capillaries
        • Tubing, Connections, & Tools
        • 1260 Infinity Isocratic LC Supplies
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      • Cartridge for Opti-Max Housing
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  • Chrom Tech Pumps
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    • Single Piston Isocratic Pumps
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      • Low Profile M1 Class
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    • Agilent Consumables
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      • Bulk GC Supplies
      • Inlet Septa
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      • Gold Seals for Split/Splitless Inlets
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    • Agilent Instrument Replacement Parts
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      • Headspace Vials
      • 7890B/7890A/6890 GC Supplies Quick Pick
      • 7890/6890/6850 Split/Splitless Inlet Supplies
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    • Restek Consumables
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      • Topaz Inlet Liners
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        • For Agilent
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      • Restek Gas Leak Detectors
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        • Polyarc Systems
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    • Hamilton Manual Syringes
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        • 7000 Series
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      • MicroLab Dual Syringe Diluters
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      • Regis Sulfonates (S-Series) For Basic Compounds
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      • Amino Acid Analysis Kits
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      • Safety Waste Containment Kit
      • HPLC Waste to Drum Kit
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      • COVID-19 Antibody Research
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  • LC and GC Supplies in Stock
  • Autosampler Chromatography Vials & Accessories
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      • 8-425 Standard Screw Thread Vials
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    HPLC Column Care and Maintenance

    Friday, December 18, 2020

    HPLC Column Care and Maintenance

    Your analytical column is a consumable in your chromatography system, and it must ultimately be replaced. To maximize your column’s lifetime, it is important to take care of and maintain your column. Evidence that your HPLC problems could potentially point to your HPLC column include:

    • High System Pressure
    • Undesirable Peak Shape (splitting/tailing)
    • Changes in Retention/Selectivity

    Please note, the above listed problems may also be associated with instrument and method issues, but, in our experience, the column is typically the first to be blamed.

    Large System Pressure Change, Excess Back Pressure

    When your HPLC system sees a significant increase in pressure, potential problems could be a plugged inlet frit, column contamination, or blockage in the system. It is important to first check the pressure with and without the column in place, as many pressure problems are due to blockages elsewhere in the system. If the excessive pressure is isolated to the column, rinse the column (remove detector from flow path) and/or clean the column. It is best to follow your column manufacturer’s guidelines for column cleaning, but, in general, you flush with stronger solvents than your mobile phase (making sure detector is NOT in flow path).

    For reversed phase columns, here is a general guideline:

    Use at least ten times column volume of each solvent for analytical columns in order of increasing strength

    1. Mobile phase without buffer salts (water/organic)
    2. 100% organic (MeOH or ACN)
    3. Is pressure back in normal range? If not, discard column or consider more drastic conditions:
      1. 75% Acetonitrile:25% Isopropanol, then
      2. 100% Isopropanol
      3. 100% Methylene Methylene Chloride* Chloride*
      4. 100% Hexane*
        *When using either Hexane or Methylene Chloride, the column must be flushed with Isopropanol before returning to your reversed phase mobile phase Isopropanol before returning to your reversed-phase mobile phase.

    In order to prevent column back pressure problems, it is recommended to filter non-HPLC grade solvents, filter buffered solutions, and install an in-line filter between injector and column (trap system debris and particulates before they get to the analytical column). Chrom Tech offers these chromatography supplies, and we are happy to assist with recommending the appropriate HPLC products for your lab.

    Undesirable Peak Shape and Changes in Retention/Selectivity

    Split peaks can be caused by column contamination, a partially plugged inlet frit, column void in packing bed, and incompatible injection solvent effects. To minimize plugging, when analyzing samples with complex matrices, filtering solvents and good sample preparation are important for HPLC Column Care. Also, it is always recommended that the injection solvent is similar in strength (and composition) to the initial mobile phase conditions.

    Changes in retention on the same column, over time, can be attributed directly to the column or other system issues: aging column, column contamination, insufficient column equilibrium, change in pump performance (mobile phase percentage, flow rate), change in column temperature, or other instrument issues.

    Summary

    In summary, for HPLC Column Care and Maintenance, it is important to follow your column manufacturer’s guidelines. In general, to maximize column lifetime, it is recommended to follow good sample preparation techniques, use HPLC grade solvents, filter buffered solvents, and use inline filters to trap particulates before they reach your column. If your column shows signs (high back pressure/poor chromatography) of aging, back flushing is an option. But, even with the best care and maintenance, at some point, every chromatographer will need to make the decision on when to retire their column, and when that time comes, please consider Chrom Tech as your preferred chromatography supplier.

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