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    This document discusses the basic principles of high performance liquid chromatography (HPLC). HPLC is used to separate and analyze compounds based on their relative solubility between a mobile liquid phase that carries the sample through a stationary phase within the column. There are two main modes - normal phase uses a polar stationary phase and non-polar mobile phase, while reverse phase uses a non-polar stationary phase like C18 bonded silica and a polar mobile phase like water or acetonitrile. Key instrumentation includes the pump, injector, column, detector, and gradient controller. Common detectors are UV, fluorescence, electrochemical and mass spectrometry. EPA and SW-846 methods are listed that use HPLC to analyze compounds like acids, explosives

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    A good piece of powerpoint presentation on HPLC

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    This document discusses the basic principles of high performance liquid chromatography (HPLC). HPLC is used to separate and analyze compounds based on their relative solubility between a mobile liquid phase that carries the sample through a stationary phase within the column. There are two main modes - normal phase uses a polar stationary phase and non-polar mobile phase, while reverse phase uses a non-polar stationary phase like C18 bonded silica and a polar mobile phase like water or acetonitrile. Key instrumentation includes the pump, injector, column, detector, and gradient controller. Common detectors are UV, fluorescence, electrochemical and mass spectrometry. EPA and SW-846 methods are listed that use HPLC to analyze compounds like acids, explosives

    Copyright:

    © All Rights Reserved
    Download as PPT, PDF, TXT or read online from Scribd
    Flag for inappropriate content
    14 views14 pages

    HPLC Basic Principles

    Uploaded by

    xbnisha
    This document discusses the basic principles of high performance liquid chromatography (HPLC). HPLC is used to separate and analyze compounds based on their relative solubility between a mobile liquid phase that carries the sample through a stationary phase within the column. There are two main modes - normal phase uses a polar stationary phase and non-polar mobile phase, while reverse phase uses a non-polar stationary phase like C18 bonded silica and a polar mobile phase like water or acetonitrile. Key instrumentation includes the pump, injector, column, detector, and gradient controller. Common detectors are UV, fluorescence, electrochemical and mass spectrometry. EPA and SW-846 methods are listed that use HPLC to analyze compounds like acids, explosives

    Copyright:

    © All Rights Reserved
    Download as PPT, PDF, TXT or read online from Scribd
    Flag for inappropriate content
    of 14

    Basic Principles of HPLC

    High

    Performance
    L iquid

    C hromatography

    High

    Pressure
    L iquid

    C hromatography

    High

    Priced

    L iquid

    C hromatography

    HPLC Methods
    Parameter Group Method

    SDW05.23000s

    EPA 555

    Compounds
    Cl-PhenoxyAcids

    WPP05.06000s

    EPA 605

    Benzidines

    WPP05.13000s

    EPA 610

    PAHs

    SHW06.26000s

    SW-846 8316

    Acrylics

    SHW06.28000s

    SW-846 8330s

    Explosives

    SW-846 8325

    Benzidines and
    N- Pesticides

    SHW07.06000s

    Compounds
    2,4,5-T

    Benzidine

    Fluorene

    Compounds
    TNT (2,4,6-Trinitrotoluene)

    H2C=CH-CN

    Acrylonitrile

    Carbaryl

    Partitioning
    Separation is based on the analytes relative
    solubility between two liquid phases

    Mobile Phase

    Stationary Phase

    Solvent

    Bonded Phase

    HPLC - Modes
    Normal Phase.
    - Polar stationary phase and
    non-polar solvent.
    Reverse Phase.
    - Non-polar stationary phase and a polar
    solvent.

    Common Reverse Phase


    Solvents
    Methanol

    CH3OH

    Acetonitrile

    CH3CN

    Tetrahydrofuran
    Water

    H2O

    Columns
    Solid Support - Backbone for bonded phases.
    Usually 10, 5 or 3 silica or polymeric
    particles.
    Bonded Phases - Functional groups firmly linked
    (chemically bound) to the solid support.
    Extremely stable
    Reproducible
    Guard - Protects the analytical column:
    Particles
    Interferences
    Analytical
    - Performs
    theof
    separation.
    Prolongs
    the life
    the analytical column

    Bonded Phases
    C-2
    C-8
    C-18
    CN

    Ethyl Silyl

    -Si-CH2-CH3

    Octyl Silyl

    -Si-(CH2)7-CH3

    Octadecyl Silyl

    -Si-(CH2)17-CH3

    Cyanopropyl Silyl

    -Si-(CH2)3-CN

    Instrumentation
    Gradient
    Controller

    Column

    Pump
    Mobile Phases

    Injector

    Detector

    Detectors
    UV
    Single wavelength (filter) [610, 8330]
    Variable wavelength (monochromator)
    [8316, 8325]

    Multiple wavelengths (PDA) [555]


    Fluorescence [610]
    Electrochemical [605]
    Mass Spectrometric [8325]

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