Scribd Logo
Upload

Welcome to Scribd!

  • 2K views

    Basic Principles of HPLC

    Uploaded by

    shahzad1840
    This document provides an overview of high performance liquid chromatography (HPLC), including its basic principles, components, types of HPLC, popularity and advantages. It describes what HPLC is, the components used including mobile phases, pumps, injectors, columns and detectors. It also discusses reverse phase, normal phase and ion-exchange chromatography and explains why HPLC is widely used.

    Copyright:

    Attribution Non-Commercial (BY-NC)
    Download as PPT, PDF, TXT or read online from Scribd

    Basic Principles of HPLC

    Uploaded by

    shahzad1840
    2K views18 pages
    This document provides an overview of high performance liquid chromatography (HPLC), including its basic principles, components, types of HPLC, popularity and advantages. It describes what HPLC is, the components used including mobile phases, pumps, injectors, columns and detectors. It also discusses reverse phase, normal phase and ion-exchange chromatography and explains why HPLC is widely used.

    Original Description:

    High Performance Liquid Charomatography. A simple and Comprihensive ppt

    Copyright

    © Attribution Non-Commercial (BY-NC)

    Available Formats

    PPT, PDF, TXT or read online from Scribd

    Did you find this document useful?

    Is this content inappropriate?

    This document provides an overview of high performance liquid chromatography (HPLC), including its basic principles, components, types of HPLC, popularity and advantages. It describes what HPLC is, the components used including mobile phases, pumps, injectors, columns and detectors. It also discusses reverse phase, normal phase and ion-exchange chromatography and explains why HPLC is widely used.

    Copyright:

    Attribution Non-Commercial (BY-NC)
    Download as PPT, PDF, TXT or read online from Scribd
    Download as ppt, pdf, or txt
    2K views18 pages

    Basic Principles of HPLC

    Uploaded by

    shahzad1840
    This document provides an overview of high performance liquid chromatography (HPLC), including its basic principles, components, types of HPLC, popularity and advantages. It describes what HPLC is, the components used including mobile phases, pumps, injectors, columns and detectors. It also discusses reverse phase, normal phase and ion-exchange chromatography and explains why HPLC is widely used.

    Copyright:

    Attribution Non-Commercial (BY-NC)
    Download as PPT, PDF, TXT or read online from Scribd
    Download as ppt, pdf, or txt
    of 18

    Basic Principles of HPLC

    Shahzad Bashir
    2009-ag-1231
    Center for Agricultural biochemistry and biotechnology
    UNIVERSITY OF AGRICULTURE
    FAISLABAD
    Outline
    • What is HPLC?

    • Types of HPLC
    H igh
    Performance
    L iquid
    C hromatography
    H igh
    Pressure
    L iquid
    C hromatography
    What is HPLC?
    • The most widely used analytical
    separations technique
    • Utilizes a liquid mobile phase to
    separate components of mixture
    • uses high pressure to push solvent
    through the column
    Components
    • Mobile phase reservoirs
    • HPLC Pump(s)
    • Sample injector (manual or auto)
    • Column
    • Detector
    • Mobile phase waste container
    Mobile phases
    • Water
    • Methanol
    • Acetonitrile
    • Additives, salts, acids, bases
    HPLC Pump(s)
    • A high pressure pump is used to generate a
    specific flow rate to mobile phase, typically
    millimeter per minute.

    Sample injector

    • An injector is able to introduce the sample into


    the continuously flowing mobile phase stream
    that carries the sample into HPLC column.
    Columns
    • Length (5-15 cm)
    • The column contains the chromatographic
    packing material needed to effect the
    separation.
    • This packing material is called the
    stationary phase.
    Detectors

    • UV
    • Fluorescence
    • Electrochemical
    • Mass Spectrometric
    Chromatograms

    Restek® ULTRA C-18 and CN Columns (250mm x 4.6mm, 5µ),


    Mobile Phase: (1:1 Methanol:Water), 1.5 mL/min.
    Types of HPLC
    • Reverse-phase (polar mobile phase/non-polar
    stationary phase/somewhat polar analytes)
    • Normal Phase (non-polar mobile phase/polar
    stationary phase/non-polar analytes)
    • Ion-Exchange (salts/ionic stationary phase)
    • Size-exclusion (aqueous/gel for large MW
    solutes, >104)
    Popularity
    Sensitivity
    Accurate quantitative determination
    suitability for separating nonvolatile and
    thermally sensitive species
    widespread applicability in industry, to many
    fields of science, and to the public
     Ideally suited for proteomic analysis.
    Advantages to HPLC

    Higher resolution and speed of


    analysis
    HPLC columns can be reused without
    repacking or regeneration
    Greater reproducibility of results.
    Easy to operation and data analysis
    Adaptability to large-scale
    Advantages of HPLC are result of 2
    major advances

    Stationary supports with very small


    particle sizes and large surface
    areas
    Appliancation of high pressure to
    solvent flow
    THANKS

    You might also like