際際滷

際際滷Share a Scribd company logo

High performance liquid chromatography

Download as PPTX, PDF
K
Khushboo Arya

This document provides an overview of high performance liquid chromatography (HPLC). It begins by defining HPLC as a chromatographic technique used to separate mixtures of compounds. The document then discusses the basic principles of chromatography, describing the stationary and mobile phases. It explains that HPLC uses high pressure to force a liquid mobile phase through a column packed with solid particles or porous layers. The document provides a brief history of HPLC and describes the basic components of an HPLC instrument, including the pump, injector, column, and detector. It concludes by discussing some applications of HPLC and its advantages over gas chromatography.

1 of 30
Downloaded 14 times
HIGH PERFORMANCE LIQUID CHROMATOGRAPHY By : Khushboo Arya
Contents Introduction Chromatography Type of chromatography What is HPLC ? History Principle Instrumentation Working Application
Introduction HPLC stands for high performance liquid chromatography or high pressure liquid chromatography . HPLC is a chromatographic technique used to separate a mixture of compounds in analytical chemistry and biochemistry with the purpose to identifying, quantifying, the purifying the individual component of the mixture . It is having high resolution and separation capacity or can be used both as qualitative and quantitative analysis.
Chromatography Chromatography is a physical method of separation of components between two phases : Stationary phase : the substance on which adsorption of analyte ( the substance to be separated during chromatography ) take place .It can be a solid gel or a solid liquid combination. Mobile phase : the solvent which carry analyte ( a liquid or a gas )
Type of Chromatography On the basis of physical state of mobile phase there are two type of chromatography:- I. Gas chromatography II. Liquid chromatography
HPLC HPLC is a type of liquid chromatography where the sample is forced through the column that is packed with stationary phase composed of irregularity or spherically shaped particle , a porous monolithic layer or a porous membrane by a liquid ( mobile phase) at high pressure.
HISTORY
Principle
High performance liquid chromatography
High performance liquid chromatography
Classification Depending upon the phase system (stationary phase and mobile phase ) HPLC is classified into following types: 1. Normal Phase HPLC: It has polar stationary phase and non polar mobile phase. The stationary phase is usually silica and typical mobile phase are hexane, methylene chloride , chloroform, diethyl ether and mixture of these. Polar samples are thus retained on the polar surface of the column packing longer than les polar materials.
2) Reverse Phase HPLC The stationary phase is non-polar (hydrophobic) in nature, while the mobile phase is a polar liquid , such as mixtures of water and methanol or acetonitrile. It works on the principle of hydrophobic interaction hence the more non-polar the material is, the longer it will be retained.
High performance liquid chromatography
High performance liquid chromatography
High performance liquid chromatography
High performance liquid chromatography
Solvent reservoir : it holds the solvent called mobile phase which moves through the various parts of the system. Pumps: the role of the pump is to force the liquid ( called the mobile phase) through the liquid chromatography at a specific flow rate , expressed in mililitres per min (ml/min). Normal flow rate in HPLC are in the 1 to 2ml/min range. Typical pumps can reach pressures in the range of 6000-9000 psi (400to 600bar).
During the chromatography experiment a pump can deliver a constant mobile phase composition (isocratic) or an increasing mobile phase composition (gradient). Sample injector :- the injector serves to introduce the liquid sample into the flow stream of the mobile phase. Typical sample volumes are 5 to -20 , microlitre. The injector must also be able to withstand the high pressure of he liquid system.
High performance liquid chromatography
High performance liquid chromatography
High performance liquid chromatography
High performance liquid chromatography
High performance liquid chromatography
High performance liquid chromatography
High performance liquid chromatography
High performance liquid chromatography
High performance liquid chromatography
WHY HPLC BETTER THAN GC
High performance liquid chromatography
High performance liquid chromatography

More Related Content

High performance liquid chromatography

  • 2. Contents Introduction Chromatography Type of chromatography What is HPLC ? History Principle Instrumentation Working Application
  • 3. Introduction HPLC stands for high performance liquid chromatography or high pressure liquid chromatography . HPLC is a chromatographic technique used to separate a mixture of compounds in analytical chemistry and biochemistry with the purpose to identifying, quantifying, the purifying the individual component of the mixture . It is having high resolution and separation capacity or can be used both as qualitative and quantitative analysis.
  • 4. Chromatography Chromatography is a physical method of separation of components between two phases : Stationary phase : the substance on which adsorption of analyte ( the substance to be separated during chromatography ) take place .It can be a solid gel or a solid liquid combination. Mobile phase : the solvent which carry analyte ( a liquid or a gas )
  • 5. Type of Chromatography On the basis of physical state of mobile phase there are two type of chromatography:- I. Gas chromatography II. Liquid chromatography
  • 6. HPLC HPLC is a type of liquid chromatography where the sample is forced through the column that is packed with stationary phase composed of irregularity or spherically shaped particle , a porous monolithic layer or a porous membrane by a liquid ( mobile phase) at high pressure.
  • 11. Classification Depending upon the phase system (stationary phase and mobile phase ) HPLC is classified into following types: 1. Normal Phase HPLC: It has polar stationary phase and non polar mobile phase. The stationary phase is usually silica and typical mobile phase are hexane, methylene chloride , chloroform, diethyl ether and mixture of these. Polar samples are thus retained on the polar surface of the column packing longer than les polar materials.
  • 12. 2) Reverse Phase HPLC The stationary phase is non-polar (hydrophobic) in nature, while the mobile phase is a polar liquid , such as mixtures of water and methanol or acetonitrile. It works on the principle of hydrophobic interaction hence the more non-polar the material is, the longer it will be retained.
  • 17. Solvent reservoir : it holds the solvent called mobile phase which moves through the various parts of the system. Pumps: the role of the pump is to force the liquid ( called the mobile phase) through the liquid chromatography at a specific flow rate , expressed in mililitres per min (ml/min). Normal flow rate in HPLC are in the 1 to 2ml/min range. Typical pumps can reach pressures in the range of 6000-9000 psi (400to 600bar).
  • 18. During the chromatography experiment a pump can deliver a constant mobile phase composition (isocratic) or an increasing mobile phase composition (gradient). Sample injector :- the injector serves to introduce the liquid sample into the flow stream of the mobile phase. Typical sample volumes are 5 to -20 , microlitre. The injector must also be able to withstand the high pressure of he liquid system.
  • 28. WHY HPLC BETTER THAN GC