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Related Concept Videos

High-Performance Liquid Chromatography: Introduction01:11

High-Performance Liquid Chromatography: Introduction

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High-performance liquid chromatography(HPLC), formerly referred to as High-pressure liquid chromatography, is a powerful technique used to separate, identify, and quantify components in complex mixtures. The term "high pressure" refers to using high pressure to push the liquid mobile phase through the tightly packed columns.
In HPLC, two phases play a critical role in the separation process:
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High-Performance Liquid Chromatography: Instrumentation00:57

High-Performance Liquid Chromatography: Instrumentation

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High-performance liquid chromatography, or HPLC, is an analytical technique that separates liquid samples under high pressures. An HPLC instrument consists of glass bottles for storing solvents called mobile phase reservoirs. HPLC-grade solvents are used to maintain high purity, and the dissolved gases are removed using a degasser, such as a vacuum pumping system or sparging with helium. The solvents are then pumped into the analytical column using a screw-driven syringe or reciprocating pumps.
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High-Performance Liquid Chromatography: Elution Process01:05

High-Performance Liquid Chromatography: Elution Process

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In High-Performance Liquid Chromatography (HPLC), the elution process is critical to the separation of analytes and the quality of chromatographic results. Elution describes how compounds move through the column and separate based on their interactions with the mobile and stationary phases. This process determines the resolution, peak shape, and retention times in the chromatogram, which are essential for identifying and quantifying components in complex mixtures. Understanding the elution...
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High-Performance Liquid Chromatography: Types of Detectors01:15

High-Performance Liquid Chromatography: Types of Detectors

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The role of the detectors in High-Performance Liquid Chromatography (HPLC) is to analyze the solutes as they exit from the chromatographic column. The detector recognizes the solute's property and generates corresponding electrical signals, which are converted into a readable graph of the detector's response versus elution time called a chromatogram at the computer. There are several types of HPLC detectors, each with its own advantages and limitations, depending on the analyte...
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Gas Chromatography: Types of Columns and Stationary Phases01:17

Gas Chromatography: Types of Columns and Stationary Phases

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Gas chromatography (GC) relies on stationary phases to separate and analyze components in a sample. There are two main types of stationary phases: liquid and solid. Liquid stationary phases are non-volatile, thermally stable, and chemically inert liquids coated onto the column. Solid stationary phases are particles of adsorbent material, such as silica gel or molecular sieves.
For an analyte to remain on the column for a sufficient amount of time, it must exhibit some level of compatibility (or...
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Types Of Column Chromatography01:29

Types Of Column Chromatography

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The stability and compatibility of column material with samples are crucial for efficient purification in chromatographic techniques. Various operating parameters such as pH, temperature, or solvent affect the packing of the column material, thereby determining the purification efficiency. The choice of column material also plays an essential role in deciding the operating parameters and can be modified based on the proteins that need to be purified.
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Post Column Derivatization Using Reaction Flow High Performance Liquid Chromatography Columns
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Column Characterization and Selection Systems in Reversed-Phase High-Performance Liquid Chromatography.

Petar Žuvela1, Magdalena Skoczylas2, J Jay Liu3

  • 1Department of Chemistry , National University of Singapore , Singapore 117543 , Singapore.

Chemical Reviews
|January 4, 2019
PubMed
Summary

Choosing the right reversed-phase high-performance liquid chromatography (RP-HPLC) column is critical for method development due to significant selectivity differences. This review offers a comprehensive guide to RP-HPLC column selection and characterization.

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Area of Science:

  • Analytical Chemistry
  • Chromatography

Background:

  • Reversed-phase high-performance liquid chromatography (RP-HPLC) is the dominant separation mode (>90%) due to its simplicity, cost-effectiveness, and automation.
  • The vast array (>600) of available RP-HPLC columns exhibit significant differences in selectivity and quality, complicating column selection.
  • Increasing sample complexity necessitates careful column selection for successful RP-HPLC method development.

Purpose of the Study:

  • To provide a comprehensive, authoritative, and critical review of RP-HPLC column selection and characterization.
  • To consolidate relevant publications from the past four decades.
  • To propose future perspectives for an "experiment-free" column selection methodology.

Main Methods:

  • Review of existing literature on RP-HPLC column selection and characterization.
  • Analysis of factors influencing column selectivity, including intermolecular interactions (dispersive, polar, hydrogen bonding, donor-acceptor).
  • Consideration of column properties such as packing type, residual silanols, end-capping, ligand density, and pore size.

Main Results:

  • RP-HPLC column performance is highly dependent on the chosen column due to variations in selectivity and physicochemical properties.
  • No universal RP-HPLC column exists, requiring analytical laboratories to maintain a diverse column inventory.
  • Objective comparison and selection of RP-HPLC columns remain challenging despite extensive available knowledge.

Conclusions:

  • Effective RP-HPLC method development hinges on rational column selection.
  • A comprehensive understanding of column characteristics is essential for optimizing separations.
  • Future research should focus on integrating computational simulations with minimal experiments for streamlined column selection.