Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Concept Videos

Optimizing Chromatographic Separations01:15

Optimizing Chromatographic Separations

Optimizing chromatographic separations is crucial for obtaining clean separations in a minimum amount of time. Optimization is required for several factors, including kinetic effects related to band broadening, plate height, capacity factor, and separation factor.
Band broadening refers to spreading solute bands as they travel through the column. This broadening can impact resolution. Plate height (H) represents the length required for one theoretical plate. A lower plate height corresponds to...
High-Performance Liquid Chromatography: Introduction01:11

High-Performance Liquid Chromatography: Introduction

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:
High-Performance Liquid Chromatography: Elution Process01:05

High-Performance Liquid Chromatography: Elution Process

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...
Principles Of Column Chromatography01:13

Principles Of Column Chromatography

The chromatography technique was first invented in 1901 by Michael S. Tswett, a Russian botanist, to separate plant pigments using organic solvents. Further, in 1941, Archer John Porter Martin and R. L. M. Synge modified the technique by packing silica gel into a column. A mixture of amino acids was then separated on the packed column using chloroform and water mixture as the mobile phase. This was the first report on column chromatography. At present, column chromatography is a widely used...
High-Performance Liquid Chromatography: Instrumentation00:57

High-Performance Liquid Chromatography: Instrumentation

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.
Affinity Chromatography01:03

Affinity Chromatography

Affinity chromatography is a powerful technique extensively utilized for separating and purifying specific biomolecules from complex mixtures. It capitalizes on the highly selective binding between an analyte and its counterpart, such as antibody-antigen interactions. The counterpart is immobilized on the stationary phase, forming an affinity column. The stationary phase typically consists of solid support, such as agarose or porous glass beads, immobilizing the affinity ligand. The mobile...

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Retention prediction using quantitative structure-retention relationships combined with the hydrophobic subtraction model in reversed-phase liquid chromatography.

Electrophoresis·2019
Same author

Retention Index Prediction Using Quantitative Structure-Retention Relationships for Improving Structure Identification in Nontargeted Metabolomics.

Analytical chemistry·2018
Same author

Retention prediction in reversed phase high performance liquid chromatography using quantitative structure-retention relationships applied to the Hydrophobic Subtraction Model.

Journal of chromatography. A·2018
Same author

Chemometric-assisted method development in hydrophilic interaction liquid chromatography: A review.

Analytica chimica acta·2018
Same author

Error measures in quantitative structure-retention relationships studies.

Journal of chromatography. A·2017
Same author

Benchmarking of Computational Methods for Creation of Retention Models in Quantitative Structure-Retention Relationships Studies.

Journal of chemical information and modeling·2017
Same journal

Stereo-sensitive modelling of gas chromatographic retention indices of mono-cycloalkanes in jet fuel range.

Journal of chromatography. A·2026
Same journal

Approaches to using retention indices with coupled column pressure tuning in gas chromatography.

Journal of chromatography. A·2026
Same journal

MOF-supported surface-imprinted polymer for hazard governance of aristolochic acids in herbal matrices: A safety-control strategy supported by multiscale simulations.

Journal of chromatography. A·2026
Same journal

Portable cold-assisted head-space solid-phase microextraction coupled with GC-MS/MS for sensitive determination of trace polychlorinated naphthalenes in water.

Journal of chromatography. A·2026
Same journal

Characterization of phosphorous impurities originating from the synthesis of Sarin.

Journal of chromatography. A·2026
Same journal

Extraction and chromatographic purification of purpurin: A scalable approach using modified dry column vacuum chromatography.

Journal of chromatography. A·2026
See all related articles

Related Experiment Video

Updated: May 10, 2026

Curtain Flow Column: Optimization of Efficiency and Sensitivity
06:44

Curtain Flow Column: Optimization of Efficiency and Sensitivity

Published on: June 12, 2016

Optimizing selectivity during reversed-phase high performance liquid chromatography method development: prioritizing

Lloyd R Snyder1, John W Dolan

  • 1LC Resources, Walnut Creek, CA, USA.

Journal of Chromatography. A
|July 9, 2013
PubMed
Summary
This summary is machine-generated.

Adjusting temperature or mobile phase composition can replicate changes in reversed-phase chromatography (RPC) selectivity typically achieved by altering the column or pH. This finding suggests rethinking standard method development strategies.

Keywords:
Experimental conditionsExperimental designHPLCMethod developmentOptimizationSelectivity

More Related Videos

Untargeted Metabolomics from Biological Sources Using Ultraperformance Liquid Chromatography-High Resolution Mass Spectrometry (UPLC-HRMS)
11:00

Untargeted Metabolomics from Biological Sources Using Ultraperformance Liquid Chromatography-High Resolution Mass Spectrometry (UPLC-HRMS)

Published on: May 20, 2013

Multi-step Preparation Technique to Recover Multiple Metabolite Compound Classes for In-depth and Informative Metabolomic Analysis
11:25

Multi-step Preparation Technique to Recover Multiple Metabolite Compound Classes for In-depth and Informative Metabolomic Analysis

Published on: July 11, 2014

Related Experiment Videos

Last Updated: May 10, 2026

Curtain Flow Column: Optimization of Efficiency and Sensitivity
06:44

Curtain Flow Column: Optimization of Efficiency and Sensitivity

Published on: June 12, 2016

Untargeted Metabolomics from Biological Sources Using Ultraperformance Liquid Chromatography-High Resolution Mass Spectrometry (UPLC-HRMS)
11:00

Untargeted Metabolomics from Biological Sources Using Ultraperformance Liquid Chromatography-High Resolution Mass Spectrometry (UPLC-HRMS)

Published on: May 20, 2013

Multi-step Preparation Technique to Recover Multiple Metabolite Compound Classes for In-depth and Informative Metabolomic Analysis
11:25

Multi-step Preparation Technique to Recover Multiple Metabolite Compound Classes for In-depth and Informative Metabolomic Analysis

Published on: July 11, 2014

Area of Science:

  • Analytical Chemistry
  • Chromatography Science

Background:

  • Reversed-phase chromatography (RPC) is a key separation technique.
  • Method development in RPC often involves optimizing column choice and mobile phase pH for selectivity.

Purpose of the Study:

  • To reexamine literature data on factors affecting RPC selectivity.
  • To propose alternative parameters for method development.

Main Methods:

  • Literature data analysis.
  • Comparative assessment of selectivity factors in RPC.

Main Results:

  • Changes in RPC selectivity induced by column or pH variations can be mimicked by adjusting temperature.
  • Mobile phase composition, specifically acetonitrile or methanol concentrations, also significantly impacts selectivity.
  • Temperature and mobile phase composition offer alternative routes to achieve desired selectivity.

Conclusions:

  • Mobile phase pH and column selection may not be the sole critical factors for optimizing RPC selectivity.
  • Temperature and mobile phase composition present viable and potentially simpler alternatives for method development in RPC.