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Related Concept Videos

High-Performance Liquid Chromatography: Types of Detectors01:15

High-Performance Liquid Chromatography: Types of Detectors

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 properties and...
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: 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.
Thin-Layer Chromatography (TLC): Overview01:11

Thin-Layer Chromatography (TLC): Overview

Thin-layer chromatography (TLC) is a chromatography technique that separates compounds based on their polarity. TLC typically uses polar silica gel, a form of silicon dioxide, as the stationary phase. The silica gel contains hydroxyl (OH) groups on its surface, which form hydrogen bonds with polar compounds, influencing their adhesion to the stationary phase.
To begin the analysis, a mixture of compounds is spotted on the starting line on the TLC plate using a thin capillary. The bottom of the...
Gas Chromatography: Types of Detectors-I01:21

Gas Chromatography: Types of Detectors-I

There are different types of detectors used in gas chromatography, each with its own specific properties that make it suitable for detecting certain types of analytes. The most commonly used detectors in GC are thermal conductivity detector (TCD), flame ionization detector (FID), and electron capture detector (ECD).
TCD is the earliest and most widely used detector that operates by measuring the changes in the thermal conductivity of the carrier gas. When a sample compound enters the detector,...
Gas Chromatography: Types of Detectors-II01:19

Gas Chromatography: Types of Detectors-II

In gas chromatography, different detectors are employed to meet specific analytical needs. These detectors are often categorized based on their detection mechanisms and the types of compounds they are best suited to analyze. Thermal Conductivity Detectors (TCD), Flame Ionization Detectors (FID), and Electron Capture Detectors (ECD) represent common categories, each with unique operating principles and applications. However, beyond these, several other detectors are designed for more specialized...

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Fluorescence detection methods for microfluidic droplet platforms
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Published on: December 10, 2011

Fluorescence detection by intensity changes for high-performance thin-layer chromatography separation of lipids using

Vicente L Cebolla1, Carmen Jarne, Pilar Domingo

  • 1Instituto de Carboquímica, Consejo Superior de Investigaciones Cientificas, C/Miguel Luesma, 4, 50018 Zaragoza, Spain. vcebolla@icb.csic.es

Journal of Chromatography. A
|December 15, 2010
PubMed
Summary

Fluorescence detection by intensity changes (FDIC) combined with HPTLC/AMD effectively detects and quantifies lipids. This method offers a new way to analyze lipid families with high sensitivity, even for cholesterol.

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Area of Science:

  • Analytical Chemistry
  • Lipidomics
  • Chromatography

Background:

  • Lipid analysis is crucial in various scientific fields.
  • Existing methods for lipid detection and quantification have limitations.
  • Berberine cation fluorescence changes offer a novel detection principle.

Purpose of the Study:

  • To develop and validate a method for lipid detection and quantification using fluorescence detection by intensity changes (FDIC) and HPTLC/AMD.
  • To separate and quantify various lipid classes, including neutral lipids, steryl glycosides, and sphingolipids.
  • To rationalize the observed fluorescent responses based on molecular interactions and computational modeling.

Main Methods:

  • Utilized fluorescence scanning densitometry with berberine cation for lipid detection.
  • Employed automated multiple development (HPTLC/AMD) for gradient-based lipid separation.
  • Developed three distinct HPTLC/AMD gradient schemes for different lipid families.
  • Applied molecular mechanics for computational analysis of lipid-fluorophore interactions.

Main Results:

  • Successfully separated and quantified neutral lipids, steryl glycosides, sphingolipids, and sphingosine-sphinganine mixtures.
  • Demonstrated a limit of detection (LOD) of 5 ng for cholesterol.
  • Rationalized fluorescent molar responses based on ion-induced dipole interactions and computational modeling.
  • Explained variations in FDIC response among different lipid classes.

Conclusions:

  • FDIC combined with HPTLC/AMD provides a sensitive and effective method for lipid analysis.
  • The method allows for the separation and quantification of diverse lipid families.
  • Understanding the underlying molecular interactions enhances the application of FDIC in lipidomics.