Chromatography Techniques and Terms

Study Notes

A technique separating mixture components based on their travel time through a stationary phase using a mobile phase.
Consists of two phases:
- Mobile phase: Liquid or gaseous, flows over or through the stationary phase.
- Stationary phase: Solid, liquid, or a solid/liquid mixture, immobilized.

Analyte: Substance being separated during chromatography.
Immobilized phase: Stationary phase attached to support particles or column tubing.
Mobile phase: Moving phase, includes the sample and solvent carrying it through the column.
Effluent: Mobile phase exiting the column.

An advanced form of liquid chromatography.
Uses tightly packed columns, high pressure (up to 5,000 psi) to force eluent through.
Utilizes smaller particle size packing material for greater surface area, improving separation quality.
HPLC Technique:
- Liquid mobile phase separates the mixture.
- Analytes are dissolved in a solvent, then pushed through the column under high pressure (up to 400 atm).
- Mixture is separated into its components within the column.
- Separation time is often 5-10 minutes compared to hours or days for conventional methods.
HPLC System Components:
- Pump: Delivers the mobile phase at a stable flow rate, preventing pulsations even when mobile phase composition varies.
- Injector: Introduces the sample under pressure, minimizing band broadening and flow disturbances.
- Column: Smooth-bore stainless steel or heavy-walled glass tubing containing the packed stationary phase.
- Detector: Monitors the elute as it leaves the column, producing an electronic signal proportional to component concentrations.
  - Crucial for trace analysis
  - High sensitivity
  - Fast response
  - Insensitive to changes in solvent type, flow rate, and temperature

Two main types:
- Valve type: Connected to the column via a pipe, with valves for opening and closing, frequently used.
- Syringe type: Directly connected to the column, used for sample delivery with a syringe.