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Grshmal Btech 4th Sem

HPLC, or high-performance liquid chromatography, is a technique used to separate and analyze components in a liquid mixture. It involves pumping a sample through a column using a mobile phase and stationary phase, separating components through interactions with these phases. HPLC instrumentation consists of several key components, including a pump to deliver the mobile phase at constant flow, an injector to introduce samples, a column for separation, a detector to analyze separated components, and a data system to record and analyze results. The document then discusses various HPLC techniques, applications, and an overview of its importance in analytical chemistry.

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24 views11 pages

Grshmal Btech 4th Sem

HPLC, or high-performance liquid chromatography, is a technique used to separate and analyze components in a liquid mixture. It involves pumping a sample through a column using a mobile phase and stationary phase, separating components through interactions with these phases. HPLC instrumentation consists of several key components, including a pump to deliver the mobile phase at constant flow, an injector to introduce samples, a column for separation, a detector to analyze separated components, and a data system to record and analyze results. The document then discusses various HPLC techniques, applications, and an overview of its importance in analytical chemistry.

Uploaded by

prototypes6341
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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HPLC

2/17/2024
Grishma timalsina
INTRODUCTION
IHPLC, or High-Performance Liquid
Introduction
Chromatography, to HPLC
is a technique used to
separate and analyze components in a liquid
mixture. It involves pumping the sample
through a column with a stationary phase and
a mobile phase, leading to the separation
based on interactions with these phases.
HPLC is widely used in analytical chemistry for
its precision and sensitivity
. In this presentation, we will
explore the
and of HPLC instrumentation,
providing a comprehensive understanding of
this powerful analytical technique.

2/17/2024
Components of HPLC Instrumentation
1. Pump: Responsible for delivering the mobile phase (solvent) at a constant flow rate through
the chromatographic column.

2. Injector: Used to introduce the sample into the HPLC system. The sample is typically a liquid
and is injected into the mobile phase flow.

3. Column: A stainless steel or glass tube packed with a stationary phase. The separation of
components in the sample occurs within the column.

4. Detector: Monitors the eluent (the liquid leaving the column) and detects the separated
components. Common types of detectors include UV-Visible spectrophotometers,
fluorescence detectors, and refractive index detectors.

2/17/2024
Components of HPLC Instrumentation
5. Data Acquisition System: Collects and processes the signals from the detector, providing a
chromatogram that represents the separation of components over time.

6. Column Oven: Maintains a constant temperature for the column, as temperature can affect
separation efficiency.

7. Mobile Phase Reservoir: Contains the solvent or mobile phase that is pumped through the
system.

8. Waste Collector: Collects the waste generated during the analysis.

9. Chromatography Software: Manages and analyzes the data collected by the detector, aiding
in the interpretation of results.

2/17/2024
Components of HPLC Instrumentation

2/17/2024
Principles of HPLC
• A separation column separates the
stationary and mobile phases during
purification.

• In a separation column, the stationary


phase is a granular substance with very
small porous particles.

• The mobile phase is a solvent or solvent


combination that is pushed through the
separation column under high pressure.

• A chromatogram is generated in the


HPLC software at the conclusion of this
operation/run.
2/17/2024
Applications of HPLC

• Drug evaluation
• Synthetic polymer analysis
• Pollution analysis in environmental analytics
• Drug determination in biological matrices
• Isolation of high-value goods

2/17/2024
Advanced Techniques in
HPLC
Advanced HPLC techniques include
(mass spectrometry),
(ultraviolet detection), and
(evaporative light scattering
detection), expanding the
capabilities of
HPLC analysis.

2/17/2024
Uses of HPLC
• Purification of water.
• Impurity detection in the pharmaceutical
industry.
HPLC plays a pivotal role
• Trace components are pre-concentrated.
in of products
• Chromatography acrossbased on ligand exchange.
• Proteinindustries, ensuring compliance
chromatography via ion exchange.
with and
• Carbohydrate and oligosaccharide
specifications. anion-
It provides accurate
exchange chromatography
and reliable at high pH.
data for decision-
making.

2/17/2024
Conclusion
HPLC (high performance liquid chromatography) is a type of column liquid
chromatography that has been greatly improved. Instead of allowing a
dissolvable to flow through a column under gravity, it is forced to do so under
heavyweights of up to 400 climates. As a result, it is substantially faster.

2/17/2024
Thank You So Much
😊

2/17/2024

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