Why is HPLC the most popular technique in analytical laboratories today?

Modern HPLC

Modern HPLC Systems

HPLC is a separation technique which is extensively used to separate, identify and quantify components of mixtures of organic compounds selectively retained on the stationary phase inside the column using liquid as the carrier phase. It has found widespread applications in academic research as well as industrial monitoring and quality control.

If you were to walk into a modern pharmaceutical laboratory you’re likely to come across a number of analytical instruments such as a UV-Visible Spectrophotometer, Atomic Absorption Spectrophotometer, 5-6 GC systems, HPTLC, Particle Size Analyzers but you’re sure to come across scores of HPLC systems. Undoubtedly HPLC has achieved the dominating position in laboratories and is bound to retain this enviable position in the coming time.

Have you ever wondered as to what factors have contributed to the ever increasing popularity of HPLC. The answer lies in its applications and unmatched flexibility offered to the analytical scientist

Applications

HPLC has found widespread applications in the following key areas:

  • Pharmaceuticals
  • Foods
  • Polymers
  • Environmental monitoring
  • Forensics
  • Biochemical and Biotechnology laboratories

Flexibility

The key reason for the popularity of HPLC is the ease with which it can be adapted to the needs of the analyst

  • Choice of mobile phase solvents and Isocratic or Gradient elution helps isolate and identify compounds of varying polarities
  • The vast scale of mass separations is difficult to come across in a single technique

Analytical – pg-ng
Semi Preparative – μg – mg
Preparative – mg-gms

It is also possible to cover large range of compounds having molecular weights ranging from 50 to over 1 million dalton

  • Operational modes

Normal phase for separation of nonpolar compounds using combination of non-polar mobile phases

Reverse phase for separation of polar molecules using aqueous/organic mobile phase mixtures

Ion exchange uses ion-exchange columns for separation of charged molecules, proteins, nucleotides, etc

Gel permeation chromatography for separation on basis of molecular size of separating compounds and has been used for separation of proteins and polymers

  • Choice of stationary phases

Majority of stationary phases are silica-based but recent advances in new materials has even further expanded the application range

  • Pressure range

Majority of applications are conducted using normal pressures of 2000 – 6000 psi. Ultra high-pressure chromatography or UHPLC uses pressure ranges from 15,000 – 18,000 psi using sub 2-μm particles for greater speed and resolution

  • Choice of detectors

The large choice of detectors offers numerous options for detection of a large number of compounds having different structural properties :

  1. UV-Visible detector
  2. Photo Diode Array detector
  3. Fluorescence detector
  4. Reflective index detector
  5. Electrochemical detector
  6. Conductivity detector
  7. Mass Sensitive detector
  8. Light Scattering detectors
  • Recent advances in HPLC technology

LC – MS offers an unmatched combination of separation by HPLC coupled with high sensitivity of the mass spectrometer detection

Core shell particle technology provides advantages offered by UHPLC without resorting to high pressures

Monolith and zirconia based packings provide benefits of extended pH operation and higher than ambient temperatures

  • Choice of column sizes

Selection of column length and diameter can optimize speed of analysis with high resolution. With narrow bore columns it is possible to achieve separations in a matter of a few minutes or seconds as compared to conventional columns

You will agree that the flexibility of adaptation provides unlimited options to the analytical scientist and HPLC is finding more and more applications in different laboratories each day. The global share of HPLC market is around US$3 billion. Pharmaceutical industry alone accounts for 35% of the market share. US and Europe currently account for 70% of the global market share. Asia Pacific region is set to achieve the fastest growth with the compound growth rate of 7.5% projected from 2000 – 2015.

Please do share your opinion and leave your valuable comments on the article.

About Dr. Deepak Bhanot

Dr Deepak Bhanot is a seasoned professional having nearly 30 years expertise beginning from sales and product support of analytical instruments. After completing his graduation and post graduation from Delhi University and IIT Delhi he went on to Loughborough University of Technology, UK for doctorate research in analytical chemistry. His mission is to develop training programs on analytical techniques and share his experiences with broad spectrum of users ranging from professionals engaged in analytical development and research as well as young enthusiasts fresh from academics who wish to embark upon a career in analytical industry.

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