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Principles Of Column Chromatography01:13

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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...
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High-Performance Liquid Chromatography: Introduction01:11

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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.
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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...
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High-Performance Liquid Chromatography: Types of Detectors01:15

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Biopharmaceutical Analysis by HPLC: Practices and Challenges.

Golshan Mahdi1, Ashkan Alamdary2, Hooman Kaghazian2

  • 1Pharmaceutical Sciences Branch, Islamic Azad University, Tehran, Iran.

Chonnam Medical Journal
|February 17, 2025
PubMed
Summary

High-Performance Liquid Chromatography (HPLC) is vital for biopharmaceutical analysis, ensuring product quality and regulatory approval. Advancements like UHPLC and hyphenated techniques enhance its capabilities for complex biomolecules.

Keywords:
Biological ProductsChromatographic SystemsChromatography, High Pressure Liquid

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Area of Science:

  • Analytical Chemistry
  • Biopharmaceutical Analysis
  • Chromatographic Techniques

Background:

  • High-Performance Liquid Chromatography (HPLC) is a cornerstone analytical technique in the biopharmaceutical sector.
  • It is indispensable for the separation, identification, and quantification of complex biological molecules, including monoclonal antibodies and recombinant proteins.
  • HPLC is critical for evaluating the purity, potency, and stability of biopharmaceutical products, essential for regulatory submission and approval.

Purpose of the Study:

  • To highlight the critical role and applications of HPLC in the biopharmaceutical industry.
  • To discuss the inherent challenges and limitations associated with HPLC method development and validation.
  • To explore recent technological advancements and their impact on enhancing HPLC's performance in biopharmaceutical analysis.

Main Methods:

  • Review of established HPLC modes: reversed-phase, ion-exchange, size-exclusion, and affinity chromatography.
  • Identification of common challenges: stationary phase selection, peak resolution, matrix effects, and parameter optimization.
  • Examination of recent technological innovations, including Ultra-High-Performance Liquid Chromatography (UHPLC) and HPLC-Mass Spectrometry (MS) hyphenation.

Main Results:

  • HPLC provides high resolution and sensitivity for analyzing complex biopharmaceutical samples.
  • Challenges in method development, standardization, and validation require careful consideration for reproducible and accurate results.
  • Emerging technologies like UHPLC and HPLC-MS significantly improve analytical speed, sensitivity, and resolution.

Conclusions:

  • HPLC remains an indispensable tool for biopharmaceutical quality control and regulatory compliance.
  • Ongoing research and development are crucial for adapting HPLC to increasingly complex biopharmaceutical products.
  • Technological advancements are continuously expanding the capabilities and efficiency of HPLC in biopharmaceutical analysis.