Designing Chromatographic Columns Around the Sample: The Future of Purpose-Driven HPLC
Author - Deepti Khanvilkar, Lead - Analytical Services, Venture Center
High-performance liquid chromatography (HPLC) has long been one of the most versatile tools for separating, identifying, and quantifying components in complex mixtures. From pharmaceutical analysis and quality control to food, environmental, forensic, and biotech applications, HPLC chromatography continues to play an important role in analytical science.
But as analytical challenges become more complex, the question is no longer simply how fast an HPLC system can run or how sensitive its detector can be.
Increasingly, the focus is shifting to something fundamental to the separation itself: the chromatographic column and its stationary phase. Let’s dig a little deeper...
What Is HPLC?
High-performance liquid chromatography, or HPLC, is a chromatographic technique used to separate and analyse components of a mixture based on their interactions with the mobile phase and the stationary phase inside a column.
In an HPLC system, the sample is introduced into a flowing liquid mobile phase and passed through a chromatographic column. Different molecules interact differently with the stationary phase, causing them to travel through the column at different rates. This difference in retention enables individual components to be separated and subsequently detected.
An HPLC test can therefore provide qualitative or quantitative information about compounds in a sample, depending on the analytical objective and method used.
HPLC chromatography finds applications across a wide range of areas, including pharmaceuticals, food testing, environmental analysis, forensics, biotechnology, and chemical research. It can be used for applications ranging from compound analysis and purity assessment to process-related studies and quality control.
Why the HPLC Column Matters
The stationary phase is the material packed inside an HPLC column. It determines how sample molecules interact with the column and, consequently, how effectively they can be separated.
For many years, C18 columns have served as a versatile workhorse in liquid chromatography. Their broad applicability has made them a common starting point for a wide range of analytical methods.
However, not every analytical problem can be addressed equally well with a general-purpose stationary phase.
The molecules being analysed today are increasingly complex. Biopharmaceuticals, glycans, oligonucleotides, metabolites, natural products, and complex botanical extracts can present separation challenges that are very different from those associated with the smaller molecules for which conventional reversed-phase approaches were originally developed.
This is driving an important shift in HPLC method development: instead of expecting one column chemistry to solve every separation problem, researchers are increasingly looking at stationary phases designed around the analytical challenge itself.
From General-Purpose HPLC Columns to Purpose-Designed Stationary Phases
The development of new stationary phases is being supported by advances in materials science, a deeper understanding of molecular interactions, computational modelling, and increasingly, artificial intelligence.
These developments make it possible to think about chromatographic column performance more deliberately, with the properties of the sample and the required separation considered from the beginning.
The result is a move away from entirely empirical column selection towards a more rational and purpose-driven approach to chromatography.
Rather than repeatedly changing mobile-phase conditions, gradients, or other method parameters to force a difficult separation, the stationary phase itself can become an important part of the solution.
This is particularly relevant when the molecules of interest are structurally similar and conventional HPLC methods struggle to provide sufficient selectivity.
Why Complex Samples Need More Selective HPLC Chromatography
For relatively straightforward samples, a widely used stationary phase may provide the required separation with limited method optimization.
More challenging samples are different.
Biopharmaceutical molecules, glycans, peptides, oligonucleotides, metabolites, and natural products can contain closely related components that differ in subtle structural characteristics. When these components behave similarly during chromatographic separation, obtaining clear resolution can require extensive method development.
This is where stationary-phase selectivity becomes particularly important.
A column that provides better discrimination between closely related molecules from the outset could potentially reduce the amount of optimization required to arrive at a usable analytical method.
Purpose-Designed Stationary Phases for Glycan Analysis
Glycan analysis provides a useful example of why stationary-phase development matters.
Glycan isomers can be difficult to separate using conventional reversed-phase HPLC columns because the molecules may differ in subtle structural features such as sugar linkages and branching patterns.
A newly engineered porous graphitized carbon (PGC) stationary phase has demonstrated improved baseline separation of glycan isomers, along with excellent reproducibility across different column batches.
The significance of this approach goes beyond one specific application.
When a stationary phase is developed with a particular separation challenge in mind, it can provide selectivity that may be difficult to achieve using a more general-purpose column. The same principle can be relevant to other challenging analytical targets where closely related molecular species need to be distinguished.
What Purpose-Designed HPLC Columns Could Mean for Method Development
For analytical scientists, one of the practical benefits of improved stationary-phase selectivity could be a reduction in method development effort.
Purpose-designed stationary phases could potentially support the separation of:
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Closely related protein variants
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Peptides
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Oligonucleotides
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Glycan isomers
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Natural products
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Other challenging molecules
With greater selectivity built into the chromatographic column, analytical methods may require less repeated optimization.
This could translate into:
Shorter method development cycles
Better separation from the beginning could reduce the number of iterations required to establish suitable chromatographic conditions.
Less repeated optimization
A more appropriate stationary phase may reduce the need to continually adjust conditions to compensate for inadequate selectivity.
Improved analytical efficiency
When separation is achieved more effectively, the overall analytical workflow can become more streamlined.
The choice of HPLC column, therefore, is not simply an equipment decision. It can be an important part of the analytical strategy.
The Role of Computational Modelling and AI in Chromatographic Development
Another important development is the growing role of computational approaches in chromatographic research.
Advances in computational modelling and AI are opening possibilities for predicting chromatographic behaviour before a column is manufactured.
Combined with advances in materials science and an improved understanding of molecular interactions, these approaches support a more targeted way of thinking about stationary-phase development.
The broader direction is clear: chromatography is moving towards increasingly informed column selection and stationary-phase development rather than relying solely on empirical trial and error.
Will C18 Columns Become Obsolete?
Not at all.
C18 columns will continue to have an important role in routine HPLC analysis because of their versatility and broad applicability.
The emerging trend is not about replacing C18 columns altogether. Instead, it is about recognizing that increasingly complex analytical questions may require more specialized stationary phases.
A general-purpose column can remain an effective choice for many applications while purpose-designed stationary phases provide additional options when conventional approaches encounter limitations.
The future of HPLC chromatography is therefore likely to involve both: versatile, widely applicable chemistries alongside stationary phases developed with specific molecule classes or analytical challenges in mind.
The Future of HPLC: Designing Around the Analytical Challenge
As analytical science moves towards increasingly complex molecules and samples, chromatographic separation will need to become more selective and more purpose-driven.
The evolution of stationary phases reflects this broader shift.
Instead of asking only which HPLC column is most commonly used, analytical scientists can increasingly consider a more fundamental question:
What does this sample require from the separation?
For challenging molecules, answering that question may influence everything from stationary-phase selection to method development and analytical efficiency.
The continued evolution of materials science, molecular-level understanding, computational modelling, and AI is likely to make this approach increasingly important in the development of next-generation chromatographic columns.
High Performance Liquid Chromatography (HPLC) at Venture Center, Pune
At Venture Center, chromatography services support scientific and industrial applications across pharmaceuticals, food testing, environmental analysis, forensics, biotechnology, and other areas. The facility provides access to HPLC systems equipped with autosamplers and different detection capabilities, including PDA, fluorescence, and UV-Vis detection. Available column chemistries include C18, C4, and HILIC.
Venture Center’s analytical team also stays abreast of the latest developments and trends in chromatography, supporting researchers and industry teams with HPLC analysis, method understanding, troubleshooting, and analytical workflows suited to their requirements. The facility has also conducted hands-on HPLC training covering separation principles, instrumentation, method development, troubleshooting, data interpretation, and recent developments in chromatography.
Whether you are evaluating a new material, developing a pharmaceutical product, analysing a complex sample, or looking for reliable separation and quantification, HPLC at Venture Center provides access to chromatography infrastructure and technical expertise to support your analytical requirements.
Book our HPLC service or consultation here.
For enquiries, contact: rutuja.patil@venturecenter.co.in or +91 8956677542