The Role of HPLC Columns in Shaping a Greener Analytical Future

As the field of analytical chemistry embraces sustainability, minimizing the environmental footprint of high-performance liquid chromatography (HPLC) analysis has emerged as a pressing priority. This article delves into advancements in green chromatography, focusing on how innovative HPLC column design can drive eco-conscious practices. Key factors such as column geometry, particle size, particle architecture, and stationary phase chemistry are evaluated for their impact on reducing reliance on hazardous and non-renewable solvents. By optimizing these parameters and utilizing sensible practices, researchers can achieve greener methods without sacrificing analytical performance. This exploration bridges cutting-edge technology with the growing imperative for sustainable practices in modern analytical laboratories.

The push for sustainability in analytical laboratories reflects a larger global trend toward reducing environmental footprints. At recent high performance liquid chromatography (HPLC) conferences, sustainability has risen to prominence, with researchers and industry leaders spotlighting ways to “green” analytical workflows (1,2). Traditional HPLC methods rely heavily on solvents, many of which are hazardous, derived from nonrenewable resources,and difficult to dispose of safely. Therefore, solvent consumption emerges as an area where laboratories can make meaningful environmental improvements.

Multiple strategies exist to achieve this, such as adopting narrow-bore columns, leveraging advanced high-efficiency particle technologies, and utilizing predictive software tools to optimize method development. Moreover, selecting the appropriate tools and modes of chromatography and understanding the true separation requirements of a given analysis can enhance both efficiency and environmental friendliness. These tools and approaches are readily available, yet their widespread adoption hinges on greater awareness and a deliberate commitment to sustainable practices.

Reduction: Optimized Hardware and Particle Design to Lower Solvent Use

One of the most effective means of reducing the environmental impact of HPLC is by minimizing solvent consumption through optimized column design. As shown in Figure 1, narrow-bore columns, such as those with a 2.1-mm internal diameter (i.d.), significantly reduce solvent flow rates compared to standard 4.6-mm i.d. columns. Transitioning to these smaller columns results in an 80% reduction in solvent usage for continuous 24-h operation. This not only lowers the cost and environmental burden of solvent procurement and disposal, but it also decreases energy consumption.

FIGURE 1: Impact of column internal diameter (i.d.) on solvent consumption (reproduced from reference [3]).

The benefits of narrow-bore columns are further amplified by advancements in particle size and particle architecture. By transitioning from traditional 5-µm particles to sub-2-µm ultrahigh-pressure liquid chromatography (UHPLC) particles, analysts can achieve dramatic improvements in efficiency. For example, James and co-workers demonstrated that a method running on a column with 5-µm particles might take 30 min, whereas the same method can be completed in under 5 min using a UHPLC column with 1.7-µm particles, resulting in 85% solvent savings and significant reductions in analysis time (3). Particle architecture is also shown by the authors to provide favorable impacts on solvent reduction. As shown in Figure 2, a 5-µm superficially porous particle (SPP) can reduce the solvent usage over a fully porous particle (FPP) of the same size by more than 50%.

FIGURE 2: Impact of particle architecture on solvent consumption (reproduced from reference [3]).

Each of these examples demonstrates how the intelligent use of available column hardware and particle technologies can yield significant ecological and operational benefits.

Software Tools: In Silico Method Optimization for Sustainability

Advancements in computational tools provide a powerful mechanism for reducing solvent use by minimizing the number of physical experiments conducted. Predictive software platforms streamline method development, enabling chromatographers to explore various parameters without consuming laboratory resources. Online software solutions allow practitioners to model method conditions virtually, replacing traditional trial-and-error experimentation. For instance, converting a separation method from acetonitrile to methanol can be complex because of differences in solvent properties, but in silico modeling enables chromatographers to predict outcomes, optimize parameters, and identify viable alternatives without expending solvents or labor hours.

Moreover, predictive and modeling software tools help to avoid costly experimental errors. Failed experiments, which waste time, resources, and solvents, can be preemptively eliminated by using software to identify unworkable conditions. In the example presented in Figure 3, a simple substitution of methanol for acetonitrile is shown to provide an inadequate separation using an online modeling tool. However, further study using the same software revealed that changing the column chemistry alongside solvent substitution promises to result in the desired separation. By utilizing the software tools, failed laboratory experiments can be avoided and, in some cases, alternative solutions can be found.

FIGURE 3: Reducing laboratory experiments through modeling.

The efficacy of these tools depends on the quality and comprehensiveness of underlying data sets. Expanding these databases to include green solvents, such as ethanol, will enhance their utility, driving broader adoption of sustainable practices. By prioritizing virtual experimentation, laboratories can significantly reduce their environmental footprint while also fostering innovation.

Selectivity: Leveraging Stationary Phases for Optimized Separation

Using the right tool for the job often results in a reduction of time, energy, and cost of completing the task. This is often true in chromatographic separations as well. For example, most HPLC scientists will start method development with a C18 stationary phase; as a result, they will often complete the task with that stationary phase. However, is C18 the best tool? In many cases, it is not. Again, citing some work from James and coworkers (3), Figure 4 shows a reduction in run time upon moving from a 150 mm x 4.6 mm, 5 µm C18 to a 50 mm x 2.1 mm, 1.7-µm column. However, the resolution of peaks 2 and 3 are reduced to a point where separation may not be adequate. By substituting a C18-perfluorophenyl (PFP) phase for the C18, a further reduction in column length was possible while maintaining or even increasing resolution. The C18-PFP phase is more selective than the C18 phase.

FIGURE 4: What about selectivity? (reproduced from reference [3])

Selectivity is the most impactful term in the resolution equation; however, it is often neglected when it comes to optimizing methods. There are many cases where alternative stationary phases produce more selective, and thus more efficient, separations than the ubiquitous C18. Understanding the alternatives and questioning standard practices may prove advantageous for developing modern sustainable and green analytical methods.

Green HILIC: Challenges and Opportunities

Hydrophilic interaction liquid chromatography (HILIC) is a powerful technique for separating polar compounds, yet it presents unique sustainability challenges because of its reliance on acetonitrile. Various attempts have been made to replace acetonitrile with greener solvents, such as ethanol or methanol, but they have achieved limited success (4,5). The unique interactions of acetonitrile with water are essential for forming the water layer on the polar stationary phase, making direct substitution problematic.

As substitution is not a likely route to more environmentally friendly HILIC, alternative modes or solvent reduction are left to aid the situation. One means to reduce the impact of HILIC is to avoid it altogether by using alternative modes of chromatography. For example, in cases where ion-exchange interactions dominate the HILIC separation, traditional ion-exchange (IEX) chromatography may serve as a viable substitute. Because ion-exchange methods often use predominantly aqueous mobile phases, they may offer a greener solution. When HILIC remains the best choice, adopting narrow-bore columns, shorter column lengths, and advanced particle technologies can help mitigate its environmental footprint, allowing laboratories to leverage HILIC’s strengths while minimizing its ecological impact.

Fit-for-Purpose Methods: Reassessing Analytical Needs

Many HPLC methods are initially overengineered to ensure robustness during product development, but, as these methods transition to routine use, their performance requirements often change. Reassessing methods to align with current needs can reveal opportunities for solvent reduction and workflow simplification. Welch and colleagues recognized this, and challenged the idea that the added performance of acetonitrile is needed, compared to other more sustainable solvents (6). Less demanding applications may allow for the substitution of acetonitrile with greener solvents such as ethanol or methanol, yielding satisfactory results with a reduced environmental footprint. Such re-evaluations may result in a significant reduction in the overall environmental impact of analytical methods.

Conclusions

Reductions in solvent, energy, and time by greater than 80% have been demonstrated by simply reducing column hardware dimensions and utilizing modern particle technologies. Even further reduction is promised through commercially available capillary columns and instruments designed to exploit capillary dimensions. Software tools for both method development and translation are available. These tools can be used judiciously to design efficient method development scenarios and test hypotheses without excessively or unnecessarily entering the laboratory. The solvent, energy, and time savings gained through the elimination of unnecessary experiments is a powerful means of reducing the environmental impact of scientific endeavors.

Selectivity has also been shown to be a significant, often forgotten, parameter that can used to improve method efficiency and effectiveness. Using the right tool for the job typically results in more efficient and more environmentally friendly processes.

There are some modes of chromatography that are difficult to render environmentally friendly. For example, HILIC separations are generally best achieved using high proportions of acetonitrile in the mobile phase. Although not favored from a sustainability standpoint, effective substitution for acetonitrile has been elusive. Alternative modes of separation, such as ion-exchange, may be a suitable substitution for HILIC in some cases. However, where HILIC is the most appropriate mode, reduction in solvent usage similar to those proposed for reversed-phase (RP) separations is encouraged.

Finally, the methods commonly used currently often overperform at the cost of environmental friendliness. The re-evaluation of methods for sustainability while still maintaining enough performance for their intended use is deemed an area of great potential for the reduction of analytical laboratory environmental impact.

There are many tools available to the chromatographer to lower the environmental impact of the analytical laboratory. Intentional use of modern hardware, particles, and software tools along with a steadfast mindset to develop and use chromatographic methods in a manner that reduces negative impact are not just future considerations, they can be accomplished today.

References

(1) Bell, D. S. Highlights from the 51st International Symposium on High Performance Liquid Phase Separations and Related Techniques (HPLC 2023) LCGC North Am. 2023, 41 (8), 322–327.DOI: 10.56530/lcgc.na.hq8476g7

(2) Muraco, C. Highlights from the 52nd International Symposium on High Performance Liquid Phase Separations and Related Techniques (HPLC 2024) LCGC International 2024, 1 (8), 12–17.

(3) James, M.; Edge, A.; Soliven, A. Applying Sustainability Concepts to Modern Liquid Chromatography. LCGC Supplements: Recent Developments in HPLC 2024, 1 (6s), 20–25.

(4) Bell, D. S., Toward Green and Sustainable Hydrophilic Interaction Liquid Chromatography (HILIC) Workflows: Can Wwe Replace Acetonitrile? in HPLC 2023., 2023, Duesseldorf, Germany.

(5) Min, L.; Ostovic, J.; Chen E. X. et al. Hydrophilic Interaction Liquid Chromatography with Alcohol as a Weak Eluent, J. Chrom. A 2009, 1216 (12), 2362–2370. DOI: 10.1016/j.chroma.2009.01.012

(6) Welch, C. J.; Brkovic, T.; Schafer, W.; Gong, X. Performance to Burn? Re-Evaluating the Choice of Acetonitrile as the Platform Solvent for Analytical HPLC. Green Chem. 2009, 11 (8), 1232–1238. DOI: 10.1039/b906215g

About the Column Editor

David S. Bell is the Lead Consultant and Owner at ASKkPrime, LLC, specializes in separation science consultancy, is Editor of the “Column Watch” series of articles and serves on the Editorial Advisory Board for LCGC International. With over 30 years of experience, he has contributed significantly to chromatography advancements, focusing on stationary phase design, device development, and molecular interaction research. Dave’s work spans gas chromatography, liquid chromatography, sample preparation, and pharmaceutical analytical method development. He holds a PhD in Chemistry from The Pennsylvania State University, has presented research globally, and has authored more than 95 peer-reviewed and trade magazine articles.

askkprime@gmail.com