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Techniques & Tools Gas Chromatography, Liquid Chromatography

Who’s Afraid of SFC?

Overall, how would you characterize the current state of SFC? 

Overall, there has been a good deal of interest in applying SFC in a variety of ways, but there remains a degree of resistance. To be honest, I think that is largely due to people not wanting to change. If someone has been using GC or LC for years, it’s not easy to convince them to learn a completely new technique and invest in a (more) expensive instrument. In my experience, the younger generation are more willing to give SFC a go – largely because they’re less likely to be completely invested in another technique. If SFC is to reach its significant potential, it will almost certainly be the next generation of scientists driving it forwards.

I should also note that there are some legitimate technical barriers to implementing SFC. For example, UV sensitivity is half of that found in LC UV detection – and that’s simply not sensitive enough for many applications, such as for quantifying low-level impurities in a pharmaceutical drug. 

How evolved are SFC instruments? 

In the early 2010s, Agilent and Waters released their SFC systems, which represented significant improvements compared with what we had before. Shimadzu’s instrument, which came out a few years later, is also worth noting for the simplicity it brought to the hyphenation of supercritical fluid extractions, which made methods more reproducible in general. 

I’d say the vendors did a great job – and their efforts generated plenty of excitement around the technique. Unfortunately, for whatever reason, we’re still using pretty much the same instruments 10 years later... And that’s a shame because I believe there is a great deal of scope to improve SFC instrumentation given what we’ve learned about the technique in the past 10–15 years. 

So what have we learned about SFC over the past decade or so? 

Quite a lot actually – and it was overdue. In the beginning, SFC was used only with pure fluids – mostly CO2 in the mobile phase. Over time, we progressively introduced small portions of co-solvents, to improve analyte solubility and peak shape, but also to have more flexibility on method optimization. Co-solvents are now used in large quantities. For example, we can now do very wide gradients starting with 100 percent CO2 and finishing in 100 percent co-solvent conditions, especially to analyze complex samples with a wide range of polarities. Admittedly, we still need to better understand diffusion coefficients and how they relate to changes in viscosity in the system. But our overall understanding of the technique is much improved and we’ve seen it applied in a number of new application areas. I don’t want to be too hard on the instrument developers, but there’s certainly room for improvement!

What would be on your wishlist for a new SFC instrument? 

Besides the UV sensitivity issue that I mentioned, I would like more simplicity in terms of the design and function. From an outsider’s perspective, SFC can look complicated – there are certainly more parameters to optimize than with GC. If the instruments operated more akin to an LC system, I think that would help adoption – especially, as an SFC instrument already looks like the typical LC “hifi tower.” I’d also like more flexibility. It isn’t particularly easy to change from one mode to another. Let’s say I want to do a very wide gradient going from 100 percent CO2 to 100 percent co-solvent, I can push the instrument to achieve that goal – but it isn’t how the instrument is designed to be used. When the co-solvent is a mixture (e.g. solvent and water), it also isn’t possible to change the proportion of solvent and water during the gradient, because the pumps are only two way, not three way. MS hyphenation also needs to be simplified. 

Such wishlist requests would expand what’s possible in a single experiment (rather than needing two separate instruments), opening up new possibilities in terms of applications. For example, there’s a great deal of interest in applying the technique to biomolecules, such as proteins or nucleic acids, but this is tricky with the current generation of instruments. 

Where is SFC most widely applied right now?  

The number one application for SFC today is natural products, which has overtaken small molecule pharmaceuticals. The main drivers are the rise of Chinese traditional medicine and foodomics, which generally don't require the same sensitivity or resolution as pharmaceuticals. We also see many applications in bioanalysis – this wasn’t the case 10 years ago and is a result of better hyphenation to mass spectrometry, which aids in the detection of minor metabolites. Again, this relates to the issue of the wider gradient; if you’re looking for polar metabolites, sensing with 100 percent CO2 won’t work – you’ll need something else in your mobile phase. Environmental analysis is another growing area – again, this is an application are that generally isn’t constrained by sensitivity or method validation to the same extent as pharma.

What are the main benefits of SFC? 

SFC is very versatile. You can use it to detect non-polar, chiral, achiral, small molecules, “big” molecules, such as synthetic polymers – though we are restricted when it comes to the large biomolecules. It’s also very complementary to other techniques. Even if your reverse phase LC is working well, you may need to confirm your results with orthogonal analysis. And in areas such as traditional Chinese medicine, the aim is to gain a comprehensive fingerprint of the product, which requires several techniques.  

What are the common misunderstandings? 

There is a common misconception that, because we mostly use CO2, SFC won’t work for polar molecules. But, as we’ve discussed, advances over the past decade have overcome this issue. I also think that people often overestimate the complexity of SFC, fearing that they won’t be able to manage the system and get it working properly – especially if they’re looking for a system suitable for non-expert users (so they can simply place their sample in an automatic injector and leave it to run). In fairness, I used to think this too – that you need some knowledge to use the technique properly – but I’ve been proven wrong in recent years. If you have some standard methods established, it really is possible for a non-expert to use SFC today. So my main message is to reassure people: SFC isn’t so frightening! 

How optimistic are you about the future of SFC? 

I’m realistic. SFC is never going to replace the better-established (notably, they’re not actually much older!) techniques of GC and LC. My hope is that SFC will be better accepted as a useful complement. Personally, I think it’s brilliant – that’s why I’ve been promoting it for the past two decades! It wasn’t so long ago that few people were listening. The instruments 10 years ago really changed that. And I think it’ll take innovation on the instrument side to kickstart SFC again.

Caroline West is an Associate Professor at the Institut de Chimie Organique et Analytique (ICOA), CNRS UMR 7311, University of Orleans, France

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About the Author
James Strachan

Over the course of my Biomedical Sciences degree it dawned on me that my goal of becoming a scientist didn’t quite mesh with my lack of affinity for lab work. Thinking on my decision to pursue biology rather than English at age 15 – despite an aptitude for the latter – I realized that science writing was a way to combine what I loved with what I was good at.

From there I set out to gather as much freelancing experience as I could, spending 2 years developing scientific content for International Innovation, before completing an MSc in Science Communication. After gaining invaluable experience in supporting the communications efforts of CERN and IN-PART, I joined Texere – where I am focused on producing consistently engaging, cutting-edge and innovative content for our specialist audiences around the world.


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