HTC-19 Insights: RPLC×HILIC–cIMS for Data-independent Profiling of Phenolics: Potential Challenges and Future Developments

At HTC-19 in Leuven, Belgium, LCGC International spoke with Nikoline Juul Nielsen from the University of Copenhagen, Denmark on her presentation ‘Data-independent Profiling of Phenolic Constituents in Plant Extracts by Comprehensive Two-dimensional Liquid Chromatography (RPLC×HILIC)-Cyclic ion Mobility-Time-of-Flight Mass Spectrometry”.

In this video interview Nielsen answers the following questions:

• How significant is the challenge of deprotomer formation for accurate CCS prediction and metabolite identification?
• What developments are required for workflows like this to be suitable for routine use?

In her presentation, Nikoline demonstrated how combining comprehensive two-dimensional liquid chromatography (LC×LC), cyclic ion mobility spectrometry (cIMS), high-resolution mass spectrometry (HRMS), and data-independent acquisition (DIA) improves the profiling of plant secondary metabolites. She also introduced a script-based workflow for processing the large multidimensional datasets generated by this platform.

Using shea (Vitellaria paradoxa) kernels and shells, she identified 50 mainly phenolic metabolites and showed that LC×LC, cIMS, and DIA improved compound annotation by enhancing chromatographic separation, spectral purity, and confidence in identification. The study also revealed that shea shells contain valuable bioactive compounds despite being considered a waste product. In wheat (Triticum spp.) flag leaves, LC×LC provided improved separation of phenolic classes, while cIMS combined with DIA substantially increased spectral purity for low-abundance metabolites. However, ion mobility alone showed limited ability to resolve most isomeric compounds.

Overall, the presentation highlighted that LC×LC, cIMS, and DIA provide complementary analytical benefits for plant metabolite profiling, although specialized computational tools are currently required to process these complex datasets.

See the full interview series here:

1. Why Combine RPLC×HILIC, cIMS, and DIA-HRMS?

2. Practical Aspects of cIMS and Potential for Routine Analysis

3. Potential Challenges and Future Developments

References

1. Nielsen N.J. Data-independent Profiling of Phenolic Constituents in Plant Extracts by Comprehensive Two-dimensional Liquid Chromatography (RPLC×HILIC)-Cyclic ion Mobility-Time-of-Flight Mass Spectrometr. Presented at HTC-19 2026, in Leuven, Belgium. https://htc-19.com/programme/( accessed 2026-06-04).
2. Nielsen, N. J.; Kronik, O. M.; Forte Neran, R. A.; Christensen, J. H.; Ravn, T. K.; de Villiers, A. Data-Independent Profiling of Phenolic Constituents in Shea Using Comprehensive Two-Dimensional Liquid Chromatography (RPLC × HILIC) Hyphenated to Cyclic Ion Mobility–Quadrupole-Time-of-Flight Mass Spectrometry. J. Chromatogr. A 2025, 1760, 466092. DOI: https://doi.org/10.1016/j.chroma.2025.466092.

Biography

Nikoline Juul Nielsen is an Associate Professor at the Department of Plant and Environmental Science at University of Copenhagen (UCPH). She has a BSc in Agronomy, an MSc in Environmental Chemistry and did her PhD on analytical and data processing strategies in natural products research. She specializes in fundamental and advanced analytical chemistry, with focus on multidimensional separations built on chromatography and mass spectrometry techniques. She profiles complex environmental and biological samples, and extracts information through targeted or non-targeted approaches. At UCPH, she is the head of studies for the MSc program in Environmental Science, and serves as educator and mentor, having co-founded the first Danish MSc-specialization in Analytical Chemistry, developed several specialized MSc and PhD courses and supervised graduate students and researchers. Recently, and with four colleagues, she founded the spin-out company NTS Analytica. The company assists in planning, executes and interprets customized non-target screening investigations.