Transformation products from per- and polyfluoroalkyl substances (PFAS) are now evident across the water cycle. Trifluoroacetic acid (TFA), the smallest and most mobile of them, has been measured at microgram-per-liter levels in precipitation and tap water, with recent studies warning of an irreversible global rise.

Analyzing these ultra-short PFAS is especially difficult. They evade retention on standard columns and contaminate even clean systems. A recent Phenomenex webinar showcased two chromatographic designs that make ultra-short PFAS measurable within routine workflows. Separation Science spoke with Sam Lodge, business development manager, and Luigi Margarucci, senior scientist, about how system design can separate true signal from persistent background.

Challenges in Measuring Ultra-Short PFAS

Perfluoropropionic acid (PFPrA) has joined TFA as a focus of new research, though most monitoring data still center on TFA. Both compounds have short biological half-lives that once suggested low risk, but ongoing studies indicate that continuous exposure may still lead to long-term effects.

“There’s significant interest in ultra-short PFAS across government, academia, and industry, even though regulatory and health data remain limited,” says Lodge. “Many labs already include them in their PFAS panels or run them as separate methods.”

As interest has grown, so has recognition of how hard these compounds are to measure. Lodge points to three main challenges:

• Retention and matrix effects. Their extreme polarity gives them little to no retention under reversed-phase conditions, and matrix ions can shift response through suppression or enhancement.
• Limited MS transitions. Most ultra-short PFAS yield only one strong transition and a weak qualifier, restricting confirmation and sensitivity at trace levels.
• Laboratory contamination. Reagents such as TFA are highly mobile and appear even in controlled systems, creating background peaks that obscure true signal.

Phenomenex certifies many PFAS-free consumables to one-third of reporting limits for C4–C18 compounds, but those safeguards don’t cover the ultra-shorts, notes Lodge. Contamination can stem from plasticware, caps, fittings, or even solvent and lab water. Even in controlled workflows, trace residues appear in the chromatogram and can raise apparent results.

Managing Background with a Delay Column

Phenomenex’s first solution tackles contamination using a delay column—a short, inert LC column placed between the mixer and injector. This additional step holds back PFAS from the mobile phase and hardware so they elute several minutes after the analytes, rather than on top of them. By shifting background peaks away from target retention times, the delay column prevents false positives without changing gradient conditions or run time.

The delay column, built on Luna Omega PS C18 chemistry, features a polar-embedded C18 phase with a polar group positioned near the silica surface. This structure delivers:

• Balanced retention for both polar and non-polar compounds.
• Stable wetting under highly aqueous or acidic conditions, preventing de-wetting common to traditional C18 phases.
• Broader selectivity that delays very short, polar PFAS (C1–C5) while still accommodating longer chains.

In the webinar, Lodge presented results showing that two delay columns paired with a Luna Omega PS C18 analytical column under acidic conditions achieved more than five minutes of retention for TFA. The setup produced over a two-minute gap between the true sample peak and any delayed signal from mobile-phase contamination, effectively eliminating early-eluting interferences and minimizing ion-suppression effects.

“The delay column is essential for removing positive bias,” says Lodge. “PFAS from the mobile phase or hardware can add directly to the analyte signal. Shifting that background out of the analyte window ensures the concentration you measure is real.”

Strengthening Retention with Polar Pesticide LC

Margarucci’s team took a different route—improving retention instead of delaying background. Their goal was to keep the ultra-short PFAS on-column long enough for clear separation and quantitation. Using the Phenomenex Luna Polar Pesticide column, a proprietary stationary phase with a well-balanced reversed-phase and hydrophilic interaction. This column achieved a stable retention for polar acids that elute too early on conventional C18 media.

Margarucci explains that the stationary phase was originally designed for polar pesticides—compounds that, like TFA and PFPrA, are small, highly ionic, and prone to early elution. “The Polar Pesticide column can retain both anionic and cationic polar compounds through a combination of hydrophobic and polar mechanisms,” he says. “That dual behavior makes it ideal for the shortest PFAS, which would otherwise elute with the solvent front.”

The same column serves two functions. In reversed-phase mode, it resolves C1–C5 PFAS within ten minutes. Under hydrophilic interaction conditions, the elution order reverses, providing built-in confirmation of identity. Because difluoroacetic acid—chemically similar to TFA—is already measured on this column in food analysis, Margarucci considered it a suitable platform for extending the method to ultra-short PFAS. A recent Phenomenex technical note demonstrated this same approach, showing reliable detection of TFA in tomato products using the Luna Polar Pesticide column and a delay column to reduce system interference.

Margarucci's results demonstrated that the Polar Pesticide design maintained stable retention and clean peak shapes for TFA, PFPrA, and related short-chain acids in various water matrices. Using 0.3 percent formic acid kept matrix effects low and reproducibility within a few percent RSD over repeated runs.

Selecting the Right LC Approach

Each design, the Phenomenex team explains, addresses a different interference. The delay column separates solvent- and hardware-derived peaks from the analyte window, while the Polar Pesticide phase retains ultra-short compounds long enough for confirmation. Used together, they sharpen signal and extend coverage without changing core LC–MS conditions.

Together they show that chromatographic control—not added sensitivity—makes the smallest compounds measurable. To see both approaches in action and hear discussions of broader strategies for PFAS analysis, watch the full Phenomenex webinar.

Meet the Experts:

Sam Lodge Environmental Senior Business Development Manager, Phenomenex Sam Lodge is the Phenomenex Senior Business Development Manager for the Environmental industry. He has been with Phenomenex for more than 30 years in roles including Key Account Manager and Senior International Sales Manager, and is a product expert in SPE and HPLC chemistries. Over the past eight years, he has worked extensively with commercial and government laboratories on PFAS analytical methods, including the validation of EPA 533, the MLV for DM 1633, and multiple internal (MOD) methods.

Luigi Margarucci, PhD Senior Scientist and Technical Trainer, Phenomenex Luigi Margarucci, PhD, joined Phenomenex in 2015 and now leads seminars and application training for EMEA (Europe, Middle East, and Africa). With a background in pharmaceutical sciences and research at the University of Salerno and the University of Utrecht, he has authored numerous publications and led educational programs in chromatography, LC-MS, GC, and SPE.