News from LabRulezLCMS Library - Week 30, 2025

Our Library never stops expanding. What are the most recent contributions to LabRulezLCMS Library in the week of 21st July 2025? Check out new documents from the field of liquid phase, especially HPLC and LC/MS techniques!

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This week we bring you posters by Agilent Technologies / ASMS, Shimadzu / ASMS, Thermo Fisher Scientific / ASMS, Waters Corporation / ASMS and application note by Metrohm!

1. Agilent Technologies / ASMS: De Novo PFAS Annotation and Classification Using Highly Accurate Formula Prediction and Kaufmann Algorithms Embedded in FluoroMatchSuite

• Poster
• Full PDF for download

FluoroMatch Suite MS1 Only Algorithms: FluoroMatch Suite covers the entire non-targeted PFAS workflow (Figure 1) using MS/MS algorithms and the largest fragmentation database to date providing a <5% false positive rate. However, MS/MS coverage is often incomplete, and DIA methods often provide lower quality spectra after deconvolution. Therefore, we developed MS1 only algorithms for providing highly accurate formula predictions (Figure 2) and for classifying features as PFAS or not PFAS using Kaufmann analysis (Figure 3) 

Validation: Algorithms were validated on NIST A and NIST C samples from the NIST PFAS interlaboratory study. NIST A consisted of known PFAS chemical standards, and NIST C consisted of an aqueous fire fighting foam (AFFF) contaminated soil sample. Formula prediction with homologous series voting (taking the most common repeating formula) was extremely accurate for both samples with a 0% false positive rate, and without homologous series voting the false positive rate was between 17-26% (Table 1). Kaufmann Analysis was highly specific for classifying PFAS with only 4% of non-PFAS being classified as PFAS with our Kaufmann based model, and only 6% of PFAS not be classified as PFAS according to our model for NIST C (Table 2).

2. Metrohm: Detection of heavy metals with differential pulse voltammetry

• Application note
• Full PDF for download

DPV, alongside square wave voltammetry (SWV), already discussed in AN-SENS-001, is another example of a pulsed voltammetric technique. DPV was actually developed first and is often linked to the field of polarography. This pulsed voltammetric technique is typically used with a dropping mercury electrode, although the general principle can be applied to static electrodes as well. In the case of DPV, a short pulse with constant height is superimposed at the end of every step of a staircase linear sweep voltammetry (LSV) determination (Figure 1). During the experiment, the current is sampled at two points: before (i1 ) and after (i2 ) the pulse. The difference (i1 -i2 ) vs. the potential is the signal which is plotted in DPV. Sometimes i1 is referred to as the base current and i2 as the pulse current. Figure 1. Typical DPV waveform for the forward direction (e.g., from 0 to 1 V). 

Like SWV, this approach allows the separation of faradaic and capacitive current, boosting the sensitivity of the technique compared to normal staircase voltammetry. In INTELLO, the i1 and i2 signals can optionally be sampled in the signals tab of the DPV command. In NOVA, custom plots of either signal can be produced in the same tab. The waveform is characterized by the height of the pulse (ΔE) and the step height (ΔEs ). The scan rate is determined only by step height and step duration of the underlying LSV. The pulse duration should not be set longer than the step duration in order to avoid artifacts, with typical values for the step being around 100–500 ms, and around 50 ms for the pulse. In comparison to SWV, DPV is considered less applicable to a wider range of systems due to the interference of oxygen and the slower scan rates required for DPV. In some cases it can produce better separation of closely positioned peaks and sharper peaks in general. The following is an example of a typical DPV measurement made with a PGSTAT302N and the Metrohm 663 VA stand, with the same measurement also possible with the next-generation VIONIC powered by INTELLO.

SAMPLE AND EXPERIMENTAL DETAILS 

A PGSTAT302N equipped with a Metrohm 663 VA stand (2.663.0020) was used for this study. The working electrode (WE) was a Multi-Mode Electrode pro (6.1246.120) in the hanging dropping mercury electrode (HDME) configuration. The reference electrode (RE) was a double junction Ag/AgCl reference electrode (6.0728.120). All potential values mentioned in this Application Note are referred to the potential of this electrode. The electrolyte was composed of 10 mL H2 O + 0.5 mL of acetate buffer, made of 1 mol/L ammonium acetate + 1 mol/L acetic acid. 

The concentration of heavy metal ions was quantified by making two standard additions. For the first addition, the solution was composed as follows: 10 mL H2 O, 500 μL acetate buffer, 100 μL Pb standard solution (1 mg/L), and 100 μL Cd standard solution (1 mg/L). For the second addition, the solution was composed as follows: 10 mL H2 O, 500 μL acetate buffer, 200 μL Pb standard solution (1 mg/L), and 200 μL Cd standard solution (1 mg/L). 

Each measurement consisted of a step to precondition the Hg electrode, where nitrogen was purged in the stirring solution and a new Hg drop was formed. Then, a reduction potential of -0.9 V was applied at the Hg drop in order to reduce the Pb and Cd cations at the Hg drop surface under stirring conditions and accumulate them at the Hg drop. The stirrer was then switched off and the DPV measurement was performed by oxidizing the previously accumulated Pb and Cd in a positive potential sweep from -0.9 to -0.2 V. The DPV measurement was repeated twice. 

The peak heights were determined from the current vs. potential plot of the three experimental iterations (i.e., sample, sample with first standard addition, and sample with second standard addition). Peak height vs. analyte concentration plots were created for both Pb and Cd. The analyte concentration in the sample can be calculated from the intersection of the regression line with the x-axis.

3. Shimadzu / ASMS: EPA 533 and 537.1 with the Novel LCMS-8050RX: Demonstration of Instrument and Method Performance to meet PFAS regulatory levels 

• Poster
• Full PDF for download

The harmful impacts of per- and polyfluorinated alkyl substances (PFAS) continue to afflict the environment. Water utilities require sensitive and robust analytical methods to assess the current and ongoing burden of PFAS from source waters to meet regulatory demands in treated water and to protect public health. Improvements in sensitivity and/or runtime without sacrificing robustness can aid laboratories by increasing sample throughput to better meet the significant demand for PFAS sample analysis. The novel RX probe was developed to yield better heat distribution and more consistent nebulization to improve sensitivity and reproducibility. Chromatographic improvements compared to existing EPA methods were also performed to decrease runtime while maintaining the integrity of sample analysis.

Methods

PFAS were separated by a Nexera LC and quantified by a Shimadzu LCMS-8050RX for EPA methods 533 and 537.1 (EPAM533 and EPAM537.1). Neat standards were analyzed in the required composition (i.e., 80% methanol for EPAM533 and 96% methanol for EPAM537.1). Standards were analyzed twenty times to determine the instrument detection limits.

Results and Conclusion

• All PFAS were sufficiently analyzed by the respective methods and passed regulatory criteria at a reduced runtime (19 minutes shorter).
• Superb precision and reproducibility were demonstrated with the RX probe, which resulted in an analytical blueprint that greatly exceeds the Final PFAS National Primary Drinking Water Regulation promulgated in 2024.

4. Thermo Fisher Scientific / ASMS: Comprehensive and high-throughput plasma proteome profiling for biomarker discovery using the Orbitrap Astral Zoom Mass Spectrometer and the Seer Proteograph ONE workflow

• Poster
• Full PDF for download

Plasma proteomics, the study of proteins in blood plasma, is crucial for biomarker discovery and understanding global signaling and immune responses1, aiding disease diagnosis, monitoring, and personalized medicine. However, the plasma proteome's complexity and dynamic range—spanning up to 12 orders of magnitude²—pose challenges, especially for detecting low-abundance proteins. Additionally, preanalytical, analytical, and biological variables complicate reliable biomarker identification, necessitating optimized techniques³. Liquid chromatography-mass spectrometry (LC-MS) excels in plasma proteomics due to its high analytical dynamic range and capability to measure peptides and characterize posttranslational modifications (PTMs). Advances in sample preparation, separation, MS, and data analysis are improving method sensitivity and robustness. We demonstrate the Thermo ScientificTM OrbitrapTM AstralTM Zoom Mass Spectrometer's high sensitivity and reproducibility in detecting a comprehensive plasma proteome across neat, depleted, and plasma processed with the Seer® Proteograph® ONE workflow, facilitating potential biomarker discovery¹.

Experimental Procedure

Liquid chromatography-mass spectrometry & data analysis

All LC-MS runs for neat, depleted, and Proteograph ONE processed peptides were separated and analyzed using a Thermo Scientific Vanquish Neo UHPLC System in trap and elute configuration, paired with an Orbitrap Astral Zoom Mass Spectrometer. Peptides were separated on the Vanquish Neo UHPLC System using Thermo Scientific EASY-Spray PepMap analytical columns (ES906, 2µm C18, 150µm × 15 cm for 100 and 60 SPD and ES75500PN 2µm C18, 75µm × 50 cm for 24SPD) and 12.6-, 22.35-, and 54-minute chromatographic gradients, respectively, were formed using 0.1% formic acid in water for mobile phase A and 0.1% formic acid in 80% acetonitrile for mobile phase B. Samples were injected with 2 loading mass amounts (200/500 ng for 100/60 SPD; 500/1250 ng for 24 SPD) in triplicate. Liquid chromatography parameters are shown below in Table 1A. Mass spectrometer scan parameters can be found below in Table 1B. All data was processed using the Proteograph® Analysis Suite (PAS) (Seer). Output files were imported to Rstudio (2023.09.0 Build 463) with R (v4.3.1) for downstream data analysis and visualization.

Conclusions 

• With the Orbitrap Astral Zoom Mass Spectrometer, neat plasma proteome coverage exceeds 1,000 protein groups in a single pooled sample, providing exceptional quantitative precision without additional sample handling steps. 
• High Select Top 14 abundant protein depletion enhances plasma proteome depth by ~2x across 100, 60, and 24 SPD throughputs with comparable or superior analytical measurement precision. 
• Seer Proteograph ONE enhances plasma proteome depth by 7.9x (100SPD), 6.9x (60SPD), and 6.2x (24SPD) with comparable or superior analytical measurement precision and no compromise on sample preparation throughput. 
• The combination of the Orbitrap Astral Zoom Mass Spectrometer and the Proteograph ONE Workflow allows for over 7,000 protein groups to be identified from a single injection. 
• The speed, sensitivity, and robustness of the Orbitrap Astral Zoom Mass Spectrometer enables in-depth biological discovery, paving the way for novel findings and advancements in clinical cohort and population-scale translational research.

5. Waters Corporation / ASMS: PFASt and Furious: A Targeted and Non-Targeted LC-MS/MS Study on the Environmental Impact of Using Fluorinated Ski Waxes 

• Poster
• Full PDF for download

This study by Waters Corporation explores the environmental impact of fluorinated ski waxes—specifically per- and polyfluoroalkyl substances (PFAS)—on soil and water quality at competitive ski sites in New England. Using both targeted LC-MS/MS and non-targeted high-resolution mass spectrometry, the research team analyzed samples from slalom and cross-country racing courses to determine the extent of PFAS contamination.

PFAS were found in all collected samples, with soil showing significantly higher concentrations than water. Long-chain perfluorocarboxylic acids, commonly used in ski waxes, dominated the contaminant profile. At slalom courses, the highest concentrations were found at the wax application zones near the start line, decreasing progressively downhill. At cross-country sites, similar patterns emerged near waxing and testing areas, with alarming migration of PFAS into adjacent ponds and even a vegetable garden.

The study employed automated sample preparation systems (CEM EDGE and Promochrom SPE-03) to ensure efficiency and reproducibility. Advanced LC-MS/MS analysis with Waters’ ACQUITY Premier and Xevo TQ Absolute platforms enabled the detection of common PFAS, while cyclic ion mobility HRMS revealed additional, untargeted long-chain species (e.g., C15/C16 carboxylic acids) not typically included in regulatory workflows.

The findings highlight the long-term environmental risks posed by legacy ski wax formulations and underscore the need for continued monitoring. Notably, unexpected PFOS levels in garden soil suggest multiple PFAS sources may be contributing to contamination, warranting further investigation.