Valérie Pichon

The nano-LC-HRMS method previously developed for the analysis of human chorionic gonadotropin (hCG) glycoforms at the intact level involved trifluoroacetic acid (TFA) as ion-pairing agent in the LC mobile phase to favor separation, but resulted in both a significant decrease in the mass spectrometry (MS) signal and in complex mass spectra due to the presence of numerous adduct forms leading to a time-consuming manual data treatment and contributing to the MS signal decrease. To find the best balance between LC resolution, ESI ionization, and absence of adduct detection, two main strategies were here explored for the first time for hCG characterization. To enhance MS signal and reduce adduct formation, a decrease from 0.05 to 0.025% of TFA concentration was first assayed. It led to an efficient increase in MS signal intensities, but high glycoform/adduct ratios were still detected. The switch from TFA to difluoroacetic acid (DFA) 0.05% significantly improved the intensity of the signal in MS and significantly decreased adduct ratios (from 30 to 10%), despite some co-elution of hCG alpha (hCGα) and beta (hCGβ) glycoforms during LC separation. Keeping DFA as an ion-pairing agent, the All Ion Fragmentation mode of the Orbitrap mass spectrometer efficiently dissociated the remaining adducts leading to an increase in MS signal intensity by a factor 3 to 11 for hCGα glycoforms and 6 to 23 for hCGβ glycoforms. With these conditions, 76 hCGα and 200 hCGβ glycoforms were detected, resulting in a significant increase in the number of glycoforms detected compared with the initial method: a twofold increase for hCGα and a sixfold increase for hCGβ, thereby enabling a more detailed characterization of hCG glycosylation. Graphical abstract The alternative text for this image may have been generated using AI.

An oligonucleotide-based sorbent was developed to enable the selective extraction of lead ions from serum and water samples. Two oligonucleotide sequences, previously reported in the literature for their high affinity toward Pb(II), were grafted onto CNBr-activated Sepharose, producing sorbents suitable for solid-phase extraction. Grafting efficiencies ranged from approximately 50 to 70%. Subsequent optimization focused on adjusting the percolation and washing conditions to ensure preferential retention of Pb(II) on the specific aptamer-modified supports, followed by controlled elution. The elution kinetic was studied to minimize the elution volume. Under optimized conditions, high selectivity was obtained, with mean extraction recoveries of 83 ± 2.7% and 79 ± 8% ( n = 3) for the specific sorbents, compared to only 7–10% for the control sorbents. Other cations showed recoveries below 15%, confirming the strong specificity of the developed sorbents. The capacity of the most efficient sorbent was estimated at approximately 600 ng per 30 mg of support (≈30 µg/g), which is suitable for the selective extraction of trace amounts of Pb(II) from serum or water samples. Applying this support to real samples made it possible to determine Pb(II) levels of 6.3 µg/L for tap water, 2.7 µg/L for the Seine River water, and 87 µg/L for digested serum samples, while reducing matrix effects and minimizing ICP-MS instrument clogging. Graphical abstract

Adduction on protein nucleophile sites by mustard agents can be monitored to assess detection of retrospective exposure to these agents. Cysteine 34 (Cys34) on human serum albumin was selected as the target of choice. This work targets di‐ and tripeptides adducted on Cys34 by sulfur mustard, sesquimustard, and nitrogen mustards separated in hydrophilic liquid chromatography (HILIC) and Reversed–Phase (RP) mode. The effect of several mobile phase additives on the mass spectrometry (MS) and MS/MS signal and on LC retention profile was studied. A mix of ammonium acetate and acetic acid offered satisfactory results in terms of MS sensitivity. Screening of HILIC columns was performed, and ZIC‐HILIC stationary phase was selected for HILIC mode, and C₁₈ stationary phase was used for RP analysis. Negative ionization mode leads to a higher S/N ratio compared to positive ionization mode. Adducted tripeptides were selected for the monitoring of mustard agents’ exposure, allowing better sensitivity than their dipeptide homologues. The two developed chromatographic methods have similar sensitivities with LOQs ranging from 1.9 to 20.5 ng/mL for the reversed‐phase liquid chromatography (RPLC)–ESI–(−)–MS/MS method and from 1.7 to 43.3 ng/mL for the HILIC–ESI–(−)–MS/MS method. The monitoring method should be selected based on the targeted mustard agent, and the remaining method can be a confirmation tool.

The aim of this work is the development of a biomimetic strategy involving a molecular recognition mechanism using aptamers immobilized on a solid support for the analysis of the mycotoxin zearalenone (ZEA) and two of its derivatives in human urine: alpha-zearelenol (α-ZEL) and beta-zearelenol (β-ZEL). Three oligonucleotide sequences reported in the literature as being specific to ZEA were thus covalently grafted onto activated sepharose, and a thorough study of the percolation and washing conditions was performed to promote the selective retention of the three targeted compounds. With the optimized extraction procedure, a strong and selective retention was obtained for ZEA and to a lesser extent α-ZEL and β-ZEL, with extraction recoveries of 88±9%, 77±15%, and 45±12% respectively, in standard solutions. Application of this procedure to spiked human urine strongly highlighted the efficiency of the clean-up effect resulting from the use of this selective sorbent. Limits of quantification of the whole analytical procedure including extraction on oligosorbent and LC-MS analysis were 0.18 and 0.24 ng mL⁻¹, for ZEA and α-ZEL, respectively, thus demonstrating clearly the potential of the developed method for monitoring human dietary exposure to these compounds. Graphical abstract

Pregnant women and their fetuses, a very fragile population, are exposed to polycyclic aromatic hydrocarbons (PAHs) and subject to potentially serious health risks. Consequently, the determination of PAHs in maternal serum and umbilical cord blood is necessary but challenging, due to their highly varied physico-chemical properties and trace presence. The aim of this project was to develop a method enabling, for the first time, the simultaneous determination of 22 listed PAHs from small volumes (100 μ L) of maternal serum and umbilical cord blood, including a sample pre-treatment reducing as far as possible the quantity of solvents used. A solid-phase extraction step on C18-grafted silica was optimized by studying the effect of different parameters (nature and proportion of organic solvent added to the sample, washing and elution conditions) on extraction yields. The evaporation step was carefully controlled to limit the loss of the most volatile PAHs. Then, to break any interactions between PAHs and serum proteins, a precipitation step prior to solid-phase extraction was optimized using a design of experiments. The final sample processing procedure led to extraction yields of between 27 and 57% and between 34 and 69% for 22 PAHs in serum and umbilical cord blood, respectively, with standard deviation values of less than 11%. Limits of quantification between 0.2 and 3.1 μ g/L were achieved for PAHs fluorescing in spiked real samples, close to the concentrations expected in biological samples.

Monitoring a synthesis reaction in real time could allow not only the detection of the intermediates involved in the synthesis, to better understand its mechanisms, but also the impurities. Spectroscopic methods could be performed but are not so performant when analyzing complex mixtures and could require specific properties for the detection of the molecules of interest, the presence of a chromophore moiety for example. Mass spectrometry (MS) may overcome these limitations and is able to reach the accuracy and sensitivity required to efficiently detect, quantify, identify, and characterize the reagents and species produced during the synthesis. This is why the hyphenation of a microreactor with MS has already allowed synthesis processes to be monitored, but most of the time it targets a specific reaction or compounds and involves solvents compatible with MS. In this study, a universal setup for the hyphenation of a microreactor with MS and based on two valves has been developed. This two-valve setup has proven itself for the analysis of molecules of different nature and hydrophilicity, soluble in a large number of solvents even in non-MS-compatible ones. The developed setup evidenced a good repeatability and a linear response for the detection of the studied compounds. In addition, the dilution step included in the two-valve setup allows the MS monitoring of compounds initially synthesized at different concentrations. Finally, it was successfully used to study an amination reaction allowing the detection of the reaction products in 4 min with good repeatability as RSD values of MS signals were lower than 17%. Graphical Abstract