Development of 19F NMR-based reproducible detection methods based on intramolecular internal standard strategy and its application

Abstract

Numerous nuclear magnetic resonance (NMR)-based sensors have been developed for detection of various analytes utilizing NMR spectroscopy, which is advantageous and compelling as an analytical technique. Especially, NMR-based sensors enable simultaneous recognition of multiple analytes, demonstrating a single readout comprising of individual signals for multiple variations within the same analytical frame. Among various nuclei applied in NMR-based sensor, fluorine (19F) nucleus has fascinating features like 100% natural abundance, little interference, and high sensitivity to slight changes in the regional environment. In addition, 19F NMR exhibited excellent signal intensity when compared to 1H NMR, and demonstrated a broad range for measurement. Based on these unique features, various 19F NMR-based sensors have intensively attracted attention. However, recently, inherent irreproducibility of 19F NMR spectroscopy has been reported. Thus, the development of new methods to improve the limitations found in 19F NMR-based measurements is highly necessary. In general, standard materials, which are used as the independent signal, provide accurate analyses and consistent results. Several referencing methods utilizing standard materials, such as a calibration protocol for traditional fluorinated compounds and development of new fluorinated materials as references, have been developed for improving reproducibility in 19F NMR spectroscopy. While these efforts have led to some improvements in reproducibility, these methods still require additional calibration or sampling processes for standard materials, reducing the efficiency in the aspect of sensor development. Consequently, several studies for efficient internal standard strategy based on the self-calibration concept have been mostly reported in optical sensor. However, this internal standard strategy has never been utilized in 19F NMR-based studies. In this study, the intramolecular internal standard strategy was first devised for reproducible detection of diols and pH using 19F NMR spectroscopy. Diol-containing analytes such as glucose are significant biomarkers, but most sensors for detecting diols have been developed as platforms that are selective for a specific analyte. As alternatives considering advantages of 19F NMR-based techniques, several 19F NMR-based sensors for exploration of various diols have been rarely reported. Herein, a fluorinated phenylboronic acid derivative for detection of diols was developed as proof-of-concept for new intramolecular internal standard strategy using 19F NMR spectroscopy. This strategy for reproducible detection of diols was demonstrated to be a dependable and consistent referencing technique, exhibiting acceptable deviation across multiple NMR spectrometers located at distinct institutions. Specifically, this strategy enables trustworthy fingerprint recognition of analytes through their unique characteristics, and facilitates concurrent qualitative and quantitative analysis of mixtures. Furthermore, the considerable recovery rates observed for D-glucose in human serum propose its prospective applicability in a wide spectrum of uses, including diagnostics related to diabetes. pH is a universal indicator that covers a wide range from strongly acidic to strongly basic, and pH measurements play an important role in understanding chemical and biological reactions. In addition, strongly acidic or basic pH can be observed in certain environments, such as the dye industry and wastewater treatment facilities, and extreme or out-of-normal ranges of pH can cause disease and ecosystem disruption. Consequently, numerous techniques have been documented for pH detection, encompassing pH meters and optical sensors. Notwithstanding their user-friendliness and precision, pH meters are vulnerable to inaccuracies in the extremely acidic or alkaline pH ranges, and mandate an internal reference solution. Meanwhile, most of optical sensors possess a restricted range of pH measurement, and are susceptible to interference from turbid samples that disrupt their photophysical properties, thereby inducing imprecise pH measurement. In this study, considering the unique features of 19F NMR spectroscopy, 19F NMR-based pH sensor with intramolecular internal standard was proposed for reproducible detection of strong acids and bases. Its 19F NMR response exhibited reliable results of deviation for the internal standard signal, and reversibility, including high stability to potential interfering factors. Furthermore, the low absolute difference observed for actual samples, such as diet Coke, indicates its promising applicability in diverse acidic beverages. Metal ions are abundant in nature and play a pivotal role in understanding biological processes for many phenomena. Among them, calcium ions are representative biomarkers, and their abnormal concentration levels in the blood are closely related to diseases such as hypercalcemia or tumors. Consequently, many techniques have been developed to detect calcium ions, including optical sensors such as fluorescent or colorimetric sensors. Despite their fast response and intuitive characteristics, optical sensor-based detection method for calcium ions have been limited by low sensitivity and susceptibility to interference from magnesium ions. To overcome, cofunctionalization strategies have been proposed, but there are synthetic inefficiencies due to the need to modify various functional groups, and simultaneous detection for various metal ions is difficult due to their characteristics related to selectivity. In this study, a 19F NMR-based metal sensor using intramolecular internal standard strategy was proposed utilizing various advantages of 19F NMR spectroscopy. In particular, by introducing a ligand with a carboxyl group that can effectively detect calcium ions, the development of an analytical system with high reproducibility in selective discrimination of calcium ions from real samples and simultaneous detection of various metal ions was performed.