Application of thymine-based copolymers in the quantification of organic pollutants in water by sensitized fluorescence
Introduction
The development of inexpensive analytical techniques for rapid on-site detection and determination of water organic pollutants is a long-lasting topic since the availability of drinking water is a complicated problem that concerns both developed and emerging countries.
Polycyclic aromatic hydrocarbons (PAHs) include a huge number of compounds, consisting on fused aromatic rings that do not contain heteroatoms or substituents. PAHs are present in food, water, soil, and air, mostly from incomplete combustion of carbon-containing materials such as oil, wood, coal or garbage [1]. Based on their source they can be classified in biogenic, pyrogenic or petrogenic [2]. For decades, special attention was given to identify the sources and modes of transportation throughout the environment considering the low biodegradability, high lipophilicity, and potential carcinogenic or mutagenic effects [3], [4], [5]. Due to the toxicity, both the Environmental Protection Agency (EPA) and the International Organization for Standardization (ISO), established robust methods for the PAHs accurate determination in different sample types [6,7]. Time-consuming chromatographic systems with sample pre-processing consisting of several extractions (liquid-liquid, solid-liquid) are used to develop these methods. But these procedures involve the use of large amounts of organic solvents like hexane or dichloromethane that can cause serious harm to the environment.
A sensitive, selective, fast, versatile, and simple analytical technique that offers remarkable analytical advantages in the determination of organic pollutants in water, is the molecular fluorescence spectroscopy sensitized by fluorescent sensors [8,9]. Among the main advantages, an enhancement in sensitivity values can be highlighted, which can become several orders of magnitude higher than the standard methods [10,11]. Hence, the design of fluorescent sensors is a research field in permanent evolution.
From a broad perspective a fluorescent sensor is a device capable to provide information about the presence of a specific analyte in a sample through the interaction with it, transforming either qualitative or quantitative chemical information into an analytically useful fluorescence signal [12,13]. There is already a wide choice of fluorescent sensors for particular applications, and many of them are commercially available. However, there is a need for sensors that, in addition to improving the analytical figures of merit of the method, will have a low impact on the probed microenvironment.
In recent years, sensitized fluorescence spectroscopy (SFS) technique have used the sensors mostly in two ways: as a powdered solid (small-sized particles, finely dissolved in solution or as solid phase), or as membranes where the compounds, analytes or reaction products are immobilized by physical or chemical procedures [14]. Fluorescence emission did not significantly change upon being emitted from solid support or a solution, although the quantum yield tends to be higher in solid. Docking a potentially fluorescent analyte on a sensor limits its motion to a rigid environment and reduces less-radiant deactivation, increasing the quantum yield. Therefore, SFS method enables improvements in the photophysical emission process while reaching up high pre-concentration factors. The possibility to generate multidimensional data (spectral, lifetime, polarization, and other simultaneous measurements) with sub-nanometric spatial resolution, sub-millisecond temporal resolution, and submicron visualization has an undeniable analytical interest [15,16]. Due to such advantages, several SFS setups have emerged, which enable their use in numerous analytical problems within different scientific fields.
The key to SFS methods is to achieve an adequate immobilization of the analyte to facilitate its fluorescent emission. One of the most used immobilization technique is the retention of the analytes on a solid phase at room temperature. Once the analyte is docked, the fluorescence phenomenon is produced from the surface of the system, resulting in a diffuse luminescence due to multiple scattering by particles of the solid phase. A simpler way to achieve the analytes protection is to dissolve the sensor as a powder into the water sample that contains them. There are different ways to implement this strategy. It is possible the use of surfactants, at concentration equal or higher than its critical micellar concentration; or alternatively, is feasible the use of water-soluble polymers, since they give structural rigidity to the analyte molecules. . . The two approaches share the same basic idea, and both are capable of being automatized. The powdered solid supports are packed into flow-through cells and used in flow analysis systems. On the other hand, membranes are implemented as probe-type systems, either in batch or continuous mode.
Regarding the application of SFS methods for the quantification of PAHs in different environmental samples, there is a wide range of strategies that have been applied. From the pioneering works so far, the use of different solid-phase extraction membranes, some soluble fluorescence enhancers and even in situ optical devices have been reported. The continuous development and improvement of room temperature SFS methods for the analysis of PAHs show the permanent interest of this topic [10,[17], [18], [19], [20], [21], [22], [23], [24], [25]].
In a recent work, the existence of non-covalent interactions between a bio-inspired thymine-based copolymer and benzo(a)pyrene (BaP) was confirmed through two analytical methods: fluorescence spectroscopy and scanning electron microscopy [26]. Accordingly, the fluorescence of BaP in water solutions of a copolymer formed by 1-(4 vinyl benzyl) thymine (VBT) and vinyl phenyl sulfonate (VPS), significantly increases. Optimized geometries obtained through a computational study suggested that two forms of π interaction exist in the system, hydrogen polar-π interaction (Hp-π) and π-π "stacking" interaction. The high affinity between the VBT-VPS copolymer and BaP can be explained by Hp-π interaction, which arises from hydrogen bonded to nitrogen at the thymine moiety of the copolymer and the external aromatic ring of BaP. Based on the structural similarities that heavy PAHs have, it is reasonable to assume that similar interactions can occur with other family compounds, such as pyrene (Pyr).
The VBT-VPS copolymer has key features to become a suitable luminescence sensor [27], [28], [29], [30], [31]. As indicated, it is capable of immobilizing some PAHs and enhance their fluorescence, and at that time the analytical signals can be related to the concentration of the analytes in the sample The copolymer can be incorporated as optical sensor in flow methodologies, making it an excellent alternative [32]. Some practical applications of VBT from hair-styling products to recyclable plastics have been reported [33], [34], [35], [36], [37]. This wide variety of applications are achieved by fine-tuning the balance between solubility and non-covalent interactions of VBT monomer [38].
The use of the VBT-VPS copolymer allows achieving the required analytical sensitivity; however, the similarity of BaP and Pyr fluorescence spectra makes complicated their simultaneous determination. In addition, the selectivity of each analyte can decrease due to the presence of interference, preventing the acquisition of accurate information. The use of a second-order calibration based on parallel factor analysis (PARAFAC) model, allows a mathematical data treatment which is able to isolate the signals related to the analytes, separating them from the interferences [39]. This valuable property, namely “second-order advantage” [40], avoids the requirement of either the interference removal (as in zeroth-order calibration) or the creation of a large and representative calibration set (as in the first-order calibration). In the present report, second-order data were obtained through excitation-emission fluorescence matrices (EEFMs).
It should be noted that our proposed method suits within the framework of Green Analytical Chemistry (GAC) methods, stating that the detection and quantification of environmental pollutants should not pollute the environment more than the analyte to be determined [41,42]. Some considerations have been made to develop environmentally friendly alternatives for the first step of the analytical process (like sample preparation step), but only a few to improve green approaches for analyzing the analytical signal [9]. The acquisition of second-order data during the measurement step minimize the negative effects of the sample treatment, without modifying the quality of the results [9]. To highlight the role of multivariate calibration as a useful tool to achieve the pursued goal, the indicator Analytical Eco-Scale was used for assessing the greenness of our analytical procedures, as proposed in ref. [41].
This work presents a simple method for the simultaneous determination of BaP and Pyr in drinkable water in the presence of interferent species, based on sensitized fluorescence spectroscopy and chemometric analysis. The key to the enhanced fluorescent signal is the use of a thymine-based copolymer solubilized in the water sample. The VBT-VPS copolymer combines an adequate solubility and a demonstrated non-covalent interaction at room temperature with the analytes, being this interaction responsible for significantly increasing the analyte fluorescence. The ability of VBT-VPS copolymers to exacerbate the analytical properties of the contaminant is then combined with a well-established PARAFAC analysis of excitation-emission fluorescence data, exploiting in this way the second-order advantage.
Section snippets
Reagents and solutions
Pyrene (Pyr) was purchased from Sigma-Aldrich (Milwaukee, WI, USA) and benzo(a)pyrene (BaP), acenaphthylene (ACE) and anthracene (ANT) were obtained from Sigma-Aldrich (Milwaukee, WI, USA). All reagents were of high-purity grade and used as received.
Stock solutions of Pyr (4 × 10−4 g mL–1), BaP (1.4 × 10−3 g mL–1), ACE (4.4 × 10−4 g mL–1), and ANT (3 × 10−4 g mL–1) were prepared in acetonitrile. From these solutions, more diluted aqueous solutions [Pyr (40 ng mL–1), BaP (280 ng mL–1), ACE
Theory
PARAFAC theory is well established and has been duly documented [39], only a brief description is presented here. PARAFAC achieves a unique decomposition of three-dimensional data arrays, allowing concentrations and spectral profiles of sample components to be resolved. The first step is to build a three-way array M of dimensions I × J × K for each EEFM, where I, J and K are the number of samples, number of emission wavelengths and number of excitation wavelengths, respectively. The algorithm
Results and discussion
To find the best sensitizer-analytes ratio, fluorescent signal saturation curves were made for every analyte separately by successive addition of amounts of VBT:VPS to BaP or Pyr water solutions (5 ng mL–1 and 1 ng mL–1, respectively). Fig. 1 shows the obtained results.
For BaP a signal saturation was observed when 1 × 10–2 g of copolymer were added (Fig. 1, white triangles). The signal enhancement factor is ~ 6 calculated as the ratio between enhanced signal and signal of the analyte solution
5. Conclusions
In the present work, a new method combining sensitized fluorescence spectroscopy and multivariate analyze was performed. Conclusions can be summarized as follows: (1) the VBT-VPS copolymer enhanced the BaP and Pyr fluorescence signal making it possible to reliably quantify them at levels below those required (i.e., 0.11 ng mL−1 for BaP and 0.06 ng mL−1 for Pyr), (2) analytical selectivity was achieved using a PARAFAC algorithm to process second-order excitation-emission fluorescence data, (3)
Declaration of Competing Interest
The authors declare no conflicts of interest.
Acknowledgments
The following institutions are gratefully acknowledged for financial support: Universidad Nacional de Rosario and CONICET (Consejo Nacional de Investigaciones Científicas y Técnicas), Argentina.
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2021, Environmental PollutionCitation Excerpt :These methods require large samples and lengthy, labor-intensive sample preparation steps that await the results and possible action for several days (Du and Jing, 2011). These methods also require large amounts of organic solvents like hexane or dichloromethane that are toxic to the human body and our environment (Ledesma et al., 2020). The limitation of these methods led to the development of other analytical tools such as visible near infra-red diffuse reflectance spectroscopy and surface enhanced Raman (SERS) spectroscopy (Chakraborty et al., 2010, 2014, 2015; Okparanma et al., 2014; Okparanma and Mouazen, 2013; Wijewardane et al., 2020; Xie et al., 2011) which are considered as rapid, non-destructive, and proximal sensing techniques for detection of total petroleum hydrocarbons.
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2021, Journal of Electroanalytical ChemistryCitation Excerpt :For example, it has been shown that, due to its hydrophobic-hydrophilic balance, VBT/VBA polyelectrolyte improves the conductive properties and stability of enzymatic bioelectrodes of lactate oxidase (Lox) [19], glucose oxidase (Gox) [20], and horseradish peroxidase (HRP) [21]. The anionic copolymer formed by 1-(4 vinylbenzyl) thymine (VBT) and 4-vinylphenyl sulfonate (VPS) has been used for the quantification of organic pollutants in water based on non-covalent interactions between the polymer and pyrene and benzo pyrene molecules [22]. Electrostatic interaction between the polycation [(VBT)1:(VBA)4]4+ and the polyanion [(VBT)1: (VPS)4]4− was recently used to obtain microcapsules of the polyelectrolytes assembled layer-by-layer on microspheres of CaCO3 as removable molds [23].
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Corresponding author at: Instituto de Química Rosario (CONICET-UNR), Suipacha 570 (S2002LRL) Rosario, Argentina.