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Melnikov, P.;  Bobrov, A.;  Marfin, Y. Optical Polymer-Based Sensors in Environmental and Biological Systems. Encyclopedia. Available online: (accessed on 17 June 2024).
Melnikov P,  Bobrov A,  Marfin Y. Optical Polymer-Based Sensors in Environmental and Biological Systems. Encyclopedia. Available at: Accessed June 17, 2024.
Melnikov, Pavel, Alexander Bobrov, Yuriy Marfin. "Optical Polymer-Based Sensors in Environmental and Biological Systems" Encyclopedia, (accessed June 17, 2024).
Melnikov, P.,  Bobrov, A., & Marfin, Y. (2022, October 25). Optical Polymer-Based Sensors in Environmental and Biological Systems. In Encyclopedia.
Melnikov, Pavel, et al. "Optical Polymer-Based Sensors in Environmental and Biological Systems." Encyclopedia. Web. 25 October, 2022.
Optical Polymer-Based Sensors in Environmental and Biological Systems

Polymers are widely used in many areas, but often their individual properties are not sufficient for use in certain applications. One of the solutions is the creation of polymer-based composites and nanocomposites. In such materials, in order to improve their properties, nanoscale particles (at least in one dimension) are dispersed in the polymer matrix. These properties include increased mechanical strength and durability, the ability to create a developed inner surface, adjustable thermal and electrical conductivity, and many others. The materials created can have a wide range of applications, such as biomimetic materials and technologies, smart materials, renewable energy sources, packaging, etc.

polymer-based nanocomposites polymer matrix enhancement optical sensor fluorescence biocomposite materials molecular design absorption

1. Introduction

Measurement and control of the composition of aquatic environments, soil and air, as well as the quality assurance of various products produced by human is impossible without the use of analytical systems, and their improvement is a necessary permanent task. The latest trend is the development of smart materials that respond to given influences from external sources through physical or chemical reaction, which leads to a change in the characteristics of the material [1]. The smart polymer-based materials play an increasingly important role [2]. Among the possible sources of an analytical signal, one can distinguish pH, enzyme, redox, temperature, electric and magnetic fields, etc. At the same time, electrochemical and optical measuring systems occupy an overwhelming position among practical applications of the transducer type [3]. It is important to note that, for example, when measuring oxygen, there is a competition between the mentioned detection methods, and optical systems are gradually replacing electrochemical ones [4]. The main reason is that the optical method is not limited to measurements at a single point but allows imaging and intracellular measurements by means of nanoparticles with indicator dye, and fiber optic microsensors greatly facilitate their integration into research or industrial equipment. Among the practical applications are plant and animal physiology research, marine sciences, clinical chemistry, biotechnology and chemical industry [4].
Polymer-based sensors are of increasing interest because the ingredients used could be easily processed and could be matched for biocompatibility [5][6][7]. The steady increase in publications on the topic and investment in this area of research are proof that such technologies are among the most promising at the moment [1][8]. Conducting polymers, hydrogels, molecularly imprinted polymers (MIP), and composites and nanocomposites are the common polymer-based materials used in sensing devices [9][10][11][12]. The latter are the most interesting, as they have the widest scope for customizing the properties of the system being created [13][14][15]. On the one hand, the creation of such polymer-based composite systems can improve the molecular recognition, where the polymer serves as the matrix for the immobilization of various functional groups (dyes, luminescent nanofillers, etc.) and makes it possible to detect target analytes [16][17]. Another important advantage of polymer-based sensors is the ability to tune their physical/chemical properties so that their resistance to degradation, biodegradability and flexibility can be modified [1][18]. The increasing opportunities for recycling and upcycling of polymers should also be mentioned [19][20].
This section considers relevant practical examples of the use of the researched technologies for creating optical sensors. Undoubtedly, the most demanded is the measurement of physiologically important analytes such as O2, CO2, pH in biological systems, environmental and industrial objects. However, the range of possible analytes is not limited to these three substances, and such recent examples are given in the section.

2. Measurent of O2, pH and CO2

2.1. Food Packaging

The modern food industry is an actively developing area, in particular, intensive progress is underway in the field of new food packaging technologies. The main efforts are aimed not only at the introduction of new areas of knowledge (such as biotechnology or nanotechnology), but also take into account the environment awareness. An important growing trend is the development of “smart” packaging that can control the safety and quality of products, extend food shelf-life, as well as taking care of the environment. Their implementation requires, on the one hand, changes in production technologies, but on the other hand, it can change the practice of selling and the lifestyle of consumers [21].
One of these technologies measures oxygen gas concentrations inside food packaging [22]. Unlike other methods, the measurement does not require opening the package. It is enough to place a small sensitive foil into it at the production stage. Of course, the sensing material must be inert to the main product [23], and the use of composites described in the previous section facilitates this task. It is important that for practical measurements in a store, the user can even use a smartphone with the appropriate software. In this case, the flash of the smartphone, together with an optical filter, is used to excite the luminescent membrane, and the rear camera is used to register the analytical signal [24]. The presence of oxygen in the analyzed medium leads to quenching of the luminescence. Representation of an analytical signal as a ratio is the most convenient in terms of stability and accuracy of measured readings. In this case, the measured intensities are compared in two different wavelength ranges corresponding to the emission and absorption peaks of the sensitive membrane or the emission of the oxygen-sensitive and reference dyes. It is important to note that the thermal dependence of modern sensor materials is significantly reduced, as is the cost of their production [25][26]. The described approach shows that modern sensory materials can be easily integrated into smart packaging, and the current state of the pocket electronics makes it possible to avoid the need for a special measuring equipment.

2.2. Microparticles-Based Ink

This technology is a simple and inexpensive way to fabricate sensor arrays suitable for both point-to-point measurements and oxygen distribution mapping. For example, the possibility to print multiple sensor patches at ones was demonstrated [27]. The researchers used a polyvinylidene chloride film as a substrate and ink mixture prepared using the monodisperse polystyrene microparticles with immobilized oxygen-sensitive PtTFPP dye. A 50% mixture of ethanol and water with suspended high molecular weight polyvinylpyrrolidone was used as a dispersion medium for an ink mixture. The polymer used provides enhanced adhesion and compatibility with the most common polymer substrates. The obtained patches exhibited reproducible and fast response in a wide range of oxygen concentrations (0–21%) with linear Stern-Volmer plot (R2 > 0.99) and sufficient sensitivity (I0/I21% > 1.55). It was found that the intensity of the printed material luminescence mainly depends on the concentration of the fluorophore and, to a lesser extent, on the size of the polystyrene particles. It was shown that patches are suitable for measurement with both a multi-frequency phase fluorometer and a smartphone. The good uniformity of the resulting printed surfaces made it possible to carry out two-dimensional measurements with mapping of the oxygen distribution.
Non-fluorescent color indicators can also be used for ink-printed sensor production. For example, a quantitative colorimetric oxygen sensor was produced using the conversion of bisphenalenyls (PQPLs) into aromatic endoperoxides (EPOs) as the indicator [28]. One of the key features of the PQPLs is the ability to generate singlet oxygen, i.e., they possess self-sensitizing reactivity and do not require an external photosensitizer. The rate of photooxygenation depends on the electron-donating ability of PQPLs substituents, and it allows an easy way to tune the sensitivity. The produced EPOs are stable under ambient conditions but could be converted back to PQPLs by the thermal action. The produced sensing films exhibit visible to the naked eye rapid red-to-colorless transition in the presence of oxygen with a very low LOD of <5 ppm O2. The reproducibility of the characteristics for the material was shown.

2.3. Cementitious Materials

Cementitious media are one of the examples where the introduction of optical methods for pH and O2 monitoring allowed measurements to be taken at a qualitatively new level [29][30]. The key benefit of optical methods in comparison with “wet” chemistry traditionally used in this field is that they allow continuous, non-contact measurements at intervals down to seconds. For example, pH change was monitored in calcium aluminate, calcium sulfoaluminate and OPC/slag cements [29]. It turned out that the measured dependence correlated with the heat of hydration. Thus, it is possible to monitor the formation or aging of solid phases in situ in a non-destructive way. Highly reproducible data obtained for hydration and consolidation reactions in highly alkaline cementing systems, for example, pastes, slurries, and hardening materials, opens up new possibilities for experimental investigation.
Optical sensors can help in solving another important problem—the search for leaks from cement structures that perform retaining functions. Such leakages can occur when cracks appear in the cementitious material, and they may have devastating environmental consequences. The use of self-healing concretes has been proposed as a solution. The principle of operation of such materials is based on the addition of bacteria during manufacture, capable of inducing the precipitation of CaCO3, which in turn leads to healing of the resulting cracks even before leakage occurs [30]. The key thing in the development of such materials is the biocompatibility of the host material with the biological object, and the pH of the medium plays a decisive role. This was confirmed practically in the study of biological activity in cement by introducing superabsorbent polymers with encapsulated endospores of Bacillus alkalinitrilicus into cracks. Measurements of the respire activity were made using oxygen optodes. It was shown that lowering of the pH of the cement from >11 to <10 by incorporating fly ash, as well as increasing the hydration time, resulted in a significant increase in bacterial activity.

2.4. Marine Environments

One of the requests in oceanography is the ability to measure and map the content of various substances, primarily associated with physiological processes, such as dissolved O2, CO2 and pH [31]. The sensors are used both for continuous in situ observation using various autonomous platforms, as well as for profiling using underwater vehicles and ships.
Optical methods in the described tasks have a number of advantages. For example, a unified optoelectronic platform can be developed that is customizable for the measurement of a specific analyte by replacing the excitation source and/or light filters [32][33]. This approach significantly reduces development and maintenance costs. Practical tests were performed for the developed platform, which differed in the conditions for performing measurements. They differed in the duration of the measurements: long-term monitoring from 5 days to 8 weeks and profiling (several hours); environmental conditions were also varied, including temperature 9–25 °C and salinity: 6–33 PSS. Both stationary variants and fastening on mobile vehicles have been tested. Practical experiments have clearly shown the critical importance of choosing the right calibration strategy, otherwise systematic distortion of the results can occur, which is unacceptable, especially for long-term measurements. The importance of choosing an anti-biofouling strategy to extend measurements in high biomass environments has also been demonstrated.
Another interesting example of the use of optical sensors is the integration of monitoring systems and vehicles capable of detecting and mapping CO2 leaks from offshore carbon capture and storage (CCS) sites [34]. The evaluation of the effectiveness of such a vault requires the development of a reliable and cost-effective control system since the CO2 storage is considered as one of the main ways at the current stage of the fight against climate change. A method has been demonstrated to create a plume model based on spatial pH distribution data. They were obtained using a remotely controlled underwater vehicle with a spatial orientation system and a fixed optical sensor.
It is important to note that, for the simultaneous measurement of the parameters in the examples given, separate sensors are currently used, which complicates and increases the cost of the design. As shown in the previous section, the combination of different sensor materials in one foil is possible and is achievable only when using composite materials. The main requirement in this case is non-overlapping fluorescence spectra of the indicators used. For example, simultaneous detection of oxygen and carbon dioxide was reported [35]. Simultaneous detection was provided by the combination of well-known PtTFPP as the O2-sensitive dye, phenol red combined with CdSe/ZnS quantum dots as the CO2-sensitive dye, and unmodified CdSe/ZnS QDs as the reference signal. Poly(isobutyl methacrylate) (PolyIBM) was used as the host matrix for the immobilization. All dyes used are capable of absorption in the near UV region, so a 380 nm LED was used for excitation. An increase in the O2 concentration leads to quenching of the fluorescence of the Pt(II) complex, and the appearance of CO2 leads to an increase in the QDs fluorescence intensity. The response and recovery times were 10 s/35 s for O2, and 20 s/60 s for CO2, respectively. The sensitivities of the ratiometric dual sensor were approximately 13 for O2 and 144 for CO2, respectively.

2.5. High Pressure Measurement

The ability to perform measurements under high hydrostatic pressure is one of the clear advantages of optical sensors over electrochemical ones, such as pH electrodes. However, such harsh conditions can significantly change the properties of the analytical system, and an assessment of such an impact on various types of sensory materials has been carried out recently [36]. The reseahrcers immobilized commonly used indicators into porous and nonporous matrix materials to evaluate the influence of the structure on the observed dependencies. A custom chamber was made that allowed cyclic pressure stepwise change up to 200 bar. The reseahrcers  used Pt(II) benzoporphyrin and Ru(II) polypyridyl complexes as indicators. They were immobilized in different microparticles, such as crosslinked polystyrene, poly(phenylsilsesquioxane), silica gels of different porosities, ZIF-8 and UiO-66 MOFs. Composite sensor was obtained by distribution of the microparticles with dyes into gas permeable Hyflon AD and silicone matrices. A comparison was made with homogeneous polystyrene and hydrogel films and nanospheres of non-porous polystyrene directly dispersed in water. All of the listed materials proved to be stable at high hydrostatic pressure, but their response to change was different. Sensors with porous materials showed a decrease in the observed concentration of oxygen, although the actual content did not change. The difference was from −0.02 to −0.45 mg O2 L−1 H2O per 100 bar. Kinetic experiments with high temporal resolution showed that during the first seconds after the change in pressure, spikes were observed due to the redistribution of oxygen between two solid phases of the composite. The changes were fully reversible. The “positive” or “negative” direction of the spike in the dependence of the observed concentration depends on which of the materials is more ductile—the porous particles with the dye or the host polymer.

2.6. Plant Roots System

An aeration system is necessary for root growth and plant survival, which is an important problem in flooded and swampy soils [37]. Special barriers are formed by many wetland species to restrict radial O2 loss (ROL) from roots to the rhizosphere. Such structures significantly enhance the diffusion of O2 in the longitudinal direction, i.e., from basal parts towards the root tip. It also prevents the entry of phytotoxic compounds into the root. However, ROL from roots provides an essential source of oxygen for the oxidation of toxic compounds in rhizosphere [38].
There are commercially available systems with planar optode suitable for the direct measurement of O2, CO2, and pH in rhizosphere of wetland plants [39]. The setup consists of a water aquarium with a fixed plant root and a sensing foil applied to the top. They are isolated from external illumination using a special box. A software-controlled LED is used to excite the luminescence of the indicator and the data is collected by a special camera in the form of images enabling mapping of the spatial distribution of oxygen.

3. Other Analytes

3.1. Uric Acid

Optical detection of uric acid in solutions was demonstrated by means of hybrid nanomaterials consisting of 5,10,15,20-tetrakis(4-amino-phenyl)-porphyrin (TAmPP) doped with copper nanoparticles (CuNPs), platinum nanoparticles (PtNPs), or both types (Pt@CuNPs) [40]. The addition of nanoparticles, as in the case of other luminescent sensors, made it possible to improve the range of measurable concentrations. UV-Vis spectrophotometry was used for detection, and the morphology was analyzed by means of atomic force microscopy (AFM). The most stable response with the highest sensitivity was demonstrated by a composite with porphyrin and PtNPs. The uric acid detection range was found to be 6.20 × 10−6–1.58 × 10−5 M, and the substances present in the human environment did not affect the measurement even in a very high concentration.

3.2. Hypochlorous Acid/Hypochorite

Combination of sodium alginate (SA) and RhB-AC placed into the interpenetrating polymer network (IPN) hydrogels allowed to obtain a biocompatible material for HClO/ClO determination [41]. This material is suitable for use as smart wound dressings or as a drug scaffold with controlled drug release. The in vitro addition of hypochlorous acid resulted in a significant change in the fluorescence of the hydrogel, confirming its ability to detect the analyte. Good biocompatibility of the material was confirmed by cell culture and toxicity tests. Both the hydrogel drug scaffold and the material itself exhibited excellent healing effects when they were used for wound healing in vivo.
Another system for the determination of HClO/ClO was demonstrated using a novel near-infrared-emitting aza-BODIPY derivative [42]. The fluorescent probe with two tellurium atoms at two upper benzyl rings showed fluorescent “turn-on” effect at 738 nm and high selectivity [42]. The photoinduced electron transfer caused by the presence of two tellurium atoms leads to significant quenching of the fluorescence of this probe. However, the exposure to HClO/ClO resulted in the oxidation of both electron-rich tellurium atoms, and a strong fluorescence emission was observed at 738 nm with the quantum yield of 0.11. Calibration plot was linear in HClO/ClO-concentration range of 0–30 μM; LOD was 0.09 μM in acetonitrile aqueous solution. Ability to work over a wide pH range (from 2 to 10) should also be mentioned. Applicability for endogenous and exogenous detection and imaging of HClO/ClO in living cells was confirmed by the experiments in RAW264.7 cells.

3.3. Ammonia

The possibility of performing a simultaneous measurement of gaseous O2 and NH3 was demonstrated using a sensor in the form of cellulose fibers impregnated on one side with the PtTFPP dye, and on the other with eosin-Y [43]. Both indicators were initially distributed in mesoporous sol-gel matrix, which was further applied to the support fibers. The concentration of each gas is determined by the quenching of the fluorescence intensity of the corresponding dye. Each gas, ideally, only affects one of the indicators. However, the synchronized measurement of several analytes using the same sensor material often suffers from cross-sensitivity effects. In the case of the created sensor, it turned out that the fluorescent peak associated with the O2 measurement is also quenched by NH3 and vice versa. The reseahrcers  proposed a new analysis method to eliminate such interference. They systematically studied the mutual influence of analytes on the recorded signal and proposed an improved algorithm for calculating the true concentration, taking into account the observed cross-sensitivity. The proposed version of the data processing made it possible to reduce the error of oxygen determination from −11.4% ± 34.3% to 2.0% ± 10.2% in a complex media.

3.4. 1-Anthraquinonsulfonic Acid

1-anthraquinonsulfonic acid (AQ) is an important marker in medical investigations of congestive and cancer diseases, pancreatic fibrosis, microbial tests and in diabetes. Possibility of measurement was demonstrated by means of the porphyrin-based composite material [44]. The reseahrcers synthesized new 5,15-bis-(3-hydroxyphenyl)-10,20-bis(3-methoxyphenyl)-porphyrin (trans-A2B2-porphyrin) and further received its Pt(II) derivative, namely Pt-trans-A2B2-porphyrin. The indicator molecule was linked to gold nanoparticles (AuNPs) to improve optical properties (see Section 3.2 for an explanation). The obtained material showed the possibility of measuring AQ in the concentration range from 2.419 × 10−8 M to 2.5 × 10−7 M which corresponds to the desired physiological range. An assessment of the crosstalk effect was performed, since the analyzed physiological samples have a complex composition, which may be the source of the interfering factors. It turned out that there is no strong influence on the AQ measurement process, even at a 50-fold excess of the impurity concentration relative to the target analyte.

3.5. Manganese Ions

The development of multifunctional materials capable of monitoring toxic metal ions in water is a significant task, necessary to maintain a clean and sustainable environment. To control the content of manganese ions, the new carboxyl-substituted A3B porphyrin, 5-(4-carboxy-phenyl)-10,15,20-tris-(4-methyl-phenyl)–porphyrin, was synthesized and a novel composite material was created [45]. The functionality toward Mn2+ detection from polluted waters and from medical samples was evaluated. The plasmonic porphyrin-k-carrageenan-AuNPs material detected Mn2+ in the concentration range from 4.56 × 10−5 M to 9.39 × 10−5 M (5–11 mg/L). Such concentrations may be useful for monitoring the health of people who have been exposed to contaminated water sources or who have consumed large amounts of manganese in their diet.

3.6. Hydrogen Peroxide

Continuous precise detection of hydrogen peroxide (H2O2) at low concentrations is essential for clinical, pharmaceutical, biological, and environmental analysis. One of the most convenient and adaptable methods for a specific application is optical detection using probes and nanomaterials [46]. An example of such composite material with fast and reversible response has been reported recently [47]. The fibrous silica particles (KCC-1) were used for in situ growth of the platinum nanoparticles (PtNPs) inside their pores at the first stage of the sensor fabrication. The obtained nanocomposite was further embedded into a hydrogel matrix with an oxygen sensing dye PtTFPP. The measurement principle is as follows: hydrogen peroxide is catalytically converted into molecular oxygen by immobilized PtNPs, the latter being determined by the optical sensor based on phosphorescence quenching. The sensor demonstrates a fast response (less than a minute), which is completely reversible, due to the use of a highly porous KCC-1 structure, that imparts good permeability and at the same time allows creating a high local concentration of nanoparticles. Thus, the advantage of composite structures in optical sensors was practically demonstrated. The measurement could be done in a wide range from 1.0 µM to 10.0 mM, while the sensor requires only 200 µL of sample for analysis. Thus, the presented sensor, due to the described advantages, can meet all industry requirements for real-time measurement and fill a vacancy in the market.

4. Biomolecular Applications

Optical sensors allow continuous monitoring of various biochemical substances. The resulting data provide insight into physiology and health. However, there are known difficulties in the development of sensors for such measurements, associated with the need to achieve biocompatibility, non-toxicity and compatibility with aqueous media, since indicators are often insoluble in water. For these reasons, silk protein-based biomaterials are particularly useful as a scaffold material for optical sensors. They possess an exceptional amphiphilic chemistry that leads to the stabilization of protein and indicator dye in an aquatic environment. A practical example of the implementation of this approach was demonstrated using an optical oxygen sensor [48]. The reseahrcers used silk films with water-insoluble dye Pd(II)tetramethacrylated benzoporphyrin (PdBMAP) and tested this material in vitro and in vivo. The stabilization of the composite was observed due to the self-assembling of physically cross-linked protein network. A twofold quenching of the lifetime of excited state of the dye was observed at a dissolved oxygen content of ≈31 μM, which suggests the applicability of the material in the physiological range of concentrations. In vitro test has shown cytocompatibility of the material, while in vivo implantation in rats confirmed the biocompatibility and displayed those mechanical properties are suitable for subcutaneous implantation. In addition, the applicability of the sensor material to real-time measurements under various physiological conditions (normoxia, hyperoxia and hypoxia) was shown.
Another approach to the development of a targeted measurement system is the chemical modification of an indicator molecule in order to give it the ability to bind to a specific target or to penetrate the lipid membrane in a biological sample under study. For example, meso-substituted BODIPY with the butanoic acid residue was covalently binded to the thioterpene moiety was performed to examine the membranotropic effect to erythrocytes and to evaluate the practical application of the conjugate in bioimaging [49]. The obtained dye exhibited high fluorescence quantum yield at 514–519 nm and high photostability. Moreover, the absence of erythrotoxicity was confirmed despite the fact that the indicator effectively penetrates the membrane of erythrocyte. Thus, it can be concluded that conjugation of the hydrophobic dyes with a thiotherpenoid is the right way to impart the affinity to biostructures, e.g., blood components.
Biocatalytic processes represent another field where optical sensors are indispensable and can provide unique information. O2-dependent biocatalytic reactions could be mentioned an example. If the process takes place in a bulk liquid, then the supply of the required amount of oxygen can be simply organized by aeration with air. However, customized O2 supply solutions are required when biocatalysis occurs in a spatially limited microstructure of the solid support. Release of O2 through controlled decomposition of hydrogen peroxide is considered as one of the most promising options, however this requires a means for continuous spatiotemporal measurement. An example of such a system, which uses optical sensing of soluble O2 formed from H2O2 in a porous carrier by immobilized catalase, was shown recently [50]. The obtained O2 was consumed by the oxidation reaction of D-methionine that was co-immobilized in the same carrier with the catalase. The optical sensor made it possible to determine that the reaction rate exhibits linear dependence on the internal O2 concentration up to the level of the air saturated solution. In addition, it was practically shown that such an oxygen generation system makes it possible to achieve a 1.5-fold acceleration of the reaction compared to air aeration. Taken together, these results show how the unique spatial measurement capabilities provided by optical sensors can be used to develop a method for the controlled delivery of O2 from H2O2 to enzymes immobilized in a carrier. Such an integrated strategy will be especially useful in biomolecular engineering to overcome the limitations of O2 supply through gas-liquid transfer.

5. Organ-On-Chip, Lab-On-Chip and Microfluidics

Compact organ-on-chip systems represent encouraging methods for in vitro research in biology, medicine and pharmaceuticals. Their main advantage over conventional cell culture platforms is the improved resemblance of the culture environment [51]. The ability to control both the biological responses of the cultured cells and the cell culture conditions is the most important aspect of these systems. On-chip integration of in situ analysis methods provides improved temporal resolution, faster readouts and continuous measurements of such properties as the metabolic activity, the release of particular molecules, rate of proliferation and differentiation [52]. Therefore, the developed models of organs and diseases could provide more information. Many sensor types have been integrated into lab-on-a-chip systems for chemical analysis, including electrical, electrochemical and optical sensors. Recently, some of these measurements and monitoring principles have also been applied to organ-on-a-chip systems.
Successful integration of a sensor into a microfluidic system requires compatibility with the fabrication process. At the same time, in the case of optical sensors, it is necessary to take into account such specific features as ensuring long-term stability despite the sensor degradation, biological fouling, and peculiarities of manufacturing processes [53]. For example, a manufacturing technology for an optical sensor of dissolved oxygen based on platinum octaethylporphyrin (PtOEP) immobilized in a polydimethylsiloxane (PDMS) membrane was presented [54]. The choice of matrix was justified by the fact that microfluidic devices are usually made from PDMS, and therefore the reseahrcers investigated the in-detail features of the fabrication process using standard microfluidic materials and technologies. The outstanding chemical and mechanical properties of PDMS, such as anti-biofouling characteristics and high oxygen permeability, made it possible to achieve sensors with better sensitivity compared to other matrix materials. The achieved detection range was from 0.5% up to 20% in a gas media, and from 0.5 mg/L up to 3.3 mg/L in liquid media at 1 atm, 25 °C.
In addition to optical fluorescent sensors, chemiluminescent (CL) systems also take an important place in microfluidic systems. For example, chemiluminescence (CL) of luminol catalyzed by AuNPs could represent an easily implemented alternative to enzyme-based methods. The aggregation of NPs leads to significantly enhanced CL signals, therefore substances disturbing such a process will lead to weakening of the signal that could be easily detected. An example of the use of such a concept was demonstrated in the detection of sulfadimethoxine (SDM) using the CL microfluidic flow-injection platform with homogeneous aptamer-based assay measurement [52]. The most important for the successful implementation of the method is the efficient mixing of the components, i.e., AuNPs, aptamers, and analyte. Two-dimensional (2D) and three-dimensional (3D) mixer designs were examined. It turned out that the first one could not provide sufficient mixing and a laminated 3D 5-layer microfluidic mixer was developed. The device has been optimized to not only improve the quality of mixing but also to ensure the image acquisition by a CCD camera. The comparison of luminol with its derivative m-carboxy luminol, which is more hydrophilic, showed that the latter exhibits a tenfold increase in the signal and more reliable readouts. The system demonstrates an extraordinary range of detectable concentrations of SDM: 0.01–1000 ng/mL (5 orders of magnitude), and LOD of 4 pg/mL. These impressive results show not only the superiority of the method in determining SDM but also that the underlying method can be applied to other analytical tasks in the food industry and in environmental control.
Lab-on-chip systems could also be used in the analysis of environmental objects. For example, they are capable to perform simultaneous measurements of water chemistry parameters with high accuracy [55]. At the same time, they can be used even in extremal conditions, for example, at depths down to 6000 m. The latter system is produced under license by Clearwater Sensors Ltd. (Southampton, UK) and utilizes the in situ lab-on-chip technology from the University of Southampton and the National Oceanography Centre (NOC). More than 200 successful cases are known, including analysis at a depth of about 4800 m, in muddy estuaries and rivers, as well as for up to a year in seasonally ice-covered regions of the Arctic. A set of methods is used combining optics with microfluidics and electromechanical valves and pumps which allowed to mix water samples with reagents and detect the response. Chemical and biological parameters could be monitored simultaneously, including Nitrate, Nitrite, Phosphate, Silicate, Iron, and pH.

6. Imaging

One of the unique features provided by optical sensors is the ability to obtain the spatial distribution of the analyte concentration, i.e., mapping. Such information makes it possible to establish, for example, the structure of biofilms, as well as to monitor their metabolic processes in dynamics [56]. All this requires tools with the necessary measurement sensitivity and resolution to detect these changes, which were lacking until recently. An example of the possibilities offered by optical composite sensors are nanosensors that include pH-sensitive fluorophores covalently encapsulated with a reference pH-insensitive dye in an inert matrix of polyacrylamide nanoparticles [57]. They can be used for real time monitoring of three-dimensional changes in pH for biofilm formers. The potential application of nanosensors for detection of sugar metabolism in real time gives a chance to improve oral health through the therapeutic solutions.
Another important area of imaging application is the study of tissue oxygenation, which plays an important role in tumor development and treatment [58]. Optical methods for measuring, for example, oxygen using phosphorescence quenching have been known for a long time, and there are a number of commercially available systems, including those for planar measurements. However, in the case of bioimaging in live systems, the creation of biocompatible phosphorescent complexes is still a difficult task [59]. As an example of a molecule suitable for the described requirements, one can mention meso-tetra(sulfophenyl)tetrabenzoporphyrin Pd(II) (TBP) [60]. In vivo experiments with S37 tumor showed that oxygen level in tumors was lower compared to normal tissues where TBP phosphorescence was completely quenched. Photodynamic therapy in the solid tumor and in the muscle resulted in the increase in TBP phosphorescence lifetimes that confirms oxygen consumption during treatment and probably the blood flow stops.
Spherical microprobes with phosphorescent dyes are well suited for the tissue engineering to monitor the oxygen concentration in close proximity to the cells during their growth phase. A fluorescence microscope could be involved for excitation and image acquisition when the standard seed trays like glass dishes are used [61]. However, the imaging of large biological samples by means of such laser scanning fluorescence lifetime imaging (FLIM) is practically unrealizable due to the small field of view (typically less than 1 mm). However, a system was also shown that allows FLIM to be performed on macroscopic objects up to 18 mm in size, with a lateral resolution of 15 μm [62].
Oxygen mapping in a variety of biological milieu could also be performed by boron nanoparticles (BNPs) that can be fabricated from dye-poly(lactic acid) (PLA) materials [63]. These nanoparticles, on the one hand, have a rigid matrix, and, on the other hand, possess double luminescence: oxygen-sensitive phosphorescence and oxygen-insensitive fluorescence as reference. These spectral properties allow real-time ratiometric determination of oxygen with spatial resolution at micron-level. The probe was tested in different conditions from hypoxic to normoxic and had shown a good response and stability even when increasing K+ concentration was used for the neuronal activity stimulation. Thus, such probes could provide fundamental insights into excitability studies and neural mechanisms.
Another recent example of using the ratiometric approach represents a sensor based on a near-infrared (NIR) oxygen-quenched luminophore Pt(II) octaethylporphine ketone (PtOEPK), and a stable reference dye dioctadecyldicarbocyanine (DiD) [64]. The advantage of this nanosensor system is that it provides an approach to overcome problems, such as optical scattering and autofluorescence in the visible wavelength range observed in biological systems. The encapsulation of dyes in a polymer host matrix makes it possible to keep their ratio constant in biological samples and eliminates the need for complex synthetic paths. When the condition of maintaining a constant ratio of the concentration of fluorophores is met, the analytical signal can be represented as the ratio of the intensities of two signals, taking into account the artifacts of the concentration of the nanosensor in the measurements. It is important that the developed nanosensors are reversible, making it possible to perform measurements in systems where the concentration of dissolved oxygen both increases and decreases. The monitoring of Saccharomyces cerevisiae (brewing yeast) in a 96-well was performed to evaluate the sensor performance. The nanosensors were added directly to the wells and incubation was monitored by the fluorescence plate reader for 60 h. The system allowed to track metabolic activity and dynamic changes that occur due to the change in cell concentration or because of toxic effects. Thus, it was shown that this system can become a platform for high-throughput screening of various microorganism species with an unknown metabolic rate in response to external stimuli (metabolites, antimicrobials, etc.).

7. Conclusions and Prospects

Optical sensing systems have proven to be very powerful in terms of physical scalability, from the tips of optical fibers to a large planar system [253]. However, fluorophore-based sensor films are still limited to a fairly small number of analytes. Thus, synthetics are working hard to expand the range of available sensors. Besides, optical sensors are subject to degradation through photobleaching and fouling. To some extent, these problems can be solved by using replaceable sensing elements, but this only applies to sufficiently large-scale systems, and not to those based on single optical fibers.
The works of recent years show a trend towards the use of various fillers forming composites, which, on the one hand, make it possible to increase the fluorescent response, and hence the measurement accuracy, and, on the other hand, to lower the detection limit [108,233].
Another important achievement of composite materials is the possibility of combine in one material either several cooperating molecules, as in the case of the determination of hydrogen peroxide [234,235,238], or several mutually insoluble analytical systems [206,208]. Moreover, newapproaches to controlwettability and surface biofouling are presented [107,186].
More and more works are published devoted to bioimaging and the study of spatial gradients of various substances during the course of various processes. The measurement principle is based on the determination of the fluorescence lifetime, which is correlated with the phase difference between the excitation light and the fluorescence output. In this case, it is important to take into account the possible nonlinear effects of the fluorescent material, since in this case the accuracy of the results will depend on the modulation frequency of the excitation light [254]. Variation in the modulation frequency showed that it mainly affects the sensitivity and phase resolution ratio of the system. A good degree of fitting with R2 value of 0.9981 and a small relative error of 0.79% was observed for the system with the optimal modulation frequency.
It is important to note that, while monitoring of individual substances using optical sensors has proven to be successful; it remains a challenging task to monitor the concentrations of multiple analytes at the same time, especially in mapping and imaging. The main problem is related to the superposition of the emission spectra of dyes, which causes a crosstalk effect of the intensities of the color channels in the recording equipment (RGB or RGB-NIR cameras). However, an original solution has recently been presented that can “free the hands” of experimenters and significantly expand the possibilities for synchronous measurement of concentrations of interest.
Recently, a new approach has been proposed that involves hyperspectral imaging combined with signal deconvolution of overlapping emission spectra of several fluorescent indicator dyes [255]. This method greatly simplifies practical measurements, since reduces the requirements for the absence of overlap in the emission spectra of the dyes used. The deconvolution algorithm to decode the superimposed sensor signals uses a linear combination model. It runs a sequential least-squares fit to determine the contribution of the individual indicator dyes to the total measured signal. As a proof of concept, the algorithm was used to analyze the measured response of an O2 sensor composed of red-emitting Pd(II)/Pt(II) porphyrins and NIR-emitting Pd(II)/Pt(II) benzoporphyrins with different sensitivities. The results showed that such a combination allowed imaging of O2 over a wide dynamic range (0–950 hPa) with a hyperspectral camera system (470–900 nm). The roots of the aquatic plant Littorella uniflora were used to demonstrate the advantage of the novel method by imaging the O2 distribution in the heterogeneous microenvironment. The reported approach of combining hyperspectral sensing with signal deconvolution is flexible and can easily be adapted for the use of various multi-indicator- or evenmultianalyte-based optical sensors with different spectral characteristics, enabling high-resolution simultaneous imaging of multiple analytes.
The combination of described new instrumental approach, and the possibilities achieved by composite polymer-based optical sensors, in a opinion, can significantly enrich the knowledge about the surrounding world in the near future.


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