DOI: 10.1002/asia.201500444
Focus Review
Biosensors
Boronic Acid-Based Carbohydrate Sensing Wenlei Zhai,[a] Xiaolong Sun,[b] Tony D. James,[b] and John S. Fossey*[a]
Chem. Asian J. 2015, 10, 1836 – 1848
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Focus Review Abstract: The covalent boron–diol interaction enables elaborate design of boronic acid-based saccharide sensors. Over the last decade, this research topic has been well developed thanks to the integration of boronic acid chemistry with a range of techniques, including supramolecular chemistry, materials chemistry, surface modification, and nanotechnology. New sensing strategies and platforms have been intro-
duced and remarkable progress has been achieved to fully utilize the unique property of boron–diol interaction and to improve the binding affinity towards different targets, especially under physiological conditions. In this review, the latest progress over the past 30 months (from late 2012 to early 2015) is highlighted and discussed to shed light on this versatile and promising platform for saccharide sensing.
1. Introduction Carbohydrates (saccharides) extensively participate in many essential life activities such as metabolism and cell recognition. According to the number of the saccharide units, they are divided into different saccharides such as mono-, di-, trisaccharides, medium-sized oligosaccharides and large polysaccharides. Each plays different roles in maintaining the basic activities of all living organisms. For example, glucose is one of the most well-known monosaccharides mainly because it serves as the primary form of energy for tissues and cells. Due to the huge potential benefits of accurate detection of carbohydrate, enormous effort has been invested in the development of new techniques for this task. Up until now, there are several available techniques for carbohydrate detection.[1] For example, enzymatic-based blood glucose meters have prevailed for decades. However, issues that need to be addressed to reduce suffering and improve the experience of patients remain. On the other hand, some of the newly developed approaches have already showed favorable features for applications in various areas. Among them, boronic acid-based sensors have attracted lots of attention in the past two decades. It has gradually become an active and important research topic in the search for effective saccharide sensors. Generally, most boronic acid sensors are developed on the basis of the reversible, boronic acid-forming, covalent bondforming interactions between boronic acid and diols. The reaction results in the formation of five- or six-membered cyclic esters. The initial study of this unique property dates back more than half a century, when Lorand and Edwards reported different binding constants between phenylboronic acid (PBA) and a variety of diols according to the observation of a pH drop.[2] As presented in Scheme 1, the binding between boronic acid and diol is an equilibrium process of the trigonal planar boronic acid form (Ktrig) and the tetrahedral boronate anion [a] W. Zhai, Dr. J. S. Fossey School of Chemistry University of Birmingham Birmingham, West Midlands, B15 2TT (UK) E-mail: [email protected] [b] X. Sun, Prof. Dr. T. D. James Department of Chemistry University of Bath Bath, BA2 7AY (UK) ORCID(s) from the author(s) for this article is/are available on the WWW under http://dx.doi.org/10.1002/asia.201500444. Chem. Asian J. 2015, 10, 1836 – 1848
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Scheme 1. The relationship between boronic acid and its diol ester.
form (Ktet). Therefore, both processes should be considered in the calculation of the general binding constant between boronic acid and diol. It is worth noting that the boronate anion in general has stronger binding strength towards diol. Therefore, higher pH is favorable in sugar binding as most of the boron-containing species will then exist in their anion form. For practical application, it is required that the sensing system should perform well under physiological conditions (pH 7.4). As a result, a great deal of effort has been made to make new boronic acid sensors with utility at lower pKa. As an active research area, many new sensory systems based on different strategies have been proposed and demonstrated. Most of the basic principles and major breakthroughs are well covered by several previous reviews.[3] Herein, we focus on recent progress of boronic acid-based saccharide sensors. According to the different sensory systems and employed techniques, they are divided into five major streams: i) boronic acid-appended molecular sensors; ii) boronic acid-appended polymers; iii) boronic acid-functionalized surface; iv) boronic acid-modified nanosensors; and v) boronic acid-based electrochemical sensors and electrophoresis. For each topic, the outstanding achievements in the past three years are summarized. The major merits of each sensing system are highlighted, and the remaining shortcomings are discussed as well.
2. Boronic Acid-Appended Molecular Sensors Generally, two types of molecular sensors have been designed and developed for saccharide detection. The first is based on the non-covalent interactions between saccharide and the sensor molecules. For example, Davis and co-workers reported 1837
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Focus Review several types of sensors with cavities exquisitely engineered to recognize the target saccharide through synergy between hydrogen bonding and hydrophobic interactions.[4] On the other hand, there are reports of covalent sensing using boronic acidbased saccharide sensing. Due to the unique property of reversibly covalent bond formation with diols in aqueous media, boronic acid can serve as an ideal receptor for saccharide. A classic sensory system was introduced by Shinkai et al., based on photoinduced electron transfer (PET).[5] In Figure 1, compound 1functions as a fluores-
Figure 1. Chemical structure of fructose sensor 1 and glucose sensors 2 and 3 based on N–B interaction.
cent fructose sensor, first reported in 1994. They exploited the interactions between o-methylphenylboronic acid and proximal tertiary amines. This Lewis acid–Lewis base interaction has two advantages. First, it allows the recognition of saccharide to occur at physiological pH, since the interaction between boronic acid and adjacent amine helps to lower the pKa of the neighboring boronic acid. Second, it enables the system to operate as an “off-on” sensor through PET effect. Therefore, this design has been widely used in boronic acid-based saccharide sensor development. Although boronic acid-mediated sugar sensing showed encouraging results in early studies, the lack of selectivity still needs to be addressed by rational design of the molecular structure of the sensor. Mono-boronic acid molecules are found to have higher affinity for fructose rather than glucose under physiological conditions. For the purpose of increasing the binding affinity to glucose, a more selective receptor is required. This was achieved by Shinkai and co-workers through introducing two boronic acid groups to help to recognize an additional pair of diols from glucose molecule.[6] As shown in Figure 1, they synthesized compound 2 with two appropriately spaced boronic acid receptors. The binding strength between bis-boronic acids and glucose, which is measured by the binding constant, is stronger than that to other monosaccharides. Chem. Asian J. 2015, 10, 1836 – 1848
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John S. Fossey is Senior Lecturer at the University of Birmingham, UK. He obtained a MChem degree from Cardiff University of Wales in 2000, and then obtained a PhD, in early 2004, working with Dr C. J. Richards at Queen Mary University of London. After a JSPS postdoctoral position with Professor S. Kobayashi at the University of Tokyo he joined the University of Bath as a temporary faculty member. Next he took up his present post at the University of Birmingham where he was recently promoted to Senior Lecturer. He holds a number of visiting positions including a guest Professor at East China University of Science and Technology (ECUST). He enjoys working at the interface of catalysis and sensing and hopes to develop smart chemical systems in the future for self-reporting synthesis. Tony D. James is a Professor at the University of Bath; in 1986 he obtained a BSc from UEA, in 1991 a PhD from the University of Victoria and from 1991 to 1995 worked as a PDRF in Japan with Professor Seiji Shinkai. He was a Royal Society Research Fellow from 1995 to 2000 at the School of Chemistry in the University of Birmingham, and in September 2000 moved to the Department of Chemistry in the University of Bath. He has held numerous visiting positions including visiting professor at Osaka, Tsukuba and Kyushu Universities and holds a guest Professorship at East China University of Science and Technology (ECUST), Shandong Normal University, Xiamen University, Nanjing University and is a Hai-Tian (Sea-Sky) Scholar at Dalian University of Technology. His research interests include wide ranging aspects of supramolecular chemistry, including molecular recognition, self-assembly and chemosensors, with a particular emphasis on boronic acids as receptors for saccharide sensing. Xiaolong Sun obtained his MSc from East China University of Science and Technology (ECUST) in 2012 under the guidance of Professor Xuhong Qian. He then joined the Chemosensor group in Department of Chemistry, University of Bath, led by Professor Tony D. James, as a PhD student. His wide ranging interests include synthesis of small molecules that can be used to interrogate and utilize biological systems, primarily via fluorescent detection of reactive oxygen and nitrogen species and carbohydrates, that is, monosaccharides. Wenlei Zhai obtained his BSc degree from China Pharmaceutical University and MSc degree from East China University of Science and Technology (ECUST), working with Professor Yi-Tao Long in developing new micro- and nanosensors for pollutants detection. In 2012, he started his PhD study in the research group of Dr J. S. Fossey at the University of Birmingham, where his project involves the use of so-called click chemistry to assemble multifunctional chemosensors.
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Focus Review In further study, they systematically studied the effect of linker length using compounds 4 (n = 3–8), and the result showed that hexamethylene and heptamethylene linkers presented high selectivity to d-glucose, while longer linkers are more favorable for fructose.[7] Inspired by this classic sensing system, many bis-boronic acid molecules have been designed and synthesized for glucose detection. For example, Guo and co-workers recently reported the one-step synthesis of a bisboronic acid sensor (Scheme 2), which showed higher binding
between boronic acid binding, p-stacking, and aggregation lie at the heart of the observed selectivity. Moreover, an enhanced signal could be obtained by “knock-out” of the fructose response by addition of PBA. Since PBA has a higher binding affinity for fructose, it helps to mop up or “knock-out” the effect of fructose in this system. Lee et al. also reported a ratiometric sensing system based on an aggregated peptide containing PBA.[11] The formation of a covalent bond between PBA and sugars resulted in the disassembly of the aggregate into monomer. As a result, a decrease in excimer emission and increase in monomer emission can be observed in aqueous medium. In a recent study Hansen et al. investigated the effect of orthosubstituents in a boronic acid Scheme 2. Synthetic route for a bis-boronic acid. Reagents and conditions: (i) MeOH; (ii) MeOH, NaBH4. Reprosensor with a BODIPY fluoroduced with permission from ref. [8]. Copyright 2015 Royal Society of Chemistry phore.[12] Interestingly, the orthofluorinated molecule showed a strong binding affinity to dglucose and d-fructose in different buffer systems. In contrast, the ortho-methylated analogue displayed a significant decrease in binding affinity to both fructose and glucose, thus reiterating the importance of ortho-substituents in both sugar binding and fluorescence response processes. PBA-based chiral sensors have been developed and the effects of chirality towards saccharide binding have been investigated over the past few years. A study in 2012 carried out by Zhu and James et al. emphasized the effect of different chiral sensors on carbohydrate sensing.[13] Bisboronic acid sensors (R)-5 and Figure 2. Cartoon illustration of highly structured aggregates formed with d-glucose (1:2 complex) and amor(S)-5(Figure 3) were synthesized, phous aggregates formed with d-fructose (1:1 complex). Reproduced with permission from ref. [10]. Copyright and their fluorescence was en2013 American Chemical Society. hanced in the presence of saccharides including d-glucose and affinity to glucose than to other tested monosaccharides.[8] In d-galactose. In the fluorescence study, (R)-5 exhibited a stronanother study, Yu and co-workers developed a BINOL-based ger response and higher binding constants with the studied fluorescent bis-boronic acid sensor for the detection of glusaccharides than (S)-5. This work demonstrated that the chiralicose.[9] ty of the sensor molecules, and the diastereomeric interaction The difficulties related to the higher binding capacity of between analyte and sensor, also has an impact on saccharide mono-boronic acids towards d-fructose versus d-glucose were sensing and it is possible to either enhance or reduce the resolved in work of Jianget al. (Figure 2) in 2013.[10] Enhanced binding with a specific saccharide by changing the chirality of a sensor. selectivity of a boronic acid for glucose was realized by using the preferred 2:1 ratio of boronic acid to glucose binding, Apart from using boronic acids as the sugar receptor, enzyversus the 1:1 ratio observed for fructose binding. A boronic matic approaches have also been employed to achieve the acid bearing a pyrene unit formed 1:1 amorphous conjugates sensitive and selective detection of glucose using a fluorescent with d-fructose. Whilst well ordered, fluorescent 2:1 adducts boronic acid derivative.[14] An elegant example of this sensing were obtained upon addition of d-glucose. Cooperative effects process, a highly fluorescent boronic acid was converted into Chem. Asian J. 2015, 10, 1836 – 1848
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Figure 3. Chemical structure of the chiral bis-boronic acid sensors 5.[13]
a low-fluorescent phenol by hydrogen peroxide, which was generated in the glucose oxidase (GOx)-catalyzed oxidation of glucose. Therefore, the concentration of glucose can be determined by measuring the decrease of the fluorescence intensity in the sensing system. The development of fluorescent boronic acid sensors could also provide valuable information for cancer diagnosis. For example, Cho examined a two-proton turn-on probe (Scheme 3) which can be used to visualize glucose uptake and the changes of intracellular glucose concentration in living cells and tissue using two-photon microscopy.[15] It is likely that progress in fluorescent sensors could be applied to the development of better probes for bio-imaging.
with boronic acid moieties.[17] These studies could help to shed light on the development of cancer cell diagnosis, since the the intracellular concentration of ROS is related to certain types of cancers. Since the first report, a large number of fluorescent boronic acid sensors have been designed and synthesized. Many of them showed good selectivity and sensitivity towards the target sugars. Previous studies focused on the detection of monosaccharides, particularly for glucose, in order to develop new methods for the monitoring of blood glucose levels. Although some complicated and synthetically challenging structures have been designed to achieve good selectivity, more work has been carried out focusing on simplifying the synthetic procedure by employing reactions that require less synthetic effort.[8–10, 15] The attempt to reduce the cost and improve the overall yield is an important step to transfer basic research into real-life applications, and make an impact in the competitive healthcare market. In addition, as we can tell from the reports published in recent years, more effort has been given to make molecular sensors targeting oligosaccharides, glycoproteins, bacteria, and cancer cells based on various sensing strategies.[3c, 15, 17] Benefitting from the knowledge and experience of previous studies, the scope of the analytes is expanding rapidly, and some of the sensors present excellent recognition capacities to their targets under physiological conditions. Since the importance of glycosylation has been highlighted in numerous biological processes, it is reasonable to expect more research interest to be focused on this area in the coming years.
3. Boronic Acid-Appended Polymers
Scheme 3. Synthetic route for a two-photon turn-on probe.
Molecular sensors based on host–guest chemistry have become a hot topic. There are several reports of constructing cyclodextrin–azo host–guest complexes as molecular sensors. For example, Jiang et al. proposed a sensing system using stilbeneboronic acid (STDBA) derivatives, where two STDBA molecules could enter the cavity of g-cyclodextrin (g-CD) to create a fluorescent 2:2 STDBA:g-CD assembly (Figure 4).[16] The system is selective and sensitive towards glucose in aqueous solutions, and was successfully applied to the detection of glucose in artificial samples of urine. It is noteworthy that the potential application of boronic acid-based fluorescent sensors is not limited to the abovementioned areas. There are also studies focused on the detection of reactive oxygen species (ROS) using fluorescent molecules Chem. Asian J. 2015, 10, 1836 – 1848
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Apart from molecular sensors, polymers containing boronic acid moiety have been used for sugar sensing for quite some time. By attaching boronic acid containing functional groups (mainly PBA) to the backbone of polymers, different forms of materials were prepared, such as hydrogels, micelles, self-assembled films, peptides, photonic crystals and so on. The binding affinity to the target sugar units was measured in many different ways, such as the volume change of the hydrogel or the color change through dye displacement. The results provide convincing evidence that boronic acid-modified polymers can serve as saccharide sensors by allowing multivalent interactions with the target sugars. 3.1. New methodologies for polymer functionalization With the development of polymer science, some new approaches have been introduced to make boronic acid-modified polymers. For example, atom transfer radical polymerization (ATRP) is well known as a robust method for facile polymer construction, and accurate control of the reaction can be achieved in this process. By employing ATRP, Ye et al. prepared a material that can be used for the separation of saccharides.[18] They used copper-catalyzed “click reactions” to attach fluorescent boronic acid monomers with propargyl acrylate and then anchored them on the surface of silica particles. The synthe-
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Figure 4. Interaction of the 2:2 STDBA-g-CD complex with glucose and fructose. Reproduced with permission from ref. [16]. Copyright 2012 Royal Society of Chemistry.
sized material showed a fluorescence response to fructose and glucose at physiological pH. The authors furthermore demonstrated the binding capacity of a large glycoprotein. Xu et al. also reported the synthesis of a fluorescent boronic acid-functionalized polymer grafted onto silica particles using “click reactions”.[19] The generated material showed fluorescence enhancements in the presence of fructose and glucose. In another example, Hoogenboom and co-workers reported a micellar sugar sensor using reversible addition–fragmentation chain transfer (RAFT) polymerization of commercially available 4-vinylphenylboronic acid monomer.[20] Dynamic light scattering (DLS) and transmission electron microscopy (TEM) studies proved that the prepared polymer self-assembled upon both pH changes and glucose inputs. They also reported the controlled polymerization of the monomer by nitroxide-mediated polymerization.[21] The obtained material is responsive to glucose between pH 9 and 10, and was examined as a pH sensor by incorporating solvatochromic dyes. Davidson and James et al. also synthesized boronic acid-terminated polylactides by using a boronic acid-functionalized ring opening polymerization initiator.[22] Alizarin Red S (ARS) was used in the dye displacement assay to demonstrate the recognition ability towards diols. 3.2. Hydrogels Boronic acid have been extensively used in the modification of hydrogels. Changes of the gel diameter (either swelling or shrinking) caused by interaction of immobilized boronic acids with sugars can be used for detection and drug release. A detailed study was carried out by Braun et al. in order to investigate the relationship between volumetric responses of the hyChem. Asian J. 2015, 10, 1836 – 1848
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drogels to glucose and different classes of PBAs used for the modification (Figure 5).[23] In this study, various structural factors including different substituents on PBAs and different linkers to the hydrogel backbone, as well as different positions of the appended boronic acids on the phenyl ring were considered and examined. It was found that the magnitude of response depended on the exact chemical structure of the PBA. However, the response rate can be enhanced by introducing electron-withdrawing substituents on PBAs. Also, a methylene linker between the PBA and the hydrogel backbone significantly decreased the response magnitude. Subsequently, a library of PBAs was established so that the most suitable
Figure 5. Concentration-dependent shrinking or swelling of the polymer. Reproduced with permission from ref. [23]. Copyright 2013 American Chemical Society.
structure can be selected based on the volumetric changes to glucose. This systematic study provided broad view for better understanding and designing boronic acid-functionalized hydrogels. In 2013 Zhou et al. presented a fluorescent glucose sensor based on dye-complexed microgels.[24] A gel containing boronic acid and fluorescent dye, Bordeaux R, was described. The Bordeaux R dye interacts with amides in the acrylamide gel and shrinks the microgel. Upon addition of d-glucose to the gel, the polymer swells and this alters the p–p stacking of dye molecules within the gel, thus in turn altering the fluorescence intensity of the dye in the gel. Recently, bioresponsive hydrogels have attracted a lot of attention. They hold potential to be used to make self-regulated carriers for drug delivery. Considering the diol-binding properties of boronic acids, some researches have focused on preparing boronic acid-functionalized polymers which can respond to a change in sugar concentration. In one study by Zhang et al., a microgel was prepared from 2-acrylamidophenylboronic
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Focus Review acid.[25] They proved that the size of the synthesized microgel was reduced after exposure to glucose. The formation of a 1:2 glucose-phenylboronate complex was proposed as the mechanism due to the observed response to glucose. They also reported a glucose-induced swelling of an ultrathin hydrogel film, which was prepared via the layer-by-layer (LbL) assembly of boronic acid-bearing polymer and poly(vinyl alcohol) (PVA).[26] 3.3. Micelles Micelles are considered as an attractive material for the design of reliable and well-controlled drug-release systems. To achieve this goal, the designed micelle must be able to respond to changes in the outside environment (pH, concentration, light, etc). Recently, several studies have focused on the synthesis of PBA-containing block copolymers, which can form self-assembled micelles at physiological pH. Chen et al. developed a self-regulated insulin release system. In this work, three PBA-functionalized block copolymers were synthesized by modifying mPEG-b-PGA with 3-aminophenylboronic acid (APBA).[27] The copolymers can form self-assembled micelles in phosphate buffers at physiological pH. The micelles responded to glucose by increasing the hydrodynamic radii (Rh). Therefore, insulin was loaded into the micelles and the system was tested as a self-regulated insulin release system triggered by glucose. Another study by Yang et al. prepared a phenylborate estercontaining polymer via ATRP. The synthesized polymer can self-assemble into nanoparticles and serve as a glucose-responsive drug carrier at neutral pH.[28]
Moreover, Shi and co-workers also reported another PBAcontaining block copolymer which can self-assemble into micelles in the presence of a glycopolymer.[29]The glucose-sensing strategy is illustrated in Figure 6. Light scattering intensity and dye displacement assays were employed to investigate the response of the resultant micelles towards glucose. The resulting material showed an enhanced response to glucose when compared with the simple PBA copolymer micelles. Certain types of glycoproteins have been found to be highly expressed on the surface of cancer cells. Therefore, it may be possible to develop an intelligent drug-delivery system to target these glycoproteins and reduce the side effects during the treatment of cancers using these strategies. In a recent study carried out by Zhuo and Zhong et al., boronic acid-modified polymers have been employed to achieve such a task.[30] They introduced a PBA-functionalized polymeric micelle system targeting HepG2 cells. The synthesized material, characterized by multiple methods and observed by confocal microscopy, was able to recognize HepG2 cells and promote drug uptake. 3.4. Molecularly imprinted polymers It is worth noting that molecularly imprinted polymers (MIP) have been used as sensors. As a small functional group with specific recognition capabilities, boronic acid units are ideally suited for this technique. Liu et al. established a new approach to prepare boronate affinity-based controllable oriented surface imprinting.[31] A glycoprotein template was covalently anchored onto the surface of a boronic acid-functionalized substrate through binding
Figure 6. Schematic illustration of the formation and glucose-responsive disintegration of PEG-b-P(AA-co-APBA)/P(AA-co-AGA) complex micelles. Reproduced with permission from ref. [29]. Copyright 2012 American Chemical Society. Chem. Asian J. 2015, 10, 1836 – 1848
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Figure 7. Schematic of boronate affinity-based controllable oriented surface imprinting of glycoproteins. Reproduced with permission from ref. [32] Copyright 2013 Royal Society of Chemistry.
with the sugar units. The substrate surface was then deposited with a thickness-controllable imprinting coating generated by an in-water self-copolymerization of dopamine and APBA. Three-dimensional cavities were formed in the imprinting layer upon removal of the glycoprotein template (Figure 7). Further study showed that non-specific binding was prevented. In addition, the binding strength towards the target glycoprotein could be tuned by adjusting the pH. Furthermore, by employing a similar strategy, they established a MIP-based ELISA method by preparing the MIP layer in a 96-well microplate.[32] The analysis of a-fetoprotein demonstrated that the resultant product had several favorable features and could be applied as a useful platform in clinical diagnosis. In another study, the detection of mannose and immunoglobulin M (IgM) was also demonstrated by coating MIP on a quartz crystal microbalance (QCM) electrode.[33] Methacryloylamidophenylboronic acid was used as a monomer and mannose was used as a template. The imprinted film was coated on the QCM electrode surface under UV light using ethylene glycol dimethacrylate as a crosslinking agent and azobisisobutyronitrile (AIBN) as a radical initiator. MIP technology was also used for the preparation of photonic crystal technology with sensitivity towards glucose.[34] 3.5. Modified peptides and other systems Schepartz and co-workers introduced a b-boronopeptide bundle with the unique property of being able to interact with different kinds of monosaccharides simultaneously.[35] As shown in Figure 8, the boronic acid moiety was introduced on the outside surface of a b-peptide through a palladium-catalyzed borylation reaction. Isothermal titration calorimetry (ITC) showed that this modified peptide was able to bind to polyol metabolites such as dopamine and sorbitol in neutral solution. Considering the three-dimensional structure and good compatibility of peptides in physiological environments, it may be possible to develop similar complexes as vehicles for polyol binding. Photonic crystals containing an acrylamide and styrene scaffold were appended with boronic acids for the determination of glucose in tear fluid, which relates closely to blood glucose Chem. Asian J. 2015, 10, 1836 – 1848
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Figure 8. Structure and synthesis of b-boronopeptides EYBK, EBBK, and EBYK. Reproduced with permission from ref. [35] Copyright 2013 American Chemical Society.
levels, in the form of non-invasive contact lenses.[36] The material showed promising potential for future application in noninvasive glucose monitoring, especially for patients suffering from diabetes. A dual glucose and oxygen sensor was developed by Tian and co-workers.[37] A bis-boronic acid fluorescent compound was immobilized in a polyacrylaminde matrix as a glucose probe, while another fluorophore was also employed as an oxygen probe. The prepared sensor was used to monitor glu-
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Focus Review cose and oxygen consumptions by bacteria and mammalian cells in real time. 3.6. Summary of boronic acid-appended polymers Numerous boronic acid-functionalized polymers have been developed in the past few years. Different forms, such as hydrogels, micelles, photonic crystals, and polymer nanoparticles have been designed and synthesized through various strategies, including ATRP, PAFT, “click chemistry”, etc. Some of the studies introduced new methods for the preparation of boronic acid-modified polymers with unique properties, while others provided important information on the ability to understand the relationship between polymer structures and their response towards changes in the outside environment. Accordingly, one may expect more polymer-based materials to be developed which not only function as sensors but also can serve as intelligent drug-delivery systems with good specificity and biocompatibility.
sensing of saccharides was demonstrated using fluorescence microscopy imaging.[38a] Such spheres could find application in the development of sensor arrays. SPR has been often used to measure molecular adsorption in many bio- and lab-on-a-chip sensors. Recently, a d-glucose selective SPR sensor (Figure 10) was designed and synthesized incorporating a glucose-selective bis-boronic acid and a thiolic part, permitting assembly on a surface of gold.[38b] The modified gold substrate was tested for glucose detection with good selectivity.
4. Boronic Acid-Functionalized Surfaces Surface sensing techniques such as surface plasmon resonance (SPR), surface-enhanced Raman spectroscopy (SERS), and QCM have been employed to develop sugar sensors by attaching boronic acid motifs on the sensing surface. Normally, these techniques are sensitive enough that only a small amount of sample is required. On the other hand, the specificity of the sensing system depends on the functional motif immobilized on the surface. Some recent work utilized the advantages of these techniques and combined them with boronic acid-based sensing systems. Sensitive and selective detection of saccharides and glycoproteins were achieved at the solid/liquid interface.[38] Fluorescence is most often used in homogeneous sensor systems but has also been used in heterogeneous systems. For example, a sensor was assembled on a gold-streptavidin surface by using a fluorophore linked to a boronic acid and biotin unit (FLAB). As shown in Figure 9, the design features included a biotin for attachment to streptavidin bound to the surface, a boronic acid part, and a commercially available fluorophore (Alexa-Fluor 647, Invitrogen, maximum emission at 647 nm). A quencher-diol molecule was prepared from a known quencher for Alexa-Fluor 647 (BHQ-3, Biosearch Tech).[39] In further study, the system was combined with polystyrene microspheres to make fluorescent sensors with boronic acid receptors. The
Figure 10. SPR sensor for glucose detection using a bis-boronic acid derivative and TEGT-terminated thiol-modified surface (left); the control without the phenylboronic acid moieties (right). Reproduced with permission from ref. [38b]). Copyright 2013 Royal Society of Chemistry.
SERS is regarded as a powerful analytical technique with single-molecule sensitivity. Boronic acids have also been used for a SERS monosaccharide assay.[40] As shown in Figure 11, 4mercaptophenylboronic acid (4-MPBA) served as a capturing agent and a 4-MPBA-triosmium carbonyl cluster conjugate (OsBA) was also used in a sandwich assay. In other studies, Yu and co-workers modified 4-MPBA on a quasi-3D plasmonic nanostructure array (Q3D-PNA) SERS substrate to achieve the detection of fructose.[41] On the recorded SERS spectra, the area ratio between symmetric and non-totally symmetric ring modes is changed after binding with fructose, which breaks the symmetry of the immobilized 4-MPBA molecule. Moreover, a multifunctional chip was recently developed by He et al. using a silicon wafer decorated with silver nanoparticles and modified with 4-mercaptophenylboronic acid. High efficacy for simultaneously capturing, sensing, and inactivating bacteria was demonstrated with this new platform.[42]
5. Boronic Acid-Modified Nanosensors
Figure 9. Chemical structure of FLAB and BHQ-diol.[39] Chem. Asian J. 2015, 10, 1836 – 1848
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In addition to immobilizing boronic acid derivatives on surfaces, there are some recent studies devoted to developing boronic acid-functionalized nanosensors. This aim is mainly achieved by incorporating boronic acid compounds into the surface of nanomaterials, such as graphene nanosheets, carbon nanotubes, dendrimeric polymers, and gold nanoparticles 1844
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Figure 12. Fluorogenic BA@GO sensors for the selective detection of monosaccharides. Reproduced with permission from ref. [43]. Copyright 2014 American Chemical Society.
Figure 11. SERS sensors for monosaccharides utilising on boronic acids. Selectivity for glucose is achieved by using 2:1 sandwich-like interaction. Reproduced with permission from ref. [40]. Copyright 2013 American Chemical Society.
(AuNPs) through interactions like p–p stacking or thiol–gold interactions. The binding between boronic acids and saccharides can be detected via multiple methods, such as recording the change of fluorescence signal or visualizing the color change of the sensing system. James and He et al. have developed nanomaterials for saccharide sensing by the functionalization of graphene oxide (GO) with fluorescence boronic acid-based probes (Figure 12).[43] The strong fluorescence of boronic acid probes is quenched in the presence of GO via FRET. Subsequently, by adding saccharide (fructose), the BA@GO sensor elicits a “turnon” fluorescence signal. Liu et al. reported the fabrication of dendrimeric boronic acid-functionalized magnetic nanoparticles, which were used for glycoprotein enrichment.[44] Menon et al. reported a new method for the colorimetric detection of glucose using calyx[4]arene/PBA-functionalized AuNPs.[45] The nanosensor was successfully applied to estimate the glucose concentration in human blood serum samples. Another glucose sensor was developed based on the functionalization of carbon nanotube transistors with pyrene-1-boronic acid.[46] The formation of boronate anions after binding with saccharides modulated the electronic transport properties of the carbon nanotube transistors, which was found to be closely related to the concentration of glucose over a certain range. The device showed better sensitivity than commercially available blood glucose meters. In another study, boronopyridinium amphiphile species were employed to create a water-soluble vesicle.[47] A dye displacement assay using ARS suggested that the synthesized vesicle has a higher affinity to fructose than to glucose. Chem. Asian J. 2015, 10, 1836 – 1848
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Compared to classic molecular sensors, boronic acid hybrid nanosensors are regarded as a relatively new research area. As can be seen from recently published works, these sensing systems are generated using simple and straightforward strategies, which require less synthetic effort and provide alternative methods for saccharide detection. With the rapid development of nanotechnology, it is envisioned that boronic acidbased nanosensors will continue to draw more attention in the future.
6. Boronic Acid-Based Electrochemical Sensors and Electrophoresis Boronic acids are also employed in constructing electrochemical sensors for various targets, including saccharides, glycoproteins, bacteria and so on. Similar to the fluorescent sensors system, electrochemical sensors normally contain an electroactive functional group that serves as a reporter. Therefore, many synthetic methods have been developed to combine boronic acids with electrophores. Among them, ferrocene is probably the most used electro-active group for this purpose. In a recent study, 4-[(ferrocenylamino)methyl]thiophene-3boronic acid (FcTBA) was synthesized and self-assembled on the surface of a gold electrode.[48] Cyclic voltammetry experiments showed that this modified electrode can be used for sensing of cis-diols such as sorbitol. Another sensing strategy was reported based on the displacement of ferrocene boronic acid derivatives, which served as an electrochemical reporter.[49] As illustrated in Figure 13, a gold electrode was first modified with thiolated mannose, and the reporter was attached to the mannose. Upon addition of the analytes, the reporter was displaced and a decrease of the electrochemical signal can be observed. The sensing of saccharide binding proteins and E. coli. was demonstrated in this work. Interestingly, a series of ferrocene–boron–carbohydrate hybrids were synthesized and screened for antibacterial activi-
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Focus Review a specially designed multipath capillary electrophoresis (CE) chip.[55] Lin et al. presented a microelectromechanical systems (MEMS) differential viscometric sensor which effectively reduced environmental disturbances by introducing a reference chamber inside the sensor. The reference chamber is filled with polymer that does not interact with glucose, while the sample chamber contains boronic acid-functionalized polymer. Accurate measurement of glucose was achieved in both in vitro and in vivo studies.[56]
7. Conclusion and Perspective Here we have summarized and discussed some recent progress in the area of boronic acid-based Figure 13. Schematic illustration of an electrochemical displacement assay for carbohydrate-binding protein and saccharide sensors. Some reE. coli. detection. Reproduced with permission from ref. [49]). Copyright 2014 Elsevier. markable achievements over the last three years have been highty.[50] At least one candidate exhibited good activity against the lighted in order to update knowledge of this rapidly developscreened microorganisms, especially E. coli. ing field and inspire others to exploit new sensing strategies. It As abovementioned, a bis-boronic acid compound was is worth noting that the potential application of these sensors modified on a gold surface to construct a SPR sensor (Figis not limited to saccharide detection. Actually, due to the imure 10).[38b] After conjugation of the same compound to a gold portance of saccharides in many biological processes, it is electrode, a sensitive and selective electrochemical sensor was likely that boronic acid-based sensing systems will be applied also developed and studied by cyclic voltammetry (CV) and in multiple analytical tasks in the future, including point-ofelectrochemical impedance spectroscopy (EIS).[51] care diagnostics for diseases,[57] bacteria detection,[42, 49] protein In another work, gold electrodes were functionalized by two modifications,[58] and bioimaging of glycoproteins/cancer approaches. One is the self-assembly of thiolated PBA, while cells[59] . At the same time, some studies are focused on develthe other method is modification of the surface with terminal oping a new generation of functional materials for other chalcarboxylates groups followed by conjugation with APBA. The lenging areas, such as the development of intelligent drug-deresult suggested that both modified electrodes can act as eleclivery systems. Moreover, some interesting studies are focused trochemical sensors for fructose and fluoride anions.[52] on exploiting the reversible covalent binding property of orgaAnother boronic acid-modified electrode for use as sacchanoboron compounds in catalysis.[60] ride sensor was fabricated by electrochemical polymerization In the early stages of studying boronic acid-based sensing, of APBA in an anodic alumina oxide membrane.[53] The eleca great deal of effort has been devoted to synthesizing sensors trode surface was characterized as highly ordered poly(aniline with good selectivity, especially for glucose. Established sensboronic acid) nanotubes, which showed high sensitivity to gluing systems and the knowledge accumulated in the process of cose and fructose. They also achieved similar performance by their development permit the design of more complex and soelectrochemically co-polymerizing phenol and 3-hydroxyphephisticated structures, thus enabling enhanced selectivity tonylboronic acid on a glassy carbon electrode. wards a wider range of important targets. More importantly, Liu et al. reported the study of an affinity capillary electrorapid developments in several areas provide new tools to furphoresis (ACE) method to investigate the interactions between ther improve the sensing performance and expand the scope boronic acids and diol-containing biomolecules.[54] Remarkably, of current strategies. Selectivity issues could be addressed and the method was found to be applicable to a wide range of greater accuracy and sensitivity may be achieved when differboth biomolecules and boronic acids. ent analytical techniques are combined. Cao and Zhang et al. reported a method of moving supraIt is also worth noting that more effort has been given to molecular boundary (MSB) fluorescent focusing for sensitive employing boronic acids in supramolecular sensing systems. detection of monosaccharides and glycoproteins, based on Instead of modulating the fluorescent properties of a single Chem. Asian J. 2015, 10, 1836 – 1848
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Focus Review sensor molecule through intramolecular mechanisms, the supramolecular sensors rely on the switch between different aggregate forms upon addition of the target saccharide, pioneered by Jiang and co-workers.[61] Moreover, some simple to perform synthetic approaches have been introduced to make a libraries of boronic acid derivatives in fewer steps and higher yields. The synthesized compounds can be screened for saccharide binding using high-throughput assays. Therefore, the capacity and efficiency for developing new sensors for various targets should be enhanced. These studies create meaningful impact and carry this research topic forwards towards future clinical use. Although further effort is still ongoing to address some common issues, such as selectivity and biocompatibility, it is reasonable to believe that future development of boronic acid-based saccharide sensors will overcome these problems and play an active role in biological research, clinical diagnosis, and even treatment of diseases.
Acknowledgements WZ, XS, TDJ and JSF thank The Catalysis and Sensing for our Environment (CASE) group for networking opportunities.[62] China Scholarship Council (CSC), the University of Birmingham and the University of Bath are also thanked for providing studentship support (WZ&XS). JSF thanks the University of Birmingham for support, the Royal Society for an Industrial Fellowship and the EPSRC for funding (EP/J003220/1). TDJ thanks the University of Bath for support. JSF and TDJ are grateful for past collaborative support (DT/F00267X/1). Keywords: boronic acid · functionalized polymers · molecular sensors · saccaride sensing · sensors [1] R. Gifford, ChemPhysChem 2013, 14, 2032. [2] J. P. Lorand, J. O. Edwards, J. Org. Chem. 1959, 24, 769. [3] a) J. Fossey, T. James in Reviews in Fluorescence 2007, Vol. 2007, Springer, New York, pp. 103; b) J. S. Fossey, F. D’Hooge, J. M. H. van den Elsen, M. P. P. Morais, S. I. Pascu, S. D. Bull, F. Marken, A. T. A. Jenkins, Y. B. Jiang, T. D. James, Chem. Rec. 2012, 12, 464; c) S. D. Bull, M. G. Davidson, J. M. H. Van den Elsen, J. S. Fossey, A. T. A. Jenkins, Y.-B. Jiang, Y. Kubo, F. Marken, K. Sakurai, J. Zhao, T. D. James, Acc. Chem. Res. 2013, 46, 312; d) Y. Kubo, R. Nishiyabu, T. D. James, Chem. Commun. 2015, 51, 2005; e) H.-C. Wang, A.-R. Lee, J. Food Drug Anal. 2015, 23, 191. [4] C. Ke, H. Destecroix, M. P. Crump, A. P. Davis, Nat. Chem. 2012, 4, 718. [5] T. D. James, K. R. A. S. Sandanayake, S. Shinkai, J. Chem. Soc. Chem. Commun. 1994, 477. [6] T. D. James, K. R. A. S. Sandanayake, R. Iguchi, S. Shinkai, J. Am. Chem. Soc. 1995, 117, 8982. [7] S. Arimori, M. L. Bell, C. S. Oh, K. A. Frimat, T. D. James, J. Chem. Soc. Perkin Trans. 1 2002, 803. [8] H. Chen, L. Li, H. Guo, X. Wang, W. Qin, RSC Adv. 2015, 5, 13805. [9] B. Xu, J. Hou, K. Li, Z. Lu, X. Yu, Chin. J. Chem. 2015, 33, 101. [10] Y.-J. Huang, W.-J. Ouyang, X. Wu, Z. Li, J. S. Fossey, T. D. James, Y.-B. Jiang, J. Am. Chem. Soc. 2013, 135, 1700. [11] L. N. Neupane, S. Y. Han, K.-H. Lee, Chem. Commun. 2014, 50, 5854. [12] J. S. Hansen, M. Ficker, J. F. Petersen, J. B. Christensen, T. Hoeg-Jensen, Tetrahedron Lett. 2013, 54, 1849. [13] Z. Xing, H.-C. Wang, Y. Cheng, C. Zhu, T. D. James, J. Zhao, Eur. J. Org. Chem. 2012, 1223. [14] K. Wannajuk, M. Jamkatoke, T. Tuntulani, B. Tomapatanaget, Tetrahedron 2012, 68, 8899. Chem. Asian J. 2015, 10, 1836 – 1848
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Manuscript received: April 29, 2015 Accepted Article published: July 14, 2015 Final Article published: August 6, 2015
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Focus Review
Biosensors
Boronic Acid-Based Carbohydrate Sensing Wenlei Zhai,[a] Xiaolong Sun,[b] Tony D. James,[b] and John S. Fossey*[a]
Chem. Asian J. 2015, 10, 1836 – 1848
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Focus Review Abstract: The covalent boron–diol interaction enables elaborate design of boronic acid-based saccharide sensors. Over the last decade, this research topic has been well developed thanks to the integration of boronic acid chemistry with a range of techniques, including supramolecular chemistry, materials chemistry, surface modification, and nanotechnology. New sensing strategies and platforms have been intro-
duced and remarkable progress has been achieved to fully utilize the unique property of boron–diol interaction and to improve the binding affinity towards different targets, especially under physiological conditions. In this review, the latest progress over the past 30 months (from late 2012 to early 2015) is highlighted and discussed to shed light on this versatile and promising platform for saccharide sensing.
1. Introduction Carbohydrates (saccharides) extensively participate in many essential life activities such as metabolism and cell recognition. According to the number of the saccharide units, they are divided into different saccharides such as mono-, di-, trisaccharides, medium-sized oligosaccharides and large polysaccharides. Each plays different roles in maintaining the basic activities of all living organisms. For example, glucose is one of the most well-known monosaccharides mainly because it serves as the primary form of energy for tissues and cells. Due to the huge potential benefits of accurate detection of carbohydrate, enormous effort has been invested in the development of new techniques for this task. Up until now, there are several available techniques for carbohydrate detection.[1] For example, enzymatic-based blood glucose meters have prevailed for decades. However, issues that need to be addressed to reduce suffering and improve the experience of patients remain. On the other hand, some of the newly developed approaches have already showed favorable features for applications in various areas. Among them, boronic acid-based sensors have attracted lots of attention in the past two decades. It has gradually become an active and important research topic in the search for effective saccharide sensors. Generally, most boronic acid sensors are developed on the basis of the reversible, boronic acid-forming, covalent bondforming interactions between boronic acid and diols. The reaction results in the formation of five- or six-membered cyclic esters. The initial study of this unique property dates back more than half a century, when Lorand and Edwards reported different binding constants between phenylboronic acid (PBA) and a variety of diols according to the observation of a pH drop.[2] As presented in Scheme 1, the binding between boronic acid and diol is an equilibrium process of the trigonal planar boronic acid form (Ktrig) and the tetrahedral boronate anion [a] W. Zhai, Dr. J. S. Fossey School of Chemistry University of Birmingham Birmingham, West Midlands, B15 2TT (UK) E-mail: [email protected] [b] X. Sun, Prof. Dr. T. D. James Department of Chemistry University of Bath Bath, BA2 7AY (UK) ORCID(s) from the author(s) for this article is/are available on the WWW under http://dx.doi.org/10.1002/asia.201500444. Chem. Asian J. 2015, 10, 1836 – 1848
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Scheme 1. The relationship between boronic acid and its diol ester.
form (Ktet). Therefore, both processes should be considered in the calculation of the general binding constant between boronic acid and diol. It is worth noting that the boronate anion in general has stronger binding strength towards diol. Therefore, higher pH is favorable in sugar binding as most of the boron-containing species will then exist in their anion form. For practical application, it is required that the sensing system should perform well under physiological conditions (pH 7.4). As a result, a great deal of effort has been made to make new boronic acid sensors with utility at lower pKa. As an active research area, many new sensory systems based on different strategies have been proposed and demonstrated. Most of the basic principles and major breakthroughs are well covered by several previous reviews.[3] Herein, we focus on recent progress of boronic acid-based saccharide sensors. According to the different sensory systems and employed techniques, they are divided into five major streams: i) boronic acid-appended molecular sensors; ii) boronic acid-appended polymers; iii) boronic acid-functionalized surface; iv) boronic acid-modified nanosensors; and v) boronic acid-based electrochemical sensors and electrophoresis. For each topic, the outstanding achievements in the past three years are summarized. The major merits of each sensing system are highlighted, and the remaining shortcomings are discussed as well.
2. Boronic Acid-Appended Molecular Sensors Generally, two types of molecular sensors have been designed and developed for saccharide detection. The first is based on the non-covalent interactions between saccharide and the sensor molecules. For example, Davis and co-workers reported 1837
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Focus Review several types of sensors with cavities exquisitely engineered to recognize the target saccharide through synergy between hydrogen bonding and hydrophobic interactions.[4] On the other hand, there are reports of covalent sensing using boronic acidbased saccharide sensing. Due to the unique property of reversibly covalent bond formation with diols in aqueous media, boronic acid can serve as an ideal receptor for saccharide. A classic sensory system was introduced by Shinkai et al., based on photoinduced electron transfer (PET).[5] In Figure 1, compound 1functions as a fluores-
Figure 1. Chemical structure of fructose sensor 1 and glucose sensors 2 and 3 based on N–B interaction.
cent fructose sensor, first reported in 1994. They exploited the interactions between o-methylphenylboronic acid and proximal tertiary amines. This Lewis acid–Lewis base interaction has two advantages. First, it allows the recognition of saccharide to occur at physiological pH, since the interaction between boronic acid and adjacent amine helps to lower the pKa of the neighboring boronic acid. Second, it enables the system to operate as an “off-on” sensor through PET effect. Therefore, this design has been widely used in boronic acid-based saccharide sensor development. Although boronic acid-mediated sugar sensing showed encouraging results in early studies, the lack of selectivity still needs to be addressed by rational design of the molecular structure of the sensor. Mono-boronic acid molecules are found to have higher affinity for fructose rather than glucose under physiological conditions. For the purpose of increasing the binding affinity to glucose, a more selective receptor is required. This was achieved by Shinkai and co-workers through introducing two boronic acid groups to help to recognize an additional pair of diols from glucose molecule.[6] As shown in Figure 1, they synthesized compound 2 with two appropriately spaced boronic acid receptors. The binding strength between bis-boronic acids and glucose, which is measured by the binding constant, is stronger than that to other monosaccharides. Chem. Asian J. 2015, 10, 1836 – 1848
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John S. Fossey is Senior Lecturer at the University of Birmingham, UK. He obtained a MChem degree from Cardiff University of Wales in 2000, and then obtained a PhD, in early 2004, working with Dr C. J. Richards at Queen Mary University of London. After a JSPS postdoctoral position with Professor S. Kobayashi at the University of Tokyo he joined the University of Bath as a temporary faculty member. Next he took up his present post at the University of Birmingham where he was recently promoted to Senior Lecturer. He holds a number of visiting positions including a guest Professor at East China University of Science and Technology (ECUST). He enjoys working at the interface of catalysis and sensing and hopes to develop smart chemical systems in the future for self-reporting synthesis. Tony D. James is a Professor at the University of Bath; in 1986 he obtained a BSc from UEA, in 1991 a PhD from the University of Victoria and from 1991 to 1995 worked as a PDRF in Japan with Professor Seiji Shinkai. He was a Royal Society Research Fellow from 1995 to 2000 at the School of Chemistry in the University of Birmingham, and in September 2000 moved to the Department of Chemistry in the University of Bath. He has held numerous visiting positions including visiting professor at Osaka, Tsukuba and Kyushu Universities and holds a guest Professorship at East China University of Science and Technology (ECUST), Shandong Normal University, Xiamen University, Nanjing University and is a Hai-Tian (Sea-Sky) Scholar at Dalian University of Technology. His research interests include wide ranging aspects of supramolecular chemistry, including molecular recognition, self-assembly and chemosensors, with a particular emphasis on boronic acids as receptors for saccharide sensing. Xiaolong Sun obtained his MSc from East China University of Science and Technology (ECUST) in 2012 under the guidance of Professor Xuhong Qian. He then joined the Chemosensor group in Department of Chemistry, University of Bath, led by Professor Tony D. James, as a PhD student. His wide ranging interests include synthesis of small molecules that can be used to interrogate and utilize biological systems, primarily via fluorescent detection of reactive oxygen and nitrogen species and carbohydrates, that is, monosaccharides. Wenlei Zhai obtained his BSc degree from China Pharmaceutical University and MSc degree from East China University of Science and Technology (ECUST), working with Professor Yi-Tao Long in developing new micro- and nanosensors for pollutants detection. In 2012, he started his PhD study in the research group of Dr J. S. Fossey at the University of Birmingham, where his project involves the use of so-called click chemistry to assemble multifunctional chemosensors.
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Focus Review In further study, they systematically studied the effect of linker length using compounds 4 (n = 3–8), and the result showed that hexamethylene and heptamethylene linkers presented high selectivity to d-glucose, while longer linkers are more favorable for fructose.[7] Inspired by this classic sensing system, many bis-boronic acid molecules have been designed and synthesized for glucose detection. For example, Guo and co-workers recently reported the one-step synthesis of a bisboronic acid sensor (Scheme 2), which showed higher binding
between boronic acid binding, p-stacking, and aggregation lie at the heart of the observed selectivity. Moreover, an enhanced signal could be obtained by “knock-out” of the fructose response by addition of PBA. Since PBA has a higher binding affinity for fructose, it helps to mop up or “knock-out” the effect of fructose in this system. Lee et al. also reported a ratiometric sensing system based on an aggregated peptide containing PBA.[11] The formation of a covalent bond between PBA and sugars resulted in the disassembly of the aggregate into monomer. As a result, a decrease in excimer emission and increase in monomer emission can be observed in aqueous medium. In a recent study Hansen et al. investigated the effect of orthosubstituents in a boronic acid Scheme 2. Synthetic route for a bis-boronic acid. Reagents and conditions: (i) MeOH; (ii) MeOH, NaBH4. Reprosensor with a BODIPY fluoroduced with permission from ref. [8]. Copyright 2015 Royal Society of Chemistry phore.[12] Interestingly, the orthofluorinated molecule showed a strong binding affinity to dglucose and d-fructose in different buffer systems. In contrast, the ortho-methylated analogue displayed a significant decrease in binding affinity to both fructose and glucose, thus reiterating the importance of ortho-substituents in both sugar binding and fluorescence response processes. PBA-based chiral sensors have been developed and the effects of chirality towards saccharide binding have been investigated over the past few years. A study in 2012 carried out by Zhu and James et al. emphasized the effect of different chiral sensors on carbohydrate sensing.[13] Bisboronic acid sensors (R)-5 and Figure 2. Cartoon illustration of highly structured aggregates formed with d-glucose (1:2 complex) and amor(S)-5(Figure 3) were synthesized, phous aggregates formed with d-fructose (1:1 complex). Reproduced with permission from ref. [10]. Copyright and their fluorescence was en2013 American Chemical Society. hanced in the presence of saccharides including d-glucose and affinity to glucose than to other tested monosaccharides.[8] In d-galactose. In the fluorescence study, (R)-5 exhibited a stronanother study, Yu and co-workers developed a BINOL-based ger response and higher binding constants with the studied fluorescent bis-boronic acid sensor for the detection of glusaccharides than (S)-5. This work demonstrated that the chiralicose.[9] ty of the sensor molecules, and the diastereomeric interaction The difficulties related to the higher binding capacity of between analyte and sensor, also has an impact on saccharide mono-boronic acids towards d-fructose versus d-glucose were sensing and it is possible to either enhance or reduce the resolved in work of Jianget al. (Figure 2) in 2013.[10] Enhanced binding with a specific saccharide by changing the chirality of a sensor. selectivity of a boronic acid for glucose was realized by using the preferred 2:1 ratio of boronic acid to glucose binding, Apart from using boronic acids as the sugar receptor, enzyversus the 1:1 ratio observed for fructose binding. A boronic matic approaches have also been employed to achieve the acid bearing a pyrene unit formed 1:1 amorphous conjugates sensitive and selective detection of glucose using a fluorescent with d-fructose. Whilst well ordered, fluorescent 2:1 adducts boronic acid derivative.[14] An elegant example of this sensing were obtained upon addition of d-glucose. Cooperative effects process, a highly fluorescent boronic acid was converted into Chem. Asian J. 2015, 10, 1836 – 1848
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Figure 3. Chemical structure of the chiral bis-boronic acid sensors 5.[13]
a low-fluorescent phenol by hydrogen peroxide, which was generated in the glucose oxidase (GOx)-catalyzed oxidation of glucose. Therefore, the concentration of glucose can be determined by measuring the decrease of the fluorescence intensity in the sensing system. The development of fluorescent boronic acid sensors could also provide valuable information for cancer diagnosis. For example, Cho examined a two-proton turn-on probe (Scheme 3) which can be used to visualize glucose uptake and the changes of intracellular glucose concentration in living cells and tissue using two-photon microscopy.[15] It is likely that progress in fluorescent sensors could be applied to the development of better probes for bio-imaging.
with boronic acid moieties.[17] These studies could help to shed light on the development of cancer cell diagnosis, since the the intracellular concentration of ROS is related to certain types of cancers. Since the first report, a large number of fluorescent boronic acid sensors have been designed and synthesized. Many of them showed good selectivity and sensitivity towards the target sugars. Previous studies focused on the detection of monosaccharides, particularly for glucose, in order to develop new methods for the monitoring of blood glucose levels. Although some complicated and synthetically challenging structures have been designed to achieve good selectivity, more work has been carried out focusing on simplifying the synthetic procedure by employing reactions that require less synthetic effort.[8–10, 15] The attempt to reduce the cost and improve the overall yield is an important step to transfer basic research into real-life applications, and make an impact in the competitive healthcare market. In addition, as we can tell from the reports published in recent years, more effort has been given to make molecular sensors targeting oligosaccharides, glycoproteins, bacteria, and cancer cells based on various sensing strategies.[3c, 15, 17] Benefitting from the knowledge and experience of previous studies, the scope of the analytes is expanding rapidly, and some of the sensors present excellent recognition capacities to their targets under physiological conditions. Since the importance of glycosylation has been highlighted in numerous biological processes, it is reasonable to expect more research interest to be focused on this area in the coming years.
3. Boronic Acid-Appended Polymers
Scheme 3. Synthetic route for a two-photon turn-on probe.
Molecular sensors based on host–guest chemistry have become a hot topic. There are several reports of constructing cyclodextrin–azo host–guest complexes as molecular sensors. For example, Jiang et al. proposed a sensing system using stilbeneboronic acid (STDBA) derivatives, where two STDBA molecules could enter the cavity of g-cyclodextrin (g-CD) to create a fluorescent 2:2 STDBA:g-CD assembly (Figure 4).[16] The system is selective and sensitive towards glucose in aqueous solutions, and was successfully applied to the detection of glucose in artificial samples of urine. It is noteworthy that the potential application of boronic acid-based fluorescent sensors is not limited to the abovementioned areas. There are also studies focused on the detection of reactive oxygen species (ROS) using fluorescent molecules Chem. Asian J. 2015, 10, 1836 – 1848
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Apart from molecular sensors, polymers containing boronic acid moiety have been used for sugar sensing for quite some time. By attaching boronic acid containing functional groups (mainly PBA) to the backbone of polymers, different forms of materials were prepared, such as hydrogels, micelles, self-assembled films, peptides, photonic crystals and so on. The binding affinity to the target sugar units was measured in many different ways, such as the volume change of the hydrogel or the color change through dye displacement. The results provide convincing evidence that boronic acid-modified polymers can serve as saccharide sensors by allowing multivalent interactions with the target sugars. 3.1. New methodologies for polymer functionalization With the development of polymer science, some new approaches have been introduced to make boronic acid-modified polymers. For example, atom transfer radical polymerization (ATRP) is well known as a robust method for facile polymer construction, and accurate control of the reaction can be achieved in this process. By employing ATRP, Ye et al. prepared a material that can be used for the separation of saccharides.[18] They used copper-catalyzed “click reactions” to attach fluorescent boronic acid monomers with propargyl acrylate and then anchored them on the surface of silica particles. The synthe-
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Figure 4. Interaction of the 2:2 STDBA-g-CD complex with glucose and fructose. Reproduced with permission from ref. [16]. Copyright 2012 Royal Society of Chemistry.
sized material showed a fluorescence response to fructose and glucose at physiological pH. The authors furthermore demonstrated the binding capacity of a large glycoprotein. Xu et al. also reported the synthesis of a fluorescent boronic acid-functionalized polymer grafted onto silica particles using “click reactions”.[19] The generated material showed fluorescence enhancements in the presence of fructose and glucose. In another example, Hoogenboom and co-workers reported a micellar sugar sensor using reversible addition–fragmentation chain transfer (RAFT) polymerization of commercially available 4-vinylphenylboronic acid monomer.[20] Dynamic light scattering (DLS) and transmission electron microscopy (TEM) studies proved that the prepared polymer self-assembled upon both pH changes and glucose inputs. They also reported the controlled polymerization of the monomer by nitroxide-mediated polymerization.[21] The obtained material is responsive to glucose between pH 9 and 10, and was examined as a pH sensor by incorporating solvatochromic dyes. Davidson and James et al. also synthesized boronic acid-terminated polylactides by using a boronic acid-functionalized ring opening polymerization initiator.[22] Alizarin Red S (ARS) was used in the dye displacement assay to demonstrate the recognition ability towards diols. 3.2. Hydrogels Boronic acid have been extensively used in the modification of hydrogels. Changes of the gel diameter (either swelling or shrinking) caused by interaction of immobilized boronic acids with sugars can be used for detection and drug release. A detailed study was carried out by Braun et al. in order to investigate the relationship between volumetric responses of the hyChem. Asian J. 2015, 10, 1836 – 1848
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drogels to glucose and different classes of PBAs used for the modification (Figure 5).[23] In this study, various structural factors including different substituents on PBAs and different linkers to the hydrogel backbone, as well as different positions of the appended boronic acids on the phenyl ring were considered and examined. It was found that the magnitude of response depended on the exact chemical structure of the PBA. However, the response rate can be enhanced by introducing electron-withdrawing substituents on PBAs. Also, a methylene linker between the PBA and the hydrogel backbone significantly decreased the response magnitude. Subsequently, a library of PBAs was established so that the most suitable
Figure 5. Concentration-dependent shrinking or swelling of the polymer. Reproduced with permission from ref. [23]. Copyright 2013 American Chemical Society.
structure can be selected based on the volumetric changes to glucose. This systematic study provided broad view for better understanding and designing boronic acid-functionalized hydrogels. In 2013 Zhou et al. presented a fluorescent glucose sensor based on dye-complexed microgels.[24] A gel containing boronic acid and fluorescent dye, Bordeaux R, was described. The Bordeaux R dye interacts with amides in the acrylamide gel and shrinks the microgel. Upon addition of d-glucose to the gel, the polymer swells and this alters the p–p stacking of dye molecules within the gel, thus in turn altering the fluorescence intensity of the dye in the gel. Recently, bioresponsive hydrogels have attracted a lot of attention. They hold potential to be used to make self-regulated carriers for drug delivery. Considering the diol-binding properties of boronic acids, some researches have focused on preparing boronic acid-functionalized polymers which can respond to a change in sugar concentration. In one study by Zhang et al., a microgel was prepared from 2-acrylamidophenylboronic
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Focus Review acid.[25] They proved that the size of the synthesized microgel was reduced after exposure to glucose. The formation of a 1:2 glucose-phenylboronate complex was proposed as the mechanism due to the observed response to glucose. They also reported a glucose-induced swelling of an ultrathin hydrogel film, which was prepared via the layer-by-layer (LbL) assembly of boronic acid-bearing polymer and poly(vinyl alcohol) (PVA).[26] 3.3. Micelles Micelles are considered as an attractive material for the design of reliable and well-controlled drug-release systems. To achieve this goal, the designed micelle must be able to respond to changes in the outside environment (pH, concentration, light, etc). Recently, several studies have focused on the synthesis of PBA-containing block copolymers, which can form self-assembled micelles at physiological pH. Chen et al. developed a self-regulated insulin release system. In this work, three PBA-functionalized block copolymers were synthesized by modifying mPEG-b-PGA with 3-aminophenylboronic acid (APBA).[27] The copolymers can form self-assembled micelles in phosphate buffers at physiological pH. The micelles responded to glucose by increasing the hydrodynamic radii (Rh). Therefore, insulin was loaded into the micelles and the system was tested as a self-regulated insulin release system triggered by glucose. Another study by Yang et al. prepared a phenylborate estercontaining polymer via ATRP. The synthesized polymer can self-assemble into nanoparticles and serve as a glucose-responsive drug carrier at neutral pH.[28]
Moreover, Shi and co-workers also reported another PBAcontaining block copolymer which can self-assemble into micelles in the presence of a glycopolymer.[29]The glucose-sensing strategy is illustrated in Figure 6. Light scattering intensity and dye displacement assays were employed to investigate the response of the resultant micelles towards glucose. The resulting material showed an enhanced response to glucose when compared with the simple PBA copolymer micelles. Certain types of glycoproteins have been found to be highly expressed on the surface of cancer cells. Therefore, it may be possible to develop an intelligent drug-delivery system to target these glycoproteins and reduce the side effects during the treatment of cancers using these strategies. In a recent study carried out by Zhuo and Zhong et al., boronic acid-modified polymers have been employed to achieve such a task.[30] They introduced a PBA-functionalized polymeric micelle system targeting HepG2 cells. The synthesized material, characterized by multiple methods and observed by confocal microscopy, was able to recognize HepG2 cells and promote drug uptake. 3.4. Molecularly imprinted polymers It is worth noting that molecularly imprinted polymers (MIP) have been used as sensors. As a small functional group with specific recognition capabilities, boronic acid units are ideally suited for this technique. Liu et al. established a new approach to prepare boronate affinity-based controllable oriented surface imprinting.[31] A glycoprotein template was covalently anchored onto the surface of a boronic acid-functionalized substrate through binding
Figure 6. Schematic illustration of the formation and glucose-responsive disintegration of PEG-b-P(AA-co-APBA)/P(AA-co-AGA) complex micelles. Reproduced with permission from ref. [29]. Copyright 2012 American Chemical Society. Chem. Asian J. 2015, 10, 1836 – 1848
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Figure 7. Schematic of boronate affinity-based controllable oriented surface imprinting of glycoproteins. Reproduced with permission from ref. [32] Copyright 2013 Royal Society of Chemistry.
with the sugar units. The substrate surface was then deposited with a thickness-controllable imprinting coating generated by an in-water self-copolymerization of dopamine and APBA. Three-dimensional cavities were formed in the imprinting layer upon removal of the glycoprotein template (Figure 7). Further study showed that non-specific binding was prevented. In addition, the binding strength towards the target glycoprotein could be tuned by adjusting the pH. Furthermore, by employing a similar strategy, they established a MIP-based ELISA method by preparing the MIP layer in a 96-well microplate.[32] The analysis of a-fetoprotein demonstrated that the resultant product had several favorable features and could be applied as a useful platform in clinical diagnosis. In another study, the detection of mannose and immunoglobulin M (IgM) was also demonstrated by coating MIP on a quartz crystal microbalance (QCM) electrode.[33] Methacryloylamidophenylboronic acid was used as a monomer and mannose was used as a template. The imprinted film was coated on the QCM electrode surface under UV light using ethylene glycol dimethacrylate as a crosslinking agent and azobisisobutyronitrile (AIBN) as a radical initiator. MIP technology was also used for the preparation of photonic crystal technology with sensitivity towards glucose.[34] 3.5. Modified peptides and other systems Schepartz and co-workers introduced a b-boronopeptide bundle with the unique property of being able to interact with different kinds of monosaccharides simultaneously.[35] As shown in Figure 8, the boronic acid moiety was introduced on the outside surface of a b-peptide through a palladium-catalyzed borylation reaction. Isothermal titration calorimetry (ITC) showed that this modified peptide was able to bind to polyol metabolites such as dopamine and sorbitol in neutral solution. Considering the three-dimensional structure and good compatibility of peptides in physiological environments, it may be possible to develop similar complexes as vehicles for polyol binding. Photonic crystals containing an acrylamide and styrene scaffold were appended with boronic acids for the determination of glucose in tear fluid, which relates closely to blood glucose Chem. Asian J. 2015, 10, 1836 – 1848
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Figure 8. Structure and synthesis of b-boronopeptides EYBK, EBBK, and EBYK. Reproduced with permission from ref. [35] Copyright 2013 American Chemical Society.
levels, in the form of non-invasive contact lenses.[36] The material showed promising potential for future application in noninvasive glucose monitoring, especially for patients suffering from diabetes. A dual glucose and oxygen sensor was developed by Tian and co-workers.[37] A bis-boronic acid fluorescent compound was immobilized in a polyacrylaminde matrix as a glucose probe, while another fluorophore was also employed as an oxygen probe. The prepared sensor was used to monitor glu-
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Focus Review cose and oxygen consumptions by bacteria and mammalian cells in real time. 3.6. Summary of boronic acid-appended polymers Numerous boronic acid-functionalized polymers have been developed in the past few years. Different forms, such as hydrogels, micelles, photonic crystals, and polymer nanoparticles have been designed and synthesized through various strategies, including ATRP, PAFT, “click chemistry”, etc. Some of the studies introduced new methods for the preparation of boronic acid-modified polymers with unique properties, while others provided important information on the ability to understand the relationship between polymer structures and their response towards changes in the outside environment. Accordingly, one may expect more polymer-based materials to be developed which not only function as sensors but also can serve as intelligent drug-delivery systems with good specificity and biocompatibility.
sensing of saccharides was demonstrated using fluorescence microscopy imaging.[38a] Such spheres could find application in the development of sensor arrays. SPR has been often used to measure molecular adsorption in many bio- and lab-on-a-chip sensors. Recently, a d-glucose selective SPR sensor (Figure 10) was designed and synthesized incorporating a glucose-selective bis-boronic acid and a thiolic part, permitting assembly on a surface of gold.[38b] The modified gold substrate was tested for glucose detection with good selectivity.
4. Boronic Acid-Functionalized Surfaces Surface sensing techniques such as surface plasmon resonance (SPR), surface-enhanced Raman spectroscopy (SERS), and QCM have been employed to develop sugar sensors by attaching boronic acid motifs on the sensing surface. Normally, these techniques are sensitive enough that only a small amount of sample is required. On the other hand, the specificity of the sensing system depends on the functional motif immobilized on the surface. Some recent work utilized the advantages of these techniques and combined them with boronic acid-based sensing systems. Sensitive and selective detection of saccharides and glycoproteins were achieved at the solid/liquid interface.[38] Fluorescence is most often used in homogeneous sensor systems but has also been used in heterogeneous systems. For example, a sensor was assembled on a gold-streptavidin surface by using a fluorophore linked to a boronic acid and biotin unit (FLAB). As shown in Figure 9, the design features included a biotin for attachment to streptavidin bound to the surface, a boronic acid part, and a commercially available fluorophore (Alexa-Fluor 647, Invitrogen, maximum emission at 647 nm). A quencher-diol molecule was prepared from a known quencher for Alexa-Fluor 647 (BHQ-3, Biosearch Tech).[39] In further study, the system was combined with polystyrene microspheres to make fluorescent sensors with boronic acid receptors. The
Figure 10. SPR sensor for glucose detection using a bis-boronic acid derivative and TEGT-terminated thiol-modified surface (left); the control without the phenylboronic acid moieties (right). Reproduced with permission from ref. [38b]). Copyright 2013 Royal Society of Chemistry.
SERS is regarded as a powerful analytical technique with single-molecule sensitivity. Boronic acids have also been used for a SERS monosaccharide assay.[40] As shown in Figure 11, 4mercaptophenylboronic acid (4-MPBA) served as a capturing agent and a 4-MPBA-triosmium carbonyl cluster conjugate (OsBA) was also used in a sandwich assay. In other studies, Yu and co-workers modified 4-MPBA on a quasi-3D plasmonic nanostructure array (Q3D-PNA) SERS substrate to achieve the detection of fructose.[41] On the recorded SERS spectra, the area ratio between symmetric and non-totally symmetric ring modes is changed after binding with fructose, which breaks the symmetry of the immobilized 4-MPBA molecule. Moreover, a multifunctional chip was recently developed by He et al. using a silicon wafer decorated with silver nanoparticles and modified with 4-mercaptophenylboronic acid. High efficacy for simultaneously capturing, sensing, and inactivating bacteria was demonstrated with this new platform.[42]
5. Boronic Acid-Modified Nanosensors
Figure 9. Chemical structure of FLAB and BHQ-diol.[39] Chem. Asian J. 2015, 10, 1836 – 1848
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In addition to immobilizing boronic acid derivatives on surfaces, there are some recent studies devoted to developing boronic acid-functionalized nanosensors. This aim is mainly achieved by incorporating boronic acid compounds into the surface of nanomaterials, such as graphene nanosheets, carbon nanotubes, dendrimeric polymers, and gold nanoparticles 1844
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Figure 12. Fluorogenic BA@GO sensors for the selective detection of monosaccharides. Reproduced with permission from ref. [43]. Copyright 2014 American Chemical Society.
Figure 11. SERS sensors for monosaccharides utilising on boronic acids. Selectivity for glucose is achieved by using 2:1 sandwich-like interaction. Reproduced with permission from ref. [40]. Copyright 2013 American Chemical Society.
(AuNPs) through interactions like p–p stacking or thiol–gold interactions. The binding between boronic acids and saccharides can be detected via multiple methods, such as recording the change of fluorescence signal or visualizing the color change of the sensing system. James and He et al. have developed nanomaterials for saccharide sensing by the functionalization of graphene oxide (GO) with fluorescence boronic acid-based probes (Figure 12).[43] The strong fluorescence of boronic acid probes is quenched in the presence of GO via FRET. Subsequently, by adding saccharide (fructose), the BA@GO sensor elicits a “turnon” fluorescence signal. Liu et al. reported the fabrication of dendrimeric boronic acid-functionalized magnetic nanoparticles, which were used for glycoprotein enrichment.[44] Menon et al. reported a new method for the colorimetric detection of glucose using calyx[4]arene/PBA-functionalized AuNPs.[45] The nanosensor was successfully applied to estimate the glucose concentration in human blood serum samples. Another glucose sensor was developed based on the functionalization of carbon nanotube transistors with pyrene-1-boronic acid.[46] The formation of boronate anions after binding with saccharides modulated the electronic transport properties of the carbon nanotube transistors, which was found to be closely related to the concentration of glucose over a certain range. The device showed better sensitivity than commercially available blood glucose meters. In another study, boronopyridinium amphiphile species were employed to create a water-soluble vesicle.[47] A dye displacement assay using ARS suggested that the synthesized vesicle has a higher affinity to fructose than to glucose. Chem. Asian J. 2015, 10, 1836 – 1848
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Compared to classic molecular sensors, boronic acid hybrid nanosensors are regarded as a relatively new research area. As can be seen from recently published works, these sensing systems are generated using simple and straightforward strategies, which require less synthetic effort and provide alternative methods for saccharide detection. With the rapid development of nanotechnology, it is envisioned that boronic acidbased nanosensors will continue to draw more attention in the future.
6. Boronic Acid-Based Electrochemical Sensors and Electrophoresis Boronic acids are also employed in constructing electrochemical sensors for various targets, including saccharides, glycoproteins, bacteria and so on. Similar to the fluorescent sensors system, electrochemical sensors normally contain an electroactive functional group that serves as a reporter. Therefore, many synthetic methods have been developed to combine boronic acids with electrophores. Among them, ferrocene is probably the most used electro-active group for this purpose. In a recent study, 4-[(ferrocenylamino)methyl]thiophene-3boronic acid (FcTBA) was synthesized and self-assembled on the surface of a gold electrode.[48] Cyclic voltammetry experiments showed that this modified electrode can be used for sensing of cis-diols such as sorbitol. Another sensing strategy was reported based on the displacement of ferrocene boronic acid derivatives, which served as an electrochemical reporter.[49] As illustrated in Figure 13, a gold electrode was first modified with thiolated mannose, and the reporter was attached to the mannose. Upon addition of the analytes, the reporter was displaced and a decrease of the electrochemical signal can be observed. The sensing of saccharide binding proteins and E. coli. was demonstrated in this work. Interestingly, a series of ferrocene–boron–carbohydrate hybrids were synthesized and screened for antibacterial activi-
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Focus Review a specially designed multipath capillary electrophoresis (CE) chip.[55] Lin et al. presented a microelectromechanical systems (MEMS) differential viscometric sensor which effectively reduced environmental disturbances by introducing a reference chamber inside the sensor. The reference chamber is filled with polymer that does not interact with glucose, while the sample chamber contains boronic acid-functionalized polymer. Accurate measurement of glucose was achieved in both in vitro and in vivo studies.[56]
7. Conclusion and Perspective Here we have summarized and discussed some recent progress in the area of boronic acid-based Figure 13. Schematic illustration of an electrochemical displacement assay for carbohydrate-binding protein and saccharide sensors. Some reE. coli. detection. Reproduced with permission from ref. [49]). Copyright 2014 Elsevier. markable achievements over the last three years have been highty.[50] At least one candidate exhibited good activity against the lighted in order to update knowledge of this rapidly developscreened microorganisms, especially E. coli. ing field and inspire others to exploit new sensing strategies. It As abovementioned, a bis-boronic acid compound was is worth noting that the potential application of these sensors modified on a gold surface to construct a SPR sensor (Figis not limited to saccharide detection. Actually, due to the imure 10).[38b] After conjugation of the same compound to a gold portance of saccharides in many biological processes, it is electrode, a sensitive and selective electrochemical sensor was likely that boronic acid-based sensing systems will be applied also developed and studied by cyclic voltammetry (CV) and in multiple analytical tasks in the future, including point-ofelectrochemical impedance spectroscopy (EIS).[51] care diagnostics for diseases,[57] bacteria detection,[42, 49] protein In another work, gold electrodes were functionalized by two modifications,[58] and bioimaging of glycoproteins/cancer approaches. One is the self-assembly of thiolated PBA, while cells[59] . At the same time, some studies are focused on develthe other method is modification of the surface with terminal oping a new generation of functional materials for other chalcarboxylates groups followed by conjugation with APBA. The lenging areas, such as the development of intelligent drug-deresult suggested that both modified electrodes can act as eleclivery systems. Moreover, some interesting studies are focused trochemical sensors for fructose and fluoride anions.[52] on exploiting the reversible covalent binding property of orgaAnother boronic acid-modified electrode for use as sacchanoboron compounds in catalysis.[60] ride sensor was fabricated by electrochemical polymerization In the early stages of studying boronic acid-based sensing, of APBA in an anodic alumina oxide membrane.[53] The eleca great deal of effort has been devoted to synthesizing sensors trode surface was characterized as highly ordered poly(aniline with good selectivity, especially for glucose. Established sensboronic acid) nanotubes, which showed high sensitivity to gluing systems and the knowledge accumulated in the process of cose and fructose. They also achieved similar performance by their development permit the design of more complex and soelectrochemically co-polymerizing phenol and 3-hydroxyphephisticated structures, thus enabling enhanced selectivity tonylboronic acid on a glassy carbon electrode. wards a wider range of important targets. More importantly, Liu et al. reported the study of an affinity capillary electrorapid developments in several areas provide new tools to furphoresis (ACE) method to investigate the interactions between ther improve the sensing performance and expand the scope boronic acids and diol-containing biomolecules.[54] Remarkably, of current strategies. Selectivity issues could be addressed and the method was found to be applicable to a wide range of greater accuracy and sensitivity may be achieved when differboth biomolecules and boronic acids. ent analytical techniques are combined. Cao and Zhang et al. reported a method of moving supraIt is also worth noting that more effort has been given to molecular boundary (MSB) fluorescent focusing for sensitive employing boronic acids in supramolecular sensing systems. detection of monosaccharides and glycoproteins, based on Instead of modulating the fluorescent properties of a single Chem. Asian J. 2015, 10, 1836 – 1848
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Focus Review sensor molecule through intramolecular mechanisms, the supramolecular sensors rely on the switch between different aggregate forms upon addition of the target saccharide, pioneered by Jiang and co-workers.[61] Moreover, some simple to perform synthetic approaches have been introduced to make a libraries of boronic acid derivatives in fewer steps and higher yields. The synthesized compounds can be screened for saccharide binding using high-throughput assays. Therefore, the capacity and efficiency for developing new sensors for various targets should be enhanced. These studies create meaningful impact and carry this research topic forwards towards future clinical use. Although further effort is still ongoing to address some common issues, such as selectivity and biocompatibility, it is reasonable to believe that future development of boronic acid-based saccharide sensors will overcome these problems and play an active role in biological research, clinical diagnosis, and even treatment of diseases.
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Manuscript received: April 29, 2015 Accepted Article published: July 14, 2015 Final Article published: August 6, 2015
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