The design of boronic acid spectroscopic reporter compounds by taking advantage of the pK(a)-lowering effect of diol binding: nitrophenol-based color reporters for diols

The complex that forms between a boronic acid and a diol is often much more acidic than the starting boronic acid. In conditions where the solution pH is between the two pK(a) values, the boron atom will convert from a neutral trigonal form to an anionic tetrahedral form upon complexation. Such a change is likely to dramatically alter the electron density of neighboring groups. Utilizing this effect, we have designed and synthesized two nitrophenol-based boronic acid reporter compounds that change ionization states and therefore spectroscopic properties upon diol binding. Both compounds show significant UV changes upon addition of saccharides. For example, a blue shift of the absorption max from 373 to 332 nm was observed with the addition of D-fructose to 2-hydroxy-5-nitrophenylboronic acid at neutral pH. Such a reporter compound can be used as a recognition and signaling unit for the construction of polyboronic acid sensors for the selective and specific recognitions of saccharides of biological significance.     

Fluororeactands for the Detection of Saccharides Based on Hemicyanine Dyes with a Boronic Acid Receptor

Three hemicyanine dyes with boronic acid receptor functions have been synthesized in a two step procedure. These dyes are capable of forming a covalent bond between their boronic acid moiety and the diol moiety of saccharides which causes fluorescence to change. In detail, the indicator dyes exhibit absorbance maxima at around 460 nm and emission at around 600 nm, show increases in fluorescence upon exposure to saccharides and can be used in aqueous solution at physiological pH.     

新型含苯硼酸和萘双亲化合物的合成与囊泡荧光传感器的制备

合成了含有识别基团苯硼酸和荧光基团萘的新型对-[(5-十二烷氧基-1-氧基)萘]甲基苯硼酸{p-[(5-dodecyloxy-1-oxy) naphthalene] methyl-phenylboronic acid, DNMPBA}双亲化合物; 该化合物在THF/水选择性溶剂中自组织成囊泡, 囊泡的相变温度为56.8 ℃; 当向囊泡体系加糖时, DNMPBA囊泡中的萘生色基在345 nm的荧光峰强度急剧增强; 荧光强度随添加不同糖的变化趋势为果糖>葡萄糖>麦芽糖>乙二醇. 荧光强度增强可能归因于所形成的硼酸酯减弱了DNMPBA双亲化合物中一个氧原子孤对电子对萘生色基的猝灭作用而使荧光强度重新恢复. DNMPBA囊泡与糖的相互作用导致体系荧光强度变化, 使该体系有可能应用于检测生物物质如糖的化学传感器.     

Potentiometric saccharide detection based on the pK(a) changes of poly(aniline boronic acid)

A novel approach for the potentiometric detection of saccharides using poly(aniline boronic acid) is presented. A model is described in which the electrochemical potential is sensitive to the change in the pK(a) of the conducting polymer as a result of boronic acid-diol complexation. In this system, boronic acid complexation is the mode of transduction and it is manifested as changes in the electrochemical potential of the polymer with remarkable selectivity. Characteristics of both transient and steady-state response associated with the complexation are discussed. The presence of Nafion and fluoride during the electrochemical polymerization of 3-aminophenylboronic acid are shown to impact the sensitivity and the stability of the electrode response. The sensor sensitivity is improved significantly by increasing the concentration of sodium fluoride during the polymerization. Finally, the nature of the selectivity of the boronic acid-diol reaction under these conditions is explored by using molecular orbital calculations.     

Oriented molecular aggregates of porphyrin-based amphiphiles and their morphology control by a boronic acid sugar interaction

Towards the morphology control of oriented porphyrin aggregates by added saccharides, three amphiphilic porphyrins bearing boronic acid groups were synthesised. Among them, an amphiphilic tetraphenylporphyrin (4) bearing two octadecyl groups at 5,10-positions and two boronic acid groups (acting as saccharide-binding sites) at 15,20-positions has been found to act as a membrane-forming amphiphile in an aqueous system. Spectroscopic (UV–Vis and CD), light-scattering, DSC and electron micrographic studies have established that in aqueous media 4 forms stable fibrous aggregates only in the presence of saccharides, which are chirally twisted by the absolute configuration of the added saccharides. This is a novel method to control the aggregate morphology by saccharides and well imitates the morphological functions of certain cell membranes, the surfaces of which are covered by saccharides.     

Binary and ternary phenylboronic acid complexes with saccharides and Lewis bases

Cumulative formation constants for the association of three boronic acids (phenylboronic acid and its ortho-anilinomethyl and ortho-benzylaminomethyl derivatives) with four saccharides (fructose, glucose, mannitol, and sorbitol) were determined by potentiometric titration. Similarly, the constants for the formation binary complexes of the three boronic acids with (hydrogen) phosphate, (hydrogen) citrate, or imidazole were determined. Finally, the formation of ternary complexes of the boronic acids, phosphate, citrate or imidazole, and the saccharides were determined based on the determined values of the binary complexes. The previously unrecognized ternary complexes are significant in all systems investigated, and under some solution compositions, they can be the dominant species in solution over a wide pH range. A value of 15-25 kJ mol⁻¹ was determined for the energy of the B-N interaction in the benzylmethyl derivative based on the relative stabilities of the ternary phosphate complexes of the three boronic acids. The data are used to rationalize the medium dependence of stepwise formation constants and the apparent acidity constants of previous literature reports.     

Detection of adrenaline on poly(3-aminobenzylamine) ultrathin film by electrochemical-surface plasmon resonance spectroscopy

In this Article, we present a novel method to detect adrenaline on poly(3-aminobenzylamine) (PABA) ultrathin films by electrochemical-surface plasmon resonance (EC-SPR) spectroscopy. We prepared a PABA film, which specifically reacts with adrenaline, on a gold electrode by electropolymerization of 3-aminobenzylamine. The specific reaction of benzylamine within the PABA structure with adrenaline was studied by XPS, UV-vis spectroscopy, and EC-SPR techniques. Adrenaline was detected in real time by EC-SPR spectroscopy, which provides simultaneous monitoring of both optical SPR reflectivity and electrochemical current responses upon injecting adrenaline into the PABA thin film. The number of changes in both current and SPR reflectivity on the injection of adrenaline exhibited the linear relation to the concentration, and the detection limit was 100 pM. The responses were distinctive to those for uric acid and ascorbic acid, which are major interferences of adrenaline.     

Multicolor saccharide-sensing chips based on boronic acid-containing thin films showing stepwise release and binding of dyes

A novel saccharide sensor that shows a distinct color change resembling a "traffic signal" was developed. By copolymerizing a boronic acid monomer and an amine monomer on a glass plate, a boronic acid-containing thin film was obtained. After adsorbing anionic blue and yellow dyes, the thin film was immersed in aqueous saccharide solutions containing a cationic red dye. As saccharide concentration increased, the thin film changed its color from green to red via yellow. Origin of the distinct color change was attributed to a stepwise release and binding of dyes.     

Buildup of gold nanoparticle multilayer thin films based on the covalent-bonding interaction between boronic acids and polyols

A novel method for the fabrication of gold nanoparticle multilayer films based on the covalent-bonding interaction between boronic acid and polyols, poly(vinyl alcohol) (PVA), was developed. The multilayer buildup was monitored by UV-vis absorbance spectroscopy, which showed a linear increase of the film absorbance with the number of adsorbed Au layers and indicated the stepwise and uniform assembling process. The atomic force microscopy (AFM) image showed that a compact gold multilayer thin film was successfully assembled. The residual boronic acid group on the surface of thin film could incorporate glycosylated-protein horseradish peroxidase (HRP), and good catalytic activity for H2O2 could be observed.     

Synthesis of Poly(anilineboronic acid) Nanofibers for Electrochemical Detection of Glucose

Poly(anilineboronic acid) (PABA) nanofibers with U‐shaped and ring‐shaped morphologies have been synthesized successfully by chemical polymerization of 3‐aminophenylboronic acid (APBA) in the presence of cetyltrimethylammonium bromide (CTAB) and NaF. The morphologies and sizes of PABA nanofibers can be controlled by adjusting the synthetic parameters, such as the concentrations of CTAB and APBA. The U‐shaped PABA nanofibers exhibit excellent electrochemical redox activity and high sensitive detection of D ‐glucose in phosphate‐buffered saline stock solution (pH 7.4) because of their high effective surface area as well as the high density of boronic acid groups.

     

基于硼酸/二醇识别的新型糖囊泡荧光传感器

摘要 合成了含有识别基团苯硼酸、喹啉发色团的新型双亲化合物,N-硼苄基-8-16烷基溴化喹啉（N-(boronobenzyl)-8-hexadecyloxyquinolinium bromide (BHQB)）.该化合物在可选择性溶剂中自组织成囊泡,囊泡的相变温度为52.4℃;研究了BHQB囊泡的荧光性质,结果表明：当向囊泡体系加入糖时,喹啉在425nm 峰逐渐增强而508nm峰急剧减弱,变化趋势为葡萄糖>果糖.实验结果表明,BHQB囊泡可以作为可植入、连续检测血糖浓度的荧光囊泡传感器,可望用于临床实际应用.     

Enhancing effect of phenylboronic acid compounds and their interactions with the diol groups of saccharides in a capillary electrophoresis-chemiluminescence detection system.

In our previous study, we proposed molecular recognition of mono- and disaccharides making use of the interaction between their diol groups and p-iodophenylboronic acid in capillary electrophoresis with a chemiluminescence detection system. Here, to extend our knowledge of molecular recognition, we first examined the enhancing effects of four phenylboronic acid compounds other than p-iodophenylboronic acid i.e., 4-biphenylboronic acid, 4-octyloxyphenyl-boronic acid, 3-octyloxyphenylboronic acid, and 4-dodecyloxyphenylboronic acid, for luminol-hydrogen peroxide-horseradish peroxidase reaction in the capillary electrophoresis-chemiluminescence detection system. Only 4-biphenylboronic acid showed an enhancing effect similar to that of p-iodophenylboronic acid; the effect was determined over the range of 0.5-10 microM in this system. Second, we estimated the apparent stability constants between the diol groups of saccharides (1-methyl-D-glucoside, D-saccharose, and D-fructose) and the boronic acid moieties of the two enhancers, p-iodophenylboronic acid and 4-biphenylboronic acid. The apparent binding constants obtained here provided insight to confirm the principle of molecular recognition for the saccharides examined here.     

Dual optical and electrochemical saccharide detection based on a dipyrido[3,2-a:2′3′-c]phenazine (DPPZ) ligand

We report a new molecular sensor based on dipyrido[3,2-a:2′3′-c]phenazine (DPPZ) functionalized with boronic acid groups. The sensor binds to saccharides and upon binding results in changes in fluorescence intensity as well as cathodic shifts in the sensor’s formal potential. The ability of the new DPPZ-based sensor to provide both electrochemical and optical signal outputs demonstrates the viability of this family of molecules to be developed as dual-signal detectors. Measurements to determine the stability constants with four saccharides are shown.     

Quartz crystal biosensor for detection of sugars and glycated hemoglobin

3-Aminophenylboronic acid (APBA) was used for construction of affinity mass sensors for determination of saccharides and glycated hemoglobin using complexation reaction with diol groups. Two approaches were tested for the bioligand layer fabrication—incorporation of APBA inside a thicker matrix and immobilization as a self-assembled monolayer, respectively. The direct affinity sensor with APBA embeded in the structure of glutaraldehyde-crosslinked bovine serum albumine provided linear response to mono- and dissacharides in the range from 0.1 to 15.0 mg/ml. This biosensor was also used for kinetic analysis of the interaction of boronic acid with diols; the values of association and dissociation constants were determined. The sensors with a monolayer of boronic groups were better suited for binding of glycated hemoglobin, probably due to improved steric access to the ligand.     

Highly Efficient Solid‐Phase Labeling of Saccharides within Boronic Acid Functionalized Mesoporous Silica Nanoparticles

Labeling is critical for the detection, quantitation, and structural identification of saccharides. However, conventional liquid‐phase labeling suffers from apparent disadvantages, such as time‐consuming, the presence of excessive labeling reagent, and high applicable saccharide concentration. A solid‐phase approach is presented for highly efficient labeling of saccharides, using boronic acid functionalized mesoporous silica nanoparticles (MSNs) as a selective extraction sorbent and nanoscale reactor. The solid‐phase labeling approach exhibited several significant advantages, including: much faster reaction speed (taking only 2 min), high product purity, and much lower applicable saccharide concentration (four orders of magnitude lower than that of liquid‐phase labeling). Thus, this labeling approach opens up new avenues to the facile and efficient labeling of saccharides.     

Direct and simultaneous high-performance liquid chromatographic assay for the determination of p-aminobenzoic acid and its conjugates in human urine

Procedures based on high-performance liquid chromatography (HPLC) were developed for identifying and measuring p-aminobenzoic acid (PABA) and its conjugate metabolites in human urine after oral doses of PABA. p-Aminohippuric acid (PAH), PABA, p-acetamidohippuric acid (PAHA) and p-acetamidobenzoic acid (PADB) in urine were resolved and determined by HPLC simultaneously and directly without extraction. A mobile phase consisting of 3% (v/v) acetonitrile in distilled water containing 0.005 M 1-heptanesulphonic acid in glacial acetic acid (PIC-B7) at pH 3.3 was eluted at 1 ml/min through a C18 Spherisorb column, followed by UV detection at 280 nm. After hydrolysis of urine samples at 37 degrees C for 3 h with beta-glucuronidase, the amounts of PABA-glucuronide and PADB-glucuronide were also determined. The retention times of PAH, a dominant peak which disappeared after hydrolysis, PABA, DABA (3,5-diaminobenzoic acid, as the internal standard), PAHA and PADB were 11.8, 14, 15, 18, 24 and 46 min, respectively. The 24-h urinary recoveries of PAH, PAHA, PADB, PADB-glucuronide, PABA and PABA-glucuronide after separate oral doses of 200 and 800 mg of PABA in one healthy subject were 43.4 and 48.1, 7 and 29.1, 11.2 and 11.8, 34.8 and 6.6, 0.2 and 0.3, and 1.0 and 2.4%, respectively. It is interesting that at high dose (800 mg) saturation of glucuronidation of PADB (N-acetylated PABA) appeared to occur, which resulted in an increase in the formation of PAHA, the glycine conjugate of PADB. Over 90% of the oral dose was accounted for by 8 h after administration.     

Boronic acid building blocks: tools for sensing and separation

In this feature article the use of boronic acids to monitor, identify and isolate analytes within physiological, environmental and industrial scenarios is discussed. Boronic acids recognise diol motifs through boronic ester formation and interact with anions generating boronates, as such they have been exploited in sensing and separation protocols for diol appended molecules such as saccharides and anions alike. Therefore robust molecular sensors with the capacity to detect chosen molecules selectively and signal their presence continues to attract substantial attention, and boronic acids have been exploited with some success to monitor the presence of various analytes. Reversible boronic acid-diol interactions have also been exploited in boron affinity chromatography realising new separation domains through the same binding events. Boronic acid diol and anion interactions pertaining to sensing and separation are surveyed.     

Elucidating the Anomalous Binding Enhancement of Isoquinoline Boronic Acid for Sialic Acid Under Acidic Conditions: Expanding Biorecognition Beyond Vicinal Diols

A zwitterionic heterocyclic boronic acid based on 4-isoquinolineboronic acid (IQBA) exhibits the highest reported binding affinity for sialic acid or N-acetylneuraminic acid (Neu5Ac, K=5390±190 m ⁻¹) through the formation of a cyclic boronate ester complex under acidic conditions (pH 3). This anomalous pH-dependent binding enhancement does not occur with common neutral saccharides (e.g., glucose, fructose, sorbitiol), because it is mediated via selective complexation to a α-hydroxycarboxylate moiety forming a stable ion pair and ternary complex with Neu5Ac in phosphate buffer. IQBA expands biorecognition beyond classical vicinal diols under neutral or alkaline buffer conditions, which enables the direct analysis of Neu5Ac by native fluorescence with sub-micromolar detection limits.     

Controlling deposition and release of polyol-stabilized latex on boronic acid-derivatized cellulose

Poly(glycerol monomethacrylate)-stabilized polystyrene (PGMA-PS) latex particles undergo specific, pH-dependent adsorption onto regenerated cellulose film bearing surface phenylboronic acid groups (cellulose-PBA). Deposition occurs at pH 10 and is driven by the boronate ester formation with the polyol latex surface coating. In contrast, no deposition occurs at pH 4, and previously deposited particles can be readily desorbed at this lower pH. In control experiments, conventional anionic sulfate-stabilized polystyrene latex did not deposit onto the hydrophilic cellulose surface. The distribution of boronate groups in the cellulose was determined by exposure to Alizarin Red S dye, which forms a fluorescent complex with phenylboronic acid; confocal microscopy was used to determine a surface density of 3 nm(2) per boronic acid group on the cellulose surface. Although the boronic acid binding constant with PGMA is relatively low (5.4 L/mol), the cooperative interactions between multiple PBA surface sites and the many PGMA chains per latex particle are sufficient to induce specific latex adsorption, providing a convenient new tool for controlling nanoparticle deposition on surfaces.     

Free and total para-aminobenzoic acid analysis in plants with high-performance liquid chromatography/tandem mass spectrometry

para-Aminobenzoic acid (PABA), a precursor in the synthesis of folates in plants, is determined by liquid chromatography/tandem mass spectrometry (LC/MS/MS). In plants PABA can be converted into its beta-D-glucopyranosyl ester (PABA-Glc) and can also exist in its free form. In this work, we developed and validated a quantitative method to study free and total PABA in plants. The total PABA (free PABA plus PABA-Glc) can be evaluated after acid hydrolysis at 80 degrees C for 2 hours. The plant material is homogenized and the PABA content is quantified using the standard addition procedure. The validated method is selective, sensitive, simple, accurate, has a recovery between 99.6 to 102.5%, is reproducible (RSD between 1.4 and 4.4%), and is linear between 2.5 and 1538 ng/mL. Free and total PABA determinations in five vegetables showed that different plant species had different amounts of free and total PABA, and that the ratios of total versus free PABA were also variable. This new method could be valuable for studies of folate synthesis in plants.