Boronic Acid Groups Research Articles

Synthesis and structure-activity relationship of boronic acid bioisosteres of combretastatin A-4 as anticancer agents

The boronic acid group plays an important role in drug discovery. Following our discovery of a boronic acid analog of combretastatin A-4 (CA-4), a series of analogs featuring a boronic acid group on the C phenyl ring of CA-4 was synthesized and evaluated for cytotoxicity, as well as for their ability to inhibit tubulin polymerization, inhibit the binding of [3H]colchicine to tubulin and cause depolymerization of cellular microtubules. Modifications on the C ring of CA-4, either eliminating the methoxy group or replacing the C phenyl ring with a pyridine ring, resulted in a reduced potency for inhibiting tubulin polymerization, colchicine binding and cytotoxic activities as compared to CA-4. Replacing the phenol group with a boronic acid group on the C ring of phenstatin led to a slight increase in cytotoxic potency but a decreased potency for inhibition of tubulin assembly and colchicine binding. Moreover, there was a significant decrease in activity by replacing the C phenyl ring with a pyridine ring. Our results indicate the critical importance of the methoxy group on the C ring as well as the importance of the C phenyl ring compared to a pyridine ring, despite the latter providing a nitrogen atom as a hydrogen bond donor/acceptor, which was predicted by molecular modeling to enhance interaction with the target. The decreased activities of our modified CA-4 boronic analogs may be attributed to weakened hydrogen bonding in our docking model based on the crystal structure of colchicine bound to αβ-tubulin. Notably, even though their effectiveness in inhibiting tubulin polymerization and colchicine binding and causing microtubule depolymerization in cells, the majority of these boronic acid analogs exhibited substantial cytotoxicity. This suggests that they may have additional cellular targets that contribute to their cytotoxicity, and this warrants further evaluation of these unique boronic acid compounds as potential anticancer agents.

A metal–organic frameworks-imprinted emulsion interface with dynamic size- and site-selective recognition for significantly enhanced flavoniods extraction

The boronic acid suspended Cr based MIL-100 (MIL-100-B) sorbents have been widely used to separation of cis-diol containing molecules. However, it’s still challenging in fabrication of high performance metal organic frameworks (MOFs) due to its irregular size and low content of functional groups. For improving identification efficiency, the emulsion microreactor were firstly designed to purify MIL-100-B prior to their use. Herein, a highly ordered MOFs-imprinted polymer microsphere was synthesized by O/W Pickering emulsion template, which can highly selective separation of MIL-100-B via the synergistic effect of boronic acid sites and imprinted cavities. The obtained purified H-MIL-100-B exhibited the uniform size and abundant boronic acid groups (B elements ratio up to 27.37 %). The emulsion imprinted microsphere (EIMs) showed the good selectivity towards MIL-100-B. Additionally, the obtained MOF-imprinted microsphere exhibited green, convenient, sustainable separation process towards the purification and recycle of MIL-100-B. The purified MOFs (H-MIL-100-B) were used to specific bind with cis-diol flavoniods naringin (NRG). Satisfactorily, H-MIL-100-B show high adsorption amount (43.69μmol/g at 308 K), fast capture kinetics (160 min), high selectivity and excellent regenerability (up to seven cycles). Remarkably, the purified NRG solutions displayed lasting antibacterial activity against Staphylococcus aureus (S. aureus) with inhibition zone of 13.47 mm. More importantly, the H-MIL-100-B could effectively promote the specific recognition ability of NRG by reducing matrix interference and improving purification efficiency (93.93 %) from complex natural extracts. A high score (0.75) was predicted via analytical GREEnness (AGREE) approaches, demonstrating environmentally friendly and greenness of separation process. The MOFs-imprinted emulsion microspheres possessed dynamic recognition properties towards target MOFs, thus opening new direction for the development of convenient and efficient microreactor for the purification of other functional MOFs.

Enhancing the Mechanical Properties of Injectable Nanocomposite Hydrogels by Adding Boronic Acid/Boronate Ester Dynamic Bonds at the Nanoparticle-Polymer Interface.

Incorporating nanoparticles into injectable hydrogels is a well-known technique for improving the mechanical properties of these materials. However, significant differences in the mechanical properties of the polymer matrix and the nanoparticles can result in localized stress concentrations at the polymer-nanoparticle interface. This situation can lead to problems such as particle-matrix debonding, void formation, and material failure. This work introduces boronic acid/boronate ester dynamic covalent bonds (DCBs) as energy dissipation sites to mitigate stress concentrations at the polymer-nanoparticle interface. Once boronic acid groups were immobilized on the surface of SiO2 nanoparticles (SiO2-BA) and incorporated into an alginate matrix, the nanocomposite hydrogels exhibited enhanced viscoelastic properties. Compared to unmodified SiO2 nanoparticles, introducing SiO2 nanoparticles with boronic acid on their surface improved the structural integrity and stability of the hydrogel. In addition, nanoparticle-reinforced hydrogels showed increased stiffness and deformation resistance compared to controls. These properties were dependent on nanoparticle concentration. Injectability tests showed shear-thinning behavior for the modified hydrogels with injection force within clinically acceptable ranges and superior recovery.

Electrochemiluminescence detection of Hg2+ using lanthanide mental organic gel based on boric acid functionalization

Recent advancements in electrochemiluminescence (ECL) have highlighted the potential of metal–organic gels (MOGs) as innovative luminescent materials for environmental monitoring. This study introduces a terbium-based Metal–Organic Gel (Tb-MOG) synthesized through a straightforward solvothermal method, exhibiting quantitative detection of mercury ions (Hg2+). The high mesoporosity of Tb-MOG facilitates co-reactant penetration and the gel does not come off easily when applied to the electrodes, enhancing the stability and efficiency of ECL signals. The organic ligand 5-borophenyl-1,3-dicarboxylic acid (5-bop) serves dual functions: it sensitizes Tb3+ luminescence and specifically recognizes Hg2+. Upon the interaction with Hg2+, the boronic acid group of 5-bop undergoes a metal transfer reaction, resulting in an enhanced ’antenna’ effect, which enhances ECL emission. The linear range for Hg2+ of the proposed method was from 1.2 μM to 24.6 μM, with a detection of limit (LOD) of 0.53 μM.

Synthesis and anti-ureolitic activity of Biginelli adducts derived from formylphenyl boronic acids

Urease is a metalloenzyme that contains two Ni(II) ions in its active site and catalyzes the hydrolysis of urea into ammonia and carbon dioxide. The development of effective urease inhibitors is crucial not only for mitigating nitrogen losses in agriculture but also for offering an alternative treatment against infections caused by resistant pathogens that utilize urease as a virulence factor. This study focuses on synthesizing and investigating the urease inhibition potential of Biginelli Adducts bearing a boric acid group. An unsubstituted or hydroxy-substituted boronic group in the Biginelli adducts structure enhances the urease inhibitory activity. Biophysical and kinetics studies revealed that the best Biginelli adduct (4e; IC50 = 132 ± 12 µmol/L) is a mixed inhibitor with higher affinity to the urease active site over an allosteric one. Docking studies confirm the interactions of 4e with residues essential for urease activity and demonstrate its potential to coordinate with the nickel atoms through the oxygen atoms of carbonyl or boronic acid groups. Overall, the Biginelli adduct 4e shows great potential as an additive for developing enhanced efficiency fertilizers and/or for medical applications.

Polydiacetylene Liposome-Based Dual-Output Optical Sensor for ppb Level Detection of Dopamine in Solution and Solid Phases.

Dopamine (DA), a neurotransmitter, plays a crucial role in regulating motor functions and emotions and can serve as a marker for several diseases. In this study, we report a highly sensitive polydiacetylenes (PDA)-based dual-output sensor for dopamine detection in both solution and solid phases that was developed by modifying PDA liposomes with boronic acid groups at the termini. This sensor exploits the high affinity between the catechol residue of dopamine and the -B(OH)2 group of the PDA-based probe (PDA-PhBA) to form boronate ester bonds, causing a stress-induced blue-to-red color change along with a steady increase in fluorescence response at λmax 622 nm. The PDA-PhBA-based sensor displays high sensitivity toward dopamine with low limit of detection of 6.2 ppb in colorimetric analysis and 0.6 ppb in fluorimetric measurements, demonstrating its dual optical output ability. The sensor works well for adrenaline, another catecholamine, with similar efficacy. Its practical applicability was validated by the successful recovery of trace level dopamine in blood serum and real water samples. Additionally, immobilizing PDA-PhBA liposomes in sodium alginate produced PDA beads for the solid-phase detection of dopamine with an limit of detection (LOD) of 59 nM (9.0 ppb) in colorimetric detection using a smartphone for capturing images and ImageJ software for analysis.

Iron-catalyzed non-directed arene C-H difluoromethylation

Practical, undirected and selective catalytic functionalization of unactivated arenes remains a challenging problem in organic synthesis. We herein report a bioinspired L-cystine-derived ligand BCPOM/Fe-enabled innate C-H difluoromethylation of unactivated arenes as limiting reagents with stable and inexpensive BrCF2CO2Et as the difluoromethylation source in high efficiency. Notably, this method uses environmentally benign H2O2 as the sole oxidant, and enables late-stage functionalization and exceptionally functional-group tolerance, even including oxidation-labile aldehyde, phenolic hydroxy, primary amine, and boronic acid groups, which is difficult to access by current means.

Boronic functionalized aptamer macroarrays with dual-recognition and isothermal amplification of lipopolysaccharide detection.

Ahighly sensitive dual-recognition fluorescence amplification method is presentedfor lipopolysaccharide (LPS) detection based on boronic functionalized aptamer macroarrays with dual-recognition and isothermal amplification. The surface of the polystyrene microplate was firstly carboxylated, and then, 3-aminophenylboronic acid was conjugated to the carboxyl groups through EDC/NHS reaction, creating boronic acid groups as the capture moiety for LPS. A recognition DNA aptamer labeled with the fluorescent dye 6-FAM, which exhibits specificity towards LPS, was selected as the signal reporting moiety. By introducing primers and Klenow enzyme, the fluorescent-labeled aptamers are released from the microplate bottom, and double-stranded structures were formed via isothermal amplification. The addition of SYBR Green I, which strongly fluoresces upon binding to the double-stranded structures, enables signal amplification and detection. This detection method exhibits a linear range of 1-10,000ng/mL and has a detection limit as low as 401.93pg/mL. This analytical approach shows high selectivity and sensitivity and may serve as a universal platform in lipopolysaccharide detection.

PH-Responsive Degradable Electro-Spun Nanofibers Crosslinked via Boronic Ester Chemistry for Smart Wound Dressings.

Recent advances in the treatment of chronic wounds have focused on the development of effective strategies for cutting-edge wound dressings based on nanostructured materials, particularly biocompatible poly(vinyl alcohol) (PVA)-based electro-spun (e-spun) nanofibers. However, PVA nanofibers need to be chemically crosslinked to ensure their dimensional stability in aqueous environment and their capability to encapsulate bioactive molecules. Herein, a robust approach for the fabrication of pH-degradable e-spun PVA nanofibers crosslinked with dynamic boronic ester (BE) linkages through a coupling reaction of PVA hydroxyl groups with the boronic acid groups of a phenyl diboronic acid crosslinker is reported. This comprehensive analysis reveals the importance of the mole ratio of boronic acid to hydroxyl group for the fabrication of well-defined BE-crosslinked fibrous mats with not only dimensional stability but also the ability to retain uniform fibrous form in aqueous solutions. These nanofibers degrade in both acidic and basic conditions that mimic wound environments, leading to controlled/enhanced release of encapsulated antimicrobial drug molecules. More importantly, drug-loaded BE-crosslinked fibers show excellent antimicrobial activities against both Gram-positive and Gram-negative bacteria, suggesting that this approach of exploring dynamic BE chemistry is amenable to the development of smart wound dressings with controlled/enhanced drug release.

Cell Membrane Engineered Polypeptide Nanonets Mimicking Macrophage Aggregates for Enhanced Antibacterial Treatment.

Drug-resistant bacterial infections and their lipopolysaccharide-related inflammatory complications continue to pose significant challenges in traditional treatments. Inspired by the rapid initiation of resident macrophages to form aggregates for efficient antibacterial action, this study proposes a multifunctional and enhanced antibacterial strategy through the construction of novel biomimetic cell membrane polypeptide nanonets (R-DPB-TA-Ce). The design involves the fusion of end-terminal lipidated polypeptides containing side-chain cationic boronic acid groups (DNPLBA) with cell membrane intercalation engineering (R-DPB), followed by coordination with the tannic acid-cerium complex (TA-Ce) to assemble into a biomimetic nanonet through boronic acid-polyphenol-metal ion interactions. In addition to the ability of RAW 264.7 macrophages cell membrane components' (R) ability to neutralize lipopolysaccharide (LPS), R-DPB-TA-Ce demonstrated enhanced capture of bacteria and its LPS, leveraging nanoconfinement-enhanced multiple interactions based on the boronic acid-polyphenol nanonets skeleton combined with polysaccharide. Utilizing these advantages, indocyanine green (ICG)is further employed as a model drug for delivery, showcasing the exceptional treatment effect of R-DPB-TA-Ce as a new biomimetic assembled drug delivery system in antibacterial, anti-inflammatory, and wound healing promotion. Thus, this strategy of mimicking macrophage aggregates is anticipated to be further applicable to various types of cell membrane engineering for enhanced antibacterial treatment.

Graphene quantum dot-based hydrogel microspheres for sensitive detection of caffeic acid

In this investigation, blue-emitting graphene quantum dots (B,N-GQDs) were synthesized via a single-step hydrothermal method in an ethanol/water mixture utilizing 3-APBA as the nitrogen and boron sources. These B,N-GQDs showcased a highly graphitized structure, complemented by surface-functionalized boronic acid and amino groups that bestowed upon them remarkable optical attributes and water solubility. Leveraging the interplay of internal filtering and dynamic quenching effects, the sensor demonstrated remarkable selectivity and sensitivity in detecting caffeic acid (CA) within the range of 0–840 μM, achieving a limit of detection (LOD) of 0.63 μM. Moreover, the fluorescent gel microspheres engineered with B,N-GQDs exhibited robust CA sensing capabilities and portability. As such, the application potential of B,N-GQDs expands to encompass drug metabolism monitoring, alongside the assessment of food and environmental safety.

Excellent Pd-Loaded Magnetic Nanocatalyst on Multicarboxyl and Boronic Acid Biligands.

An effective palladium nanocatalyst (Fe3O4@SiO2-FPBA-DTPA-Pd) was proposed and prepared, which was immobilized on magnetic silica with ethylenediamine pentaacetic acid and formylphenylboronic acid as biligands. A series of characterizations showed that Fe3O4@SiO2-FPBA-DTPA-Pd was 5-15 nm and contained 1.44 mmol/g Pd2+/Pd0. It was stable below 232.7 °C, and its saturation magnetization value was 21.17 emu/g which was easily recycled by a magnet. Its catalytic ability was evaluated through 7 Suzuki reactions and 15 Heck reactions. Results showed that the yields of 14 reactions catalyzed by Fe3O4@SiO2-FPBA-DTPA-Pd were more than 90%, while were better than those of the other two immobilized Pd catalysts on a single diethyltriamine pentaacetic acid (DTPA) group or boronic acid group. Moreover, Fe3O4@SiO2-FPBA-DTPA-Pd showed good reusability in both Suzuki and Heck reactions. In two model Suzuki and Heck reactions, after seven cycles, its yields were still above 95% without significant loss, which exceeded those of many reported catalysts; therefore, it has great potential in future large-scale industrial production.

Rapid screening of acetylcholinesterase active contaminants in water: A solid phase microextraction-based ligand fishing approach

Effect-directed analysis (EDA) has been increasingly used for screening toxic contaminants in the environment, but conventional EDA procedures are often time-consuming and labor-extensive. This challenges the use of EDA for toxicant identification in the scenarios when quick answers are demanded. Herein, a solid phase microextraction ligand fishing (SPME-LF) strategy has been proposed as a rapid EDA approach for identifying acetylcholinesterase (AChE) active compounds in water. The feasibility of ligand fishing techniques for screening AChE active chemicals from environmental mixtures was first verified by a membrane separation method. Then, SPME fibers were prepared through self-assembly of boronic acid groups with AChE via co-bonding and applied for SPME-LF. As AChE coated SPME fibers selectively enriched AChE-active compounds from water, comparing sorbing compounds by the SPME fibers with and without AChE coating can quickly distinguish AChE toxicants in mixtures. Compared with conventional EDA, SPME-LF does not require repeating sample separations and bioassays, endowing SPME-LF with the merits of low-cost, labor-saving, and user-friendly. It is believed that cost-efficient and easy-to-use SPME-LF strategy can potentially be a rapid EDA method for screening receptor-specific toxicants in aquatic environment, especially applicable in time-sensitive screening.

High-strength self-healing hydrogels based on boronate-diol complexation cross-linking with good bioavailability

In this report, new self-healing hydrogels were obtained through simple one-pot free radical polymerization of acrylamide, phenylboronic acid, and 2-hydroxyethyl acrylate as copolymer monomers, potassium persulfate (KPS) as initiator, and tetramethylethylenediamine (TEMED) as promoter under low-temperature alkaline (pH = 9) conditions. The boronic acid group of phenylboronic acid (PBA) can interact with the hydroxyl group of hydroxyethyl acrylate (HEA) to form dynamic boronic ester bonds instead of the conventional bis acrylate crosslinkers. The effect of PBA content on the mechanical strength of the hydrogels was systematically investigated. The hydrogels exhibited higher stress, elongation, and self-healing properties due to the synergistic effect of H-bonds and boronate-diol complexation. Further studies revealed that the A–H–0.2–P–0.08 hydrogels exhibited excellent resistance to compression fatigue and also showed good biocompatibility through in vitro cytotoxicity tests.

Modifying cellulose fibres with carbon dots: a promising approach for the development of antimicrobial fibres.

This study focuses on the development of antimicrobial fibres for use in medical and healthcare textile industries. Carbon dots (CDs) were designed with boronic acid groups for the attachment to cellulose fibres found in cotton textiles and to enhance their attachment to glycogens on bacterial surfaces. Boronic acid-based and curcumin-based CDs were prepared and characterized using various techniques, showing a nanoscale size and zeta potential values. The CDs inhibited the growth of both Staphylococcus epidermidis and Escherichia coli bacteria, with UV-activated CDs demonstrating improved antibacterial activity. The antimicrobial activity of the CDs was then tested, revealing strong adherence to cellulose paper fibres with no CD diffusion and potent inhibition of bacterial growth. Cytotoxicity assays on human cell lines showed no toxicity towards cells at concentrations of up to 100 µg ml-1 but exhibited increased toxicity at concentrations exceeding 1000 µg ml-1. However, CD-modified cellulose paper fibres showed no toxicity against human cell lines, highlighting the antimicrobial properties of the CD-modified cellulose fibres are safe for human use. These findings show promising potential for applications in both industrial and clinical settings.

One-pot preparation of magnetic composite containing boronic acid groups and aminated multwalled carbon nanotubes for the speciation of Se(IV) and Se(VI) in water and milk samples by combination with chromatographic quantification

Speciation of Se(IV) and Se(VI) is essential due to their significant differences in reactivity, toxicity and bioavailability. Efficient extraction is the pivotal step in the quantification of inorganic selenium species. In this work, a new magnetic nano-composite (MNC) containing boronic acid group and aminated multwalled carbon nanotubes was facilely fabricated by means of one-pot hydrothermal strategy. The prepared MNC contained abundant functional groups and satisfactory magnetic saturation value. Combining with magnetic solid phase extraction (MSPE) format, the MNC displayed satisfactory capture performance towards the complex formed by the coordination of Se(IV) and o-phenylenediamine (OPA). Adsorption isotherm and adsorption kinetics were studied in detail to investigate the adsorption procedure of Se(IV)/OPA complex on MNC. Under the optimal preparation conditions of MNC and extraction parameters, the MNC/MSPE was connected with HPLC equipped with a diode array detector (DAD) to quantify trace Se(IV) and Se(VI) species in water and milk samples. Se(VI) was reduced to Se(IV) and then the total inorganic Se was quantified by the developed method. Subtraction method was used to measure the concentration of Se(VI). The achieved limits of detection were in the ranges of 0.0082–0.013 μg/L and 0.041–0.13 μg/kg for water and milk samples, respectively. Recoveries in actual samples spiked with different amounts of analytes varied from 81.0 % and 117 %, and the RSDs for repeatability varied from 1.0 % to 10 %. In comparison with existing studies based on MSPE, the established method presents some merits such as greenness in the preparation of magnetic adsorbent, rapid extraction procedure, low cost and satisfactory sensitivity in the speciation of inorganic Se species.

Boronic acid conjugated polyacrylate coating: A strategy for material-independent surface functionalization

Most biomaterial surfaces are non-functional, which inevitably requires additional functionalization steps. However, these steps are typically material-dependent; only a few limited methods, such as catechol functionalization using polydopamine coating, have been reported as material-independent for surface modification. In this study, we developed a one-step boronic acid surface functionalization method utilizing a polyacrylate that contains both butyl and boronic acid groups. The butyl group exhibits strong interfacial adhesion due to its low glass transition temperature (Tg), which maintains its softness and tackiness, thereby facilitating attachment to various substrates. Meanwhile, the boronic acid acts as a functional group for surface modification. Various substrates, including polymers, metals, and ceramics widely used in implants, were successfully coated with the polyacrylate, bestowing boronic acid functionality on the surface. Given that boronic acid is one of the most widely applied functional groups in the biomaterials field, we anticipate that our methodology will be applicable in developing various biomedical applications such as antifouling coatings, biosensors, and bioadhesives.

Noncrystalline Zeolitic Imidazolate Frameworks Tethered with Ionic Liquids as Catalysts for CO2 Conversion into Cyclic Carbonates.

Noncrystalline zeolitic imidazolate frameworks (ZIFs) tethered with ionic liquids (ILs) were successfully employed as catalysts for mild CO2 conversion into cyclic carbonates for the first time. Notably, noncrystalline ZIFs exhibit outstanding catalytic performance in terms of activity, stability, and substrate suitability. Z3 was obtained through the simultaneous incorporation of a boronic acid group and ILs into its ZIF framework and exhibited a superior catalytic activity. A reaction mechanism for the propylene oxide-CO2 cycloaddition has been proposed, which integrates experimental findings with density functional theory calculations. The results indicate that zinc, ILs, and boronic acid play crucial roles in achieving high activity. Zinc and ILs are identified as key contributors to epoxide activation and ring opening, while boronic acid plays a crucial role in stabilizing the turnover frequency-determining transition states. The simplicity of this ZIF synthesis approach, combined with the high activity, stability, and versatility of the products, facilitates practical and efficient conversion of CO2 and epoxides into cyclic carbonates.

A chemiluminescence immunosensor for biomarker detection based on boronic acid-modified magnetic composite microspheres.

High-sensitivity detection of biomarkers in biological samples is crucial for the early diagnosis and treatment of diseases. In this paper, a versatile and flexible immobilization technique based on the specific affinity interaction between boronic acid and cis-diol groups of antibodies was developed for biomarker detection. As a model, the boronic acid-modified immunomagnetic beads were used for facile and quick immobilization of the alpha-fetoprotein (AFP) antibody due to the specific affinity interactions. Based on this new class of immunomagnetic beads, the chemiluminescence immunosensor could efficiently detect the biomarker of AFP. Under optimal conditions, the limit of detection (LOD) is as low as 8 fM (S/N = 3), showcasing superior sensitivity and detection specificity for AFP. Subsequently, the system was successfully applied to the detection of AFP in fetal bovine serum samples, and the average recovery rate is greater than 95%. Its performance surpassed that of commercial immunomagnetic beads, showcasing the potential application of this new strategy for bioanalysis and clinical diagnosis.

Effective enrichment of glycated proteome using ultrasmall gold nanoclusters functionalized with boronic acid.

Glycated proteins play a crucial role in various biological pathways and the pathogenesis of human diseases. A comprehensive analysis of glycated proteins is essential for understanding their biological significance. However, their low abundance and heterogeneity in complex biological samples necessitate an enrichment procedure prior to their detection. Current enrichment strategies primarily rely on the boronic acid (BA) affinity method combined with functional nanoparticles; however, the effectiveness of these approaches is often suboptimal. In this study, a novel nanocluster (NC)-based enrichment material was synthesized for the first time, characterized as Au22SG18 functionalized with 24 BA groups, in which SG is glutathione. The functionalized BA established a reversible covalent bond with the cis-dihydroxy group through pH adjustment, enabling selective enrichment of glycated peptides. After the optimization of the enrichment protocol, we demonstrated highly sensitive and selective enrichment of standard glycopeptides using the NC-based enrichment material, exhibiting excellent reusability. Efficient enrichment was also demonstrated for the glycated proteome from human serum. These results highlight the potential of the atomically well-defined ultrasmall Au NCs as a powerful tool for high-throughput analysis of glycated peptides.