Design Of Saccharides-based Electrochemical Chiral Sensor For Recognition Of Amino Acid Enantiomers

Chirality is one of the most common properties in natural systems,and vast majority of important biomolecules possess chiral properties.Chiral recognition is a fundamental property of many biomolecules,which has important research significance in biology,medicine and biotechnology due to the different pharmacological activities of enantiomers in living systems.Amino acids are essential and important substances for life.They play a very important role in protein synthesis,catalysis,metabolism,immunity and other processes,and can be used as important biomarkers for monitoring various metabolic diseases.Although a couple of enantiomers of any amino acid exhibit identical physical properties,different biological interactions,pharmacological activities and metabolic behaviors of the two enantiomers in biological systems could be observed.It’s worth noting that one enantiomer might even have serious side effects or toxic effects.Therefore,chiral recognition of amino acid enantiomers is not only closely related to human health and life safety,but also has a profound impact on clinical and pharmaceutical applications and other important research areas.Development of simple,rapid,time-saving,efficient and highly sensitive methods for chiral recognition of amino acid enantiomers has become a hotspot of current researches.Among the existing chiral recognition methods,electrochemical sensing has been widely used due to its advantages of facile operation,fast response and low cost.Saccharides are green organic molecules which are cheap,easy-to-obtain,biocompatible and easy-to-degrade.Most importantly,saccharides are natural optically pure chiral compounds,which can be used as chiral selectors for electrochemical chiral recognition of amino acid enantiomers.This dissertation focuses on the design of saccharide-based composites,and the construction of electrochemical chiral sensors,as well as the evaluation of their recognition performances toward different amino acid enantiomers.The main research works are as follows:（1）A novel electrochemical chiral sensor based on graphene oxide（GO）,bovine serum albumin（BSA）and an aminosaccharide-chitosan（CS）modified glassy carbon electrode（GO/BSA/CS/GCE）was constructed by a layer-to-layer drop-casting method,which was used for recognition of tryptophan（Trp）enantiomers.Differential pulse voltammetry（DPV）was employed to identify and quantify Trp enantiomers at GO/BSA/CS/GCE.Trp enantiomers concentrations showed good linear relationship with peak current values within the concentration range 1.0-8.0 m M,and the chiral recognition efficiency of D-Trp and L-Trp on GO/BSA/CS/GCE was satisfactory.（2）Based on two polysaccharides,soluble starch（SS）and CS,a self-assembled electrochemical chiral interface was fabricated and used for chiral recognition of tyrosine（Tyr）enantiomers via square wave voltammetry（SWV）.Under the optimized experimental conditions,the oxidation peak current ratio（I_L/I_D）and the difference between the two peak potentials（ΔEp=E_D-E_L）of L-Tyr and D-Tyr at SS-CS/GCE were1.38 and 12 m V,respectively.Furthermore,a good linear relationship between Tyr enantiomer concentrations and peak currents could be observed in the concentration range 0.01-1.00 m M.In addition,SS-CS/GCE showed the ability to predict the ratios of L-Tyr and D-Tyr in the enantiomeric mixtures.The proposed SS-CS/GCE chiral sensor exhibited high anti-interference ability and good repeatability as well as excellent reproducibility.（3）CS and D-（+）-Galactosamine hydrochloride（Gal N）were assembled together via hydrothermal assembly to obtain a novel composite（CS-Gal N）,and a novel and facile chiral sensing platform based on CS-Gal N modified glassy carbon electrode（CS-Gal N/GCE）was fabricated and used for electrochemical recognition of Tyr enantiomers.Under optimized experimental conditions,the oxidation peak current ratio of L-Tyr to D-Tyr（I_L/I_D）and the difference between their peak potentials（ΔEp=E_D-E_L）at CS-Gal N/GCE by SWV were 1.70and 28 m V,respectively.The peak current increased linearly with Tyr enantiomer concentration in the range 0.01-1.00 m M,and limit of detection（LOD）of 0.65μM and 0.86μM for L-Tyr and D-Tyr at CS-Gal N/GCE could be obtained,respectively.In addition,CS-Gal N/GCE possessed remarkable sensitivity,excellent stability and good reproducibility,as well as exhibited the ability to determine the percentage of D-Tyr in the enantiomeric mixtures.（4）Based on maltosyl-β-cyclodextrin（Mal-βCD）and black phosphorus nanosheets（BPNSs）,a self-assembled chiral interface for electrochemical recognization of Tyr enantiomers was constructed.BPNSs and Mal-βCD modified GCE（Mal-βCD/BPNSs/GCE）was facilely prepared by a layer-to-layer drop-casting method.Under the optimum conditions,Mal-βCD/BPNSs/GCE showed excellent recognition toward Tyr enantiomers（I_L/I_D=1.51,ΔEp=E_D-E_L=20 m V）.Furthermore,we observed that the peak currents of D-Tyr and L-Tyr at Mal-βCD/BPNSs/GCE increased linearly with an increase in Tyr enantiomer concentrations in the range 0.01-1.00 m M.Most importantly,the proposed chiral sensor could predict the percentage of D-Tyr in enantiomeric mixtures.Additionally,Mal-βCD/BPNSs exhibited good anti-interference ability,good repeatability and excellent reproducibility.The mechanism of chiral recognition of Tyr enantiomers by Mal-βCD/BPNSs/GCE was elucidated by means of different characterizations.（5）We fabricated Nafion（NF）stabilized BPNSs and Mal-βCD)composite（BPNSs-Mal-βCD）as novel electrochemical sensor platform for chiral recognition of Trp enantiomers.BPNSs-Mal-βCD composite modified GCE（BPNSs-Mal-βCD/GCE）was further coated with NF,which served as a protective film to immobilize BPNSs-Mal-βCD on the electrode surface to achieve higher stability.Under the optimum conditions,the difference oxidation peak current ratio（I_L/I_D）and between the peak potential（ΔEp=E_D-E_L）of L-Trp to D-Trp at NF/BPNSs-Mal-βCD/GCE by SWV were 1.49 and 20 m V,respectively.The chiral recognition mechanism was proposed,and the intermolecular hydrogen bonding interactions,as well as the hydrophobic-cavity-triggered penetrating effects dominated the effective chiral recognition.A linear calibration curve of peak currents versus Trp enantiomers could be obtained in the concentration range 0.01-1.00 m M,and LODs of 1.07μM and 1.71μM for L-Trp and D-Trp could be observed,respectively.NF/BPNSs-Mal-βCD/GCE also showed excellent stability,good reproducibility and acceptable anti-interference capability.Therefore,the designed chiral sensor is expected to be practically applied to the sensitive recognition of Trp enantiomers in real samples such as banana juice or egg white.Compared with the previous work,an interesting phenomenon observed is that two similar sensors fabricated by different methods showed totally different recognition ability toward either Tyr or Trp enantiomers,and the underlying mechanism was analyzed and discussed in detail.（6）Single-layer graphene oxide（SGO）and amino-β-cyclodextrin（NH₂-βCD）were integrated through an amidation reaction,and then assembled with BPNSs with puckered orthorhombic layered structure to construct a chiral composite,which was used to modify glassy carbon electrode to obtain a chiral electrochemical sensor（SGO-NH₂-βCD/BPNSs/GCE）.Compared with previous works,the covalent coupling and self-assembly methodologies for preparation of SGO-NH₂-βCD/BPNSs/GCE greatly improved its recognition efficiency for Tyr enantiomers.SGO-NH₂-βCD/BPNSs/GCE showed relatively higher affinity for D-Tyr,and lower oxidation peak potential and higher oxidation peak current could be observed.Importantly,the concentrations of Tyr enantiomers had a good linear relationship with the peak currents,and the percentages of D-Tyr in racemic Tyr mixtures could be predicted at SGO-NH₂-βCD/BPNSs/GCE,confirming its ability to detect and quantitatively analyze Tyr enantiomers.