Electrochemical recognition for tryptophan enantiomers based on 3, 4, 9, 10-perylenetetracarboxylic acid–chitosan composite film
- 88 Downloads
A novel and simple chiral sensing platform had been successfully fabricated by means of amidation reaction between 3, 4, 9, 10-perylenetetracarboxylic acid (PTCA) and chitosan (CS) to form 3, 4, 9, 10-perylenetetracarboxylic acid–chitosan (PTCA–CS) composite film. Since CS has chiral center and PTCA has excellent electrical conductivity, the PTCA–CS composite modified glassy carbon electrode (PTCA–CS/GCE) could be treated as an effective electrochemical chiral sensor and applied for chiral discrimination of tryptophan (Trp) enantiomers theoretically. PTCA–CS composite was characterized by Fourier transform infrared (FTIR) spectroscopy and cyclic voltammetry (CV). When the prepared chiral sensing interface interacted with tryptophan isomers, a higher selectivity was received from D-Trp by differential pulse voltammetry (DPV). It indicated that the PTCA–CS/GCE can be treated as an electrochemical chiral sensor for the discrimination of Trp enantiomers. Further study demonstrated that the peak currents were linearly increased with the increasing percentage of L-Trp of Trp racemic mixture. Furthermore, the enantioselective interaction of the PTCA–CS/GCE was systematically studied by other experimental factors, such as the incubation time and acidity.
KeywordsElectrochemical chiral sensor Tryptophan enantiomers Enantioselectivity recognition 3, 4, 9, 10-perylenetetracarboxylic acid–chitosan composite
Chirality is the ubiquitous phenomenon of nature, playing a vital role in living organisms. Macroscopic chirality is caused by the chiral small molecules that constitute them. L-amino acids are the foundational component of protein synthesis in humans and animals. However, the monosaccharides that make up the polysaccharides are in the D-configuration. Several studies have suggested that individual enantiomer of a drug often has different pharmacological activity and biological activity. One enantiomer is therapeutically effective, while the other may not be effective, and even cause dangerous side effects . At present, numerous analytical techniques have been applied for chiral discrimination of optically active chiral compounds, such as capillary electrophoresis [2, 3], fluorescence spectroscopy , high performance liquid chromatography [5, 6, 7], quartz crystal microbalance , and electrochemical measurements [9, 10, 11, 12, 13, 14]. A great deal of attention has been focused on electrochemical techniques for study of chiral discrimination due to their advantages of simple operation, low equipment requirements, low-cost, easy repeatability, environmentally friendly, and real-time operation [15, 16, 17, 18, 19].
3, 4, 9, 10-Perylenetetracarboxylic acid (PTCA), as an excellent aromatic organic dye, has drawn increasing attention in the fields of electrochemistry and field-effect transistors due to its brilliant chemical stability as well as good solubility [20, 21]. Moreover, PTCA has redox activity for its electronic property. Its electrical conductivity ranges from 10−1 to 10−2 S cm−1 . In addition, PTCA-based materials have been carried over into the development of electrochemical sensors for its simple membrane-forming performance [23, 24]. In the regular methods, PTCA was just used for semiconductor template to develop PTCA-based materials for electrochemical sensors. Apparently, it has great potential and prospects for construction of electrochemical sensors.
Tryptophan is one of the essential amino acids (L-Trp) related to the component of proteins, which plays a significant role in maintaining nitrogen balance in humans and animals and can also act as an anti-depressant . However, D-Trp has no apparent effect on physiological and pharmacological activity. Consequently, it is of great importance to identify optically active compounds in the field of pharmaceutical industry and biological sciences. The basic step in developing an electrochemical chiral biosensor is to fabricate a chiral interface with recognition sites for chiral recognition. Chitosan (CS), a natural polysaccharide, possesses outstanding chiral selectivity owing to the existence of large amounts of chiral sites . Benifiting from its excellent water permeability, easy film-forming ability, good adhesion, and as nontoxicity, chitosan is commonly used for constructing electrochemical sensors and biosensors [27, 28, 29]. However, the construction of electrochemical chiral sensor based on chitosan-containing suffered some restrictions owing to dielectric and insulating properties of CS. But an effective method for electrochemical identifying tryptophan (Trp) was proposed by group of Yong Kong, offering an effective evidence for electrochemical chiral recognition of Trp enantiomers with protonated CS from the viewpoint of electrochemistry .
In this article, we prepared a novel electrochemical chiral sensor based on PTCA–CS composite film electrodeposited on a glassy carbon electrode (PTCA–CS/GCE) to design a simple method for enantiorecognition of Trp enantiomers. The electrodeposited PTCA–CS chiral interface combined the corresponding advantages of each element, such as the stereoselectivity of CS and the excellent electrical conductivity of PTCA. Therefore, the chiral interface was used for electrochemical discrimination of Trp enantiomers by differential pulse voltammetry (DPV). PTCA–CS/GCE demonstrated completely different selectivity to L-Trp and D-Trp, indicating a larger current response to D-Trp.
Materials and reagents
N-(3-Dimethylaminopropyl)-N-ethylcarbodiimidehy drochloride (EDC), N-hydroxy succinimide (NHS), L-Trp, and D-Trp were purchased from Aladdin Chemistry Co., Ltd. (China). 3, 4, 9, 10-Perylenetetracarboxylic dianhydride (C24H8O6, PTCDA) was obtained from BaiYin LiangYou Chemical reagents Co., Ltd. Chitosan (CS), K4Fe(CN)6, and K3Fe(CN)6 were from Tianjin FuYu Fine Chemicals Co., Ltd. 0.1 M phosphate buffer solution (PBS) at different pH values was prepared with 0.1 M KH2PO4 and 0.1 M K2HPO4 containing 0.1 M KCl as supporting electrolyte. 5.0 mM [Fe(CN)6]4−/3− solution was prepared by K4Fe(CN)6 and K3Fe(CN)6. The supporting electrolyte was 0.1 M KCl. Ultra-pure water was applied throughout all the experiments.
Cyclic voltammetry (CV) and differential pulse voltammetry (DPV) were carried out on a CHI660 electrochemical workstation (Chen Hua Instruments Co., Shanghai, China) with a conventional three electrode system containing a saturated calomel electrode (SCE) as the reference electrode, a platinum wire as the counter electrode, and a modified glassy carbon electrode(GCE) as the working electrode. Fourier transform infrared (FT-IR) spectra of the samples were obtained from an EQUINOX-55 FTIR spectrometer.
Synthesis of PTCA-based composite
The whole procedure involved in preparing the PTCA–CS composite was displayed schematically as shown below: in short, PTCA was obtained from hydrolyzing PTCDA in KOH aqueous solution (2%) . Subsequently, 40 mg of EDC and 60 mg of NHS were added into the PTCA solution respectively to activate carboxyl sufficiently on PTCA accompanied by continuous stirring for 2 h. One hundred milligram CS powder was added in 50 mL of 0.1 M acetic acid solution with continuous stirring for 2 h, and the as-prepared CS solutions were added dropwise into the activated PTCA within 20 min. The amidation reaction between PTCA and CS was stirred continuously for 4 h at room temperature. The synthesized product (expressed as PTCA–CS) was filtered and then washed thoroughly with 0.1 M acetic acid solution and distilled water to remove unreacted PTCA and CS, finally stored at 5 °C.
Fabrication procedure of the electrochemical biosensors
Electrochemical chiral recognition of tryptophan isomers
Electrochemical chiral recognition of Trp isomers by PTCA–CS/GCE was conducted by differential pulse voltammetry. The as-prepared PTCA–CS chiral interface was immersed into 25 mL 5 mM L-Trp or D-Trp solution (pH 6.0) for 5 min, respectively. Then, the differential pulse voltammograms of these systems were compared (PTCA-L-Trp vs PTCA-D-Trp, CS-L-Trp vs CS-D-Trp; PTCA–CS-L-Trp vs PTCA–CS-D-Trp). Every differential pulse voltammetry was repeated five times to calculate for the error bars. To study the pH-sensitive properties of the PTCA–CS chiral sensor, the enantiorecognition between PTCA–CS and Trp isomers was performed at t pH range from 4.5 to 7.5.
Results and discussion
Characterization of PTCA–CS
Electrochemical properties of different modified electrodes
The PTCA–CS/GCE was repeatedly cycled in 5.0 mM [Fe (CN)6]4−/3− solution containing 0.1 M KCl. After scanning for 40 cycles continuously, the chiral biosensor had excellent reproducibility and stability with the relative standard deviation (RSD) of 3.1% (n = 13).
The interaction between PTCA–CS and the Trp enantiomers
Influence of incubation time and pH on enantiorecognition
As displayed in Fig. 5b, the electrochemical chiral recognition of Trp enantiomers was dependent on the pH of the solution. The electrochemical properties of Trp enantiomers on the PTCA–CS chiral sensing platform at pH values of 4.5, 5.0, 5.5, 6.0, 6.5, 7, and 7.5 were investigated. It is obviously that peak current ratio increased with increasing pH (4.5–6.0). However, the recognition efficiency had not risen but deteriorated with the increase of pH. The low efficiency at low pH values was most likely due to the decomposition of natural polysaccharide including CS in strong acidic medium, leading to the instability of the PTCA–CS recognition systems on GCE . It was reported that the isoelectric point of tryptophan is 5.89 . Therefore, Trp isomers were negatively charged when pH exceeded 6. The electrostatic repelling action between negatively charged L-Trp or D-Trp and a large quantity of oxygen groups on PTCA would reject the combination of Trp enantiomers with the PTCA–CS composite, resulting in reduced recognition efficiency with the PTCA–CS/GCE. Therefore, it was obviously that any interaction might undoubtedly affect enantioselectivity. It can be observed that the maximum peak current ratio appeared at pH = 6. Accordingly, pH = 6 was applied in experiments.
Current response to different concentrations of Trp enantiomers
Application of chiral biosensor
Comparison of different modified electrodes for enantiorecognition of Trp isomers
I L /I D
In summary, a simple and feasible electrochemical chiral sensor with high sensitivity for recognition of L- and D-Trp was designed in this study. The recognition efficiency at the PTCA–CS/GCE was higher than those at PTCA/GCE and CS/GCE (or bare GCE) owing to the synergistic effects between PTCA and CS. This work may open up a new application of PTCA for the fabrication of the feasible and convenient electrochemical chiral biosensors for the electrochemical chiral recognition of other optically active compounds.
This work was supported by the National Natural Science Foundation of China (51262027), the financial support of the Natural Science Foundation of Gansu Province (1104GKCA019 and 1010RJZA023), the Science and Technology Tackle Key Problem Item of Gansu Province (2GS064-A52-036-08) and the fund of the State Key Laboratory of Solidification Processing in NWPU (SKLSP201011).
- 3.Szabó ZI, Tóth G, Völgyi G, Komjáti B, Hancu G, Szente L, Noszál B (2016) Chiral separation of asenapine enantiomers by capillary electrophoresis and characterization of cyclodextrin complexes by NMR spectroscopy, mass spectrometry and molecular modeling. J Pharm Biomed 117:398–404CrossRefGoogle Scholar
- 5.Kharaishvili Q, Jibuti G, Farkas T, Chankvetadze B (2016) Further proof to the utility of polysaccharide-based chiral selectors in combination with superficially porous silica particles as effective chiral stationary phases for separation of enantiomers in high-performance liquid chromatography. J Chromatogr A 1467:163–168CrossRefGoogle Scholar