Method for identifying amino acid enantiomers by using SERS-Cha substrate

Abstract

The invention discloses a method for identifying amino acid enantiomers by using an SERS-Cha substrate. The method comprises the following steps: preparing a chiral silver nano spiral structure by using an electron beam evaporation coating instrument in combination with a liquid nitrogen low-temperature system; on the basis of the SERS-Cha theory, the chiral substrate is used for performing SERS characterization on two chiral histidine molecules, and the chiral histidine molecules with two configurations are identified according to the significant difference of SERS intensities of enantiomer molecules on the same chiral platform. The method provided by the invention is different from the traditional method for detecting the chiral enantiomer by circularly polarized light, realizes the identification of the chiral enantiomer by linearly polarized light, and is an effective and simpler method.

Description

Technical Field

[0001] The invention belongs to the field of nanotechnology, relates to a biochemical detection substrate and method, and particularly designs a method for identifying amino acid enantiomers using a SERS-Cha substrate. Background Art

[0002] Enantiomer identification is crucial in many areas of analytical chemistry, molecular biology, pharmaceutical and pesticide science. Various optical spectroscopic techniques, such as optical rotation and circular dichroism, have been used to detect and quantify enantiomers. Typically, circularly polarized light is used to identify the absolute configuration and composition of enantiomers. However, these methods cannot be used for racemates or compounds with weak optical responses. Surface-enhanced Raman spectroscopy is a powerful technique that provides ultrasensitive structural information through the vibrational "fingerprints" of trace analytes. The molecules interact with the strong electromagnetic fields around the metal nanostructures, which also contribute to the strong radiation enhancement. Surface enhanced Raman optical activity (SEROA) has also been proposed as a technique to identify enantiomers, using the difference in surface enhanced Raman scattering (SERS) intensity of left-handed and right-handed circularly polarized light. However, similar to other methods, SEROA also provides only relatively weak anisotropic signals and is ineffective for racemates.

[0003] Using non-circularly polarized light (linearly polarized light or non-polarized light) combined with a chiral substrate platform to distinguish enantiomers is an effective and universal method for identifying enantiomers, called surface-enhanced Raman scattering-Chiral anisotropy (SERS-Cha) method, which is based on the selective Raman enhancement of different configurations of enantiomers on the chiral plasma surface, that is, the enantiomer with the same chirality as the detection substrate platform will detect a higher SERS peak intensity, so the configuration can be judged based on the difference in SERS intensity of enantiomer molecules on the same chiral platform. At present, chiral inorganic nanostructures have been applied to chiral detection and sensing, and chiral structures have realized the direct identification of chiral molecules with non-circularly polarized light, but the chiral signals of synthesized inorganic nanostructures need to be further improved and enhanced. Summary of the invention

[0004] The object of the present invention is to provide a method for identifying amino acid enantiomers using a SERS-Cha substrate.

[0005] To achieve the above object, the present invention provides a method for preparing a SERS-Cha substrate and identifying amino acid enantiomers, comprising the following steps:

[0006] (1) Using low-temperature grazing angle deposition technology, setting appropriate tilt angles, sample stage rotation speeds, and rotation directions, left-handed nano helix (LNH) and right-handed nano helix (RNH) silver nanohelix structures were prepared respectively;

[0007] (2) L-Histidine (L-His) and D-Histidine (D-His) solutions of the same volume and concentration were dripped onto the substrate and incubated for a certain period of time, and the corresponding SERS spectra were obtained by sampling with a Raman spectrometer;

[0008] (3) Analyze the SERS spectral data of two histidines on different chiral substrates and combine them with the SERS-Cha method to realize the chiral identification of histidine.

[0009] Preferably, the specific steps of step (1) are as follows: placing a clean glass substrate into the vacuum chamber of a coating machine, cooling the sample stage to minus 170°C with liquid nitrogen, setting the sample stage tilt angle to 85°, the evaporation rate to 0.3 nm / s, the sample stage rotation rate to 0.142° / s, and rotating the sample stage counterclockwise and clockwise to prepare LNH and RNH, respectively.

[0010] Preferably, the specific steps of step (2) are as follows: weigh a certain amount of histidine and dissolve it in water, sonicate for five minutes to completely dissolve it, take 2 μL of L-His and D-His solutions of the same concentration and drop them on the LNH and RNH substrates respectively, place them in a -4°C environment and incubate them for 30 minutes, load them with a glass slide and place them on a 40°C heating table to heat until the liquid is completely dried. Set the laser wavelength, power, and integration time, scan the object to be measured on the substrate, and obtain its SERS spectrum.

[0011] Preferably, the specific steps of step (3) are as follows: peak searching and unblocking of the spectral data, analyzing the peak intensities of four chiral substrates and histidine combinations, namely, L-His and LNH, D-His and LNH, L-His and RNH, and D-His and RNH four amino acids and helical chiral substrate combinations, the Raman peak intensity of the same chiral combination (L-His and LNH, D-His and RNH) is about 4.3 times that of the different combinations (D-His and LNH, D-His and LNH), which is consistent with the SERS-Cha theory. The SERS intensity of the same chiral combination will be higher than that of the different chiral combinations, and the chirality of the amino acids can be identified according to the chiral properties of the helical substrate and the SERS peak intensity of the amino acids with different configurations.

[0012] Compared with the prior art, the present invention has the following beneficial effects:

[0013] 1. The substrate preparation process is simple and efficient, with nanoscale structure, obvious chiral characteristics and excellent circular dichroism.

[0014] 2. The present invention can determine the configuration based on the difference in SERS intensity of enantiomer molecules on the same chiral platform, which is an effective, universal and simple method for identifying enantiomers. BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 is a SEM picture of the chiral substrate designed by the present invention;

[0016] Figure 2 is the circular dichroism spectrum of the chiral substrate designed by the present invention;

[0017] Figure 3 The figure is a comparison of the SERS intensity differences between the chiral substrate of the present invention and different amino acid configurations. DETAILED DESCRIPTION

[0018] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0019] Example 1: Identification of histidine enantiomers based on chiral substrate and SERS-Cha theory.

[0020] Step 1: Weigh a certain amount of histidine and dissolve it in water. Ultrasonicate for five minutes, take 2 μL of L-His and D-His solutions of the same concentration and drop them on the LNH and RNH substrates respectively, incubate them at -4°C for 30 minutes, load them with a glass slide and place them on a 40°C heating table until the liquid is completely dried. Set the laser wavelength, power, and integration time, scan the object on the substrate, and obtain its SERS spectrum.

[0021] Step 2: Peak search and unblocking of the spectral data were performed to analyze the peak intensities of four chiral substrates and histidine combinations, namely, L-His and LNH, D-His and LNH, L-His and RNH, and D-His and RNH combinations of four amino acids and helical chiral substrates. The Raman peak intensity of the same chiral combination (L-His and LNH, D-His and RNH) was about 4.3 times that of the different combinations (D-His and LNH, D-His and LNH), which was consistent with the SERS-Cha theory. The SERS intensity of the same chiral combination would be higher than that of the different chiral combinations, and the chirality of the amino acids could be identified based on the chirality of the helical substrate and the peak intensity of amino acids with different configurations.

Claims

1. A method for identifying amino acid enantiomers using a SERS-Cha substrate, characterized in that: The following steps are involved: (1) Using low-temperature grazing angle deposition technology, setting appropriate tilt angles, sample stage rotation speeds, and rotation directions, left-handed and right-handed chiral silver nanohelical structures (LNH and RNH) were prepared respectively; (2) taking L-His and D-His solutions of the same volume and concentration and dropping them onto the substrate and incubating them for a certain period of time, and obtaining the corresponding SERS spectra using a Raman spectrometer; (3) Analyze the SERS spectral data of two histidines on different chiral substrates and combine them with the SERS-Cha method to realize the chiral identification of histidine.

2. The method according to claim 1, characterized in that: The specific steps of step (1) are as follows: place a clean glass substrate into the vacuum chamber of a coating machine, cool the sample stage to -170°C with liquid nitrogen, set the sample stage tilt angle to 85°, the evaporation rate to 0.3nm / s, the clockwise and counterclockwise rotation rates of the sample stage to 0.142° / s, prepare chiral helical structures with different handedness, and store them in a dry vacuum.

3. The method according to claim 1, characterized in that: The specific steps of step (2) are as follows: weigh a certain amount of histidine and dissolve it in water, sonicate for five minutes to completely dissolve it, take 2 μL of L-His and D-His solutions of the same concentration and drop them on the LNH and RNH substrates respectively, incubate them at -4°C for 30 minutes, load them with a glass slide and place them on a 40°C heating table to heat until the liquid is completely dried. Set the laser wavelength, power, and integration time, scan the object on the substrate, and obtain its SERS spectrum.

4. The method according to claim 1, characterized in that: The specific steps of step (3) are as follows: peak searching and unblocking of spectral data, analyzing the peak intensities of four chiral substrates and histidine combinations, i.e., combinations of four amino acids, L-His and LNH, D-His and LNH, L-His and RNH, and D-His and RNH, with helical chiral substrates. The Raman peak intensity of the same chiral combination (L-His and LNH, D-His and RNH) is about 4.3 times that of the different combinations (D-His and LNH, D-His and LNH), which is consistent with the SERS-Cha theory. The SERS intensity of the same chiral combination will be higher than that of the different chiral combinations, and the chirality of the amino acids can be identified based on the chiral properties of the helical substrate and the SERS peak intensity of amino acids with different configurations.

5. The method according to claims 1 to 4 realizes a chiral identification method for histidine based on a chiral substrate and SERS-Cha theory.

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