CN114252429A

CN114252429A - Capillary-based SERS substrate based on gold trioctahedron modification, preparation method and application thereof

Info

Publication number: CN114252429A
Application number: CN202111631967.6A
Authority: CN
Inventors: 卢革宇; 刘晓敏; 张美玲
Original assignee: Jilin University
Current assignee: Jilin University
Priority date: 2021-12-29
Filing date: 2021-12-29
Publication date: 2022-03-29
Anticipated expiration: 2041-12-29
Also published as: CN114252429B

Abstract

A capillary-based SERS substrate based on gold trioctahedron modification, a preparation method and application thereof in fentanyl detection belong to the technical field of biosensing detection. Soaking a capillary tube in a piranha solution to modify hydroxyl, and then soaking in an ethanol solution of 3-Aminopropyltriethoxysilane (APTES) to modify amino, thereby preparing a capillary tube with a positively charged inner wall; and then, uniformly and compactly assembling the gold trioctahedron on the inner wall of the capillary by using the gold trioctahedron coated by polyvinylpyrrolidone (PVP) as an assembling unit by utilizing electrostatic adsorption to obtain the capillary-based SERS substrate. The invention solves the problems of difficult on-site sample suction, easy pollution, poor signal repeatability and the like of the currently and generally adopted plane substrate, and achieves the purpose of further improving the SERS detection sensitivity.

Description

Capillary-based SERS substrate based on gold trioctahedron modification, preparation method and application thereof

Technical Field

The invention belongs to the technical field of biosensing detection, and particularly relates to a gold trioctahedral modification-based capillary-based SERS substrate, a preparation method and application thereof in fentanyl detection.

Background

Fentanyl, a potent narcotic analgesic, has been widely used in cancer therapy and surgical analgesia. However, excessive fentanyl intake can lead to confusion, nausea, subsequent addiction, hypotension, and ultimately death from respiratory depression. More importantly, fentanyl is also increasingly used as an adulterant for illegal drugs such as heroin. Abuse of fentanyl-like drugs has become a global public health problem. Therefore, accurate identification and quantitative analysis of fentanyl are of great significance for control and prevention of novel psychoactive substances.

Generally, the first techniques for trace analysis of fentanyl and its metabolites are typically gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS). Although these methods are accurate and sensitive, they require bulky equipment, high detection costs, cumbersome pre-processing steps and high expertise. Furthermore, immunoassays (such as ELISA) are also capable of recognizing opioids, but due to their lack of molecular specificity, further confirmation is required. Surface Enhanced Raman Scattering (SERS) is a spectroscopic technique that uses precious metal nanomaterials to form high-intensity electromagnetic hot spots to enhance the raman signal of sample molecules. The method has the characteristics of fingerprint identification, high sensitivity, no damage and no interference of water molecules, and is widely concerned in the field of forensic trace drug detection.

Currently, most SERS substrates for drug detection use planar substrates, such as paper-based substrates, tape-based substrates, and poly (dimethylsiloxane) -based substrates, most of which require complicated surface modification techniques and are more susceptible to contamination from the external environment due to exposure of sample molecules to air during detection. Therefore, in order to improve SERS-based quantitative analysis of in-situ environmental toxicology samples, it is important to design a functional SERS substrate with high sensitivity, high stability and high reproducibility.

Disclosure of Invention

The invention aims to solve the problems of difficulty in on-site sample drawing, poor detection repeatability and the like by utilizing a planar substrate. A capillary-based SERS substrate based on gold trioctahedron modification, a preparation method and application thereof in fentanyl detection are provided.

A capillary base SERS substrate based on gold trioctahedral modification is characterized in that: soaking the capillary in piranha solution to modify hydroxyl, and then soaking in ethanol solution of 3-Aminopropyltriethoxysilane (APTES) to modify amino, to prepare capillary with positive inner wall; and then, uniformly and compactly assembling the gold trioctahedron on the inner wall of the capillary by using the gold trioctahedron coated by polyvinylpyrrolidone (PVP) as an assembling unit by utilizing electrostatic adsorption to obtain the capillary-based SERS substrate.

The golden trioctahedron disclosed by the invention is marked as Au TOHs.

The 3-aminopropyl triethoxysilane provided by the invention is marked as APTES.

The piranha solution is marked as piranha solution.

The solutions according to the invention are all aqueous solutions, if not stated otherwise.

The invention relates to a preparation method of a capillary-based SERS substrate based on gold trioctahedral modification, which comprises the following steps:

(1) preparing Au TOHs by using a seed growth method: mixing 2-2.5 mL of 0.1M hexadecyltrimethylammonium chloride (CTAC) solution with 5-20 mu L of 10nm gold nanosphere seed solution, then adding 130-150 mu L of 0.1M Ascorbic Acid (AA) solution, and adding 2-2.5 mL of 0.5 multiplied by 10^-3M HAuCl₄Reacting the solution at 25-30 ℃ for 15-20 min, and centrifuging at 5500-6000 rpm for 10-15 min; dispersing the precipitate into 0.5-1 mL of polyvinylpyrrolidone (PVP) ethanol solution with the mass fraction of 1%, centrifuging at 5500-6000 rpm for 10-15 minutes, dispersing the precipitate into 0.5-1 mL of ethanol to obtain an Au TOHs ethanol solution, and storing at 4 ℃;

(2) using piranha solution and APTPreparation of amino functionalized capillary tube with ES ethanol solution: firstly, mixing the raw materials in a volume ratio of 1-3: 1, ultrasonically cleaning the glass capillary tube for 10-20 minutes by using an acetone and ethanol mixed solution, and then washing with water and drying; soaking the dried capillary tube in piranha solution (H)₂SO₄And H₂O₂The volume ratio of (A) to (B) is 12-7: 3, H₂SO₄Is 98% by mass, H₂O₂30%) for 1-3 hours, and then washed with water and dried, so that hydroxyl groups are modified on the inner wall of the capillary; finally, soaking the capillary tube with the inner wall modified with hydroxyl in an APTES ethanol solution with the volume fraction of 1-10% for 10-15 hours, and then washing with water and drying, thereby modifying amino on the inner wall of the capillary tube;

(3) preparing a capillary-based SERS substrate: concentrating the Au TOHs ethanol solution prepared in the step (1) by 50-200 times, and vertically inserting the capillary with the modified amino group on the inner wall into the concentrated Au TOHs solution for 1-15 hours to prepare the gold trioctahedron modification-based capillary SERS substrate.

Further, 2-2.5 mL of 200X 10^-3Mixing M hexadecyl trimethyl ammonium chloride solution with 40-60 mu L of hexadecyl trimethyl ammonium bromide (CTAB) coated gold nanocluster solution, then adding 1.5-3 mL of 0.1M ascorbic acid solution, and adding 2-2.5 mL of 0.5 multiplied by 10 after reactants are completely dissolved^-3M HAuCl₄The solution reacts for 15-20 minutes at 25-30 ℃, and then is centrifuged at 14000-15500 rpm for 20-40 minutes to be re-suspended in 1-2 mL of 20 multiplied by 10^-3Obtaining gold nanosphere seed solution in the M CTAC solution;

further, 0.3 to 0.4g of cetyltrimethylammonium bromide is weighed and dissolved in 4 to 6mL of water, and the mixture is stirred slightly at 25 to 30 ℃. 4-6 mL of 0.5mM HAuCl₄Adding the solution into the solution, and rapidly adding 0.5-0.8 mL of 10 multiplied by 10 at 500-900 rpm^-3M NaBH₄And (3) violently stirring the solution for 1.5-3.0 minutes, and standing at 25-30 ℃ for 2-5 hours to ensure complete reaction to obtain the CTAB coated gold nanocluster solution.

The principle of the invention is as follows: the invention creatively provides a method for preparing the capillary-based SERS substrate by taking Au TOHs as an assembly unit and utilizing electrostatic adsorption by taking the improvement of sensitivity and repeatability of SERS field detection as an entry point and combining the capability of a capillary substrate for drawing samples, solves the problems of difficulty, easy pollution, poor detection repeatability and the like of generally adopting a planar substrate for drawing samples on the field, and prepares the capillary-based SERS substrate with uniform gold trioctahedron modification.

The invention has the beneficial effects that:

(1) the gold trioctahedral assembled on the inner wall of the capillary is densely arranged and uniformly distributed, the detection uniformity can be effectively improved, and the Relative Standard Deviation (Relative Standard development) of continuous detection for 400 times in the vertical direction of the same capillary base SERS substrate can be controlled to be about 4.6%;

(2) regular hot spots exist among the gold trioctahedral assembled on the inner wall of the capillary, the sensitivity of SERS detection can be obviously enhanced, and the optimal assembly concentration is 50 mM;

(3) the larger the size of the gold trioctahedron assembled on the inner wall of the capillary is, the better the SERS enhancement effect is, and the optimal size is 75 nm;

(4) the longer the amino functionalized capillary tube is soaked in the concentrated Au TOHs solution, the higher the density of the nano particles covered on the inner wall of the capillary tube is, and the optimal soaking time is 15 hours;

(5) the granularity of the obtained gold trioctahedral can reach the nanometer level, the average size can reach about 75nm, and the particle size distribution is uniform;

(6) the obtained capillary-based SERS substrate is more convenient for drawing a sample and does not generate background interference on a detection result;

(7) the SERS trace detection has high repeatability, and the detection limit change rate does not exceed 5 percent according to the result obtained by repeating the experiment for two times.

The invention utilizes electrostatic adsorption to lead the gold nano particles assembled on the inner wall of the capillary to be densely arranged and evenly distributed, thereby effectively improving the detection uniformity. More importantly, the amino functionalized capillary is optimally soaked in the concentrated Au TOHs solution for 15 hours, the optimal concentration is 50mM, and the optimal size is 75 nm. Regular hot spots exist among Au TOHs, and meanwhile, the capillary-based SERS substrate is more convenient for drawing samples and is not easy to pollute, so that the repeatability of SERS detection is further improved.

Drawings

FIG. 1 is a transmission electron microscope image of Au TOHs obtained in example 1 of the present invention; as shown in FIG. 1, it is shown that Au TOHs have a trioctahedral structure and an average particle size of 75 nm.

FIG. 2 is a scanning electron micrograph (inner surface of a capillary) of a capillary-based SERS substrate obtained in example 3 of the present invention; as shown in FIG. 2, the inner wall of the capillary is uniformly covered with a layer of Au TOHs.

FIG. 3 is a SERS spectrum of fentanyl in serum detected by the capillary-based SERS substrate obtained in example 3 of the present invention, wherein the detection limit in serum can be as low as 40.63ng/mL (the detection limit is calculated according to the formula specified by International Union of theory and applied chemistry (IUPAC): LOD ═ kS_bM, wherein S_bStandard deviation of blank multiple measurements; m is the slope of the analytical calibration curve in the low concentration range; k is 3).

Fig. 4 is a SERS spectrogram of a fentanyl-doped heroin sample detected by the capillary-based SERS substrate obtained in example 3 of the present invention, wherein the detection limit can be as low as 0.1% by mass concentration.

Detailed Description

Example 1: preparation of gold trioctahedral (Au TOHs)

(1) Preparation of 3nm gold nanocluster solution:

0.365g of cetyltrimethylammonium bromide (CTAB) was weighed out and dissolved in 5mL of water, and stirred gently at 27 ℃.5mL of a 0.5mM chloroauric acid solution (HAuCl)₄) Added to the above solution, and then rapidly added at 600rpm with 0.6mL of 10X 10^- ³M sodium borohydride solution (NaBH)₄) Stirred for 2 minutes and placed at 27 ℃ for 3 hours to ensure complete reaction to obtain a CTAB coated gold nanocluster solution.

(2) Preparing a 10nm gold nanosphere seed solution:

2mL of a suspension (200X 10)^-3Mixing M cetyl trimethyl ammonium chloride solution (CTAC) with 50 μ L of CTAB coated gold nanocluster solution obtained in step (1), and adding 1.5mL of 0.1M ascorbic acidAcid solution (AA), 2mL, 0.5X 10^-3M HAuCl₄The solution was reacted at 27 ℃ for 15 minutes and then resuspended in 1mL of 20X 10 medium by centrifugation at 14500rpm for 30 minutes^-3Obtaining gold nanosphere seed solution with the particle size of 10nm from the M CTAC solution.

(3) Preparation of 75nm Au TOHs:

mixing 2mL of 0.1M CTAC solution with 5 μ L of 10nm gold nanosphere seed solution obtained in step (2), adding 130 μ L of 0.1M AA solution, adding 2mL of 0.5 × 10 AA solution after the reactant is completely dissolved^-3M HAuCl₄The solution is reacted for 15min at the temperature of 27 ℃, centrifuged at 6000rpm for 15min, then the precipitate is dispersed into 0.5mL of polyvinyl pyrrolidone (PVP) ethanol solution with the mass fraction of 1 percent, centrifuged at 6000rpm for 15min, and the precipitate is redispersed in 0.5mL of ethanol to obtain Au TOHs ethanol solution, and the Au TOHs ethanol solution is stored at the temperature of 4 ℃.

Example 2: preparation of amino-functionalized capillaries

(1) Modifying hydroxyl groups:

the glass capillary was first ultrasonically cleaned with a mixed solution of acetone and ethanol (volume ratio 2: 1) for 20 minutes, then rinsed with water and dried. Soaking capillary in piranha solution (H)₂SO₄：H₂O₂4: 1, volume ratio), then rinsed with water and dried, thereby modifying hydroxyl groups on the inner wall of the capillary.

(2) Modified amino group:

soaking the capillary treated in the step (1) in an ethanol solution of APTES with the volume fraction of 1% for 12 hours, then washing with water and drying, thereby modifying amino groups on the inner wall of the capillary.

Example 3: preparation and detection of capillary-based SERS substrate

(1) Preparation of capillary-based SERS substrate:

the amino-functionalized capillary tube treated in example 2 was vertically inserted into the concentrated (150-fold) Au TOHs ethanol solution prepared in example 1 for 15 hours, and the Au TOHs with weak electronegativity was uniformly assembled in the inner wall of the amino-functionalized capillary tube by electrostatic adsorption.

(2) Preparation of fentanyl samples

Fresh human blood was centrifuged at 3000rpm for 10 minutes, and serum was removed and diluted to 10% (by volume) with water. Fentanyl normal saline standard solution (50. mu.g/mL) was diluted with the serum solution to various concentrations (10. mu.g/mL, 5. mu.g/mL, 2. mu.g/mL, 1. mu.g/mL, 500ng/mL, 200ng/mL, 100ng/mL) to prepare fentanyl serum samples 1 at various concentrations.

Then, 1, 10, 50, 100, 200, 500. mu.L of a fentanyl physiological saline solution (5. mu.g/mL) and 50. mu.L of a heroin acetonitrile solution (100. mu.g/mL) were mixed to prepare fentanyl-doped heroin samples 2 having different concentrations of 0.1%, 1%, 5%, 10%, 20%, and 50% by mass fraction, respectively.

(3) Detection of capillary-based SERS substrates

And (3) respectively drawing the fentanyl serum sample 1 with different concentrations and the fentanyl-doped heroin sample 2 with different concentrations in the step (2) by using the capillary-based SERS substrate obtained in the step (1), then placing the samples on a storage rack of a micro-Raman spectrometer, and detecting the samples by using 785nm laser, wherein the results are respectively shown in fig. 3 and fig. 4.

Two strong peaks characteristic to fentanyl, located at 1002cm each, were observed in the spectrum of FIG. 3^-1And 1028cm^-1Here, the intensity of the raman signal is directly proportional to the number of fentanyl molecules adsorbed on the nanoparticles. However, with further increase in fentanyl concentration, the adsorption reached a dynamic equilibrium, while the coverage of the nanoparticles on the inner wall of the capillary remained almost unchanged, resulting in a tendency for the signal intensity to saturate.

In FIG. 4, a characteristic peak of fentanyl at 1002cm can be clearly observed^-1(1028cm^-1The characteristic peak still exists) and the characteristic peak of heroin is 623cm^-1And with the more mass of fentanyl doping, the characteristic peak of fentanyl is 1002cm^-1The characteristic peak of heroin becomes 623cm with increasing strength^-1The product becomes weaker and weaker, and can still be observed to be positioned at 1002cm in a heroin sample with the content of fentanyl doped with 0.1 percent^-1Characteristic peak of (2). Therefore, the detection limit of the SERS platform based on the capillary in serum can be as low as 40.63ng/mL, and the SERS platform can be used for detecting fentanyl in serumIdentifying the fentanyl doped with 0.1 percent of impurity in the heroin.

Claims

1. A preparation method of a capillary-based SERS substrate based on gold trioctahedron modification comprises the following steps:

(1) preparing Au TOHs by using a seed growth method: mixing 2-2.5 mL of 0.1M hexadecyltrimethylammonium chloride solution with 5-20 mu L of 10nm gold nanosphere seed solution, then adding 130-150 mu L of 0.1M ascorbic acid solution, and adding 2-2.5 mL of 0.5 multiplied by 10 after reactants are completely dissolved^-3M HAuCl₄Reacting the solution at 25-30 ℃ for 15-20 min, and centrifuging at 5500-6000 rpm for 10-15 min; dispersing the precipitate into 0.5-1 mL of polyvinylpyrrolidone ethanol solution with the mass fraction of 1%, centrifuging at 5500-6000 rpm for 10-15 minutes, dispersing the precipitate into 0.5-1 mL of ethanol to obtain Au TOHs ethanol solution, and storing at 4 ℃;

(2) amino-functionalized capillaries were prepared with piranha solution and APTES ethanol solution: firstly, mixing the raw materials in a volume ratio of 1-3: 1, ultrasonically cleaning the glass capillary tube for 10-20 minutes by using an acetone and ethanol mixed solution, and then washing with water and drying; soaking the dried capillary tube in the piranha solution for 1-3 hours, and then washing with water and drying, so that hydroxyl is modified on the inner wall of the capillary tube; finally, soaking the capillary tube with the inner wall modified with hydroxyl in an APTES ethanol solution with the volume fraction of 1-10% for 10-15 hours, and then washing with water and drying, thereby modifying amino on the inner wall of the capillary tube;

(3) preparing a capillary-based SERS substrate: concentrating the Au TOHs ethanol solution prepared in the step (1) by 50-200 times, and vertically inserting a capillary with amino groups modified on the inner wall into the concentrated Au TOHs solution for 1-15 hours to prepare a capillary-based SERS substrate modified based on gold trioctahedron;

wherein, Au TOHs is gold trioctahedral, and APTES is 3-aminopropyl triethoxysilane.

2. The preparation method of the gold trioctahedral modification-based capillary-based SERS substrate as claimed in claim 1, wherein: is 2 to 2.5mL、200×10^-3Mixing the M hexadecyl trimethyl ammonium chloride solution with 40-60 mu L of hexadecyl trimethyl ammonium bromide coated gold nanocluster solution, then adding 1.5-3 mL of 0.1M ascorbic acid solution, and adding 2-2.5 mL of 0.5 multiplied by 10 after the reactants are completely dissolved^-3M HAuCl₄The solution reacts for 15-20 minutes at 25-30 ℃, and then is centrifuged at 14000-15500 rpm for 20-40 minutes to be re-suspended in 1-2 mL of 20 multiplied by 10^-3And obtaining gold nanosphere seed solution in the M CTAC solution.

3. The preparation method of the gold trioctahedral modification-based capillary-based SERS substrate as claimed in claim 2, wherein: 0.3-0.4 g of hexadecyl trimethyl ammonium bromide is weighed and dissolved in 4-6 mL of water, and the mixture is stirred slightly at the temperature of 25-30 ℃. 4-6 mL of 0.5mM HAuCl₄Adding the solution into the solution, and rapidly adding 0.5-0.8 mL of 10 multiplied by 10 at 500-900 rpm^-3M NaBH₄And (3) violently stirring the solution for 1.5-3.0 minutes, and standing at 25-30 ℃ for 2-5 hours to ensure complete reaction to obtain the gold nanocluster solution coated by the hexadecyl trimethyl ammonium bromide.

4. A capillary base SERS substrate based on gold trioctahedral modification is characterized in that: is prepared by the method of any one of claims 1 to 3.

5. The use of a gold trioctahedral-based modified capillary-based SERS substrate according to claim 4 for the detection of fentanyl.