A surface-enhanced raman spectroscopy substrate and its preparation method and use

Abstract

The application relates to a surface enhanced Raman spectroscopy (SERS) substrate, and relates to a kit containing the substrate and a method for detecting a fentanyl compound in a sample by using the substrate, and relates to the use of the substrate and the kit for detecting the fentanyl compound in the sample.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of analytical chemistry, and relates to a surface-enhanced Raman spectroscopy (SERS) substrate, and further relates to a kit containing the substrate and a method for detecting fentanyl compounds in a sample by using the substrate, and further relates to the use of the substrate and the kit for detecting fentanyl compounds in a sample. BACKGROUND

[0002] Fentanyl is a typical opioid drug, which is widely used in the medical field due to its good analgesic effect. It is urgent to develop new rapid detection methods and new materials suitable for such detection methods, so as to be suitable for sensitive, specific and rapid detection of fentanyl compounds in complex matrices, including environmental samples and biomedical samples.

[0003] The currently developed rapid detection methods for fentanyl include electrochemical sensor methods (TrAC Trends in Analytical Chemistry 2020, 132, 116037), colorimetric methods (Anal. Chem. 2021, 93, 6544-6550), lateral flow test strip methods (Harm Reduct J 2021, 18(1), 30-39), etc. The basic principle of the electrochemical sensor method is to use voltammetry to cause the irreversible oxidative dealkylation of piperidine tertiary amine to obtain a specific "fingerprint". The electrochemical detection sensor method has the advantages of relatively low cost, easy to carry and fast analysis, but the disadvantage is that the sensor has low sensitivity, and lacks the precise amount required for identifying and accurately quantifying fentanyl compounds. The colorimetric method is based on the halogen bond and hydrogen bond interaction between the specific probe and fentanyl, which produces charge transfer to produce color change, with a sensitivity of sub-mg / mL level, suitable for the detection of water solution, diluted urine and domestic sewage samples. However, its disadvantages include narrow detection range, not obvious color change of fentanyl and poor specificity, etc. The lateral flow test strip method uses immunological analysis method to develop and use antibodies against fentanyl to carry out competitive detection (elimination method), but cannot completely avoid the problem of false positive recognition of structurally similar substances caused by cross recognition of antibodies. Compared with the above methods, surface-enhanced Raman spectroscopy (SERS) has high sensitivity, strong specificity and fast response, and is expected to play a unique advantage in the rapid detection method of fentanyl compounds. For example, Wang Kai et al. used gold nanoparticles (AuNPs) and substances providing I- ions to jointly carry out rapid SERS detection of fentanyl compounds in samples (Electrophoresis 2019, 40, 2193-2203).

[0004] SERS is a high-sensitivity, rapid vibrational spectroscopy technique, which can achieve rapid response to analytes. However, SERS generally requires special substrates and complex micro-nano processing technology, and is only suitable for detection of simple solution or gas systems. In various noble metal materials constituting the SERS substrate, the electromagnetic field strength between adjacent units is significantly enhanced due to the local plasmon resonance (LSPR), and the interval region is the SERS "hot spot" with an optimal interval of 1-2 nm. However, the ordered and precise regulation of SERS "hot spots" has been a difficult problem in the field of SERS. In addition, some typical SERS nanomaterial substrates are prone to non-specific adsorption and even aggregation in a matrix sample environment with high complexity and high complexity, which leads to weakening, instability or disappearance of the characteristic response signal of SERS to analytes. Therefore, how to prepare a high-sensitivity, anti-interference and practical SERS substrate has been a hot and frontier field of SERS research. SUMMARY

[0005] The present application provides a surface-enhanced Raman spectroscopy substrate, comprising or consisting of magnetic particles, a metal-organic framework material and gold nanoparticles (AuNPs), wherein the metal-organic framework material is coated on the surface of the magnetic particles, and the gold nanoparticles are distributed in and / or on the surface of the metal-organic framework material.

[0006] In some embodiments, the metal-organic framework material described in the present application is NH2-MIL-101(Fe), NH2-MIL-101(Cr) or NH2-MIL-101(Al).

[0007] In some embodiments, the magnetic particles described in the present application are carboxyl-modified Fe3O4(Fe3O4-COOH) particles.

[0008] In some embodiments, the magnetic particles described in the present application are spherical particles.

[0009] In some embodiments, the average particle size of the magnetic particles coated with the metal-organic framework material in the present application is 230 ± 20 am.

[0010] The present application also provides a method for preparing the surface-enhanced Raman spectroscopy substrate described in the present application, comprising:

[0011] 1) providing magnetic particles;

[0012] 2) coating a metal-organic framework material on the surface of the magnetic particles by a solvothermal method;

[0013] 3) loading the gold nanoparticles into and / or onto the metal-organic framework material coated on the surface of the magnetic particles by chloroauric acid reduction method;

[0014] In some embodiments, step 2) in the method of the present application comprises mixing the magnetic particles and the raw materials for preparing the metal-organic framework material in a solvent, and reacting.

[0015] In some embodiments, the raw materials for preparing the metal-organic framework material of the present application comprise metal compounds (e.g. FeCl3·6H2O, Cr(NO3)3·9H2O, AlCl3·6H2O), 2-amino terephthalic acid.

[0016] In some embodiments, the solvent of the present application is N,N-dimethylformamide.

[0017] In some embodiments, step 3) in the method of the present application comprises mixing the magnetic particles coated with the metal-organic framework material obtained in step 2), chloroauric acid and a reducing agent to obtain a chloroauric acid reduction system, and reacting.

[0018] In some embodiments, the reducing agent of the present application is sodium borohydride, citrate (e.g. trisodium citrate), tannic acid or ascorbic acid, preferably sodium borohydride.

[0019] In some embodiments, in the chloroauric acid reduction system of the present application, the concentration of chloroauric acid is 0.05-0.15% (w / v) (e.g. 0.08% (w / v), 0.1% (w / v), 0.12% (w / v)), and the concentration of the reducing agent is 0.3-1.6 mM (e.g. 0.5-1.5 mM, 0.6-1.4 mM, 0.7 mM, 0.8 mM, 0.9 mM, 1 mM, 1.1 mM, 1.2 mM, 1.3 mM).

[0020] The present application also provides a kit comprising the surface-enhanced Raman spectroscopy substrate of the present application.

[0021] In some embodiments, the kit of the present application further comprises an agglomeration agent. In some embodiments, the agglomeration agent is a substance capable of providing I- ions, such as NaI, KI.

[0022] In some embodiments, the kit of the present application further comprises bovine serum albumin (BSA).

[0023] In some embodiments, the kit of the present application further comprises NaOH and / or NaCI.

[0024] In some embodiments, the kit of the present application further comprises deuterated fentanyl.

[0025] In certain embodiments, the kits of the present application further comprise one or more microtiter plates.

[0026] In certain embodiments, the kits of the present application further comprise instructions.

[0027] In certain embodiments, the instructions in the kits of the present application describe the methods of the present application for detecting fentanyl compounds and / or morphine compounds.

[0028] In certain embodiments, the instructions in the kits of the present application describe the methods of the present application for detecting morphine compounds.

[0029] The present application also provides the use of the surface-enhanced Raman spectroscopy substrate of the present application for detecting fentanyl compounds in a sample.

[0030] The present application also provides the use of the surface-enhanced Raman spectroscopy substrate of the present application for preparing a kit for detecting fentanyl compounds in a sample.

[0031] The present application also provides the use of the kit of the present application for detecting fentanyl compounds in a sample.

[0032] In certain embodiments, the fentanyl compounds described in the present application can also be referred to as fentanyl substances, which refer to substances having one or more of the following conditions compared with fentanyl: 1. using other acyl groups to replace propionyl; 2. using any substituted or unsubstituted monocyclic aromatic group to replace the phenyl group directly connected to the nitrogen atom; 3. having alkyl, alkenyl, alkoxy, ester, ether, hydroxyl, halogen, halogenated alkyl, amino and nitro groups and the like on the piperidine ring; 4. using other arbitrary groups (except hydrogen) to replace phenethyl. Such compounds include: fentanyl substances listed in the Narcotic Drug List, Psychotropic Drug List and Non-Drug Narcotic and Psychotropic Drug Control List, such as acetyl-alpha-methylfentanyl, alfentanil, alpha-methylfentanyl, alpha-methylthiofentanyl, beta-hydroxyfentanyl, beta-hydroxy-3-methylfentanyl, fentanyl, 3-methylfentanyl, 3-methylthiofentanyl, para-fluorofentanyl, remifentanil, sufentanil, thiofentanyl, acetyl fentanyl (N-(1-phenethylpiperidin-4-yl)-N-phenylacetamide), butyryl fentanyl (N-(1-phenethylpiperidin-4-yl)-N-phenylbutyramide), beta-hydroxythiofentanyl (N-(1-(2-hydroxy-2-(thiophen-2-yl)ethyl)piperidin-4-yl)-N-phenylpropanamide), 4-fluorobutyryl fentanyl (N-(4-fluorophenyl)-N-(1-phenethylpiperidin-4-yl)-butyramide), isobutyryl fentanyl (N-(1-phenethylpiperidin-4-yl)-N-phenylisobutyramide), ohfentanyl (N-(1-phenethylpiperidin-4-yl)-N-phenyl-2-oxo-1-pyrrolidine-5-carboxamide), and the like.(2-Fluorophenyl)-2-methoxy-N-(1-phenethylpiperidin-4-yl)acetamide), propionylfentanyl (N-(1-Phenethylpiperidin-4-yl)-N-phenylpropionamide), carfentanyl (Methyl-4-(N-phenylpropionamido)-1-phenethylpiperidine-4-carboxylate), furanyl fentanyl (N-(1-Phenethylpiperidin-4-yl)-N-phenylfuran-2-carboxamide), valeryl fentanyl (N-(1-Phenethylpiperidin-4-yl)-N-phenylpentanamide), 4-fluoroisobutyryl fentanyl (N-(4-Fluorophenyl)-N-(1-phenethylpiperidin-4-yl)isobutyramide), tetrahydrofuryl fentanyl (N-Phenyl-N-(1-phenethylpiperidin-4-yl)tetrahydrofuran-2-carboxamide), 4-aminophenyl-N-phenethylpiperidine (4-Aminophenyl-N-phenethylpiperidine), N-phenethyl-4-piperidone (N-phenethyl-4-piperidone), and the like.

[0033] In certain embodiments, the fentanyl compound of the present application includes fentanyl, 3-methylfentanyl, carfentanyl, sufentanyl, 4-fluoroisobutyryl fentanyl.

[0034] In certain embodiments, the sample of the present application is blood plasma or urine.

[0035] In certain embodiments, the sample of the present application contains a morphine compound.

[0036] In certain embodiments, the morphine compound of the present application includes morphine, heroin.

[0037] The present application also provides a method for detecting a fentanyl compound in a sample, comprising:

[0038] 1) adding the surface enhanced Raman spectroscopy substrate of any one of claims 1-4 and a group aggregating agent (preferably a substance capable of providing I"ions, such as NaI, KI) to the sample to obtain a mixture;

[0039] 2) collecting the surface enhanced Raman spectroscopic pattern of the mixture obtained in step 1).

[0040] In certain embodiments, the method for detecting fentanyl compounds in a sample according to the present application further comprises: determining whether the fentanyl compounds are present in the sample if the surface enhanced Raman spectroscopic pattern of the mixture has a characteristic peak at a wave number of about 995-1005 cm -1 In certain embodiments, the method for detecting fentanyl compounds in a sample according to the present application further comprises: determining whether the fentanyl compounds are present in the sample if the surface enhanced Raman spectroscopic pattern of the mixture has a characteristic peak at a wave number of about 995-1005 cm

[0041] In certain embodiments, the method for detecting fentanyl compounds in a sample according to the present application further comprises: quantifying the fentanyl compounds in the sample by using the intensity of the characteristic peak.

[0042] In certain embodiments, the method for detecting fentanyl compounds in a sample according to the present application further comprises: adding an internal standard (e.g. deuterated fentanyl) to the sample in step 1). Preferably, the fentanyl compounds are quantified by using the ratio of the intensity of the characteristic peak of the fentanyl compounds to that of the deuterated fentanyl, wherein the characteristic peak of the deuterated fentanyl is at a wave number of about 953-993 cm -1 In certain embodiments, the method for detecting fentanyl compounds in a sample according to the present application further comprises: adding an internal standard (e.g. deuterated fentanyl) to the sample in step 1). Preferably, the fentanyl compounds are quantified by using the ratio of the intensity of the characteristic peak of the fentanyl compounds to that of the deuterated fentanyl, wherein the characteristic peak of the deuterated fentanyl is at a wave number of about 953-993 cm

[0043] In certain embodiments, the method for detecting fentanyl compounds in a sample according to the present application further comprises: pretreating the sample before step 1).

[0044] In certain embodiments, the method for detecting fentanyl compounds in a sample according to the present application, wherein the sample is plasma, further comprises: adding bovine serum albumin (BSA) to the sample in step 1).

[0045] In certain embodiments, the method for detecting fentanyl compounds in a sample according to the present application, wherein the sample is urine, further comprises: pretreating the sample before step 1). Preferably, the pretreatment comprises: adding sodium hydroxide and sodium chloride to the urine, mixing, solid-liquid separation, and taking the supernatant as the sample to be detected.

[0046] The present application also provides a method for detecting fentanyl compounds and / or morphine compounds in a sample, comprising:

[0047] 1) adding the surface enhanced Raman spectroscopic substrate and the aggregating agent (preferably a substance capable of providing I" ions, such as NaI, KI) according to any one of claims 1-4 to the sample to obtain a mixture;

[0048] 2) collecting the surface enhanced Raman spectroscopic pattern of the mixture obtained in step 1).

[0049] In certain embodiments, the method for detecting fentanyl compounds and / or morphine compounds in a sample according to the present application further comprises: determining whether the sample contains fentanyl compounds if the surface-enhanced Raman spectrum of the mixture exhibits a characteristic peak at a wave number of about 995-1005 cm -1 In certain embodiments, the method for detecting fentanyl compounds and / or morphine compounds in a sample according to the present application further comprises: determining whether the sample contains morphine compounds if the surface-enhanced Raman spectrum of the mixture exhibits a characteristic peak at a wave number of about 620-635 cm -1 In certain embodiments, the method for detecting fentanyl compounds and / or morphine compounds in a sample according to the present application further comprises: determining whether the sample contains morphine compounds if the surface-enhanced Raman spectrum of the mixture exhibits a characteristic peak at a wave number of about 620-635 cm

[0050] In certain embodiments, the method for detecting fentanyl compounds and / or morphine compounds in a sample according to the present application further comprises: quantifying the fentanyl compounds and / or morphine compounds in the sample by using the intensity of the characteristic peaks.

[0051] In certain embodiments, the method for detecting fentanyl compounds and / or morphine compounds in a sample according to the present application further comprises: adding an internal standard (e.g. deuterated fentanyl) to the sample in step 1). Preferably, the fentanyl compounds and / or morphine compounds are quantified by using the intensity ratio of the characteristic peaks of the fentanyl compounds and / or morphine compounds to the characteristic peak of the deuterated fentanyl, wherein the characteristic peak of the deuterated fentanyl is at a wave number of about 953-993 cm -1 In certain embodiments, the method for detecting fentanyl compounds and / or morphine compounds in a sample according to the present application further comprises: adding an internal standard (e.g. deuterated fentanyl) to the sample in step 1). Preferably, the fentanyl compounds and / or morphine compounds are quantified by using the intensity ratio of the characteristic peaks of the fentanyl compounds and / or morphine compounds to the characteristic peak of the deuterated fentanyl, wherein the characteristic peak of the deuterated fentanyl is at a wave number of about 953-993 cm

[0052] In certain embodiments, the method for detecting fentanyl compounds and / or morphine compounds in a sample according to the present application further comprises: pretreating the sample before step 1).

[0053] In certain embodiments, the method for detecting fentanyl compounds and / or morphine compounds in a sample according to the present application, wherein the sample is plasma, further comprises: adding bovine serum albumin (BSA) to the sample in step 1).

[0054] In certain embodiments, the method for detecting fentanyl compounds and / or morphine compounds in a sample according to the present application, wherein the sample is urine, further comprises: pretreating the sample before step 1). Preferably, the pretreatment comprises: adding sodium hydroxide and sodium chloride to the urine, mixing, solid-liquid separation, and taking the supernatant as the sample to be detected.

[0055] In the present application, the unit "% (w / v)" means mass percentage, for example, 0.08% (w / v) means 0.08 g of solute per 100 mL of solution.

[0056] In the present invention, the term "and / or" when used in a list of two or more items, means that any one of the listed items can be employed by itself, or any combination of two or more of the listed items can be employed. For example, if a composition is described as containing or not containing components A, B, and / or C, the composition can contain or not contain A alone; B alone; C alone; a combination of A and B; a combination of A and C; a combination of B and C; or a combination of A, B, and C. As another example, "nanogold particles are distributed in and / or on the surface of the metal organic framework material" means that the nanogold particles can be distributed in the metal organic framework material, on the surface of the metal organic framework material, or both in the metal organic framework material and on the surface of the metal organic framework material.

[0057] In the present invention, the term "surface" includes inner and outer surfaces, for example, "the surface of the metal organic framework material" includes the outer surface and / or the inner surface of the metal organic framework material.

[0058] In the present invention, the term "about" is understood to be within the normal tolerance range of the art, for example, about can be understood to be within 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.1%, 0.05%, or 0.01% of the stated value. In addition, unless otherwise apparent from context, all numerical values provided herein are modified by the term "about."

[0059] Advantages of the present application

[0060] The SERS substrate provided by the present invention has one or more of the following advantages:

[0061] 1) The SERS substrate provided by the present invention can be used for qualitative or quantitative detection of fentanyl compounds, with short detection time (e.g., the detection time can be shortened to 1-2 min) and high detection efficiency;

[0062] 2) The SERS substrate provided by the present invention can be used for qualitative or quantitative detection of trace amounts of fentanyl compounds in morphine compound (including morphine and heroin) samples;

[0063] 3) The SERS substrate provided by the present invention can be used for qualitative or quantitative detection of fentanyl compounds in blood plasma and urine;

[0064] 4) The SERS substrate provided by the present invention is resistant to acid and alkali, and can remain stable in samples with a pH value of 4-12, with no significant change in SERS signal intensity when used for detection of fentanyl compounds;

[0065] 5) The SERS substrate provided by the application is salt-resistant, can remain stable in a sodium chloride sample with a concentration ranging from 10nM to 1000nM, and has no obvious change in SERS signal intensity when used for detecting fentanyl compounds;

[0066] 6) The SERS substrate provided by the application can remain stable within 24 hours, and has no obvious change in SERS signal intensity when used for detecting fentanyl compounds;

[0067] 7) The SERS substrate provided by the application can be applied to the detection of fentanyl compounds in complex biological samples.

[0068] The detection method provided by the application is sensitive, rapid, easy to operate, and can be used in many fields such as on-site rapid detection, poison detection, and forensic identification. BRIEF DESCRIPTION OF DRAWINGS

[0069] Figure 1-A A transmission electron microscope (TEM) image of Fe3O4-COOH prepared for Example 1 of the application;

[0070] Figure 1-B A transmission electron microscope (TEM) image of Fe3O4-COOH@NH2-MIL-101 prepared for Example 1 of the application; s

[0071] Figure 1-C A transmission electron microscope (TEM) image of Fe3O4-COOH@NH2-MIL-101@AuNPs prepared for Example 1 of the application;

[0072] Figure 2 An X-ray diffraction (XRD) spectrum of the SERS substrate material prepared for Example 1 of the application;

[0073] Figure 3 A Fourier infrared (FT-IR) spectrum of the SERS substrate material prepared for Example 1 of the application;

[0074] Figure 4 A hysteresis curve of the SERS substrate material prepared for Example 1 of the application measured by a vibrating sample magnetometer (VSM);

[0075] Figure 5 A SERS spectrum of the SERS substrate material prepared by using different concentrations of sodium borohydride for detecting fentanyl;

[0076] Figure 6-A SERS signal enhancement effects of different types of aggregating agents on fentanyl;

[0077] Figure 6-B SERS signal enhancement effects of different concentrations of NaI on fentanyl; ​

[0078] Figure 7 SERS spectra of fentanyl detected by using the SERS substrate material prepared in Example 1 of the present application;

[0079] Figure 8-A SERS spectra of trace fentanyl in a complex system with heroin as the main compound;

[0080] Figure 8-B SERS spectra of trace fentanyl in a complex system with morphine as the main compound;

[0081] Figure 9-A SERS spectra of fentanyl C Raman SERS spectra of 100 μg / mL;

[0082] Figure 9-B SERS spectra of fentanyl C SERS SERS spectra of 20 ng / mL;

[0083] Figure 10-A SERS spectra of fentanyl detected by using the SERS substrate material prepared in Example 1 of the present application under different pH environments;

[0084] Figure 10-B SERS spectra of fentanyl detected by using the SERS substrate material prepared in Example 1 of the present application under different concentrations of sodium chloride;

[0085] Figure 11 Intra-batch stability test results of the SERS substrate material prepared in Example 1 of the present application;

[0086] Figure 12 Linear standard curve of fentanyl in plasma when deuterated fentanyl is used as an internal standard;

[0087] Figure 13 Linear standard curve of fentanyl in urine when deuterated fentanyl is used as an internal standard;

[0088] Figure 14 Comparison results of the SERS substrate material prepared in Example 1 of the present application and 60 nm AuNPs. DETAILED DESCRIPTION

[0089] The essential content of the present application will be further described in combination with specific examples of the present application. It should be understood that the following examples are only used to illustrate the present application, but do not limit the protection scope of the present application. The specific conditions not mentioned in the following examples are carried out according to the conventional conditions or the manufacturer's recommendations. The drugs or reagents not mentioned by the manufacturer are all conventional products that can be obtained by purchase.

[0090] Although many of the materials and methods used in the following examples are well known in the art, the present application is described in as much detail as possible without unnecessarily obscuring the application. It will be apparent to those skilled in the art that materials and procedures of the following examples are well known in the art, unless otherwise indicated.

[0091] The portable Raman spectrometer used in the following examples is a model i-Raman portable Raman spectrometer (B&W TEK, DE, USA).

[0092] Example 1 Preparation of SERS substrate material

[0093] The SERS substrate material prepared in this example is named Fe3O4-COOH@NH2- MIL-101@AuNPs, and the preparation method steps are as follows:

[0094] (1) Synthesis of Fe3O4-COOH: Fe3O4-6H2O (1.0812 g), trisodium citrate dihydrate (0.2 g) were weighed and dissolved in 20 mL of ethylene glycol, and ultrasonic mixing was performed for 30 min until the reactants were completely dissolved. Anhydrous sodium acetate (1.2 g) was then added, and ultrasonic mixing was performed for 30 min until the solution became a dark yellow viscous liquid. The solution was then transferred to a high-pressure reaction kettle, and the reaction temperature was set to 200°C for 8 h. After the reaction cooled to room temperature, it was transferred to a 250 mL beaker, and the reaction liquid was washed three times with anhydrous ethanol and ultrapure water under the action of a magnet. The product was collected after magnetic separation. The obtained black-brown liquid was placed in a vacuum drying oven at 60°C for 6 h to obtain Fe3O4-COOH, which is a carboxyl-modified Fe3O4.

[0095] (2) Synthesis of Fe3O4-COOH@NH2-MIL-101: Fe3O4-COOH (200 mg), FeCl3-6H2O (0.8448 g), and 2-amino terephthalic acid (0.543 g) were dissolved in N,N-dimethylformamide (20 mL) and ultrasonic mixing was performed for 20 min until complete dissolution. The reaction was then transferred to a 100 mL polytetrafluoroethylene-lined reaction kettle, and the reaction was carried out at 110°C in an oven for 20 h. After the reaction cooled to room temperature, it was washed twice with N,N-dimethylacetamide, ultrapure water, and anhydrous ethanol under the action of a magnet. The product was collected after magnetic separation, and vacuum drying was performed at 60°C for 6 h to obtain a dark brown Fe3O4-COOH@NH2-MIL-101 powder.

[0096] (3) Synthesis of Fe3O4-COOH@NH2-MIL-101@AuNPs: 30 mg of Fe3O4-COOH@NH2-MIL-101 was suspended in aqueous tetrachloroauric acid trihydrate solution (30 mL, 0.08% (w / v, based on tetrachloroauric acid trihydrate)), ultrasonically mixed for 15 min, stirred at 0 °C for 1 h, 500 μL of freshly prepared aqueous sodium borohydride (80 mM) solution was added, and the stirring was continued in an ice bath for 1 h, and the solution color changed to purple red. The synthesized Fe3O4-COOH@NH2-MIL-101@AuNPs nanoparticles were centrifuged and washed with ultrapure water for three times under the action of a magnet, and then redissolved in 30 mL of ultrapure water and placed at room temperature for use.

[0097] Example 2 Characterization of SERS substrate material

[0098] In this example, the microstructure, crystal lattice, and physical and chemical properties of the SERS substrate material prepared in Example 1 were characterized by Fourier infrared spectroscopy (XRD), transmission electron microscopy (TEM), X-ray diffraction (XRD), and vibrating sample magnetometer (VSM).

[0099] 2.1 Electron microscopy characterization

[0100] The microstructure of the sample was observed using a cryo-TEM (Talos L120C G2) transmission electron microscope. After ultrasonic dispersion, the sample was dropped onto a 200-mesh carbon-coated copper grid, and the operating voltage was 120 kV. After vacuum drying, TEM testing was performed. The microstructure of Fe3O4-COOH, Fe3O4-COOH@NH2-MIL-101, and Fe3O4-COOH@NH2-MIL-101@AuNPs was obtained, as shown in Figure 1-A , 1-B , 1-C.

[0101] The TEM image shows that the Fe3O4-COOH (carboxyl-functionalized Fe3O4) nanoparticles are regular spheres with a diameter of about 160 nm, as shown in Figure 1-A . After loading NH2-MIL-101 on Fe3O4-COOH, the average diameter of Fe3O4-COOH@NH2-MIL-101 increased to about 220 + / - 20 nm, and the shell thickness was about 30 nm, as shown in Figure 1-B . After loading AuNPs, uniformly sized and dispersed Au nanoparticles were found to be embedded in NH2-MIL-101, as shown in Figure 1-C .

[0102] 2.2 Powder X-ray diffraction characterization

[0103] The crystal structure of the SERS substrate material prepared in Example 1 was tested by a Rigaku Smart Lab 3kW X-ray diffractometer to analyze the material composition, sample structure and morphology, the tube voltage was 40 kV, the tube current was 40 mA, the scanning speed was 8° / min, the scanning step was 0.02, and the 2θ was 3°-80°.

[0104] The XRD patterns of Fe3O4-COOH, Fe3O4-COOH@NH2-MIL-101 and Fe3O4-COOH@NH2-MIL-101@AuNPs are shown in FIG. 2. Figure 2 The diffraction peaks of the carboxyl-functionalized Fe3O4 microspheres at 2θ = 30.07°, 35.39°, 43.14°, 53.47°, 57.01°, 62.55° were in good agreement with JCPDS 75-1609 (Fe3O4). After loading NH2-MIL-101, the characteristic peaks of NH2-MIL-101 appeared. After loading AuNPs, the diffraction peaks at 2θ = 38.07°, 44.34°, 64.51° and 77.57° corresponded to the crystal faces of Au(111), Au(200), Au(220) and Au(311) respectively, proving the successful synthesis of the magnetic substrate.

[0105] 2.3 Fourier infrared characterization

[0106] The functional group information of the SERS substrate material prepared in Example 1 was analyzed by infrared characterization using VERTEX70. After drying at 60°C, the sample was mixed with potassium bromide and pressed into a semitransparent wafer. The blank potassium bromide wafer was used as a blank, and the test wave number range was 4000cm -1 -450cm -1 .

[0107] The FT-IR of Fe3O4-COOH, Fe3O4-COOH@NH2-MIL-101 and Fe3O4-COOH@NH2-MIL-101@AuNPs is shown in FIG. 3. Figure 3 The peak at 582cm -1 was attributed to Fe-O stretching vibration, the peaks at 1390cm -1 and 1049cm -1 were the vibration peaks of -COOH, proving that a large number of carboxyl groups were modified on the surface of Fe3O4 microspheres. After loading NH2-MIL-101, the peak at 1576cm -1 corresponded to C=C vibration, the peak at 1380cm -1 corresponded to -O-C-O- vibration of carboxyl, and the peak at 1253cm -1C-N stretching vibration peak, indicating that NH2-MIL-101 was successfully loaded on Fe3O4-COOH. After loading gold nanoparticles, the peak position of the material was not found to be changed, proving that loading gold nanoparticles on the composite material would not affect the original framework of the material.

[0108] 2.4 Magnetic property characterization

[0109] The magnetic properties of the SERS substrate material prepared in Example 1 were studied using a vibrating sample magnetometer (VSM) model 7404. The magnetic hysteresis curve of the sample was tested at 300 K, as shown in Figure 4 .

[0110] The magnetic hysteresis curves of Fe3O4-COOH, Fe3O4-COOH@NH2-MIL-101 and Fe3O4-COOH@NH2-MIL-101@AuNPs are shown in Figure 4 . The corresponding saturation magnetization (MS) of Fe3O4-COOH and Fe3O4-COOH@NH2-MIL-101@AuNPs were 56.65, 48.44 and 21.35 emu / g, respectively, showing obvious magnetic properties. Compared with Fe3O4-COOH, the MS value of COOH@NH2-MIL-101@AuNPs was lower, which might be due to the presence of metal-organic framework (MOF) shell and AuNPs. Under the action of an external magnetic field, Fe3O4-COOH@NH2-MIL-101@AuNPs could be quickly magnetically separated within 30 s.

[0111] Example 3 Effect of SERS substrate materials prepared with different concentrations of reducing agent on the SERS signal intensity of fentanyl

[0112] Referring to the method of Example 1, and adjusting the concentration of sodium borohydride solution in step (3), SERS substrate materials were prepared, and the SERS signal intensity of fentanyl was detected using the obtained SERS substrate materials.

[0113] The detection method was as follows: 200 μL of SERS substrate material solution prepared in Example 1 (concentration about 1 mg / mL) was taken, the supernatant was discarded after magnetic separation, 180 μL of fentanyl aqueous solution with a concentration of 500 ng / mL was added, and 20 μL of NaI aqueous solution (2 M) was added after mixing, so that the final concentration of NaI in the sample was 200 mM, and the sample was mixed. SERS detection was performed using a portable Raman spectrometer, the laser power was set to 90 mW, the integration time was set to 10 s, and each sample was averaged for 3 parallel acquisitions (n = 3).

[0114] The SERS signals generated when SERS substrate materials prepared with different concentrations of sodium borohydride solution were used to detect fentanyl are shown inFigure 5 The results show that the prepared SERS substrate material can produce fentanyl signal when the concentration of sodium borohydride is 20-100 mM, and the prepared SERS substrate material produces the strongest fentanyl signal when the concentration of sodium borohydride is 80 mM.

[0115] Example 4: Influence of inorganic salt type and concentration on fentanyl SERS signal enhancement effect

[0116] Take 200 μL of the SERS substrate material solution prepared in Example 1 (the concentration is about 1 mg / mL), discard the supernatant after magnetic separation, and obtain about 0.2 mg of SERS substrate material for standby. Take 9 portions of fentanyl aqueous solution with a concentration of 500 ng / mL, each with a volume of 180 μL, and add about 0.2 mg of the above SERS substrate material to each sample, mix, and then add KCl, MgCl2, NaCl, MgSO4, Na2SO4, Na2S, NaBr, KI, and NaI aqueous solutions to each sample, respectively. The final concentrations of KCl, MgCl2, NaCl, MgSO4, Na2SO4, Na2S, NaBr, KI, and NaI in each sample are 200 mM, respectively, mix, and use a portable Raman spectrometer for SERS detection. The laser power is set to 90 mW, the integration time is set to 10 s, and each sample is averaged and collected in parallel for 3 times (n=3).

[0117] The aggregation of metal nanoparticles induced by inorganic salts can produce Raman signal enhancement. The SERS enhancement effect of 9 kinds of inorganic salts on fentanyl is shown in Figure 6-A The results show that the introduction of I- can quickly enhance the response of fentanyl. After the introduction of iodine ions, Au-I bonds can be quickly adsorbed on the Au surface. At the same time, since the fentanyl molecule has two tertiary amine structures, it makes fentanyl closer to the negatively charged Au surface more quickly, and at the same time avoids the interference of other analytes.

[0118] Take 200 μL of the SERS substrate material solution prepared in Example 1 (the concentration is about 1 mg / mL), discard the supernatant after magnetic separation, and obtain about 0.2 mg of SERS substrate material for standby. Take 5 portions of fentanyl aqueous solution with a concentration of 500 ng / mL, each with a volume of 180 μL, and add about 0.2 mg of SERS substrate material to each sample, mix, and then add NaI aqueous solution to each sample to make the final concentration of NaI in each detection sample be 50 mM, 100 mM, 200 mM, 300 mM, or 400 mM, respectively, mix, and use a portable Raman spectrometer for SERS detection. The laser power is set to 90 mW, the integration time is set to 10 s, and each sample is collected in parallel for 3 times (n=3). The detection results are shown in Figure 6-BAs shown in the figure, the intensity of the Raman peak of fentanyl is the highest when the concentration of NaI in the sample is 200 mM.

[0119] Example 5 Detection of fentanyl compounds by the SERS substrate material prepared in the present application

[0120] This example verifies that the SERS substrate material prepared in the present application can also be used for detection of fentanyl compounds. The analytes include fentanyl compounds: fentanyl, 3-methylfentanyl, carfentanyl, sufentanyl, 4-fluoroisobutyrylfentanyl.

[0121] Take 200 μL of the SERS substrate material solution prepared in Example 1 (the concentration is about 1 mg / mL), and discard the supernatant after magnetic separation to obtain about 0.2 mg of the SERS substrate material for standby. Take 180 μL of the analyte aqueous solution with a concentration of 500 ng / mL, add about 0.2 mg of the SERS substrate material, and then add NaI aqueous solution to each sample to make the final concentration of NaI in each detection sample about 200 mM, mix well, and use a portable Raman spectrometer for SERS detection. The laser power is set to 90 mW, the integration time is set to 10 s, and each sample is collected in parallel for 3 times (n=3). The results are shown in Figure 7 .

[0122] The results show that the electrostatic and π-π interactions between the SERS substrate material prepared in the present application and the analyte can quickly position the target analyte in the hotspot area to produce signal enhancement.

[0123] Example 6 Detection of trace fentanyl in a complex system by the SERS substrate material prepared in the present application

[0124] This example verifies that the SERS substrate material prepared in the present application can be used for SERS selective detection of trace fentanyl when heroin and morphine are the main components.

[0125] The concentration of fentanyl is 50 ng / mL, and the concentration ratio of fentanyl to heroin or morphine is 1:1, 1:5, 1:10, 1:50, 1:100, and 1:500. Add about 0.2 mg of the SERS substrate material prepared in Example 1 to 180 μL of the fentanyl / heroin or morphine mixture aqueous solution, and then add NaI aqueous solution to make the final concentration of NaI in the detection sample about 200 mM, mix well, and use a portable Raman spectrometer for SERS detection to collect the spectrum. The laser power is set to 90 mW, the integration time is set to 10 s, and each sample is collected in parallel for 3 times (n=3). The results are shown in Figure 8-A and 8-B .

[0126] Since fentanyl is often adulterated in heroin and morphine, this example tests for trace amounts of fentanyl in heroin and morphine. As shown in Figure 3, while the presence of heroin inhibits detection of the fentanyl SERS signal, as low as 0.2% fentanyl can be detected in a heroin matrix. Figure 8-A As shown in Figure 4, in fentanyl detection in a morphine matrix, as low as 0.05% fentanyl can be detected. These results demonstrate that the SERS substrate material of the present application can be used to detect trace amounts of fentanyl in complex systems. Figure 8-B As shown in Figure 4, in fentanyl detection in a morphine matrix, as low as 0.05% fentanyl can be detected. These results demonstrate that the SERS substrate material of the present application can be used to detect trace amounts of fentanyl in complex systems.

[0127] Example 7 Analytical Enhancement Factor (AEF) of the SERS substrate material prepared in the present application

[0128] The AEF of the SERS substrate material prepared in Example 1 was calculated using the formula Under the same experimental conditions, the ratio between the SERS intensity (I SERS = 20 ng / mL; C Raman = 100 μg / mL) and the normal Raman intensity (I Figure 9-B ) of fentanyl was calculated. Figure 9-A

[0129] To 180 μL of the sample to be tested (fentanyl concentration of 20 ng / mL), about 0.2 mg of the SERS substrate material prepared in Example 1 was added, and an aqueous NaI solution was added to make the final concentration of NaI in the sample 200 mM, and mixed well. The SERS detection was performed using a portable Raman spectrometer. To 180 μL of the sample to be tested (fentanyl concentration of 100 μg / mL), an aqueous NaI solution was added to make the final concentration of NaI in the sample 200 mM, and mixed well. The normal Raman detection was performed directly using a portable Raman spectrometer. The results are shown in Figures 5 and 6. Figure 9-A 9-B It was calculated that the AEF of fentanyl at 1000 cm -1 was 2.58 x 10 5 using the SERS substrate material prepared in Example 1.

[0130] Example 8 Acid, base and salt resistance of the SERS substrate material prepared in the present application

[0131] Since fentanyl detection is often required in complex field environments, and gold nanoparticles are prone to aggregation in acid, base and salt solutions, which can result in unstable SERS signals, in this experiment, 60 nm AuNPs were used as a control to test the fentanyl SERS signals of the SERS substrate material prepared in Example 3 in different pH and different concentrations of sodium chloride salt solutions.

[0132] ​​The detection method is as follows: 9 portions of 500 ng / mL fentanyl solution with pH of 4, 5, 6, 7, 8, 9, 10, 11, and 12 are taken, each portion has a volume of 180 μL, about 0.2 mg of the SERS substrate material prepared in Example 1 is added to each sample, and then the samples are mixed, 20 μL of NaI aqueous solution (2M) is added to each sample, and then the samples are mixed, SERS detection is performed by using a portable Raman spectrometer, the laser power is set to 90 mW, the integration time is set to 10 s, and each sample is collected in parallel for 3 times (n = 3). The detection results are shown in Figure 10-A .

[0133] 8 portions of 500 ng / mL fentanyl sodium chloride solution with sodium chloride concentrations of 10 mM, 20 mM, 50 mM, 100 mM, 140 mM, 300 mM, 700 mM, and 1000 mM are taken, each portion has a volume of 180 μL, about 0.2 mg of the SERS substrate material prepared in Example 1 is added to each sample, and then the samples are mixed, 20 μL of NaI aqueous solution (2M) is added to each sample, and then the samples are mixed, SERS detection is performed by using a portable Raman spectrometer at 0 min and 30 min, respectively, the laser power is set to 90 mW, the integration time is set to 10 s, and each sample is collected in parallel for 3 times (n = 3). The detection results are shown in Figure 10-B .

[0134] As shown in Figure 10-A , when the pH is 4-12, the SERS substrate material prepared by the application can detect the characteristic peak signal of the analyte. As shown in Figure 10-B , the SERS substrate material prepared by the application can remain stable in different concentrations of sodium chloride solution (10 mM, 20 mM, 50 mM, 100 mM, 140 mM, 300 mM, 700 mM, and 1000 mM) within 30 min, and has no obvious effect on the SERS signal enhancement of fentanyl, which indicates that the material can be applied to complex biological detection.

[0135] Example 9 Batch stability of the SERS substrate material prepared by the application

[0136] The stability and reproducibility of the Raman substrate are directly related to the reliability of SERS analysis. In this embodiment, the repeated SERS spectra of the same batch of SERS substrate material to the same concentration of fentanyl are tested, as shown in Figure 11 , the relative standard deviation (RSD) is 6.12% < 15%. The reliability of the detection results is proved.

[0137] Example 10 Drawing of a linear standard curve of fentanyl in a plasma sample

[0138] In the quantitative detection of samples, due to uncontrollable influences such as laser fluctuation, on-site environment, and physicochemical properties of the substrate, quantitative detection of analysis becomes difficult. The addition of an internal standard can effectively avoid the interference of complex factors on the analysis results. Deuterated fentanyl was selected as the internal standard to establish a SERS method for detecting fentanyl. The characteristic peak of deuterated fentanyl at 958 cm -1 was used as the relative intensity of the characteristic peak intensity ratio of fentanyl at 1000 cm -1 , and a standard curve was drawn.

[0139] Because the plasma contains macromolecular substances such as proteins and salts, bovine serum albumin (BSA) was added to avoid the interference of macromolecules in the plasma.

[0140] Fentanyl stock solutions with different concentration gradients were taken, 70 μL of 1 μL / mL deuterated fentanyl solution, 35 μL of 10% (w / v) BSA solution, 200 μL of fentanyl plasma samples with different concentrations were prepared by adding different volumes of plasma, 180 μL of fentanyl plasma samples with different concentration gradients were taken, 20 μL of 2M NaI aqueous solution was added, and the mixture was mixed well. SERS detection was performed using a portable Raman spectrometer, the laser power was set to 90 mW, the integration time was set to 10 s, and each sample was collected in parallel for 3 times (n = 3).

[0141] The relative intensity of the characteristic peak of deuterated fentanyl at 958 cm -1 was used as the relative intensity of the characteristic peak intensity ratio of fentanyl at 1000 cm -1 , and a standard curve was drawn, as shown in Figure 12 , the linear range was 20 ng / mL-500 ng / mL, the correlation coefficient (R2) was 0.9823. The recovery rate was 83.23%-133.12%, and all RSD values were less than 15%.

[0142] Example 11: Preparation of a linear standard curve for fentanyl in urine samples

[0143] 70 μL of Wistar male mouse urine was taken, 35 μL of 10 μL / mL deuterated fentanyl solution, 70 μL of 10% (w / v) NaOH solution, and 70 μL of 5M NaCl solution were added. 700 μL of fentanyl urine samples with different concentration gradients were prepared by adding different volumes of ultrapure water, and centrifuged at 14000 rpm for 10 min. The supernatant was taken for standby.

[0144] Take 180 μL supernatant sample, add about 0.2 mg of SERS substrate material prepared in Example 1 (the concentration of SERS substrate material solution is about 1 mg / mL, take 200 μL supernatant after magnetic separation and discard the supernatant to leave the precipitate), mix well, add NaI aqueous solution to make the final concentration 200 mM, mix well, use portable Raman spectrometer for SERS measurement, set the laser power to 90 mW, set the integration time to 10 s, collect SERS spectrum, collect 3 times for each sample in parallel (n=3).

[0145] Because urine contains a large amount of urea, which greatly interferes with the signal peak of fentanyl, dilute the urine ten times as an analyte, and use sodium hydroxide and sodium chloride solution to pretreat the urine. The relative intensity of the characteristic peak of deuterated fentanyl at 958 cm -1 to the characteristic peak of fentanyl at 1000 cm -1 is the ordinate, and the fentanyl concentration is the abscissa. Draw the standard curve as shown in Figure 13 , the linear range is 20 ng / mL-500 ng / mL, and the correlation coefficient (R 2 ) is 0.9633. The recovery rate is 88.77%-139.06%, and all RSD values are less than 15%.

[0146] Example 12 Influence of SERS substrate material prepared in the application and AuNPs substrate on the SERS detection intensity of fentanyl

[0147] Because typical SERS substrates such as AuNPs are prone to non-specific adsorption or even aggregation in a multi-morphology and high-complexity matrix sample environment, which leads to the weakening, instability, or disappearance of the characteristic response signal of SERS to the analyte. In this example, 60 nm AuNPs (which can be prepared according to Anal. Chem 2007, 79, 4215-4221) are used as a control to investigate the influence of the SERS detection intensity of fentanyl on the response stability of fentanyl to AuNPs substrate.

[0148] Take 1 mL of 60 nm AuNPs particle solution (the concentration is about 0.05 nM) in a centrifuge tube, centrifuge at 6000 rpm for 10 min, discard the supernatant, add 180 μL of 500 μg / mL fentanyl solution, mix well, then add 20 μL of NaI aqueous solution (2 M) to make the final concentration of NaI in the sample 200 mM, mix well, use a portable Raman spectrometer to perform SERS detection and collect spectrum at 0.5 h, 1 h, 10 h, 12 h, and 24 h, respectively, set the laser power to 90 mW, set the integration time to 10 s, collect 3 times for each sample in parallel (n=3).

[0149] Take 200 μL of the SERS substrate material prepared in Example 1 (concentration of about 1 mg / mL), after magnetic separation, discard the supernatant, add 180 μL of a fentanyl aqueous solution of 500 ng / mL, mix, then add 200 μL of a NaI aqueous solution (2M), so that the final concentration of NaI in the sample is 200 mM, mix, and use a portable Raman spectrometer to perform SERS detection at 0.5 h, 1 h, 10 h, 12 h, and 24 h, respectively, to collect the spectrum, the laser power is set to 90 mW, the integration time is set to 10 s, and each sample is averaged and collected in parallel for 3 times (n = 3).

[0150] The SERS substrate material prepared in the present application can protect the AuNPs from easy aggregation due to the presence of the MOFs shell. Figure 14 As shown in FIG. 6, compared with AuNPs, the SERS substrate material prepared in the present application can remain stable within 24 h, and the fentanyl signal in the sample is stable, and can be detected within 24 h.

[0151] Example 13 Batch stability of the SERS substrate material prepared in the present application

[0152] Three batches of SERS substrate materials were synthesized by the method of the reference example, and the material batch recovery rate was verified by the plasma standard curve in Reference Example 10. Each batch of samples was tested on the first day, the second day, and the third day, and the recovery rate was calculated, and the results are shown in the following table.

[0153]

[0154]

[0155]

[0156] The results show that the recovery rate of the three batches of samples is 85.35%-144.64%, and all RSDs are <15%, which proves the stability of the SERS substrate material prepared in the present application.

[0157] Finally, it should be noted that: the above examples are only used to illustrate the technical solutions of the present application and not to limit them; although the present application has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the specific embodiments of the present application can be modified or some technical features can be replaced by equivalent ones without departing from the spirit of the technical solutions of the present application, and they should be covered in the technical solution range of the present application.

Claims

1. A method for preparing a surface-enhanced Raman spectroscopy substrate, comprising: 1) Provide magnetic particles, wherein the magnetic particles are carboxyl-modified Fe3O4 (Fe3O4-COOH) particles; 2) A metal-organic framework material is coated on the surface of magnetic particles using a solvothermal method. The metal-organic framework material is NH2-MIL-101(Fe), NH2-MIL-101(Cr), or NH2-MIL-101(Al). 3) Mix the magnetic particles coated with metal-organic framework material obtained in step 2), chloroauric acid and reducing agent to obtain chloroauric acid reduction system, react, and load the gold nanoparticles into the metal-organic framework material coated on the surface of the magnetic particles and / or onto the surface. The reducing agent is sodium borohydride, and in the chloroauric acid reduction system, the concentration of chloroauric acid is 0.05~0.15 w / v, and the concentration of the reducing agent is 0.6~1.4 mM.

2. The method according to claim 1, wherein the magnetic particles are spherical particles.

3. The method according to claim 1, wherein the average particle size of the magnetic particles coated with metal-organic framework material obtained in step 2) is 230±20 nm.

4. The method of claim 1, wherein step 2) comprises: The magnetic particles and the raw materials for preparing metal-organic framework materials are mixed in a solvent and reacted.

5. The method according to claim 4, wherein the raw materials for preparing the metal-organic framework material include metal compounds and 2-aminoterephthalic acid.

6. The method according to claim 5, wherein the metal compound is FeCl3·6H2O, Cr(NO3)3·9H2O or AlCl3·6H2O.

7. The method according to claim 4, wherein the solvent is N,N-dimethylformamide.

8. The method according to any one of claims 1-7, wherein in the chloroauric acid reduction system, the concentration of chloroauric acid is 0.08 w / v%, 0.1 w / v%, or 0.12 w / v, and the concentration of the reducing agent is 0.7 mM, 0.8 mM, 0.9 mM, 1 mM, 1.1 mM, 1.2 mM, or 1.3 mM.

9. A surface-enhanced Raman spectroscopy substrate, prepared by the method described in any one of claims 1-8.

10. A kit comprising the surface-enhanced Raman spectroscopy substrate of claim 9.

11. The kit of claim 10, further comprising the ability to provide I - The substance.

12. The kit according to claim 11, wherein the reagent is capable of providing I - The substance is NaI or KI.

13. The kit according to claim 10, further comprising bovine serum albumin.

14. The kit according to claim 10, further comprising NaOH and / or NaCl.

15. The kit according to any one of claims 10-14, further comprising deuterated fentanyl.

16. The kit according to any one of claims 10-14, further comprising one or more titration plates.

17. The use of the surface-enhanced Raman spectroscopy substrate according to claim 9 or the kit according to any one of claims 10-16 in the detection of fentanyl-like compounds and / or morphine-like compounds in a sample.

18. The application according to claim 17, wherein the fentanyl compound refers to a substance whose chemical structure, compared to fentanyl, meets one or more of the following conditions:

1. Replace the propionyl group with other acyl groups; 2. Replace the phenyl group directly attached to the nitrogen atom with any substituted or unsubstituted monocyclic aromatic group; 3. The piperidine ring has alkyl, alkenyl, alkoxy, ester, ether, hydroxyl, halogen, haloalkyl, amino or nitro substituents. IV. Replace phenylethyl with any group other than hydrogen atoms.

19. The application according to claim 17, wherein the fentanyl compound includes fentanyl substances listed in the "List of Narcotic Drugs", "List of Psychotropic Drugs" and "Supplementary List of Controlled Non-Pharmaceutical Narcotic Drugs and Psychotropic Drugs".

20. The application according to claim 19, wherein the fentanyl compound comprises acetylalfamethylfentanyl, alfentanyl, alfamethylfentanyl, alfamethylthiofentanyl, beta-hydroxyfentanyl, beta-hydroxy-3-methylfentanyl, fentanyl, 3-methylfentanyl, 3-methylthiofentanyl, p-fluorofentanyl, remifentanyl, sufentanyl, thiofentanyl, acetylfentanyl, butyrofentanyl, β-hydroxythiofentanyl, 4-fluorobutyrofentanyl, isobutyrofentanyl, oxfentanyl, acryloylfentanyl, carfentanyl, furanylfentanyl, valerate fentanyl, 4-fluoroisobutyrofentanyl, tetrahydrofuranylfentanyl, 4-phenylamino-N-phenylethylpiperidine, and N-phenylethyl-4-piperidinone.

21. The application according to claim 20, wherein the fentanyl compound comprises fentanyl, 3-methylfentanyl, carfentanyl, sufentanyl, and 4-fluoroisobutyrylfentanyl.

22. The application according to any one of claims 17-21, wherein the sample is plasma or urine.

23. Use of the surface-enhanced Raman spectroscopy substrate of claim 9 in the preparation of a kit for detecting fentanyl-like compounds and / or morphine-like compounds in a sample.

24. The use according to claim 23, wherein the fentanyl compound refers to a substance whose chemical structure, compared to fentanyl, meets one or more of the following conditions:

1. Replace the propionyl group with other acyl groups; 2. Replace the phenyl group directly attached to the nitrogen atom with any substituted or unsubstituted monocyclic aromatic group; 3. The piperidine ring has alkyl, alkenyl, alkoxy, ester, ether, hydroxyl, halogen, haloalkyl, amino or nitro substituents. IV. Replace phenylethyl with any group other than hydrogen atoms.

25. The use according to claim 23, wherein the fentanyl compound includes fentanyl substances listed in the "List of Narcotic Drugs", "List of Psychotropic Drugs" and "Supplementary List of Controlled Non-Pharmaceutical Narcotic Drugs and Psychotropic Drugs".

26. The use according to claim 23, wherein the fentanyl compound comprises acetylalfamethylfentanyl, alfentanyl, alfamethylfentanyl, alfamethylthiofentanyl, beta-hydroxyfentanyl, beta-hydroxy-3-methylfentanyl, fentanyl, 3-methylfentanyl, 3-methylthiofentanyl, p-fluorofentanyl, remifentanyl, sufentanyl, thiofentanyl, acetylfentanyl, butyrofentanyl, β-hydroxythiofentanyl, 4-fluorobutyrofentanyl, isobutyrofentanyl, oxfentanyl, acryloylfentanyl, carfentanyl, furanylfentanyl, valerate fentanyl, 4-fluoroisobutyrofentanyl, tetrahydrofuranylfentanyl, 4-phenylamino-N-phenylethylpiperidine, and N-phenylethyl-4-piperidinone.

27. The use according to claim 26, wherein the fentanyl compound comprises fentanyl, 3-methylfentanyl, carfentanyl, sufentanyl, and 4-fluoroisobutyrylfentanyl.

28. The use according to any one of claims 23-27, wherein the sample is plasma or urine.

29. A method for detecting fentanyl-like compounds and / or morphine-like compounds in a sample, comprising: 1) Add the surface-enhanced Raman spectroscopy substrate of claim 9 and a substrate capable of providing I to the sample. - The substances were mixed to obtain a mixture. 2) Collect the surface-enhanced Raman spectrum of the mixture obtained in step 1).

30. The method of claim 29, further comprising: 3) Determine if fentanyl-like compounds are present in the sample. If the surface-enhanced Raman spectrum of the mixture is within the wavenumber range of 995-1005 cm⁻¹ -1 The presence of characteristic peaks at these locations confirms the presence of fentanyl-like compounds in the sample; if the surface-enhanced Raman spectrum of the mixture is within the wavenumber range of 620-635 cm⁻¹... -1 The presence of a characteristic peak at the location confirms the presence of morphine-like compounds in the sample.

31. The method of claim 30, further comprising: 4) Quantify fentanyl-like compounds and / or morphine-like compounds in the sample by using the intensity of characteristic peaks.

32. The method of claim 29, wherein the ability to provide I - The substance is NaI or KI.

33. The method of claim 29, wherein step 1) further comprises: Add an internal standard to the sample.

34. The method of claim 33, wherein the internal standard is deuterated fentanyl.

35. The method according to claim 34, wherein quantification is performed using the ratio of the characteristic peak intensity of fentanyl-like compounds and / or morphine-like compounds to deuterated fentanyl, the characteristic peak of deuterated fentanyl being in the wavenumber range of 953-993 cm⁻¹. -1 Place.

36. The method according to any one of claims 29-35, wherein the sample is pretreated prior to step 1).

37. The method according to any one of claims 29-35, wherein the sample is plasma or urine.

38. The method according to any one of claims 29-35, wherein the fentanyl compound refers to a substance whose chemical structure, compared to fentanyl, meets one or more of the following conditions:

1. Replace the propionyl group with other acyl groups; 2. Replace the phenyl group directly attached to the nitrogen atom with any substituted or unsubstituted monocyclic aromatic group; 3. The piperidine ring has alkyl, alkenyl, alkoxy, ester, ether, hydroxyl, halogen, haloalkyl, amino or nitro substituents. IV. Replace phenylethyl with any group other than hydrogen atoms.

39. The method according to any one of claims 29-35, wherein the fentanyl compounds include fentanyl substances listed in the "List of Narcotic Drugs", "List of Psychotropic Drugs" and "Supplementary List of Controlled Non-Pharmaceutical Narcotic Drugs and Psychotropic Drugs".

40. The method according to any one of claims 29-35, wherein the fentanyl compound comprises acetylalfamethylfentanyl, alfentanyl, alfamethylfentanyl, alfamethylthiofentanyl, beta-hydroxyfentanyl, beta-hydroxy-3-methylfentanyl, fentanyl, 3-methylfentanyl, 3-methylthiofentanyl, p-fluorofentanyl, remifentanyl, sufentanyl, thiofentanyl, acetylfentanyl, butyrofentanyl, β-hydroxythiofentanyl, 4-fluorobutyrofentanyl, isobutyrofentanyl, oxfentanyl, acryloylfentanyl, carfentanyl, furanylfentanyl, valerate fentanyl, 4-fluoroisobutyrofentanyl, tetrahydrofuranylfentanyl, 4-phenylamino-N-phenylethylpiperidine, and N-phenylethyl-4-piperidinone.

41. The method according to any one of claims 29-35, wherein the fentanyl compound comprises fentanyl, 3-methylfentanyl, carfentanyl, sufentanyl, and 4-fluoroisobutyrylfentanyl.

42. The method according to any one of claims 29-35, wherein the morphine compound includes morphine and heroin.

43. The method according to any one of claims 29-35, wherein the sample is plasma, step 1) further comprises: Bovine serum albumin was added to the sample.

44. The method according to any one of claims 29-35, wherein the sample is urine, the method further comprising: The sample is pretreated before step 1). The pretreatment includes: adding sodium hydroxide and sodium chloride to urine, mixing, separating solids and liquids, and taking the supernatant as the sample to be tested.

Images