Cucurbituril derivatives for drug detection, and methods of making and using the same

Abstract

The present application relates to the technical field of biological analysis detection, and particularly relates to cucurbituril derivatives for drug detection, a preparation method and application of the cucurbituril derivatives, a detection probe containing the cucurbituril derivatives, a drug for treating drug acute poisoning, and a method for drug detection. The derivative introduces a functional group of o-cyanobenzaldehyde (CNBA) on the basis of retaining the original supramolecular recognition characteristics of cucurbituril, and the obtained compound can realize efficient covalent capture on the primary amine groups in common drugs and metabolites (such as methyl phenylpropylamine and its metabolites, 3,4-methylenedioxy methyl phenylpropylamine, ketamine and its metabolites, fluoroamine and its metabolites, phenylcyclohexylpiperidine, and casiketone molecules), so as to realize rapid, high-sensitivity and visual detection on the above target objects, and can be used for antagonistic treatment of acute poisoning caused by related drugs and metabolites.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biological analysis detection, and particularly relates to cucurbituril derivatives for drug detection and a preparation method and application thereof. BACKGROUND

[0002] Drug abuse (methamphetamine, heroin, ketamine, etc.) directly damages the central nervous system, leading to serious physical dependence and mental dependence. Drug abuse not only leads to individual addiction, nerve damage and organ failure, but also exacerbates social problems including rising crime rates, family breakdown, posing a serious threat to public safety and social stability. Therefore, it is urgent to develop drugs with rapid on-site detection and high-efficiency detoxification functions to support clinical first aid and law enforcement applications.

[0003] At present, drug detection methods mainly include laboratory instrument analysis and rapid screening method; laboratory instrument analysis method (such as chromatography, mass spectrometry and combined technology, etc.) has high detection accuracy, but has defects such as complex pretreatment, dependence on equipment and high cost, which is difficult to meet the needs of pre-hospital emergency and grassroots law enforcement, etc.; rapid screening methods mainly include immunoassay, spectroscopy and chemical colorimetric method; immunoassay is simple to operate, but has defects such as being easily disturbed by environment, low sensitivity (methamphetamine 1000 ng / ml, heroin 300 ng / ml, ketamine 1000 ng / ml), and false positive risk; spectroscopy can non-destructively analyze different phase samples, but has defects such as dependence on standard spectrum library, easy to miss detection and difficult to guarantee the uniformity of the base material; chemical colorimetric method is mature, rapid and intuitive, but has defects such as poor specificity, easy to misjudge low content or similar structures, etc.

[0004] In the clinical treatment of drug poisoning, there is still a lack of specific antagonistic drugs at present, and mainly non-specific means such as promoting metabolism and excretion, neutralizing toxic products and symptomatic support are adopted, which is difficult to achieve efficient clearance of drugs and their active metabolites.

[0005] Cucurbit[n]uril (CB[n]) is a kind of supramolecular host macrocyclic compound with a cavity structure. Due to its excellent water solubility, controllable cavity structure, good biocompatibility and high binding capacity, it has shown wide potential in the fields of materials, environment, information and medicine. At present, a series of structures such as CB[4]-CB

[14] have been developed, and their water solubility and recognition performance have been improved by modification. Such derivatives mainly realize the recognition of drug molecules through mechanisms such as hydrophobic interaction, ionic interaction, π-π stacking and cavity matching. For example, the detection limit of CB[7] for morphine can reach 0.07 μg / L. However, most of the existing CB derivatives rely on complex devices (such as organic field effect transistors) to realize detection, which is not conducive to on-site rapid application and is not conducive to the needs of on-site rapid detection, and the applicability is limited. At the same time, the above recognition is non-specific recognition, and the recognition specificity and affinity need to be improved.

[0006] The current drug detection method generally has the problems of complex operation, high equipment requirement, insufficient sensitivity and specificity, etc., and it is difficult to meet the needs of rapid, intuitive and reliable detection in the scene of first-line law enforcement and emergency treatment. On the other hand, there is a lack of specific drugs that can efficiently remove drugs and their metabolites in the treatment aspect. Therefore, it is of great significance to develop a new type of recognition and antidote material that integrates rapid visual detection and poisoning antagonistic treatment. SUMMARY

[0007] Therefore, the present application provides a cucurbituril derivative for drug detection and a preparation method and application thereof.

[0008] The present application provides a detection probe for drug detection, and the preparation raw materials thereof include a cucurbituril derivative and a fluorescent molecule.

[0009] The cucurbituril derivative is prepared by reacting an o-cyanobenzaldehyde derivative with a non-open ring cucurbituril or an open ring cucurbituril;

[0010] The open ring cucurbituril includes at least one substituted C1-C6 terminal alkyne alkoxy on the naphthalene ring; and the tertiary carbon hydrogen atom of at least one glycosyl urea group of the non-open ring cucurbituril is substituted by a C1-C6 terminal alkyne alkoxy;

[0011] The o-cyanobenzaldehyde derivative has any one of the following structures:

[0012]

[0013] Wherein, a is 2, 3, 4 or 5.

[0014] In some embodiments, the molar ratio of the fluorescent molecule and the cucurbituril derivative is 1:1.

[0015] In some embodiments, the fluorescent molecule comprises Rhodamine 6G (R6G) and / or Rhodamine B (RhB).

[0016] The application also provides a preparation method of the detection probe, comprising: mixing the cucurbituril derivative and the fluorescent molecule in proportion to obtain the detection probe.

[0017] The application provides a cucurbituril derivative for drug detection, which is prepared by reacting an o-cyanobenzaldehyde derivative with a non-open-ring cucurbituril or an open-ring cucurbituril.

[0018] The open-ring cucurbituril comprises at least one substituted C1-C6 terminal alkyne alkoxy on a naphthalene ring; and the non-open-ring cucurbituril comprises a C1-C6 terminal alkyne alkoxy substituted on a tertiary carbon hydrogen atom of at least one glycosyl urea group.

[0019] The o-cyanobenzaldehyde derivative has any one of the following structures:

[0020]

[0021] wherein a is 2, 3, 4 or 5.

[0022] The cucurbituril derivative designed by the application integrates CNBA into the cucurbituril structure by using an artificial host (cucurbituril) with good binding performance. Through this unique molecular structure design, we use the (open-ring) cucurbituril host with conformational adaptability and negatively charged carbonyl to realize accurate recognition and efficient binding of various psychoactive drugs, and further integrate CNBA to target the primary amine metabolites, thereby significantly improving the detoxification efficiency of psychoactive drug molecules. Taking specific compounds CNBA-acCB-a, CNBA-Hex-acCB and CNBA-CB[7] as examples, the detection effect of the application on common drugs is verified, and the results show that, compared with Calabadion 2 disclosed in the prior art which only contains a methyl group in the waist, the compound obtained by adding the CNBA structure in the open-ring cucurbituril (such as CNBA-acCB-a) has a significantly improved binding capacity for the three psychoactive drugs, i.e., methamphetamine (METH), ketamine (KET) and heroin (HER) at pH 4.0.

[0023] In some embodiments, the cucurbituril derivative of the application has a structure shown in Formula I:

[0024]

[0025] Formula I;

[0026] In Formula I, each of R1, R2, R3 and R4 is independently selected from propyl sulfonate or or ;

[0027] and R1, R2, R3, and R4 are not all the same;

[0028] R5, R6, R7, and R8 are independently selected from H, C1-C6 alkyl;

[0029] R5, R6, and the structure to which they are attached are cyclic or acyclic, and R7, R8, and the structure to which they are attached are cyclic or acyclic;

[0030] m is 1 or 2.

[0031] In some embodiments, in the structure of Formula I, two of the substituents R1, R2, R3, and R4 are propyl sulfonate and the other two substituents are , or ;

[0032] wherein a is 2, 3, 4, or 5.

[0033] In some embodiments, in the structure of Formula I, R5, R6 are acyclic, and R7, R8 are acyclic. In the present application, R5, R6, R7, and R8 are independently selected from H or methyl.

[0034] In some embodiments, in the structure of Formula I, the waist of the two cucurbituril units is a cycloalkyl structure, i.e., R5, R6, and the structure to which they are attached are cyclic, forming a cycloalkyl, and R7, R8, and the structure to which they are attached are cyclic, forming a cycloalkyl. In the present application, the cycloalkyl is preferably a C3-C6 cycloalkyl. In some specific embodiments, the cycloalkyl is a cyclohexyl.

[0035] In some specific embodiments, in the structure of Formula I, m is 1.

[0036] In some embodiments, two of the substituents R1, R2, R3, and R4 are selected from or when R1and R2, R1and R3, R1and R4, R2and R3, R2and R4, R3and R4. In some specific embodiments, a is 3.

[0037] In some embodiments, the cucurbituril derivative of the present application is a non- ring-opened cucurbituril modified with o-cyanobenzaldehyde, i.e., introducing an o-cyanobenzaldehyde structure on the tertiary carbon atom of the cucurbituril group of CB. Specifically, the cucurbituril derivative has a structure shown in Formula II:

[0038]

[0039] Formula II;

[0040] In formula II, R9 is any one of the following structures:

[0041] , ;

[0042] a is 2, 3, 4 or 5;

[0043] n is 6 or 7.

[0044] In some embodiments, the cucurbituril derivative described in the present application has any one of the following structures:

[0045]

[0046] .

[0047] In formula II, R9 is any one of the following structures: .

[0048] The present application also provides a preparation method of the cucurbituril derivative described in the present application, which is an open ring cucurbituril modified by an o-cyanobenzaldehyde derivative, and the preparation method comprises:

[0049]

[0050] q is 2, 3 or 4;

[0051] carrying out an open ring reaction on compound 4 and a cucurbituril to obtain an open ring cucurbituril with an alkynyl group connected to a terminal carbonyl oxygen atom;

[0052] carrying out an addition reaction on the open ring cucurbituril and an o-cyanobenzaldehyde derivative shown in compound 2-1 or compound 2-2 to obtain the cucurbituril derivative.

[0053] In the preparation method described above, the solvent for the open ring reaction is a mixed solution of trifluoroacetic acid and acetic anhydride. In some embodiments, the volume ratio of the trifluoroacetic acid and acetic anhydride is 1:1.

[0054] In the preparation method described above, the open ring reaction comprises stirring at 70 °C for 3 hours.

[0055] In the preparation method described above, the molar ratio of compound 4 and the cucurbituril in the open ring reaction is 2:1.

[0056] In the preparation method described above, the solvent for the addition reaction is a mixed solution of dimethyl sulfoxide and water. In some embodiments, the volume ratio of the dimethyl sulfoxide and water is 1:1.

[0057] The catalyst for the addition reaction in the preparation method comprises copper sulfate pentahydrate, sodium ascorbate, or and tri[(1-benzyl-1H-1,2,3-triazol-4-yl)methyl]amine.

[0058] In the preparation method, the addition reaction is stirred at 70 °C for 20 hours.

[0059] In the preparation method, the molar ratio of the open-ring cucurbituril to the compound 2-1 (or the compound 2-2) in the addition reaction is 1:24.

[0060] The application further provides another preparation method of a cucurbituril derivative, wherein the cucurbituril derivative is a cucurbituril modified by an o-cyanobenzaldehyde derivative, and the preparation method comprises the following steps:

[0061]

[0062] The o-cyanobenzaldehyde derivative shown in the compound 2-1 or 2-2 is subjected to an addition reaction with the non-open-ring cucurbituril to obtain the cucurbituril derivative.

[0063] The tertiary carbon hydrogen atom of at least one glycosylurea group of the non-open-ring cucurbituril is substituted by a C1-C6 terminal alkyne alkoxy.

[0064] In the preparation method, the solvent for the addition reaction is a dimethyl sulfoxide aqueous solution.

[0065] The catalyst for the addition reaction in the preparation method comprises copper sulfate pentahydrate, sodium ascorbate, or and tri[(1-benzyl-1H-1,2,3-triazol-4-yl)methyl]amine.

[0066] In the preparation method, the conditions for the addition reaction are stirring at 70 °C for 20 hours.

[0067] In the preparation method, the molar ratio of the cucurbituril to the compound 2 in the addition reaction is 1:24.

[0068] The application further provides applications of the cucurbituril derivative prepared by the preparation method in any one of the following aspects:

[0069] (1) drug detection or preparation of a probe for drug detection;

[0070] (2) preparation of a drug for treating acute poisoning.

[0071] In the applications, the poisoning is acute poisoning caused by the drug.

[0072] In the applications, the drug comprises at least one of the following: methamphetamine and metabolites thereof, 3,4-methylenedioxy methamphetamine, ketamine and metabolites thereof, flumazenil and metabolites thereof, phencyclidine, and a cathinone.

[0073] The present application also provides a drug for treating drug acute poisoning, comprising the cucurbituril derivative and an excipient.

[0074] The present application also provides a method for detecting drugs, comprising: detecting drugs in a sample to be detected by using the detection probe of the present application.

[0075] Compared with the prior art, the present application has the following remarkable advantages and positive effects:

[0076] 1. The detection performance is significantly improved:

[0077] Compared with the phenyl, naphthyl open-loop cucurbituril and closed-loop cucurbit[7]uril disclosed in the prior art, the CNBA-acCB, CNBA-Hex-acCB and CNBA-CB[7] designed in the present application achieve double advantages through a unique molecular structure: (1) the hydrophobic cavity composed of the dimethyl bridge and the hexyl glycoluril of the artificial host (cucurbituril) with good binding performance can accurately recognize the psychoactive drug molecules through conformational changes; the negative charge characteristics of the two end carbonyl groups can produce electrostatic attraction with the amino cations of the drugs. The above structure can achieve selective binding of various active drug organic molecules in a water environment, thereby improving the detoxification efficiency; (2) the CNBA is integrated into the cucurbituril structure in the present application, and the detoxification effect on methamphetamine and ketamine is achieved by targeting the primary amine metabolites of drugs.

[0078] 2. Breakthrough in molecular recognition ability:

[0079] The naphthalene ring open-loop cucurbituril disclosed in the prior art (i.e. Calabadion 2) contains only a methyl group at the waist, and the binding constants for the above-mentioned methamphetamine (METH), ketamine (KET) and heroin (HER) at pH 7.4 are (2.00±0.10)×10 6 M -1 、(5.29±0.89)×10 5 M -1 、(3.7±0.47)×10 5 M -1 . The present application optimizes the effect of pH on the binding ability, and it is found that the effect at pH 4.0 is better than that at pH 7.4, so the determination of the binding constant in the present patent is at pH 4.0. Compared with Calabadion 2, the CNBA-acCB-a obtained by adding a cyanobenzaldehyde (CNBA) structure to the naphthalene ring in the present application has significantly improved binding ability for the three common drugs. The binding constants for the active drugs methamphetamine, ketamine and heroin are (2.86±0.23)×10 7 M -1 、(1.70±0.16)×106 M -1 , (7.42 ± 1.70) x 10 5 M -1 On the basis of CNBA-acCB-a, the present application increases cyclohexyl structure in its waist to obtain CNBA-Hex-acCB-a, which has binding constants of (6.27 ± 0.28) x 10 6 M -1 , (1.46 ± 1.14) x 10 5 M -1 for the above-mentioned methamphetamine and heroin, respectively, and a poor fitting relationship with ketamine. In addition, CNBA-CB[7] obtained by introducing CNBA structure into the closed ring cucurbituril CB[7] has good binding efficiency for methamphetamine, with a binding constant of (1.24 ± 0.20) x 10 7 M -1 , and a poor fitting effect with ketamine and heroin. However, overall, the binding ability of CNBA-Hex-acCB-a and CNBA-CB[7] for methamphetamine is superior to Calabadion 2.

[0080] 3. The detection limit (LOD) is better than the currently used immunocolloid gold technology

[0081] Methamphetamine: The sensitivity of CNBA-acCB-a for methamphetamine (i.e. the minimum detection limit, 0.12 μmol / L) is more than 55.83 times higher than that of immunocolloid gold (6.70 μmol / L). The sensitivity of CNBA-Hex-acCB-a for methamphetamine (0.09 μmol / L) is more than 74.4 times higher than that of immunocolloid gold (6.70 μmol / L). The sensitivity of CNBA-CB[7]-a for methamphetamine (0.16 μmol / L) is more than 41.87 times higher than that of immunocolloid gold (6.70 μmol / L).

[0082] Ketamine: The detection sensitivity of CNBA-acCB-a for ketamine (0.24 μmol / L) is 17.54 times higher than that of immunocolloid gold (4.21 μmol / L).

[0083] Heroin: The detection sensitivity of CNBA-acCB-a for heroin (0.21 μmol / L) is 3.86 times higher than that of immunocolloid gold (0.81 μmol / L).

[0084] 4. Multi-modal detection system innovation:

[0085] The developed CNBA-acCB-a⊃R6G, CNBA-Hex-acCB-a⊃R6G and CNBA-CB[7]-a⊃R6G fluorescent probes can realize quantitative detection with a limit of 1.0-2.0 μmol / L, and realize rapid and visual detection of common drug molecules on site.

[0086] 5. Detoxification effect

[0087] In vivo and in vitro experiments found that the developed CNBA-acCB-a host molecule can significantly inhibit methamphetamine-induced cell necrosis, reduce the body temperature of mice caused by methamphetamine, and reduce the behavioral sensitization of mice induced by methamphetamine, showing pharmacological effects of antagonizing acute methamphetamine poisoning. BRIEF DESCRIPTION OF DRAWINGS

[0088] Figure 1 Fig. 4 is a fluorescence spectrum and nonlinear fitting result of CNBA-acCB-a (2 μmol / L) combined with methamphetamine, ketamine and heroin; wherein A is methamphetamine; B is ketamine; C is heroin, and the upper right corner is a nonlinear fitting diagram;

[0089] Figure 2 Fig. 5 is a result of visual detection of common drug medicines by using CNBA-acCB-a:

[0090] A. 1.0 μmol / L CNBA-CB[7]⊃R6G;

[0091] B. 1.0 μmol / L CNBA-CB[7]⊃R6G + 2.0 μmol / L heroin;

[0092] C. 1.0 μmol / L CNBA-CB[7]⊃R6G + 2.0 μmol / L ketamine;

[0093] D. 1.0 μmol / L CNBA-CB[7]⊃R6G + 2.0 μmol / L methamphetamine;

[0094] Figure 3 Fig. 6 is a fluorescence spectrum and nonlinear fitting result of CNBA-Hex-acCB-a (2 μmol / L) combined with methamphetamine, ketamine and heroin; wherein A is methamphetamine; B is ketamine; C is heroin, and the upper right corner is a nonlinear fitting diagram;

[0095] Figure 4 Fig. 7 is a result of visual detection of common drug medicines by using CNBA-Hex-acCB-a:

[0096] A. 1.0 μmol / L CNBA-CB[7]⊃R6G;

[0097] B. 1.0 μmol / L CNBA-CB[7]⊃R6G + 2.0 μmol / L heroin;

[0098] C. 1.0 μmol / L CNBA-CB[7]⊃R6G + 2.0 μmol / L ketamine;

[0099] D. 1.0 μmol / L CNBA-CB[7]⊃R6G + 2.0 μmol / L methamphetamine;

[0100] Figure 5 The fluorescence spectra and nonlinear fitting results of CNBA-CB[7] (2 μmol / L) combined with methamphetamine, ketamine and heroin; Wherein A is methamphetamine; B is ketamine; C is heroin, the upper right corner of the illustration is a nonlinear fitting diagram;

[0101] Figure 6 The results of visualizing detection of common drug medicines using CNBA-CB[7]:

[0102] A. 1.0 μmol / L CNBA-CB[7]⊃R6G;

[0103] B. 1.0 μmol / L CNBA-CB[7]⊃R6G + 2.0 μmol / L heroin;

[0104] C. 1.0 μmol / L CNBA-CB[7]⊃R6G + 2.0 μmol / L ketamine;

[0105] D. 1.0 μmol / L CNBA-CB[7]⊃R6G + 2.0 μmol / L methamphetamine;

[0106] Figure 7 The results of CCK-8 method for detecting cell activity;

[0107] Figure 8 The results of the body molecule (CNBA-acCB-a) on the drug molecule induced body temperature change of mice (n=10), wherein, A is the effect of 1.0 mg / kg methamphetamine on the body temperature of mice; B is the effect of 1.0 mg / kg heroin on the body temperature of mice; C is the effect of 5.0 mg / kg ketamine on the body temperature of mice;

[0108] Figure 9To detect the influence of drug molecules on the movement ability of mice by using the open field box; A is the detection result of methamphetamine, B is the detection result of heroin, and C is the detection result of ketamine. DETAILED DESCRIPTION

[0109] The present application provides cucurbituril derivatives for drug detection, and a preparation method and application thereof. Those skilled in the art can improve the process parameters according to the content herein. It should be particularly pointed out that all similar replacements and changes are obvious to those skilled in the art, and they are all regarded as included in the present application. The method and application of the present application have been described by preferred embodiments, and relevant personnel can obviously modify or appropriately change and combine the method and application herein without departing from the content, spirit and scope of the present application, to realize and apply the present application technology.

[0110] The test materials used in the present application are all ordinary commercially available products, which can be purchased in the market.

[0111] The present application will be further described below in combination with examples:

[0112] Example 1

[0113] I. Synthesis route of o-cyanobenzaldehyde (CNBA) derivatives

[0114]

[0115] 1. Synthesis of product 1: To a solution of 4-chloro-2-formylbenzonitrile (1.65 g, 10 mmol) in toluene (10 mL) was added ethylene glycol (80 mmol). The mixture was refluxed to generate water, which was removed by a reflux water trap. After the complete reaction of the raw material was confirmed by TLC monitoring, the mixture was cooled to room temperature, quenched with water and extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and then purified by column chromatography (eluent ratio: ethyl acetate: petroleum ether = 1:100 to 1:4) to obtain white solid product 1 (1.68 g, 8.07 mmol, yield 81%).

[0116] Characterization of product 1: 1 H NMR (400 MHz, CDCl3) δ 7.69 (d, J = 2.0 Hz, 1H), 7.51(dd, J = 8.4, 2.0 Hz, 1H), 7.49 (d, J = 8.0 Hz, 1H), 5.96 (s, 1H), 4.41 (dt,J = 6.0, 2.4 Hz, 2H), 4.05 (m, 2H). 13C NMR (101 MHz, CDCl3) δ 143.0, 134.6,133.0, 131.1, 127.8, 116.7, 110.6, 101.2, 66.2. HRMS (ESI): Calcd for [M+H] + m / z = 210.0322, Found: 210.0326.

[0117] 2、CNBA derivative 2 synthesis: To a suspension of potassium carbonate (28.5 mg, 206.2 μmol) in dry dichloromethane (2 mL) was added 11-azido-3,6,9-trioxaundecan-1-amine (20.0 mg, 91.6 μmol) under nitrogen atmosphere and stirred for 10 minutes. Then, a solution of compound 1 (28.7 mg, 137.5 μmol) in dry dichloromethane (1 mL) was added and stirring was continued for 3 hours at room temperature under nitrogen atmosphere. After completion of the reaction, the solvent was removed by rotary evaporation and the crude product was purified by column chromatography (elution gradient: ethyl acetate in n-hexane 0-50%) to obtain the intermediate product as a brown viscous liquid. This was dissolved in acetonitrile and deprotection reaction was carried out by adding 2 N hydrochloric acid. After completion of the reaction, the system was quenched with saturated sodium bicarbonate solution and extracted with ethyl acetate. The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate and concentrated under reduced pressure. The product was again purified by column chromatography (elution gradient: ethyl acetate / petroleum ether = 1:100 to 1:1) to finally obtain compound CNBA derivative 2 (20 mg, 59.2 μmol, yield 64.7%).

[0118] CNBA derivative 2 characterization: 1 H NMR (400 MHz, CDCl3) δ 10.36 (s, 1H), 7.47 (d, J =7.5 Hz, 1H), 7.17 (s, 1H), 7.09 (d, J = 7.5 Hz, 1H), 3.75-3.61 (m, 12H),3.41–3.37 (m, 4H). 13 C NMR (101 MHz, CDCl3) δ 189.0, 151.1, 132.6, 130.4,117.1, 115.8, 115.0, 112.7, 70.6, 70.6, 70.5, 69.8, 69.7, 68.1, 49.3, 43.1.HRMS (ESI): Calcd for [M+H] + m / z = 348.1672, Found: 348.1666.

[0119] II. Synthesis route of propargyl functionalized cucurbituril derivatives

[0120] .

[0121] 3. Synthesis of product 3: Bromopropyne (374 mg, 3.15 mmol) was added slowly dropwise to a suspension of 1,4-dihydroxynaphthalene (480 mg, 3 mmol) and potassium carbonate (456 mg, 3.3 mmol) in acetone (30 mL). The reaction mixture was stirred at room temperature for 24 h until the starting material completely disappeared. After completion of the reaction, the volatile organic solvent was evaporated and the crude product was extracted with ethyl acetate and saturated ammonium chloride solution. The organic phase was separated and dried over anhydrous sodium sulfate and finally purified by silica gel column chromatography (elution ratio: ethyl acetate / petroleum ether = 1:5 (v / v)). Product 3 was obtained as a solid (386 mg, 1.95 mmol, 65% yield).

[0122] Characterization of product 3: 1 H NMR (400 MHz, CDCl3) δ 8.25 (m, 1H), 8.13 (m, 1H), 7.52(m, 2H), 6.96 (m, 1H), 6.84 (m, 1H), 4.84 (s, 1H), 2.54 (s, 1H). 13 C NMR (101MHz, CDCl3) δ149.2, 148.4, 128.2, 126.7, 125.9, 125.2, 123.3, 122.9, 112.7,109.5, 78.6, 77.2, 58.5. HRMS (ESI): Calcd for [M+H] + m / z = 199.0759, Found:199.0756.

[0123] 4. Synthesis of product 4: Monosubstituted naphthalene hydroquinone 3 (3.96 g, 20 mmol) and sodium hydroxide (1 g, 25 mmol) were dissolved in acetonitrile (200 mL) followed by the addition of sodium 3-bromopropane-1-sulfonate (4.5 g, 20 mmol). The reaction mixture was stirred at 80 °C for 24 h. After completion of the reaction, the precipitate was collected by filtration and washed with acetonitrile (4 x 30 mL). The precipitate was dissolved in a small amount of hot water and the solid that precipitated upon cooling the solution to room temperature was collected by centrifugation and recrystallized from water. The final product 4 was obtained (2.67 g, 7.8 mmol, 39% yield).

[0124] Characterization of product 4: 1H NMR (400 MHz, DMSO-d6) δ 8.11 - 8.17 (m, 2H), 7.53 - 7.56 (m, 2H), 6.89 (d, J = 8.4 Hz, 1H), 6.83 (d, J = 8.4 Hz, 1H), 4.76 (s, 2H), 4.18 (t, J = 6.4 Hz, 2H), 2.45 (s, 1H) 2.68 (t, J = 8.0 Hz, 2H), 2.08 - 2.18 (m, 2H) ppm; 13 C NMR (101 MHz, DMSO-d6): δ 152.1, 150.8, 128.1, 128.1, 127.7, 127.0, 123.4, 123.3, 110.0, 109.9, 78.6, 77.2, 68.0, 58.5, 52.1, 24.7. HRMS (ESI): Calcd for [M-Na] - m / z = 319.0640, Found: 319.0651.

[0125] .

[0126] 5, 5a (acCB-a) / 5b (acCB-b) synthesis: Tetramer compound (1.56 g, 2 mmol) was dissolved in trifluoroacetic acid / acetic anhydride (1 : 1, v / v, 20 mL) mixture and heated to 70 °C. Then sulfonate 4 (1.37 g, 4 mmol), i.e. product 4, was added and the reaction was stirred at 70 °C for 3 hours. After completion of the reaction, the still warm reaction was slowly poured into methanol (75 mL) and the precipitate was separated by centrifugation. The precipitate was suspended in small amounts of methanol and centrifuged several times until the supernatant was nearly colorless. The crude product was refluxed in ethanol overnight, centrifuged and dried under vacuum to give a mixture of regioisomers 5a (acCB-a) and 5b (acCB-b) 2.3 g (yield 75 %). The obtained mixture was dissolved in hot water (5 mL) and the pH was adjusted to 10-11 with 0.5 M aqueous sodium hydroxide solution. The precipitate was separated by centrifugation and washed with methanol (3 x 15 mL) and dried under vacuum to give 5a (yield 28 %). The aqueous phase was made to precipitate 5b by adding ethanol (15 mL), which was separated by centrifugation, washed with methanol (3 x 15 mL) and dried (yield 30 %).

[0127] 5a (acCB-a) characterization: 1H NMR (400 MHz, DMSO-d6): δ 8.27 (d, J = 8.2 Hz,2H), 7.81 (d, J = 8.2 Hz, 2H), 7.72 (m, 4H), 5.52 (m, 8H), 5.32–5.40 (m, 6H),4.76 (s, 4H), 4.36–4.42 (m, 4H), 4.10–4.17 (m, 8H), 3.86 (m, 2H), 2.78 (m,4H), 2.39 (s, 2H), 2.04–2.11 (m, 4H), 1.69 (m, 12H). 13 C NMR (101 MHz, DMSO-d6): δ156.7, 156.2, 155.5, 155.1, 148.7, 145.2, 128.8, 128.6, 128.4, 128.3,126.8, 122.1, 122.0, 78.6, 78.2, 77.6, 76.2, 74.1, 70.8, 70.6, 58.5, 48.6,48.3, 36.3, 35.8, 26.3, 18.2, 15.3. HRMS (ESI): Calcd for [M-Na] - m / z =1419.3924, Found: 1419.3929.

[0128] 5b (acCB-b) Characterization: 1 H NMR (400 MHz, DMSO-d6): δ 8.19 (d, J = 8.4 Hz, 2H),7.93 (d, J = 8.4 Hz, 2H), 7.75 (t, J = 8.0 Hz, 2H), 7.56 (t, J = 8.1 Hz, 2H),5.56 (d, J = 14.8 Hz, 4H), 5.42 (m, 4H,), 5.25–5.35 (m, 6H,), 4.77 (s, 4H),4.31–4.43 (m, 8H), 4.06–4.12 (m, 6H), 3.89–3.98 (m, 6H), 3.74–3.81 (m, 2H),2.80 (m, 4H), 2.39 (s, 2H), 2.07–2.21 (m, 4H), 1.73 (m, 12H). 13C NMR (101 MHz, DMSO-d6): δ 155.7, 155.3, 154.9, 153.6, 149.0, 147.0, 127.9, 127.8,127.5, 127.2, 126.7, 126.7, 122.7, 122.7, 78.2, 77.6, 76.2, 74.2, 70.9, 70.5,58.5, 48.7, 48.6, 48.3, 36.3, 35.5, 26.5, 16.0, 15.6. HRMS (ESI): Calcd for[M-Na] - m / z = 1419.3924, Found: 1419.3939.

[0129] .

[0130] 6, 6a (Hex-acCB-a) / 6b (Hex-acCB-b) synthesis: The cyclohexyl tetramer compound (1.66 g, 2 mmol) was dissolved in trifluoroacetic acid / acetic anhydride (v / v, 1:1, 20 mL) mixed solvent and heated in an oil bath at 70 °C. After the system temperature was stable, sulfonate 4 (1.37 g, 4 mmol) was added and the reaction was stirred at 70 °C for 3 h. After the reaction was completed, the reaction solution was slowly poured into pre-cooled methanol (75 mL) while white precipitate was generated. After centrifugal separation, the precipitate was collected and washed with a small amount of methanol by ultrasonic dispersion-centrifugal separation (3 x 10 mL) until the supernatant was colorless and transparent. The obtained crude product was purified by refluxing in ethanol overnight (12 h), and after centrifugation, it was dried in a vacuum oven at 40 °C for 24 h, and finally a mixture of regioisomers 6a and 6b was obtained, 0.88 g, yield 30%. The above mixture was dissolved in hot distilled water (5 mL), and 0.5 M NaOH aqueous solution was slowly added under magnetic stirring to adjust the pH to 10-11 (pH meter accurate control). At this time, a light yellow solid was precipitated, which was separated by centrifugation (4000 rpm, 10 min), and the precipitate was washed with methanol (3 x 15 mL) and dried in a vacuum drying oven at 40 °C for 6 h to obtain pure 6a (yield 15 %). The supernatant after centrifugation was collected, and anhydrous ethanol (15 mL) was added immediately to generate white flocculent precipitate, which was separated by centrifugation under the same conditions. The precipitate was also washed with methanol (3 x 15 mL), and the product was dissolved in an appropriate amount of hot water, filtered while hot, and then ethanol (V / V = 2:1) was slowly added, and slowly crystallized in a refrigerator at 4 °C for 12 h to obtain isomer 6b (yield 16 %).

[0131] 6a characterization: 1H NMR (400 MHz, DMSO-d6): δ 8.27 (d, J = 8.2 Hz, 2H), 7.81 (d, J = 8.2 Hz, 2H), 7.72 (m, 4H), 5.52 (m, 8H), 5.32–5.40 (m, 6H), 4.77 (s, 4H), 4.36–4.42 (m, 4H), 4.10–4.17 (m, 8H), 3.86 (m, 2H), 2.78 (m, 4H), 2.39 (s, 2H), 2.38-2.07 (m, 16H), 2.04–2.11 (m, 4H). 13 C NMR (101 MHz, DMSO-d6): δ 157.7, 152.1, 150.4, 148.4, 127.3, 127.2, 126.4, 122.2, 122.2, 120.8, 120.0, 78.8, 76.7, 71.0, 71.0, 70.9, 70.3, 70.3, 69.0, 69.0, 61.5, 57.5, 51.4, 50.6, 36.8, 30.1, 29.9, 25.4, 18.8. HRMS (ESI): Calcd for [M-Na] - m / z = 1457.4080, Found: 1457.4093.

[0132] 6b Characterization: 1 H NMR (400 MHz, DMSO-d6): δ 8.19 (d, J = 8.4 Hz, 2H), 7.93 (d, J = 8.4 Hz, 2H), 7.75 (t, J = 8.0 Hz, 2H), 7.56 (t, J = 8.1 Hz, 2H), 5.56 (d, J = 14.8 Hz, 4H), 5.42 (m, 4H,), 5.25–5.35 (m, 6H,), 4.79 (s, 4H), 4.31–4.43 (m, 8H), 4.06–4.12 (m, 6H), 3.89–3.98 (m, 6H), 3.74–3.81 (m, 2H), 2.80 (m, 4H), 2.36 (s, 2H), 2.38-2.07 (m, 16H), 2.07–2.21 (m, 4H). 13C NMR (101 MHz, DMSO-d6): δ 157.6, 152.1, 150.3, 149.0, 127.3, 127.1, 126.4, 122.2, 121.9, 120.8, 120.0, 78.8, 76.9, 71.1, 70.8, 70.3, 70.2, 69.1, 61.9, 57.5, 51.4, 50.9, 36.5, 30.2, 29.9, 25.6, 18.8, 18.6. HRMS (ESI): Calcd for [M-Na] - m / z = 1457.4080, Found: 1457.4076.

[0133] III. Synthesis of o-cyanobenzaldehyde (CNBA) functionalized cucurbituril derivatives

[0134]

[0135] .

[0136]

[0137] .

[0138] 7. Synthesis of CNBA-acCB-a and CNBA-acCB-b: 5a or 5b (144 mg, 0.1 mmol) was dissolved in dimethyl sulfoxide / water (1:1, v / v, 2 mL) mixed solvent under nitrogen protection and heated to 70 °C. Copper sulfate pentahydrate (0.04 mM, 0.04 equiv), sodium ascorbate (0.16 mM, 0.16 equiv), tris [(1-benzyl-1H-1,2,3-triazol-4-yl)methyl]amine (0.04 mM, 0.04 equiv) and 2 (2.4 mmol, 2.4 equiv) were added successively, and the resulting mixture was stirred at 70 °C for 20 h. After the reaction was completed, the reaction solution was slowly added to acetonitrile (20 mL), and the product was collected by centrifugation. The crude product was washed successively with methanol (3 x 15 mL) and diethyl ether (3 x 10 mL) and dried under vacuum. The solid was further purified by precipitation with ethanol after repeated dissolution in water to obtain the target product CNBA-acCB-a: (142 mg, 67 %) or CNBA-acCB-b: (159 mg, 75 %).

[0139] Characterization of CNBA-acCB-a: 1H NMR (400 MHz, DMSO-d6): δ 10.33 (s, 2H), 8.67 (s,2H), 7.99–7.94 (m, 2H), 7.70–7.47 (m, 8H), 7.21 (s, 2H), 7.05 (d, J = 7.5 Hz,2H),5.63–5.45 (m, 8H), 5.30–5.22 (m, 4H), 5.10–4.90 (m, 4H), 4.07–4.37 (m,14H), 3.96–3.89 (m, 2H), 3.75-3.37 (m, 32H), 2.81–2.77 (m, 4H), 2.14 (m, 4H),1.70 (m, 12H); 13 C NMR (101 MHz, DMSO-d6): δ 189.1, 155.8, 151.1, 148.8,147.2, 132.6, 130.4, 127.9, 127.8, 127.6, 127.3, 127.3, 126.7, 122.8, 122.0,117.1, 115.8, 112.7, 77.6, 74.1, 71.0, 70.6, 69.8, 68.1, 49.3, 43.1, 35.7,26.4, 16.1, 15.6. HRMS (ESI): Calcd for [M-2Na] 2- m / z = 1049.8477, Found:1049.8459。

[0140] CNBA-acCB-b characterization: 1 H NMR (400 MHz, DMSO-d6): δ10.41 (s, 2H), 8.92 (s,2H, H32), 7.97–7.79 (m, 4H), 7.50 –7.43 (m, 4H), 7.37–7.21 (m, 4H), 7.04 (d,J = 7.5 Hz, 2H), 5.60–5.43 (m, 8H), 5.28 (m, 6H), 4.33–4.23 (m, 6H), 4.15–4.09 (m, 6H), 3.90–3.83 (m, 2H), 3.75–3.61 (m, 24H), 3.41–3.37 (m, 8H), 2.77(m, 4H), 2.13–2.08 (m, 4H), 1.73–1.66 (m, 12H); 13C NMR (101 MHz, DMSO-d6): δ189.0, 155.8, 154.8, 151.1, 148.9, 146.4, 132.6, 130.4, 127.8, 127.7, 126.6,122.8, 122.1, 117.1, HRMS (ESI): Calcd for[M-2Na] 2- m / z = 1049.8477, Found: 1049.8479.

[0141]

[0142]

[0143] .

[0144] 8. Synthesis of CNBA-Hex-acCB-a and CNBA-Hex-acCB-b: Using 6a or 6b as raw materials, the same reaction as 5a was carried out to obtain CNBA-acCB-a (154 mg, 71%) or CNBA-acCB-b (158 mg, 73%), respectively.

[0145] CNBA-Hex-acCB-a characterization: 1 H NMR (400 MHz, DMSO-d6): δ 10.27 (s, 2H), 8.29 (d, J = 8.2 Hz, 2H), 7.82–7.73 (m, 6H), 7.26 (m, 4H), 7.11 (m, 2H), 5.52 (m,8H), 5.40–5.32 (m, 6H), 4.42–4.36 (m, 4H), 4.19–4.11 (m, 8H), 3.79–3.62 (m,26H), 3.41–3.37 (m, 8H), 2.78 (m, 4H), 2.38-2.04 (m, 20H). 13C NMR (101 MHz, DMSO-d6): δ 186.6, 155.2, 149.7, 148.7, 146.6, 130.2, 128.0, 124.9, 124.0, 119.8, 119.5, 118.4, 117.6, 114.7, 113.4, 112.6, 110.3, 69.7, 68.4, 67.9, 67.3, 66.7, 65.7, 59.5, 52.1, 49.0, 48.5, 41.7, 34.1, 27.8, 27.5, 23.2, 16.4, 16.2. HRMS (ESI): Calcd for [M-2Na] 2- m / z = 1064.3685, Found: 1064.3693.

[0146] CNBA-Hex-acCB-b characterization: 1 H NMR (400 MHz, DMSO-d6): δ 10.37 (s, 2H), 8.22 (d, J = 8.0 Hz, 2H), 7.85 - 7.56 (m, 6H), 7.47 (d, J = 7.5 Hz, 2H), 7.17 - 7.09 (m, 4H), 5.56 - 5.35 (m, 14H), 4.59 - 4.38 (m, 8H), 4.11 - 3.92 (m, 12H), 3.80 - 3.66 (m, 26H), 3.40 - 3.33 (m, 8H), 2.80 (m, 4H), 2.38 - 2.07 (m, 20H). 13 C NMR (101MHz, DMSO-d6): δ 186.6, 155.2, 149.7, 148.7, 146.6, 130.2, 128.0, 124.9, 124.0, 119.8, 119.5, 118.4, 117.6, 114.7, 113.4, 112.6, 110.3, 69.7, 68.4, 67.9, 67.3, 66.7, 65.7, 59.5, 52.1, 49.0, 48.5, 41.7, 34.1, 27.8, 27.5, 23.2, 16.4, 16.2. HRMS (ESI): Calcd for [M-2Na] 2- m / z = 1064.3685, Found: 1064.3677.

[0147] .

[0148] 9. Synthesis of CNBA-CB[7]: The cucurbit[7]uril ylidenesubstituted (135 mg, 0.1 mmol) and compound 2 were dispersed in dimethyl sulfoxide (3 mL) under nitrogen protection. Separately, sodium L-ascorbate (19.8 mg, 0.1 mmol) was added to a 55% aqueous solution of dimethyl sulfoxide (2 mL) containing copper sulfate pentahydrate (15.9 mg, 0.1 mmol). The resulting mixture was stirred at 70 °C for 20 h. After the reaction was completed, 10 mL of methanol was added to the mixture, and the precipitate was washed with methanol (3 x 35 mL) and dried under high vacuum to obtain a dark solid. The product was purified by HPLC to obtain CNBA-CB[7] (31 mg, 19%).

[0149] Characterization of CNBA-CB[7]: 1 H NMR (400 MHz, D2O): δ 8.11 (s, 1H), 7.62 (d, J = 7.5Hz, 1H), 7.21 (s, 1H), 7.12 (d, J = 7.5 Hz, 1H), 6.03–5.87 (m, 12H), 5.73 (m,14H), 5.52 (m, 12H), 4.24 (m, 14H), 3.94–3.86 (m, 16H). 13 H NMR (101 MHz,D2O): δ 189.3, 161.6, 151.4, 132.3, 130.3, 122.6, 117.1, 115.7, 115.3, 111.3,81.5, 72.3, 70.7, 70.5, 69.6, 69.4, 68.3, 61.1, 53.8, 49.2, 43.0. HRMS (ESI):Calcd for [M+H] + m / z = 1564.5213, Found: 1564.5221.

[0150] Example 2: Test and visual detection of the binding performance of CNBA-acCB-a with common drugs

[0151] 2.1 Preparation of the probe

[0152] CNBA-acCB-a was mixed with rhodamine 6G (R6G) at a 1:1 molar ratio to prepare a CNBA-acCB-a⊃R6G complex probe. The probe is pink under visible light and emits light yellow fluorescence under 365 nm ultraviolet light.

[0153] 2.2 Calculation of binding constant

[0154] At 25℃, 2.0 μmol / L CNBA-acCB-a⊃R6G probe was added to sodium acetate buffer (pH 4.0) in a quartz cuvette. Using a microsyringe, methamphetamine standard solution (0-24 μL), heroin standard solution (0-40 μL), ketamine standard solution (0-60 μL) were added in turn, and each time was balanced for 5 minutes. At an excitation wavelength of 310 nm, the change in fluorescence intensity at 350 nm was recorded. As shown in Figure 1 , with the increase of guest concentration, the fluorescence intensity showed regular enhancement. By nonlinear fitting, the binding constants Ka of CNBA-acCB-a⊃R6G with methamphetamine, ketamine and heroin were calculated to be (2.86±0.23)×10 7 M -1 , (1.70±0.16)×10 6 M -1 , (7.42±1.70)×10 5 M -1 , respectively, indicating that there was a strong interaction between the host and the guest.

[0155] 2.3 Analysis of detection sensitivity

[0156] Fluorescence spectrum analysis showed that within the range of 0-1.0 μmol / L, the fluorescence intensity of the probe CNBA-acCB-a⊃R6G at 527 nm was linearly related to the concentration of methamphetamine, ketamine and heroin, and the detection limits were 0.12 μmol / L, 0.24 μmol / L and 0.21 μmol / L, respectively. The probe system realized visual detection while maintaining high sensitivity.

[0157] Table 1 Detection limit of CNBA-acCB-a with methamphetamine, ketamine and heroin

[0158] 2.4 Method validation

[0159] The 2.0 μmol / L methamphetamine solution was determined for 6 times in parallel, and the relative standard deviation (RSD) was 1.57%, respectively. The standard addition recovery experiment showed that within the range of 0.5-1.5 μmol / L, the average recovery rate was 100.67 ± 2.29%, 104.33 ± 4.28%, 100.67 ± 6.72% (n=3), respectively, proving that the method was accurate and reliable.

[0160] Table 2 Recovery rate of CNBA-acCB-a with methamphetamine

[0161]

[0162] 2.5 Visual detection application

[0163] After adding different concentrations of methamphetamine, ketamine and heroin into the CNBA-acCB-a⊃R6G probe solution, the color of the solution changed obviously with the increase of concentration: under ultraviolet light, the fluorescence changed from light yellow to yellow Figure 2 ) fluorescence. The detection limit of naked eye can reach 1.0 μmol / L, which meets the demand of on-site rapid screening.

[0164] Example 3: Binding performance test and visual detection of CNBA-Hex-acCB-a and common products

[0165] 3.1 Preparation of probe

[0166] CNBA-Hex-acCB-a and rhodamine 6G were mixed at a molar ratio of 1:1 to prepare CNBA-Hex-acCB-a⊃R6G complex probe.

[0167] 3.2 Calculation of binding constant

[0168] At 25℃, 2.0 μmol / L CNBA-Hex-acCB-a⊃R6G probe was added to sodium acetate buffer (pH 4.0) in a quartz cuvette. Using a microsyringe, heroin standard solution (0-80 μL), ketamine standard solution (0-26 μL), and methamphetamine standard solution (0-40 μL) were added in turn, and each time was balanced for 5 minutes. At an excitation wavelength of 527 nm, the change in fluorescence intensity at 553 nm was recorded. As shown in Figure 3 , with the increase of guest concentration, the fluorescence intensity showed regular enhancement. Through nonlinear fitting, the binding constants Ka with methamphetamine and heroin were calculated to be (6.27±0.28)×10 6 M -1 and (1.46±1.14)×10 5 M -1 , respectively, indicating that there was a strong interaction between the host and the guest; while the fitting effect of CNBA-Hex-acCB-a and ketamine was poor, indicating that the host-guest interaction was low.

[0169] 3.3 Sensitivity analysis

[0170] Methamphetamine, heroin gradient solution of 0-1.5 μmol / L, respectively, with 2.0 μmol / L CNBA-Hex-acCB-a⊃R6G mixed and placed for 10 minutes. In the range of 0-0.8 μmol / L, the fluorescence intensity and concentration showed good linear relationship. According to the calculation of 3 times the signal-to-noise ratio, the detection limit was 0.09 μmol / L, 0.44 μmol / L, respectively. The sensitivity was improved by 74.4 times and 1.84 times compared with the traditional immunocolloidal gold method. The probe system realized visual detection while maintaining high sensitivity.

[0171] Table 3 Detection limit of CNBA-acCB-a with methamphetamine, ketamine and heroin

[0172]

[0173] 3.4 Method validation

[0174] The relative standard deviation (RSD) of 1.0 μmol / L methamphetamine solution was 0.39 % for 6 parallel determinations. The standard addition recovery experiment showed that the average recovery rate was 118.33 ± 2.44 %, 104.45 ± 4.60 %, and 102.50 ± 3.23 % (n=3) in the range of 0.4-0.8 μmol / L, respectively, proving that the method was accurate and reliable.

[0175] Table 4 Recovery rate of CNBA-Hex-acCB-a with methamphetamine

[0176]

[0177] 3.5 Visual detection application

[0178] After adding different concentrations of methamphetamine, ketamine and heroin to the CNBA-Hex-acCB-a⊃R6G probe solution, the color of the solution changed obviously with the increase of concentration: under ultraviolet light, the fluorescence changed from light yellow to yellow Figure 4 ) fluorescence. The detection limit that can be distinguished by naked eye reached 1.0 μmol / L, meeting the needs of on-site rapid screening.

[0179] Example 4: Test of binding performance of CNBA-CB[7] with common products and visual detection

[0180] 4.1 Preparation of probe

[0181] CNBA-CB[7] and rhodamine 6G were mixed at a molar ratio of 1:1 to prepare CNBA-CB[7]⊃R6G complex probe. The probe showed magenta color under visible light and emitted light yellow fluorescence under 365 nm ultraviolet light.

[0182] 4.2 Calculation of binding constant

[0183] At 25 °C, 1.0 μmol / L of CNBA-CB[7]·R6G was placed in a quartz cuvette with sodium acetate buffer (pH 4.0). Using a microsyringe, heroin standard solution (0-10 μL), ketamine standard solution (0-10 μL), and methamphetamine standard solution (0-32 μL) were added in turn, and allowed to equilibrate for 5 minutes after each addition. The fluorescence intensity at 557 nm was recorded at an excitation wavelength of 527 nm. As shown in Fig. A, the fluorescence intensity increased regularly with increasing guest methamphetamine concentration, while there was no significant change in fluorescence intensity after the addition of heroin and ketamine. The binding constant Ka with methamphetamine was calculated by nonlinear fitting to be (1.24 ± 0.20) x 10 Figure 5 7 M -1 , indicating that there is a strong interaction between CNBA-CB[7] and methamphetamine, while CNBA-CB[7] cannot be fitted with heroin and ketamine.

[0184] 4.3 Analysis of detection sensitivity

[0185] 0-1.5 μmol / L of methamphetamine gradient solution was prepared and mixed with 2.0 μmol / L CNBA-CB[7] and allowed to stand for 10 minutes. The fluorescence intensity showed a good linear relationship with the concentration in the range of 0-0.8 μmol / L. The detection limit was calculated to be 0.16 μmol / L according to the 3 times signal-to-noise ratio, which is 41.87 times more sensitive than the traditional immunocolloidal gold method (6.70 μmol / L).

[0186] 4.4 Method validation

[0187] 1.0 μmol / L of methamphetamine solution was determined in parallel for 6 times, and the relative standard deviation (RSD) was 0.40%, respectively. The standard addition recovery experiment showed that the average recovery rate in the range of 0.4-0.8 μmol / L was 99.17 ± 2.91 %, 92.78 ± 3.74 %, and 104.58 ± 3.65 % (n=3), respectively, proving that the method is accurate and reliable.

[0188] Table 5 Recovery rate of CNBA-CB[7] with methamphetamine

[0189] 4.5 Visual detection application

[0190] ​The color of the CNBA-CB[7]⊃R6G probe solution changed obviously with the increase of the concentration of three drugs: methamphetamine, ketamine and heroin. The color of the solution changed from light yellow fluorescence to yellow fluorescence under UV light Figure 6 ). The detection limit of naked eye reached 1.0 μmol / L, which met the needs of rapid screening on site.

[0191] Example 5: Application of acute poisoning detoxification

[0192] 5.1 CCK-8 detection of cell activity

[0193] HEK-293 cells were treated with CNBA-acCB-a at concentrations of 1, 2, 5 and 10 mm for 24 h, and the cell activity was detected by CCK-8 method. The results are shown in Figure 7 .

[0194] The results showed that the cell activity was still 90% after 24 h treatment with 10 mm, and there was no significant effect on the cell activity after 96 h treatment (90.47 ± 2.71% vs. 85.10 ± 5.29%, P = 0.051). 1.0 mm-methamphetamine, 0.1 mm-heroin and 1.0 mm-ketamine could significantly reduce the activity of HEK-293 cells, while 10.0 mm CNBA-acCB-a treatment could significantly reverse the above-mentioned drug-induced reduction of cell activity (methamphetamine: 80.31 ± 5.08% vs. 69.94 ± 3.17%, P = 0.002. Heroin: 78.97 ± 3.79% vs. 73.10 ± 4.80%, P = 0.041. Ketamine: 80.47 ± 4.15% vs. 74.94 ± 3.98%, P = 0.040).

[0195] 5.2 Detection of body temperature changes by rectal thermometer

[0196] The rectal thermometer was used to detect the body temperature changes of mice, and the results are shown in Figure 8 .

[0197] Results showed that 120 mg / kg CNBA-acCB-a had no significant effect on body temperature of mice (0.5 h: 37.66 ± 0.19 vs. 37.62 ± 0.14, P = 0.615). 1 mg methamphetamine could significantly increase body temperature of mice, and reached the peak at 0.5 h (37.66 ± 0.19 vs. 38.76 ± 0.42, P < 0.001), and then gradually decreased, and the body temperature was still significantly higher than that before the experiment at 2 h (37.80 ± 0.32 vs. 37.26 ± 0.13, P < 0.001). 120 mg / kg CNBA-acCB-a could significantly reduce the increase of body temperature of mice induced by 1 mg / kg methamphetamine, especially at 0.5 h (38.75 ± 0.42 vs. 38.09 ± 0.11, P < 0.001). The body temperature of mice in 120 mg / kg CNBA-acCB-a + 1.0 mg / kg methamphetamine group was still significantly lower than that in 1.0 mg / kg methamphetamine group at 2 h (38.75 ± 0.20 vs. 37.80 ± 0.33, P = 0.009).

[0198] 1.0 mg / kg heroin could significantly increase body temperature of mice, and reached the peak at 0.5 h (38.56 ± 0.30 vs. 37.62 ± 0.14, P < 0.001), and then gradually decreased, and the body temperature was still significantly higher than that before the experiment at 2 h (37.89 ± 0.15 vs. 37.33 ± 0.16, P < 0.001). 120 mg / kg CNBA-acCB-a could significantly reduce the increase of body temperature of mice induced by 1 mg / kg heroin, especially at 0.5 h (38.56 ± 0.30 vs. 38.23 ± 0.22, P = 0.012), and there was no statistical difference in body temperature between the two groups at 2 h (37.90 ± 0.15 vs. 37.85 ± 0.16, P = 0.012).

[0199] 5.0 mg / kg ketamine significantly reduced body temperature in mice, peaking at 0.5 h (36.35±0.28 vs. 37.62±0.14, P<0.001), and then gradually returned to normal. After 2 h, the body temperature was still significantly lower than before the experiment (37.10±0.17 vs. 37.32±0.16, P=0.006). 120 mg / kg CNBA-acCB-a significantly alleviated the hypothermia induced by 5.0 mg / kg ketamine in mice at 0.5 h (36.63±0.20 vs. 36.35±0.28, P = 0.020), with no statistically significant difference between the two groups at 1 h and 2 h (1 h: 36.72±0.24 vs. 36.66±0.28, P = 0.612; 2 h: 37.15±0.11 vs. 37.10±0.17, P = 0.442).

[0200] 5.3 Open field test to detect changes in mouse activity

[0201] Changes in mouse activity were detected using an open field box. Figure 9 ).

[0202] like Figure 9 As shown in Figure A, 1.0 mg / kg methamphetamine significantly increased the motor function of mice, with the distance traveled in the open field box at 15 min being significantly greater than that of the control group (35010.84±9325.29 cm vs. 5442.28±1470.03 cm, P<0.001). 120 mg / kg CNBA-acCB-a had no effect on the motor function of mice (6454.39±738.88 cm vs. 5442.28±1470.03 cm, P<0.068). Furthermore, 120 mg / kg CNBA-acCB-a significantly alleviated the increased viability induced by 1.0 mg / kg methamphetamine in mice (21010.54±6457.70 cm vs. 35010.84±9325.29 cm, P = 0.001).

[0203] like Figure 9 As shown in Figure B, the distance traveled in the open field box by mice in the 1.0 mg / kg heroin group at 15 min was significantly greater than that in the control group (21610.84±7199.81 cm vs. 6454.40±738.88 cm, P<0.001). 120 mg / kg CNBA-acCB-a significantly alleviated the increased activity induced by 1.0 mg / kg heroin in mice (21610.84±7199.81 cm vs. 15410.54±4219.45 cm, P = 0.030).

[0204] As Figure 9 As shown in FIG. 5C, the distance moved in the open field box was significantly lower in the 5.0 mg / kg ketamine group than in the control group (4654.40 ± 1362.91 cm vs. 6454.40 ± 738.88 cm, P = 0.002). Meanwhile, the present study showed that the distance moved was significantly higher in the 120 mg / kg CNBA-acCB-a + 5.0 mg / kg ketamine group than in the 5.0 mg / kg ketamine group (5854.40 ± 1163.26 cm vs. 4654.40 ± 1362.91 cm, P = 0.048).

[0205] The above merely describes the preferred embodiments of the present application, and it should be noted that, for those skilled in the art, some improvements and refinements can be made without departing from the principles of the present application, and these improvements and refinements should also be considered as falling within the scope of the present application.

Claims

1. A cucurbituril derivative for drug detection, characterized in that, It has any of the following structures: 。 2. The method for preparing the cucurbituril derivative for drug detection according to claim 1, characterized in that, The cucurbituril derivative is CNBA-Hex-acCB-a or CNBA-Hex-acCB-b, and its preparation method includes: q = 2; a = 3; Compound 4 and cucurbituril were subjected to a ring-opening reaction to obtain ring-opening cucurbituril with structural formulas such as 6a or 6b. The open-ring cucurbituril was subjected to an addition reaction with the o-cyanobenzaldehyde derivative shown in compound 2-1 to obtain the cucurbituril derivative CNBA-Hex-acCB-a or CNBA-Hex-acCB-b. The structural formula of the cucurbituril is as follows: 。 3. The method for preparing the cucurbituril derivative for drug detection according to claim 1, characterized in that, The cucurbituril derivatives are CNBA-acCB-a and CNBA-acCB-b, and their preparation methods include: q = 2; a = 3; Compound 4 and cucurbituril were subjected to a ring-opening reaction to obtain ring-opened cucurbituril with structural formulas such as 5a or 5b. The open-ring cucurbituril was subjected to an addition reaction with the o-cyanobenzaldehyde derivative shown in compound 2-1 to obtain the cucurbituril derivative CNBA-acCB-a or CNBA-acCB-b; The structural formula of the cucurbituril is as follows: 。 4. The preparation method according to claim 2 or 3, characterized in that, The solvent for the ring-opening reaction is a mixed solution of trifluoroacetic acid and acetic anhydride; And / or, the ring-opening reaction comprises a stirred reaction at 70 °C for 3 hours; And / or, in the ring-opening reaction, the molar ratio of compound 4 to the cucurbituril is 2:1; And / or, the solvent for the addition reaction is a mixed solution of dimethyl sulfoxide and water; And / or, the catalyst for the addition reaction includes copper sulfate pentahydrate, sodium ascorbate, and / or tris[(1-benzyl-1H-1,2,3-triazol-4-yl)methyl]amine; And / or, the addition reaction is carried out at 70°C with stirring for 20 hours; And / or, in the addition reaction, the molar ratio of the open-ring cucurbituril to compound 2-1 is 1:

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5. The method for preparing the cucurbituril derivative for drug detection according to claim 1, characterized in that, The cucurbituril derivative is CNBA-CB[7], and its preparation method includes: The o-cyanobenzaldehyde derivative and non-ring-opening cucurbituril shown in compound 2-2 were subjected to an addition reaction to obtain the cucurbituril derivative; The structural formula of the non-open-ring cucurbituril is as follows: 。 6. The preparation method according to claim 5, characterized in that, The solvent for the addition reaction is an aqueous solution of dimethyl sulfoxide; And / or, the catalyst for the addition reaction includes copper sulfate pentahydrate and / or sodium ascorbate; And / or, the conditions for the addition reaction are: stirring at 70°C for 20 hours; And / or, in the addition reaction, the molar ratio of the cucurbituril to the compound 2-2 is 1:

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7. The application of the cucurbituril derivative for drug detection according to claim 1, characterized in that, Including any of the following: (1) Application in drug detection or in the preparation of detection probes for drug detection; (2) Application in the preparation of drugs for treating acute poisoning; The poisoning is acute poisoning caused by the drug. The drugs include at least one of the following: methamphetamine, 3,4-methylenedioxymethamphetamine, ketamine, and heroin.

8. A detection probe containing a cucurbituril derivative, characterized in that, Includes the cucurbituril derivative and fluorescent molecule as described in claim 1; The fluorescent molecule is Rhodamine 6G.

9. A drug for treating acute drug poisoning, characterized in that, It includes the cucurbituril derivative for drug detection as described in claim 1, and excipients.

10. A method for drug detection, characterized in that, The drug in the sample to be tested is detected using the detection probe containing cucurbituril derivative as described in claim 8; The drugs include at least one of methamphetamine, 3,4-methylenedioxymethamphetamine, ketamine, and heroin.

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