Patent Description:
Pain is an unpleasant feeling and emotional experience caused by tissue damage or potential tissue damage. It is a warning signal sent by the body. However, severe or long-term pain causes an unbearable torture on the body and seriously affects the patient's life. Therefore, the International Pain Society has designated October <NUM> of each year as "Global Pain Day" since <NUM>.

Pain can be divided into acute pain and chronic pain based on duration time. Acute pain is nociceptive pain usually caused by tissue trauma, while chronic pain is a disease mainly dominated by neuropathic pain. Traditional analgesic drugs mainly comprise opioids and non-steroidal anti-inflammatory drugs. Opioids have strong analgesic effect, but long-term use of opioids is easy to cause tolerance, dependence and addiction, and opioids have adverse effects such as respiratory depression and central sedation. Non-steroidal anti-inflammatory drugs only have moderate analgesic effect, while having adverse effects such as gastrointestinal bleeding and cardiotoxicity, etc..

TRPA1, also known as ANKTM1, is a member of TRP ion channel superfamily. TRPA1 is mainly distributed on the primary sensory neurons of dorsal root nerve (DRG), trigeminal nerve (TG) and vagus nerve (VG), and is expressed in peptide energy neurons having rich neuropeptides CGRP, SP and neurotrophic factor receptor TrkA, and in non-peptidergic neurons co-expressing purinergic receptors P2X3, neurturin, artemin, G protein-coupled receptors in Mrg family, and GFRα<NUM>, GFRα<NUM> in GDNF receptor family). From the distribution of human system, TRPA1 is highly expressed in the peripheral nervous system, respiratory system, gastrointestinal system and urinary system. When these organs and tissues have abnormal functions, the expression and function of TRPA1 channels are usually abnormal simultaneously. TRPA1 can convert cold stimulation, chemical stimulation and mechanical stimulation into inward currents, trigger a series of physiological functions, and participate in the formation of various pain sensations.

Inflammation is the defensive response of living tissues with vascular system to injury factors. The stimulation of inflammatory mediators such as prostaglandin, serotonin, bradykinin, etc. is the main cause of local pain caused by inflammation. Inflammatory pain is common problem of certain chronic diseases, and there is still no effective treatment method in the clinic. Animal experiments have shown that TRPA1 is involved in inflammatory response and plays an important role in inflammatory pain. The use of TRA1 specific blockers can significantly reduce the inflammatory pain response in rats. The pathogenesis of asthma and cough has become more and more clear as the research continues. From the current research, TRPA1 plays an important role in the occurrence of asthma and cough. Compounds that induce asthma and cough, either cellular endogenous factors or exogenous factors, can activate TRPA1. TRPA1 antagonists can reduce asthma symptoms and block airway hyper-responsiveness.

Visceral pain, as a main visceral sensation, is often caused by viscera stimulation such as mechanical traction, spasm, ischemia, or inflammation, etc. It is confirmed that TRPA1 is involved in the regulation of visceral hypersensitivity through different visceral hypersensitivity animal models such as colitis, rectal dilatation or stress. Neuropathic pain is a pain syndrome caused by central or peripheral nervous system damage or disease, mainly manifested as allodynia, allodynia, and spontaneous pain. Different from inflammatory pain, neurogenic pain is not related to vascular response of central inflammation, but depends on the damage and dysfunction of nervous system, which is often caused by peripheral nerve damage. In recent years, more and more studies have shown that TRPA1 channel plays an important role in different neurogenic pain, such as diabetic neuropathy and neuropathy caused by chemotherapy drugs. Recent studies have also shown that TRPA1 also has a mediating role in toothache, migraine and other pains. The administration of TRPA1 antagonists can significantly alleviate the pain symptoms.

Since TRPA1 is widely distributed and expressed in the human system, the importance of its function is self-evident. In addition to the physiological functions involved by TRPA1, the development of TRPA1 inhibitor indications reported so far also involves inflammatory bowel disease, chronic obstructive pulmonary disease, antitussive, antipruritic, allergic rhinitis, ear disease, diabetic, urinary incontinence, etc. It is proved that TRPA1 is a new target for pain treatment. There is no commercial drug for TRPA1 target. Pain is a refractory disease.

<CIT> describes a compound:
<CHM>
as well as its activity to inhibit RTPA <NUM> activity and use to treat diseases related to RTP.

<CIT> describes compounds of the formulae below showing affinity and activity towards the subunit α2δ of voltage-gated calcium channels (VGCC):
<CHM>.

Z is one of the following moieties:
<CHM>.

<CIT> describes compounds of the formula below:
<CHM>.

However, there is still an urgent need in the art to develop a medicament targeting TRP, especially TRPA1, to improve therapeutic effect of diseases.

It is an object of the present invention to provide novel compounds which target TRP channel, especially TRPA1, and uses thereof.

In the first aspect of the present invention, it provides a compound of formula I, or a pharmaceutically acceptable salt thereof for use in preventing and/or treating a disease related to transient receptor potential channel protein (TRP), wherein the disease is selected from the group consisting of pain, epilepsy, inflammation, respiratory disorder, pruritus, urinary tract disorder and inflammatory bowel disease;
<CHM>
wherein,.

In another preferred embodiment, X is S.

In another preferred embodiment, the heteroaryl contains <NUM>-<NUM> heteroatoms selected from the group consisting of N, O and S.

In another preferred embodiment, the transient receptor potential channel protein (TRP) is TRPA1.

In another preferred embodiment, R<NUM> and R<NUM> are each independently hydrogen atom, or C<NUM>-C<NUM> alkyl.

In another preferred embodiment, R<NUM> is hydrogen atom, halogen, or substituted or unsubstituted C<NUM>-C<NUM> alkyl.

In another preferred embodiment, R<NUM> and R<NUM> are each independently hydrogen, or methyl,.

In another preferred embodiment, R<NUM> is hydrogen atom, chlorine atom or methyl.

In the second aspect of the present invention, it provides a compound selected from the following group, or a pharmaceutically acceptable salt thereof for use in preventing and/or treating a disease related to transient receptor potential channel protein (TRP), wherein the disease is selected from the group consisting of pain, epilepsy, inflammation, respiratory disorder, pruritus, urinary tract disorder and inflammatory bowel disease:
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>.

In another preferred embodiment, the disease related to transient receptor potential channel protein (TRP) is pain.

In another preferred embodiment, the pain comprises acute inflammatory pain, chronic inflammatory pain, visceral pain, neurogenic pain, fibromyalgia, headache, neuralgia or pain caused by cancer.

In another preferred embodiment, the headache is migraine or muscle tension pain.

In another preferred embodiment, the neuralgia is trigeminal neuralgia, diabetic pain or post-zoster neuralgia.

In another preferred embodiment, the pain is selected from the group consisting of acute pain, fibromyalgia, visceral pain, inflammatory pain, neuralgia, or a combination thereof.

In another preferred embodiment, the other pain is fibromyalgia.

In the third aspect of the present invention, it provides a compound of formula I, or a pharmaceutically acceptable salt thereof;
<CHM>
wherein,.

In another preferred embodiment, R<NUM> and R<NUM> are each independently hydrogen, or methyl.

In the fourth aspect of the present invention, it provides a compound selected from the following group or a pharmaceutically acceptable salt thereof:
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>.

In the fifth aspect of the present invention, it provides a pharmaceutical composition which comprises the compound, or a pharmaceutically acceptable salt thereof according to the third and fourth aspect of the present invention; and a pharmaceutically acceptable carrier.

In the sixth aspect of the present invention, it provides a method for preparing the compound, or a pharmaceutically acceptable salt thereof according to the third and fourth aspect of the present invention, wherein the method comprises: reacting an intermediate of formula II with R<NUM>-NH-R<NUM> in an inert solvent, thereby forming the compound,:
<CHM>
wherein X, Y, A, R<NUM>, R<NUM>, R<NUM> and "*" are as defined in the third aspect of the present invention.

In another preferred embodiment, the method comprises the following step:
<CHM>
wherein X, Y, A, R<NUM>, R<NUM>, R<NUM> and "*" are as defined in the third aspect of the present invention;.

In another preferred embodiment, the method further comprises the following step:
(c1) in an inert solvent and in presence of a condensing agent, reacting the compound of formula Ic with acetic acid to form a compound of formula ID;.

In another preferred embodiment, the method further comprises the following step:
(c2) in an inert solvent and in presence of a base, subjecting the compound of formula IE to hydrolysis reaction to form a compound of formula IF.

In another aspect of the present disclosure, it provides an intermediate of formula II or III for preparing the compounds of the present invention:
<CHM>
wherein, X, Y, A, R<NUM> and "*" are as defined in the third aspect of the present invention.

In the another aspect of the present disclosure, it provides a method for preparing the intermediate for preparing the compounds of the present invention,
<CHM>
wherein X, Y, A, R<NUM>, R<NUM>, R<NUM> and "*" are as defined in the third aspect of the present invention.

In the seventh aspect of the present invention, it provides a method for non-therapeutically and/or non-diagnostically inhibiting activity of transient receptor potential channel protein in vitro, which comprises contacting a transient receptor potential channel protein or a cell expressing the protein with the compound, or a pharmaceutically acceptable salt thereof according to the third and fourth aspect of present invention, thereby inhibiting activity of transient receptor potential channel protein.

It should be understood that, in the present invention, each of the technical features specifically described above and below (such as those in the Examples) can be combined with each other, thereby constituting new or preferred technical solutions which need not be redundantly described one-by-one, as long as they fall within the claims.

Based on an extensive and intensive research, the inventors have unexpectedly and firstly developed a compound of formula I, or a pharmaceutically acceptable salt thereof. The experimental results have shown that the compound of present invention have significant inhibitory effect on TRP channels. The compound of present invention can effectively treat a pain and the like related to TRP (especially TRPA1) targets. On this basis, the inventors have completed the present invention.

As used herein, the terms "comprise", " comprising", and "containing" are used interchangeably, which not only comprise closed definitions, but also semi-closed and open definitions. In other words, the term comprises "consisting of" and "essentially consisting of".

As used herein, "R<NUM>", "R1" and "R<NUM>" have the same meaning and can be used interchangeably. The other similar definitions have the same meaning.

As used herein, the terms "C<NUM>-C<NUM> alkyl" or "C<NUM>-C<NUM> alkyl" refer to a linear or branched chain alkyl with <NUM>-<NUM> or <NUM>-<NUM> carbon atoms, such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, or the like.

As used herein, the term "C<NUM>-C<NUM> alkoxy" refers to a linear or branched alkoxy having <NUM> to <NUM> carbon atoms, such as methoxyl, ethoxyl, propoxyl, isopropoxyl, butoxyl, isobutoxyl, sec-butoxyl, tert-butoxyl, pentoxyl, hexyloxyl, or the like.

As used herein, the term "C<NUM>-C<NUM> benzoalicyclic group" refers to a group having <NUM>-<NUM> carbon atoms, and comprises indanyl, tetrahydronaphthyl, dihydronaphthyl, or the like.

As used herein, the term "C<NUM>-C<NUM> cycloalkyl" refers to a cycloalkyl having <NUM>-<NUM> carbon atoms, and comprises monocyclic, bicyclic or polycyclic ring, such as cyclopropyl, cyclobutyl, methylcyclobutyl, cyclopentyl, cycloheptyl, or the like.

As used herein, the term "C<NUM>-C<NUM> ester group" refers to a group having C1-C5 alkyl-COO- structure or a group having -COO-C1-C5 alkyl structure, wherein the alkyl can be linear or branched chain, such as CH<NUM>COO-, C<NUM>H<NUM>COO-, C<NUM>H<NUM>COO-, (CH<NUM>)<NUM>CHCOO-, -COOCH<NUM>, -COOC<NUM>H<NUM>, -COOC<NUM>H<NUM>, or the like.

As used herein, the term "C<NUM>-C<NUM> amide group" refers to a group having C<NUM>-C<NUM> alkyl-CO-NH- structure or a group having -CO-NH-C<NUM>-C<NUM> alkyl structure, wherein the alkyl can be linear or branched, such as CH<NUM>-CO-NH-, C<NUM>H<NUM>-CO-NH-, C<NUM>H<NUM>-CO-NH-, -COOCH<NUM>, -CO-NH-C<NUM>H<NUM>, -CO-NH-C<NUM>H<NUM>, or the like.

As used herein, the term "C<NUM>-C<NUM> acyl" refers to a group having C<NUM>-C<NUM> alkyl-CO-structure, wherein the alkyl can be linear or branched, such as CH<NUM>-CO-, C<NUM>H<NUM>-CO-, C<NUM>H<NUM>-CO-, or the like.

As used herein, the term "C<NUM>-C<NUM> heterocycloalkyl" refers to a monocyclic and polycyclic heterocycle (preferably monocyclic heterocycle) having <NUM>-<NUM> ring carbon atoms and <NUM>-<NUM> heteroatoms (preferably containing one nitrogen atom, which is the nitrogen atom commonly adjacent to R<NUM> and R<NUM>), such as piperidinyl, tetrahydropyrrolyl, or the like.

As used herein, the term "<NUM>-<NUM> membered carbocyclic ring" refers to any stable <NUM>-, <NUM>- or <NUM>-membered monocyclic, bicyclic or polycyclic ring. The carbocyclic ring can be a saturated, partially unsaturated, or unsaturated ring, but cannot be an aromatic ring. Examples of carbocyclic ring comprise, but are not limited to, cyclopropyl, cyclobutyl, cyclobutenyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloheptyl, cycloheptenyl, adamantyl, cyclooctyl, cyclooctenyl, cyclooctadienyl, bicyclo[<NUM>. <NUM>]octanyl, bicyclo[<NUM>. <NUM>]nonanyl, bicyclo[<NUM>. <NUM>]decanyl, bicyclo[<NUM>. <NUM>] octanyl, fluorenyl, indanyl.

As used herein, the term "heterocyclic ring" refers to any stable monocyclic, bicyclic or polycyclic ring, for example, <NUM>-, <NUM>- or <NUM> membered ring, wherein the heterocyclic ring contains one or more (such as <NUM>-<NUM>) heteroatom selected from N, O and S, the heterocyclic ring can be a saturated, partially unsaturated, or unsaturated ring, but can not be an aromatic ring.

As used herein, the terms "C<NUM>-C<NUM> haloalkyl" and "C<NUM>-C<NUM> haloalkyl" mean that one or more hydrogen atoms of a linear or branched alkyl having <NUM>-<NUM> and <NUM>-<NUM> carbon atoms are substituted by halogen, such as monochloromethanyl, dichloroethanyl, trichloropropanyl, or the like.

As used herein, the term "C<NUM>-C<NUM> carboxyl" refers to C<NUM>-C<NUM> alkyl-COOH, wherein the alkyl can be linear or branched, such as CH<NUM>COOH, C<NUM>H<NUM>COOH, C<NUM>H<NUM>COOH, (CH<NUM>)<NUM>CHCOOH, or the like.

As used herein, the term "C<NUM>-C<NUM> aryl" refers to a monocyclic or bicyclic aromatic hydrocarbon group having <NUM> to <NUM> carbon atoms in the ring, such as phenyl, naphthyl, biphenyl, or the like.

As used herein, the term "heteroaryl" refers to an optionally substituted aromatic group (for example, <NUM>- to <NUM>-membered monocyclic ring) which contains at least one heteroatom and at least one carbon atom, for example pyrrolyl, thienyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, furanyl, imidazolyl, thiazolyl, oxazolyl, triazolyl or the like.

As used herein, the term "halogen" refers to F, Cl, Br, and I.

As used herein, the term "substituted" means that a hydrogen atom on the group is replaced by a non-hydrogen atom group, but the valence requirement must be met and the substituted compound is chemically stable. In the specification, it should be understood that each substituent is unsubstituted, unless it is expressly described as "substituted" herein.

Similarly, it should be understood that substituents can be connected to a parent group or a substrate on any atom in present invention, unless the connection violates the valence requirement; and the hydrogen atoms of parent group or substrate can be on the same atom or different atoms.

As used herein, as for a numerical range from P1 to P2, the range contains not only the endpoints P1 and P2, but also any numerical points between the endpoints P1 and P2. In addition, when both P1 and P2 are positive numbers, for an integer n, its value range contains any integer value point between the endpoints P1 and P2. For example, for an integer, when its value range is <NUM>-<NUM>, it contains <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM> and <NUM>; the value range <NUM>-<NUM> contains <NUM>, <NUM>, <NUM>, <NUM>, and <NUM>. Typically, as for a group, C<NUM>-C<NUM> contains C3, C4, C5, C6 and C7.

As used herein, the terms "compound of present invention" and "<NUM>-aryloxy-<NUM>-aryl-propylamine compound of present invention" refer to a compound or a pharmaceutically acceptable salt thereof according to the third and fourth second aspect of the present invention,. It should be understood that the term also comprises a mixture of the above components.

The compound of present invention not only has inhibitory effect on TRPA1, but also has certain inhibitory effect on other members of TRP family.

The term "pharmaceutically acceptable salt" refers to a salt formed by a compound of the present invention and an acid or a base suitable for use as a medicine. Pharmaceutically acceptable salts include inorganic salts and organic salts. A preferred type of salt is the salt formed by the compound of the present invention and an acid. Acids suitable for salt formation include (but are not limited to): inorganic acid such as hydrochloric acid, hydrobromic acid, hydrofluoric acid, sulfuric acid, nitric acid, phosphoric acid and the like; organic acid such as formic acid, acetic acid, propionic acid, oxalic acid, malonic acid, succinic acid, fumaric acid, maleic acid, lactic acid, malic acid, tartaric acid, citric acid, picric acid, methane sulfonic acid, toluene sulfonic acid, benzenesulfonic acid and the like; and acidic amino acid such as aspartic acid and glutamic acid. A preferred type of salt is a metal salt formed by the compound of the present invention and a base. Suitable bases for salt formation include (but are not limited to): inorganic base such as sodium hydroxide, potassium hydroxide, sodium carbonate, sodium bicarbonate, sodium phosphate and the like; and organic base such as ammonia, triethylamine, diethylamine and the like.

The preferred compounds of present invention comprise any compound selected from Table <NUM> (except Compounds IA-<NUM> to IA-<NUM> and IB-<NUM> to IB-<NUM>, which are provided as reference examples):.

The present disclosure further provides a method for preparing <NUM>-aryloxy-<NUM>-aryl-propylamine compounds of formula IA to IF.

The present invention further provides a method for preparing intermediates of formula II-III which are useful for preparing the above-mentioned compounds.

The specific synthesis method are as follows:.

Potassium tert-butoxide is added into dimethyl sulfoxide (DMSO) solution of (S)-<NUM>-(methylamino)-<NUM>-(thiophen-<NUM>-yl)-<NUM>-propanol, and the mixture is stirred for <NUM>-<NUM> minutes. - Quinoline or isoquinoline substituted by fluorine at different position is added, and the reaction is carried out at <NUM>-<NUM> overnight. After the reaction is completed, water is added into the reaction system, then reaction system is extracted three times with ethyl acetate, washed with water and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The residue is separated by column chromatography to obtain compounds of formula IA and IB.

<CHM>
wherein, A, X, Y, R<NUM>, R<NUM>, R<NUM> and "*" are as defined above.

The compound of the present invention can be converted into its pharmaceutically acceptable salt by conventional methods. For example, a solution of corresponding acid can be added into the solution of above compounds, and the solvent is removed under reduced pressure after the salt is formed, thereby forming the corresponding salt of the compound of the present invention.

Transient receptor potential channel protein is a protein superfamily composed of important cation channels on the cell membrane. Transient receptor potential channel protein comprises multiple subfamily, such as TRPA, TRPC, TRPM, TRPV, TRPML and TRPP subfamily.

Studies have found that TRPA1 channel protein is related to a disease such as pain, epilepsy, inflammation, respiratory disorder, pruritus, urinary tract disorder, inflammatory bowel disease, etc. TRPA1 is a target useful for treating a disease such as pain, epilepsy, inflammation, respiratory disorder, pruritus, urinary tract disorder and inflammatory bowel diseases, etc..

The present invention further provides a method for inhibiting transient receptor potential channel protein (TPR), and a method for treating a disease related to transient receptor potential channel protein.

The compound of present invention can inhibit transient receptor potential channel protein, thereby preventing or treating a disease related to transient receptor potential channel protein.

In the present invention, the disease related to transient receptor potential channel protein is selected from: pain, epilepsy, inflammation, respiratory disorder, pruritus, urinary tract disorder, and inflammatory bowel disease. Representatively, the pain comprises (but is not limited to): acute inflammatory pain, chronic inflammatory pain, visceral pain, neurogenic pain, fibromyalgia, headache (such as migraine, muscle tension pain, etc.), neuralgia (such as trigeminal neuralgia, diabetic pain, post-zoster neuralgia, etc.), or pain caused by cancer.

In another aspect, the present invention provides a method for non-therapeutically and non-diagnostically inhibiting the activity of transient receptor potential channel protein in vitro, which comprises, e.g., in an in vitro culture system, contacting a transient receptor potential channel protein or a cell expressing the protein with the compound or a pharmaceutically acceptable salt thereof of the present invention, thereby inhibiting the activity of transient receptor potential channel protein.

The invention provides a composition for inhibiting activity of transient receptor potential channel protein. The composition comprises (but is not limited to): pharmaceutical composition, food composition, dietary supplement, beverage composition, etc..

Typically, the composition is a pharmaceutical composition which comprises the compound , or a pharmaceutically acceptable salt thereof of the present invention; and a pharmaceutically acceptable carrier.

In the present invention, the dosage form of pharmaceutical composition comprises (but is not limited to) oral preparation, injection and external preparation.

Representatively, the dosage form comprises (but is not limited to): tablet, injection, infusion, ointment, gel, solution, microsphere, and film.

The term "pharmaceutically acceptable carrier" refers to one or more compatible solid, semi-solid, liquid or gel fillers, which are suitable for use in humans or animals and must have sufficient purity and sufficient low toxicity. The "compatible" means each ingredient of the pharmaceutical composition and active ingredient of the drug can be blended with each other without significantly reducing the efficacy.

It should be understood that the carrier is not particularly limited. In the present invention, the carrier can be selected from materials commonly used in the art, or can be obtained by a conventional method, or is commercially available. Some examples of pharmaceutically acceptable carriers are cellulose and its derivatives (such as methylcellulose, ethylcellulose, hydroxypropylmethylcellulose, sodium carboxymethylcellulose, etc.), gelatin, talc, solid lubricants (such as stearic acid, magnesium stearate), calcium sulfate, plant oil (such as soybean oil, sesame oil, peanut oil, olive oil, etc.), polyols (such as propylene glycol, glycerin, mannitol, sorbitol, etc.), emulsifier (such as Tween), wetting agent (such as sodium lauryl sulfate), buffer agent, chelating agent, thickener, pH regulator, transdermal enhancer, colorant, flavoring agent, stabilizer, antioxidant, preservative, bacteriostatic agent, pyrogen-free water, etc..

Representatively, in addition to the active pharmaceutical ingredient, the liquid formulations can contain inert diluents commonly used in the art, such as water or other solvents, solubilizers and emulsifiers, for example, ethanol, isopropanol, ethyl carbonate, ethyl acetate, propylene glycol, <NUM>,<NUM>-butanediol, dimethylformamide and oils, especially cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil and sesame oil or a mixture thereof. In addition to these inert diluents, the composition can also contain adjuvants such as wetting agents, emulsifiers and suspensions and the like.

The pharmaceutical preparation should be matched with the mode of administration. The formulation of present invention can also be used together with other synergistic therapeutic agents (including before, simultaneous or after administering). When a pharmaceutical composition or preparation is administered, a safe and effective dose of drug is administered to a subject in need (e.g. human or non-human mammals). The safe and effective dose is usually at least <NUM>µg/kg body weight, and does not exceed about <NUM>/kg body weight in most cases, and preferably the dose is about <NUM>µg/kg body weight to about <NUM>/kg body weight. Of course, the route of administration, patient health and other factors, should also be taken into account to determine the specific dose, which are within the ability of the skilled physicians.

The present invention will be further illustrated below with reference to the specific examples. The experimental methods with no specific conditions described in the following examples are generally performed under the conventional conditions, or according to the manufacturer's instructions. Unless indicated otherwise, parts and percentage are calculated by weight.

<NUM> of (S)-<NUM>-(methylamino)-<NUM>-(thiophen-<NUM>-yl)propan-<NUM>-ol was dissolved in <NUM> of dimethyl sulfoxide solution, <NUM> of potassium tert-butoxide was added, the mixture was stirred at room temperature for <NUM> minutes, <NUM> of <NUM>-fluoroquinoline was added, and the reaction was carried out at <NUM> overnight. After the reaction was completed, water was added into the reaction system, the mixture was extracted three times with ethyl acetate, washed with water and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated by column chromatography (methanol/dichloromethane=<NUM>:<NUM>) to afford <NUM> of the title compound as brown oil. Yield: <NUM>%.

<NUM>H-NMR (<NUM>, CDCl<NUM>) δ <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dt, J = <NUM>, <NUM>, <NUM>), <NUM> (dt, J = <NUM>, <NUM>, <NUM>), <NUM> (dq, J = <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that <NUM>-fluoroquinoline was replaced with <NUM>-fluoroquinoline, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM>. <NUM> of the title compound as brown oil was obtained. Yield: <NUM>%.

<NUM>H-NMR (<NUM>, CDCl<NUM>) δ <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> (td, J = <NUM>, <NUM>, <NUM>), <NUM> (dq, J = <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

<NUM>H-NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J= <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (ddd, J = <NUM>, <NUM>, <NUM>, <NUM>), <NUM> (ddd, J = <NUM>, <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (hept, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

<NUM>H-NMR (<NUM>, CDCl<NUM>) δ <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dt, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dtd, J = <NUM>, <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that <NUM>-fluoroquinoline was replaced with <NUM>-fluoroisoquinoline, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM>. <NUM> of the title compound as brown oil was obtained. Yield: <NUM>%.

<NUM>H-NMR (<NUM>, CDCl<NUM>) δ <NUM> (s, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (q, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

<NUM>H-NMR (<NUM>, CDCl<NUM>) δ <NUM> (s, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (q, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> (dq, J = <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that <NUM>-fluoroquinoline was replaced with <NUM>-fluoroisoquinoline, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM>, <NUM> of the title compound as brown oil was obtained. Yield: <NUM>%.

<NUM>H-NMR (<NUM>, CDCl<NUM>) δ <NUM> (s, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

<NUM> of (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propan-<NUM>-ol, <NUM> of <NUM>-hydroxybenzofuran and <NUM> of triphenylphosphine were dissolved in <NUM> of anhydrous tetrahydrofuran, and <NUM>µl of diisopropyl azodicarboxylate was slowly added dropwise into the system under ice bath condition. After the dropwise addition was completed, the system was shifted to room temperature and reacted overnight. After the reaction was completed, the system was directly spin-dried, and the residue was separated and purified by column chromatography to afford <NUM> of the title compound as colorless oil. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (t, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dt, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (ddd, J = <NUM>, <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

<NUM> of Intermediate II-<NUM> was dissolved in saturated sodium iodide solution in acetone, and the mixture was refluxed overnight. After the reaction was completed, the solvent was spin-dried, water was added into the system, and the mixture was extracted three times with ethyl acetate, washed with saturated brine, dried with anhydrous sodium sulfate, filtered, and concentrated. The residue was dissolved in <NUM> of tetrahydrofuran solution, <NUM> of <NUM> % methylamine aqueous solution was added, and the reaction was carried out overnight. After the reaction was completed, the solvent was spin-dried, sodium hydroxide aqueous solution was added into the system, then the mixture was extracted with ethyl acetate three times, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated by column chromatography (methanol/dichloromethane=<NUM>:<NUM>) to afford <NUM> of the title compound as colorless oil. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol was replaced with (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)prop-<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as yellow oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol was replaced with (R)-<NUM>-chloro-<NUM>-(furan-<NUM>-yl)prop-<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as colorless oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> (td, J = <NUM>, <NUM>, <NUM>), <NUM> (dtd, J = <NUM>, <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (ddd, J = <NUM>, <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol was replaced with (R)-<NUM>-chloro-<NUM>-(<NUM>-methylthiophen-<NUM>-yl) propyl -<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as colorless oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol was replaced with (R)-<NUM>-chloro-<NUM>-(<NUM>-chlorothiophene-<NUM>-yl)propyl-<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as colorless oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (ddd, J = <NUM>, <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol was replaced with <NUM>-chloro-<NUM>-(thiazol-<NUM>-yl)propyl-<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. After racemate of compound Ic-<NUM> was obtained, <NUM> of the title compound as yellow oil was obtained by chiral resolution. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that methylamine aqueous solution was replaced with dimethylamine, the other required raw materials, reagents and preparation method were the same as Example <NUM>. <NUM> of the title compound as colorless oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (dt, J = <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (dt, J = <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that <NUM>-hydroxybenzofuran was replaced with <NUM>-hydroxybenzofuran, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as yellow oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (td, J = <NUM>, <NUM>, <NUM>), <NUM> (dt, J = <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that <NUM>-hydroxybenzofuran was replaced with <NUM>-indanol, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as brown oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dt, J = <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (p, J = <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol was replaced with (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Example <NUM>. <NUM> of the title compound as brown oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dt, J = <NUM>, <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (ddd, J = <NUM>, <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol was replaced with (R)-<NUM>-chloro-<NUM>-(furan-<NUM>-yl)propyl-<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Example <NUM>. <NUM> of the title compound as brown oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> (td, J = <NUM>, <NUM>, <NUM>), <NUM> (ddd, J = <NUM>, <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J= <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> (td, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol was replaced with (R)-<NUM>-chloro-<NUM>-(thiazol-<NUM>-yl)propyl-<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Example <NUM>. After racemate of compound IC-<NUM> was obtained, <NUM> of the title compound as yellow oil was obtained by chiral resolution. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that methylamine aqueous solution was replaced with dimethylamine, the other required raw materials, reagents and preparation method were the same as Example <NUM>. <NUM> of the title compound as brown oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dt, J = <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that <NUM>-hydroxybenzofuran was replaced with <NUM>-indanol, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as yellow oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (dt, J = <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (q, J = <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that <NUM>-hydroxybenzofuran was replaced with tetrahydronaphthol, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as yellow oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that <NUM>-hydroxybenzofuran was replaced with <NUM>,<NUM>-dihydronaphthol, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as brown oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, DMSO) δ <NUM> (s, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that <NUM>-hydroxybenzofuran was replaced with <NUM>-hydroxy-<NUM>-methylisoindoline-<NUM>,<NUM>-dione, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as brown oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (s, <NUM>), <NUM> (s, <NUM>), <NUM> (s, <NUM>), <NUM> (s, <NUM>), <NUM> (d, J = <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that <NUM>-hydroxybenzofuran was replaced with <NUM>-hydroxyimidazo[<NUM>,<NUM>-a]pyridine, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as brown oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (d, J= <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that <NUM>-hydroxybenzofuran was replaced with <NUM>-hydroxy-<NUM>-benzo[d]imidazole-<NUM>(<NUM>)-one, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as brown oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CD<NUM>OD) δ <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that <NUM>-hydroxybenzofuran was replaced with <NUM>-fluoroquinoxaline, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as brown oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J= <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (s, <NUM>), <NUM> (s, <NUM>), <NUM> (s, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

<NUM> of Intermediate II-<NUM>, <NUM> of potassium phthalimide and <NUM> of sodium iodide were dissolved in <NUM> of N,N-dimethylformamide, and the reaction was carried out at <NUM> under nitrogen protection overnight. After the reaction was completed, water was added into the system, and the mixture was extracted three times with ethyl acetate, washed with water and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated by column chromatography (ethyl acetate/petroleum ether=<NUM>: <NUM>) to afford <NUM> of the title compound as yellow solid. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM>-<NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dt, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (td, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

<NUM> of Intermediate III-<NUM> and <NUM> of hydrazine hydrate were dissolved in <NUM> of methanol solution, and the reaction was carried out at room temperature overnight. After the reaction was completed, the solvent was spin-dried, and the residue was separated by column chromatography (methanol/dichloromethane=<NUM>:<NUM>) to afford <NUM> of the title compound as colorless oil. Yield: <NUM>%.

<NUM>H NMR (<NUM>, DMSO) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (ddt, J = <NUM>, <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol was replaced with (R)-<NUM>-chloro-<NUM>-(furan-<NUM>-yl)propyl-<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Reference Examples <NUM> and <NUM> and Example <NUM>. <NUM> of the title compound as colorless oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (ddt, J = <NUM>, <NUM>, <NUM>, <NUM>), <NUM> (dtd, J = <NUM>, <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM>(M+H)+.

<NUM> NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dq, J = <NUM>, <NUM>, <NUM>), <NUM> (qd, J = <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM>(M+H)+.

Except that (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol was replaced with (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and <NUM> and Example <NUM>. <NUM> of the title compound as colorless oil was obtained. Yield: <NUM>%.

<NUM> NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM>(M+H)+.

Except that <NUM>-hydroxybenzofuran was replaced with benzo[b]thiophen-<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as yellow oil was obtained. Yield: <NUM>%.

<NUM> NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> (tt, J = <NUM>, <NUM>, <NUM>), <NUM> (ddd, J = <NUM>, <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM>(M+H)+.

Except that (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol was replaced with (R)-<NUM>-chloro-<NUM>-(furan-<NUM>-yl)propan-<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Example <NUM>. <NUM> of the title compound as colorless oil was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (td, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM>(M+H)+.

Except that (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol was replaced with (R)-<NUM>-chloro-<NUM>-(oxazol-<NUM>-yl)propyl-<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Reference Example <NUM> and Example <NUM>. <NUM> of the title compound as yellow oil was obtained. Yield: <NUM>%.

<NUM> NMR (<NUM>, CDCl<NUM>) δ <NUM> (s, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM>(M+H)+.

<NUM> of acetic acid, <NUM> of <NUM>-(<NUM>-dimethylaminopropyl)-<NUM>-ethylcarbodiimide hydrochloride, <NUM> of <NUM>-dimethylaminopyridine and <NUM> of triethylamine were dissolved in <NUM> anhydrous dichloromethane, the mixture was stirred at room temperature for <NUM>, <NUM> of dichloromethane solution containing <NUM> of compound Ic-<NUM> was added into the system, and the reaction was carried out at room temperature overnight. After the reaction was completed, water was added into the system, the mixture was extracted three times with ethyl acetate, washed with saturated brine, dried with anhydrous sodium sulfate, filtered, and concentrated. The residue was separated by column chromatography (methanol/dichloromethane=<NUM>:<NUM>) to afford <NUM> of the title compound as yellow oil. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>). MS (ESI, m/z): <NUM>(M+H)+.

<NUM> of Intermediate II-<NUM>, <NUM> of methyl L-proline ester hydrochloride, <NUM> of potassium carbonate and <NUM> of sodium iodide were dissolved in <NUM> of acetonitrile, and the reaction was carried out at <NUM> overnight under nitrogen protection. After the reaction was completed, water was added into the system, the mixture was extracted with ethyl acetate three times, washed with water and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was separated by column chromatography (ethyl acetate/petroleum ether = <NUM>: <NUM>) to afford <NUM> of the title compound as colorless oil. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (ddd, J = <NUM>, <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (s, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (s, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (s, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

<NUM> of compound IE and <NUM> of sodium hydroxide were dissolved in <NUM> mixed solution of water and tetrahydrofuran, and the reaction was carried out at <NUM> overnight. After the reaction was completed, water was added into the system, and the reaction system was adjusted to pH=<NUM> with diluted hydrochloric acid, extracted with ethyl acetate three times, washed with water and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to afford <NUM> of the title compound as colorless oil. Yield: <NUM>%.

<NUM> NMR (<NUM>, DMSO) δ <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (td, J = <NUM>, <NUM>, <NUM>), <NUM> (ddd, J = <NUM>, <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (ddd, J = <NUM>, <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

Except that (R)-<NUM>-chloro-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-ol was replaced with (R)-<NUM>-chloro-<NUM>-phenylpropane-<NUM>-ol, the other required raw materials, reagents and preparation method were the same as Example <NUM>. <NUM> of compound C1 was obtained. Yield: <NUM>%.

<NUM>H NMR (<NUM>, CDCl<NUM>) δ <NUM> - <NUM> (m, <NUM>), <NUM> (t, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (d, J = <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, J = <NUM>, <NUM>, <NUM>), <NUM> (dt, J = <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>). MS (ESI, m/z): <NUM> (M+H)+.

The inhibitory activity of some compounds in the Examples of present invention on transient receptor potential channel protein TRPA1 was tested in this example. The compound of formula A (<CIT>) was used as a positive control compound:
<CHM>.

The method was as below:
IonWorks Barracuda (IWB) automated patch clamp detection was used as test method: HEK293 cells stably expressing TRPA1 were placed in DMEM medium containing <NUM>µg/mL Blasticidin S HCl, <NUM>µg/mL Hygromycin B and <NUM>% FBS in the T175 culture flask, and cultured in <NUM>, <NUM>% CO<NUM> incubator. When the cell density reached about <NUM>%, the culture medium was removed, rinsed with phosphate buffered saline (PBS) without calcium and magnesium. <NUM> of Trypsin was added to digest for <NUM>, <NUM> of culture medium was added to terminate the digestion. The cells were collected to <NUM> centrifuge tube and centrifuged at <NUM> rpm for <NUM>. After the supernatant was removed, the cells were added to appropriate volume of extracellular fluid for re-suspending, and the cell density was controlled at <NUM>-<NUM>×<NUM><NUM>/mL for IWB experiment. Extracellular fluid formulation (in mM): <NUM> NaCl, <NUM> KCl, <NUM> MgCl<NUM>, <NUM> HEPES, <NUM> EGTA, <NUM> Glucose (pH <NUM>); intracellular fluid formulation (in mM): <NUM> CsCl, <NUM> HEPES, <NUM> EGTA, <NUM> CaCl<NUM>, <NUM> MgCl<NUM> (pH <NUM>). <NUM>/mL of amphotericin B was freshly prepared with DMSO on the day of experiment, and then final concentration of <NUM>/mL was prepared with intracellular fluid.

Population patch clamp (PPC) plate was used in IWB experiment. The entire detection process was automatically carried out by the instrument. Extracellular fluid was added into <NUM> wells of PPC plate, and <NUM> of the intracellular fluid was added into plenum (under the PPC plate), <NUM> of cell fluid was added for sealing test, and finally the intracellular fluid in plenum was replaced with amphotericin B-containing intracellular fluid to establish a whole-cell recording mode after perforating sealed cells. The sampling frequency for recording TPRA1 current was <NUM>, the cells were clamped at <NUM> mV, the voltage stimulation command (channel protocol) was a ramp voltage from -<NUM> mV to +<NUM> mV for <NUM>. This voltage stimulation was applied every <NUM>, mTRPA1 current was induced by <NUM> AITC.

Data recording and current amplitude measurement export were carried out by IWB software (version <NUM>. <NUM>, Molecular Devices Corporation, Union City, CA). No statistics was recorded for holes with sealing impedance lower than <NUM> MΩ. The original current data was corrected by software for leakage reduction. TRPA1 current amplitude was measured at +<NUM> mV. Each PPC plate in the experiment had a dose-effect data of HC030031 as a positive control. If IC<NUM> value of HC030031 exceeded <NUM> times the average IC<NUM> value previously obtained on each plate, it would be re-tested. The dose-effect curve of compounds and IC<NUM> were fitted and calculated by GraphPad Prism <NUM> (GraphPad Software, San Diego, CA).

Some compounds of present invention were detected by IonWorks Barracuda (IWB) automated patch clamp detection method for IC<NUM> inhibitory activity. The activity data was shown in Table <NUM>, and the dose-effect relationship of some representative compounds in inhibiting TRPA1 activity was shown in <FIG>.

The results showed that the compounds of present invention exhibited potent inhibitory activity on TRPA1, wherein the IC<NUM> values of <NUM> compounds on TRPA1 were <NUM>-<NUM>. It could be seen from <FIG> that the IC<NUM> values of compounds of Ic-<NUM>, Ic-<NUM>, Ic-<NUM>, Ic-<NUM>, and Ic-<NUM> on TRPA1 were <<NUM>. Therefore, it could be concluded that the compounds of formula I of present invention had potent inhibitory activity on TRPA1.

In addition, the test results of S configuration of compound IC-<NUM> and the corresponding R configuration of compound IC-<NUM>, i.e., (R)-<NUM>-(<NUM>,<NUM>-dihydro- <NUM>-inden-<NUM>-yl)oxy)-N-methyl-<NUM>-(<NUM>-thienyl)- <NUM>-amine, and S configuration of compound Ic-<NUM> and the corresponding R configuration of compound, i.e., (R)-<NUM>-(benzofuran-<NUM>-yloxy)-<NUM>-(thiophen-<NUM>-yl)propyl-<NUM>-amine showed that the ratio of IC<NUM> of R configuration of compound to that of S configuration of compound was ≥<NUM>, which suggested that compounds of the present invention in S configuration had higher inhibitory activity on TRPA1.

In addition, the activity ratio of compound Ic-<NUM> (containing heteroaryl
<CHM>
) to compound C1 (containing phenyl), i.e., the IC<NUM> of compound Cl/the IC<NUM> of compound Ic-<NUM>, was about <NUM>-fold, which suggested that the compounds of present invention containing heteroaryl (such as compound Ic-<NUM>) had higher inhibitory activity on TRPA1.

In addition, compared with compounds with naphthalene ring as A group, such as duloxetine, the IC<NUM> values of compound Ic-<NUM> with benzoalicyclic ring as A group, compound Ic-<NUM>, compound Ic-<NUM>, and compound Ic-<NUM> with benzoheteroaryl as A group were significantly decreased. Specifically, the ratio of the IC<NUM> of S configuration of duloxetine to the IC<NUM> of any of compound Ic-<NUM>, compound Ic-<NUM>, compound Ic-<NUM> or compound Ic-<NUM> was about <NUM>-<NUM> (note: the IC<NUM> of R configuration of duloxetine was <NUM>, and the IC<NUM> of S configuration of duloxetine was <NUM>). The results suggested that the compounds of present invention with benzoalicyclyl or heteroaryl as A group had higher (about <NUM>-<NUM> times higher) inhibitory activity on TRPA1.

Similarly, the inventors also measured the TRPA1 inhibitory activity of S configuration of duloxetine and compound Ic-<NUM> by manual patch clamp detection. Similar to the test results of automatic patch clamp detection method, in the manual patch clamp detection, the ratio of IC<NUM> of the S configuration of duloxetine to the IC<NUM> of the compound Ic-<NUM> was <NUM>/<NUM>=<NUM>.

Similarly, the inventors also measured the TRPA1 inhibitory activity of compound Ic-<NUM> by manual patch clamp detection. The method was as follows:
The HEK293 cell line stably expressing human TRPA1 channel was placed in T75 culture flask with DMEM medium containing <NUM>µg/mL Blasticidin S HCl, <NUM>µg/mL Hygromycin B and <NUM>% FBS, and cultured in incubator at <NUM> and <NUM>% CO<NUM>. When the cell density reached about <NUM>%, the culture medium was removed, the residue was rinsed with phosphate buffered saline (PBS) without calcium and magnesium. <NUM> of trypsin was added to digest for <NUM> minutes, and <NUM> of culture medium was added to terminate the digestion. The cells were collected to <NUM> centrifuge tube and centrifuged at <NUM> rpm for <NUM>. After the supernatant was removed, an appropriate volume of extracellular fluid was added to re-suspend the cells.

In manual patch clamp detection, HEKA system (Patch Master software) was used together with EPC-<NUM> amplifier to record the whole cell current of TRPA1 stably transfected cell line at room temperature. intracellular fluid formulation for whole cell recording was as follows(mM): <NUM> CsCl, <NUM> HEPES, <NUM> EGTA, <NUM> CaCl<NUM>, <NUM> MgCl<NUM> (pH <NUM>, osmotic pressure <NUM>-<NUM> mOsm); Ca<NUM>+-free extracellular fluid formulation for recording was as follows (mM): <NUM> NaCl, <NUM> KCl, <NUM> EGTA, <NUM> MgCl<NUM>, <NUM> Glucose, <NUM> HEPES (pH <NUM>, osmotic pressure <NUM>-<NUM> mOsm). Glass microelectrode resistance used for patch clamp recording was <NUM>-<NUM> MΩ, the sampling frequency was <NUM>, the filter frequency was <NUM>, the cell clamp was <NUM> mV, and the voltage stimulation command (channel protocol) was a linear voltage from -<NUM> mV to +<NUM> mV for <NUM>, then restored to <NUM> mV clamping potential. The recording was performed every <NUM>. The hTRPA1 current was induced by <NUM> AITC. To ensure the accuracy of current recording, the series resistance was used for <NUM>% compensation during recording.

Data recording and current amplitude measurement export were completed by Patch Master software. Cells with sealing impedance lower than <NUM> MΩ w were not included in the statistics. The original current data were corrected by software for leakage reduction, and the hTRPA1 current amplitude was measured at +<NUM> mV. The compound dose-effect curve and IC<NUM> were fitted and calculated by GraphPad Prism <NUM> (GraphPad Software, San Diego, CA).

Similar to the results of the automatic patch clamp detection, the ratio of IC<NUM> of S configuration of duloxetine to that of compound Ic-lwas <NUM> /<NUM> = <NUM> in the manual patch clamp detection.

In this example, the method for determining hepatotoxicity and neurotoxicity of compound Ic-<NUM> and compound Ic-<NUM> were as follows:.

The results of hepatotoxicity (HepG2 cell) and neurotoxicity (SH-SY5Y) of compound Ic-<NUM> and compound Ic-<NUM> showed: the hepatotoxicity and neurotoxicity of duloxetine were <NUM> and <NUM> (IC<NUM>, µM), respectively, while the hepatotoxicity and neurotoxicity of compound Ic-<NUM> and compound Ic-<NUM> of the present invention were about <NUM>-<NUM> (IC<NUM>, µM). It suggested that the toxicity and side effects of compounds of the present invention were significantly lower, and were about <NUM>/<NUM> or <NUM>/<NUM> of toxicity and side effects of duloxetine. The results showed the compounds of the present invention had excellent safety.

In this example, analgesic activity of compound Ic-<NUM> of present invention was evaluated by mice formalin pain model. The method was as follows:
<NUM> C57BL/<NUM> mice (male, <NUM>-week aged) were randomly divided into <NUM> groups: solvent control group (vehicle, saline), duloxetine group (duloxetine, <NUM>-HT reuptake and NE reuptake inhibitor) and Ic-<NUM> group (compound Ic-<NUM> of the present invention). Before the start of experiment, the mice were allowed to adapt to the experimental environment for <NUM> without fasting. The test drug was administrated by intraperitoneal injection at a dose of <NUM>/kg, and then the mice were placed in a transparent, ventilated plexiglass cylinder for <NUM>, and then <NUM>µl of <NUM>% formalin solution was injected into the left hind plantar of mice of each group by microinjector. The claw pain response of mice was real-time recorded by miniature camera. The number of times of lifting (<NUM>/time), shaking (<NUM>/time), and licking (<NUM>/time) left claw and the time length of licking left claw were used as indicators of pain response, the cumulative scoring and licking time in two stages of <NUM>-<NUM> (phase I, acute pain phase) and <NUM>-<NUM> (phase II, inflammatory pain phase) were observed and recorded, and the statistical analysis was conducted.

The results of analgesic activity of compound Ic-<NUM> of the present invention in mice formalin pain model were shown in <FIG>. The results showed that in the statistical detection indicator of licking time, the compound Ic-<NUM> of the present invention had showed significant analgesic activity in both phase I (<NUM>-<NUM>) and phase II (<NUM>-<NUM>) at a dose of <NUM>/kg, and almost completely inhibited licking claw behavior caused by pain as compared with saline group, and had similar analgesic activity to clinical drug duloxetine.

In this example, the analgesic activity of compound Ic-<NUM>, compound Ic-<NUM>, compound Ic-<NUM> and other compounds of the present invention were evaluated by C57 mice hot plate pain model. The method was as follows:.

SPF-grade C57 male mice were placed at hot plate with constant <NUM>±<NUM>. Mice with painful response such as licking claw within <NUM>-<NUM> were selected (the mice which evade and jump were abandoned). If the pain reaction of mice was observed, the mice were taken out immediately to prevent mice from scalding.

The <NUM> selected animals were weighed and randomly divided into <NUM> groups: saline control group (blank control), duloxetine group (positive control group), gabapentin group (positive control group) and Ic-<NUM> group (compound of the present invention).

The test compounds were freshly prepared on the day of administration. <NUM>% NaCl physiological saline solution was prepared as solvent for later use. Appropriate amount of test compounds were added into required volume of physiological saline and fully suspended, and the concentration of the drug compound in preparation was <NUM>/ml. The standard volume of administration to mice was <NUM>/kg (or <NUM>/<NUM> ).

The administration mode was intraperitoneal administration. animals did not need to fast before administration. The administration volume was <NUM>/kg. The dosage of duloxetine and Ic-<NUM> was <NUM>/kg, and the dosage of gabapentin was <NUM>/kg.

Hot plate observation index: the reaction time of mice on the hot plate at <NUM>±<NUM> (Time latency). Measurement and recording were conducted at <NUM> before administration and <NUM>, <NUM>, and <NUM> after administration.

Maximum possible effect (MPE) % was used to evaluate analgesic effect of each compound, i.e., MPE%=[(Post drug latency-baseline latency)/(<NUM>-baseline latency)]x100. MPE% at different time points was recorded. The higher the value of MPE% was, the stronger the analgesic effect of the compound was.

The results of analgesic activity of compound Ic-<NUM> of the present invention in the mice hot plate pain model were shown in <FIG>. The results showed that compared with the saline control group, the compound Ic-<NUM> of the present invention showed very potent analgesic effect at a dose of <NUM>/kg with a significant difference. Compared with the positive control group, the analgesic activity of the compound Ic-<NUM> of the present invention was significantly stronger than <NUM>/kg of gabapentin, and stronger than <NUM>/kg of duloxetine within <NUM> minutes.

Moreover, the analgesic activity of compound Ic-<NUM> and compound Ic-<NUM> were stronger than those of gabapentin and duloxetine at the same dose (<NUM>/kg),.

The hot plate pain model was a classic model for evaluating the efficacy of drugs on acute pain. Therefore, the compounds of the present invention had excellent therapeutic effect on acute pain.

In this example, the pharmacokinetic properties of compounds such as duloxetine and compound Ic-<NUM> were tested in rat. The method was as follows:.

A certain amount of sample was weighed and dissolved in deionized water to prepare <NUM>/mL of solution. Male SD rats were used as test animals. The dose of single intravenous (IV) injection was <NUM>/kg, and the dose of oral (PO) administration was <NUM>/kg. Each group had three rats. The rats in oral group were fasted for <NUM>-<NUM> hours before administration, and were fed <NUM> hours after administration. Animal blood collection time points were as follows: before administration, and <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, and <NUM> after administration for intravenous administration; before administration, and <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM> and <NUM> after administration for oral administration. About <NUM> of blood was collected via the jugular vein from each animal, and heparin sodium was used for anticoagulation. After collecting the blood samples, the blood samples were placed on ice and centrifuged to separate the plasma (centrifugation conditions: <NUM> rpm, <NUM>, <NUM>). The collected plasma was stored at -<NUM> before analysis. <NUM>µL of plasma sample was taken into <NUM> of centrifuge tube, <NUM>µL of internal standard solution was added, and the same volume of methanol rather than internal standard solution was added in the blank group. The mixture was vortexed and mixed, centrifuged at <NUM>,<NUM> rpm for <NUM>, <NUM>µL of supernatant was taken into <NUM>-well sample plate, and the LC-MS/MS was used for sample analysis. In the linear regression analysis, the peak area was taken as the y-axis and the drug concentration was taken as the x-axis. The linear relationship between the peak area ratio and the concentration was expressed by the correlation coefficient (R) obtained from the regression equation of the compound. According to the blood concentration data of the drug, the pharmacokinetic calculation software WinNonlin7. <NUM> non-compartmental model was used to calculate the pharmacokinetic parameters of the test drug.

Internal standard working solution: a certain amount of tolbutamide internal standard stock solution with a concentration of <NUM>,<NUM> ng/mL was taken into a volumetric flask with a certain volume, diluted to a desired volume with methanol and mixed well to obtain the internal standard working solution with a concentration of <NUM> ng/mL.

According to average blood concentration data of drug in each group, the pharmacokinetic calculation software WinNonlin7. <NUM> non-compartmental model was used to calculate pharmacokinetic parameters of compound in each group. The results were shown in Table <NUM>.

Table <NUM> showed that after compound Ic-<NUM> was injected intravenously at a dose of <NUM>/kg, the peak concentration (Cmax, <NUM> ng/mL) was reached at the first time point of sampling (<NUM>), the elimination half-life (T1/<NUM>) ) was <NUM>, AUC(<NUM>-∞) was <NUM>*ng/mL; after compound Ic-<NUM> was orally administered at a <NUM>-fold dose (<NUM>/kg), the peak concentration (Cmax, <NUM> ng/mL) was reached at <NUM>, the elimination half-life (T1/<NUM>) was <NUM>, and AUC(<NUM>-∞) was <NUM>*ng/mL. Based on the calculation on AUC (<NUM>-∞), the oral bioavailability was <NUM>%.

After duloxetine was injected intravenously at a dose of <NUM>/kg, the peak concentration (Cmax, <NUM> ng/mL) was reached at <NUM>, the elimination half-life (T1/<NUM>) was <NUM>, and the AUC (<NUM>-∞) was <NUM>*ng/mL; after duloxetine was orally administered at a <NUM>-fold dose (<NUM>/kg), the peak concentration (Cmax, <NUM> ng/mL) was reached at <NUM>, the elimination half-life (T1/<NUM>) was <NUM>, and the AUC ( <NUM>-∞) was <NUM>*ng/mL. Based on the calculation on AUC (<NUM>-∞), the oral bioavailability was <NUM>%.

The results showed that, compared with duloxetine, the compounds of formula I of the present invention (such as compound Ic-<NUM>) had more excellent pharmacokinetic properties with a longer half-life, higher exposure in plasma, and better bioavailability, so that it was suitable for development into a medicine for oral administration, and had a good prospect of medicine.

The experimental groups were solvent control group, <NUM>/kg duloxetine group (positive control group) and <NUM>/kg compound Ic-<NUM> group (the compound prepared in Example <NUM>).

<NUM>/kg duloxetine: <NUM> of duloxetine was weighed, dissolved in saline and diluted to <NUM>. After completely dissolved, the duloxetine was administered orally. The volume of administration was <NUM>/kg.

<NUM>/kg compound Ic-<NUM>: <NUM> of compound Ic-<NUM> was weighed, dissolved in saline and diluted to <NUM>. After completely dissolved, the compound Ic-<NUM> was administered orally. The volume of administration was <NUM>/kg.

Male C57BL/<NUM> mice weighed <NUM>-<NUM> were selected at the beginning of experiment. Each cage had <NUM> mice which were allowed to feed and drink water freely. Each experimental group had <NUM> mice, and the experimental mice were labeled by tail labeling method.

Day <NUM>: At <NUM>:<NUM> pm, the mice were put in a plexiglass box with stainless steel mesh, and then the plexiglass box was put in cold environment (<NUM>±<NUM>) overnight. The mice were allowed to feed freely and drink, and agar was used to replace water.

Day <NUM>: At <NUM>:<NUM> am, the mice were delivered to room temperature (<NUM>±<NUM>) environment for <NUM>, and then the mice were delivered to cold environment for <NUM>. The above steps were repeated until <NUM>:<NUM> pm, and the mice were put in cold environment overnight.

Day <NUM>: the operation on day <NUM> was repeated.

Day <NUM>: the mice were taken out from cold environment at <NUM>:<NUM> am.

The compounds were administered orally according to the schedule of experiment, and the dosage was <NUM>/kg.

On the fourth day after modeling, the pre-administration mechanical allodynia test was performed on the animals. Animals with PWT value greater than <NUM> were excluded and were not used for experiments. On the fifth day after modeling, the animals were tested for mechanical allodynia at <NUM>, <NUM>, and <NUM> after the administration of compounds.

The test method for mechanical allodynia was as follows:
The mice was individually placed in a plexiglass box with a grid on the bottom of the box to ensure that the mice claw could be tested. The mice were allowed to adapt for <NUM> before the test. After the adaptation was completed, the test fiber was used to test the center of the left hind claw of mice. The test fiber comprised <NUM> test strengths: <NUM>(<NUM>), <NUM>(<NUM>), <NUM>(<NUM>), <NUM>(<NUM>), <NUM>(<NUM>), <NUM>(<NUM>), <NUM>(<NUM>), and <NUM>(<NUM>). During the test, the test fiber was pressed vertically against the skin and forced to bend for <NUM>-<NUM> with a <NUM> interval of test.

During the test, rapid withdrawal of animal claw was recorded as pain response. When the test fiber was removed from animal skin, the withdrawal of animal claw was also recorded as pain response. If animal moved, the pain response was not recorded, and the test was repeated. In the test, <NUM>(<NUM>) was used firstly. If animal responded to pain, the test fiber with lower strength was used in next test; if the animal did not respond to pain, test fiber with higher strength was used in next test (Chaplan et al. The maximum strength of tested fiber was <NUM>(<NUM>).

The test results were recorded in Table <NUM> below, wherein pain response was recorded as X
and no pain response was recorded as O.

Mechanical allodynia was calculated by using the following formula: <MAT>.

Excel software was used to collect data, and prism software was used to analyze data. The larger the withdrawal threshold (PWT) was, the stronger the analgesic effect of the compound was.

The results of analgesic activity of compound Ic-<NUM> in the mice ICS model were shown in Table <NUM> and <FIG>.

The Table <NUM> and <FIG> showed that compound Ic-<NUM> of the present invention had very potent analgesic effect at a dose of <NUM>/kg, and could inhibit mechanical allodynia <NUM> hour and <NUM> hours after oral administration in ICS model. Compared with the positive control group, compound Ic-<NUM> had stronger analgesic effect than that of duloxetine at <NUM>, <NUM> and <NUM>. The mice ICS model was classic model for evaluating the efficacy of drug in the treatment of fibromyalgia. Therefore, the compound Ic-<NUM> of the present invention had excellent therapeutic effect on fibromyalgia.

Evaluation of therapeutic effect of compound Ic-<NUM> on visceral pain and inflammatory pain in mice acetic acid writhing pain model.

Male ICR mice, weighed <NUM>-<NUM>, were fasted but allowed to drink water freely for <NUM> before administration. All ICR mice were weighed and grouped randomly, and the number of animals in each group was ><NUM>. The negative control group was saline group (vehicle, blank control), and the positive control group was administrated with <NUM>/kg indomethacin (a non-steroidal anti-inflammatory drug), <NUM>/kg anisodamine (an antispasmodic drug with clinically analgesic activity), <NUM>/kg duloxetine and <NUM>/kg duloxetine. The test compound was Ic-<NUM> (the compound prepared in Example <NUM>), and the administration dosages were <NUM>/kg and <NUM>/kg. The drug was administrated by gavage based on the weight of mice. <NUM>% acetic acid solution (<NUM>/<NUM>) was injected intraperitoneally <NUM> hour after administration, and the number of times of visceral pain in each group was observed within <NUM>. When concave abdomen, stretched trunk and hind claw, and high buttocks appeared, one number point was recorded. Finally, the number of appearance of the above phenomenon was counted within <NUM> minutes. After administration, the fewer visceral pains in the mice were, the stronger the analgesic effect of the compound was.

The mice acetic acid writhing pain model test results were shown in <FIG> showed that the compound Ic-<NUM> (<NUM>/kg and <NUM>/kg) of the present invention could significantly reduce the number of appearance of writhing reaction in mice caused by acetic acid with a significant difference as compared with the number of appearance of writhing reaction in mice in the saline group (vehicle, blank control) (<NUM> times). At a dosage of <NUM>/kg of compound Ic-<NUM>, the number of appearance of writhing reaction in mice was <NUM> times, which was <NUM>% lower than that (<NUM> times) in the saline control group, suggesting that the half effective dose (ED<NUM>) of compound Ic-<NUM> was less than <NUM>/kg in the model. The analgesic effect of compound Ic-<NUM> at a dosage of <NUM>/kg (<NUM> times) was stronger than that of positive drug indomethacin (<NUM> times), anisodamine (<NUM> times) and duloxetine (<NUM> times) at the same dosage. The analgesic effect of compound Ic-<NUM> at a dosage of <NUM>/kg (<NUM> times) was equivalent to that of duloxetine at a dosage of <NUM>/kg (<NUM> times).

This experiment showed that the analgesic activity of compound Ic-<NUM> of the present invention was significantly stronger than positive control drug in the mice acetic acid writhing pain model. The mice acetic acid writhing pain model was a classical model for evaluating the efficacy of drug in treating visceral pain and inflammatory pain. Therefore, the compound Ic-<NUM> of the present invention had excellent therapeutic effect on visceral pain and inflammatory pain.

Male SPF grade SD rats, weighed <NUM>-<NUM>, were selected. Aseptic operation during surgery was performed. The animals were anesthetized with sodium pentobarbital (<NUM>/kg, intraperitoneal injection). The surgical area of animal waist was shaved and the skin was disinfected three times with iodophor and <NUM>% ethanol. After the skin was dried, the operation was started. Surgical knife was used to make a longitudinal incision at the back of the sacrum of the animal waist to expose the left paraspinal muscles, and stretcher was used to separate muscle tissue to expose the spine. The left spinal nerves L5 and L6 were separated and ligated with a <NUM>-<NUM> silk thread, and the wound was sutured. After the operation, the animals were placed on electrothermal pad, and <NUM> of saline was injected subcutaneously to prevent dehydration. After the animals were fully awakened and could move around freely, the animals were put back in the cage.

After the operation, the animals were adapted in the experimental environment for <NUM>/day for <NUM> days. One day before the administration, the rats were subjected to mechanical allodynia baseline test, and the animals that did not exhibit mechanical allodynia (the withdrawal threshold was greater than <NUM>) were eliminated and the remaining rats were randomly divided into one control group and two experimental groups.

The animals were weighed to calculate the dosage. The rats in two experimental groups were administrated with <NUM>/kg gabapentin (gabapentin was currently the first-line drug for the treatment of neuralgia) and <NUM>/kg compound Ic-<NUM> (the compound prepared in Example <NUM>), and the rats in control group were administrated with equal volume of saline orally. <NUM> after administration, mechanical allodynia test was performed. The rat was individually placed in a plexiglass box with a grid on the bottom of the box to ensure that the rat claw could be tested. The rats were allowed to adapt the environment for <NUM> before test. After the adaptation was completed, the test fiber was used to test the center of the left hind claw of the rat. The test fiber comprises <NUM> test strengths: <NUM> (<NUM>), <NUM> (<NUM>), <NUM> (<NUM>), <NUM> (<NUM>), <NUM> (<NUM>), <NUM> (<NUM>), <NUM> (<NUM>), <NUM> (<NUM>). During the test, the test fiber was pressed vertically against the skin and forced to bend the fiber for <NUM>-<NUM> with a <NUM> interval of test. During the test, rapid withdrawal of animal claw was recorded as pain response. When the test fiber was removed from animal skin, the withdrawal of animal claw was also recorded as pain response. If animal moved, the pain response was not recorded and the test was repeated. In the test, <NUM> (<NUM>) was used firstly. If animal responded to pain, the test fiber with lower strength was used in next test; if the animal did not respond to pain, test fiber with higher strength was used in next test. The maximum strength of tested fiber was <NUM> (<NUM>).

Mechanical allodynia was expressed as withdrawal threshold (PWT) in rat behavioral test, which was calculated with the following formula:<MAT>.

Excel software was used to collect data, and Prism <NUM>(Graph pad software, Inc. ) software was used to analyze data. The larger the withdrawal threshold (PWT) was, the stronger the analgesic effect of the compound was.

The results of analgesic activity in the rat SNL model were shown in Table <NUM> and <FIG>.

Table <NUM> and <FIG> showed that, compared with the saline control group, the compound IC-<NUM> of the present invention had very potent analgesic effect at a dosage of <NUM>/kg with a significant difference. Compared with the positive control group, the analgesic activity of compound Ic-<NUM> of the present invention was equivalent to the analgesic effect of <NUM>/kg gabapentin within <NUM> after administration. The rat SNL model was a classical model for evaluating the efficacy of drug in the treatment of nerve pain. Therefore, the compound Ic-<NUM> of the present invention had excellent therapeutic effect on nerve pain.

Evaluation of therapeutic effect of compound IC-<NUM> on acute pain and inflammatory pain in mice formalin pain model.

<NUM> male C57BL/<NUM> mice (<NUM>-week aged), were randomly divided into <NUM> groups to evaluate analgesic activity of two compounds in the mice formalin pain model, and each group had <NUM> mice. The experimental groups were duloxetine group and the compound Ic-<NUM> group (the compound prepared in Example <NUM>), respectively. Before the start of experiment, the mice were allowed to adapt experimental environment for <NUM> with free feeding and drinking water. The test drug was administrated intraperitoneally, and the dosage was as follows:.

After administration, the mice were placed in a transparent, ventilated plexiglass cylinder, and <NUM> later, <NUM>µl of <NUM>% formalin solution was injected into the left hind plantar of mice in each group by micro-injector, and claw pain response in mice was recorded in real time by a mini-camera. The time length of licking left claw was used as an indicator of pain response, licking time in <NUM>-<NUM> (phase I) and <NUM>-<NUM> (phase II) was observed and recorded, and the statistical analysis was conducted. The half effective dose (ED<NUM>) of three compounds was calculated: ED<NUM> referred to the dose of the drug that decreased licking time by half as compared with the blank control group. The smaller the ED<NUM> value was, the lower the effective analgesic dose of the compound was and the stronger the analgesic effect was.

The claw licking time statistical results of compound Ic-<NUM> and duloxetine in the mice formalin model at different dosages (<NUM>-<NUM>) were shown in Table <NUM> and <FIG>.

Table <NUM> and <FIG> showed that the licking time of compound Ic-<NUM> of the present invention in phase II (<NUM>-<NUM>) at a dosage of <NUM>/kg had decreased by more than <NUM>% as compared with that of blank Vehicle. The analgesic effect (ED<NUM>) in phase II pain was <NUM>/kg, while the ED<NUM> of duloxetine in phase II pain was <NUM>/kg. The analgesic activity of compound Ic-<NUM> of the present invention was significantly stronger than that of duloxetine at the same dosage. From the above data, it could be seen that the compound Ic-<NUM> of the present invention showed very strong analgesic activity in the mice formalin pain model. The mice formalin model was a classical model for evaluating drug effect on acute pain and inflammatory pain. Therefore, the compound Ic-<NUM> of the present invention had excellent therapeutic effect on acute pain and inflammatory pain.

Claim 1:
A compound of formula I, or a pharmaceutically acceptable salt thereof for use in preventing and/or treating a disease caused by transient receptor potential channel protein (TRP), wherein the disease is selected from the group consisting of pain, epilepsy, inflammation, respiratory disorder, pruritus, urinary tract disorder and inflammatory bowel disease;
<CHM>
wherein,
A is benzofuranyl or indanyl;
R<NUM> and R<NUM> are each independently hydrogen, substituted or unsubstituted C<NUM>-C<NUM> alkyl; X is oxygen atom, or sulfur atom;
Y is CH or nitrogen atom;
R<NUM> is hydrogen, halogen, substituted or unsubstituted C<NUM>-C<NUM> alkyl;
n is <NUM>;
"*" represents a chiral carbon atom, and the absolute configuration of the chiral carbon atom is S type;
wherein, the "substituted" means that <NUM>-<NUM> hydrogen atoms on the group are substituted by a substituent selected from the group consisting of C<NUM>-C<NUM> alkyl, C<NUM>-C<NUM> cycloalkyl, C<NUM>-C<NUM> haloalkyl, halogen, nitro, cyano, hydroxyl, C<NUM>-C<NUM> carboxyl, C<NUM>-C<NUM> ester group, C<NUM>-C<NUM> amide group, C<NUM>-C<NUM> alkoxyl, C<NUM>-C<NUM> haloalkoxy, benzyl, <NUM>- or <NUM>- membered aryl or heteroaryl.