Opioid receptor antagonist conjugate and use thereof

Abstract

The present invention relates to an opioid receptor antagonist conjugate and a use thereof. In particular, the present invention relates to a covalent coupling conjugate of a hydrophilic polymer and an opioid receptor antagonist and the use thereof.

Claims

1. A conjugate as represented by formula (V) or a pharmaceutically acceptable salt thereof: W.sub.1—P—W.sub.2 (V), wherein W.sub.1 and W.sub.2 each independently have structure of: ##STR00013## wherein Y is ##STR00014## Z is —OH, X is selected from the group consisting of amido group, amine group, carbamate group, thioether group, ether group, urea group and methylene group, and P is a polyethylene glycol having 2 to 20 —CH.sub.2CH.sub.2O— structural unit.

2. The conjugate according to claim 1, wherein W.sub.1 and W.sub.2 are each independently: ##STR00015##

3. The conjugate according to claim 1, which is represented by formula (II) or a pharmaceutically acceptable salt thereof: ##STR00016## wherein, n is a natural number from 2 to 20.

4. The conjugate according to claim 1, which is represented by formula (III) or a pharmaceutically acceptable salt thereof: ##STR00017## wherein, n is a natural number from 2 to 20.

5. The conjugate according to claim 1, which is represented by formula (IV) or a pharmaceutically acceptable salt thereof: ##STR00018## wherein, n is a natural number from 2 to 20.

6. The conjugate according to claim 3, wherein n is 3, 4, 6, 8, or 10.

7. The conjugate according to claim 1, wherein the hydrophilic polymer is a monodisperse polyethylene glycol having 2 to 10 —CH.sub.2CH.sub.2O— structural unit and the opioid receptor antagonist prior to conjugation is naltrexone.

8. The conjugate according to claim 1, which is selected from the group consisting of PEG3-(6-α-naltrexol).sub.2-ether, PEG4 (6-α-naltrexol).sub.2-ether, PEG6-(6-α-naltrexol).sub.2-ether, PEG8-(6-α-naltrexol).sub.2-ether, PEG10-(6-α-naltrexol).sub.2-ether, PEG4 (6-β-naltrexol).sub.2-ether, PEG6-(6-β-naltrexol).sub.2-ether, and PEG8-(6-β-naltrexol).sub.2-ether.

9. A pharmaceutical composition, comprising the conjugate according to claim 1, and a pharmaceutically acceptable carrier.

10. A kit, comprising the conjugate according to claim 1.

11. A method of treating constipation induced by opioid receptors, treating pain in combination with an opioid, reducing side effect caused by an opioid and preventing opioid abuse, comprising administrating an effective amount of the conjugate according to claim 1 to a subject in need thereof.

12. The conjugate according to claim 2, wherein W.sub.1 and W.sub.2 are each independently selected from the group consisting of ##STR00019##

Description

BRIEF DESCRIPTION OF THE DRAWING

(1) The above-described and other aspects of the present disclosure will be clearly explained according to the detailed description and drawings of the present disclosure. To exemplify the present disclosure, the embodiments in the drawings are preferred are present, however, it is understood that the present disclosure is not limited to the disclosed specific embodiments.

(2) FIG. 1. A dose-response curve of Na1029 antagonizing the antidiarrheal effect of loperamide hydrochloride.

DETAILED DESCRIPTION

(3) Definitions

(4) The terms “activated form” or “derived form” (which are used interchangeably) as used herein refers to a compound in a particular position of which, a reactive group is introduced or a group is modified to a reactive functional group thereof to make the compound capable of coupling with a water soluble molecular.

(5) The “opioid receptor antagonist” mentioned herein is a class of compounds having similar structure to that of opioids, without agonistic effect on opioid receptors themselves, but antagonizing opioid analgesics and removing opioid analgesics which bind with receptors or competitively bind with analgesics and eliminate side effects such as functional bowel disorders and respiratory depression induced by the use of some opioid analgesics.

(6) The “polyethylene glycol” mentioned herein has the meaning commonly understood by those of ordinary skill in the art, including both polyethylene glycol (including bifurcation structures with different structures such as linear, branched, star) itself as well as derivatives with modified ends, unless otherwise explicitly stated. For example, the PEG is methoxypolyethylene glycol (mPEG). Herein, the terminal group of polyethylene glycol (PEG) is a hydroxyl group or other groups, unless otherwise specified. The other groups include, but are not limited to, alkoxy group, cycloalkoxy group, cycloalkyloxy group, alkenyl group, aryloxy group or aralkyloxy, group. These types of PEG molecule are known in the art and are routinely used in polypeptide modification. The PEG side chain can be linear, branched, bifurcated or consists of multiple arms, and different polyethylene glycols may have different polymeric chain lengths and polymeric structures. In a preferred embodiment of the present disclosure, the polyethylene glycol of the present disclosure is an oligo ethylene glycol, for example the repeating units are less than 20, 15, 10 and the like. More preferably, the polyethylene glycol of the present disclosure is monodispersed.

(7) The “conjugate” mentioned herein refers to a product formed by a conjugation between a biologically active molecule (e.g., an opioid receptor antagonist) and a hydrophilic polymer molecule (e.g., polyethylene glycol) via covalent bond through direct linkage or by a linker.

(8) The “pharmaceutically acceptable salt” mentioned herein may be hydrochloride, hydrobromide, sulfate, nitrate, phosphate, tartrate, fumarate, maleate, lactate, benzene sulfonate, pantothenate, ascorbate, etc., or a combination thereof. Preferably, the pharmaceutically, acceptable salt is a hydrochloride salt.

(9) The “monodisperse” mentioned herein refers to that the polymer of the present disclosure (e.g., polyethylene glycol) are homogeneous, for example, the molar percentage of non-target products is less than 10%, 8%, 5%, 3%, 2%, 1%, 0.5%, 0.1%, 0.01% or 0.001%.

(10) Terms used herein such as “including”, “containing” and “comprising” are not intended to limit. In addition, “or” means “and/or” unless otherwise stated.

(11) In addition, it should be noted that, as used in the specification, the singular form includes the plural form of the subject matter to which it refers unless it is clearly and explicitly limited to one object. And if a specific value is mentioned, it at least includes the value unless the article otherwise clearly indicates.

(12) When a numerical value indicates approximate value, it should be interpreted as that a particular value forms another embodiment. As what is used, “about X” (where X is a numerical value) refers to ±10% (inclusive) of the value listed. If exists, all ranges are included and can be combined.

(13) The term “pharmaceutical composition” used herein represents a combination of at least one medicine and optionally a pharmaceutically acceptable carrier or excipient combining together to achieve a particular purpose. In certain embodiments, the pharmaceutical composition includes combinations that are separated in time and/or space, as long as they are capable of working together to achieve the purposes of the present disclosure. For example, the components of the pharmaceutical composition (for example, the conjugate according to the present disclosure) may be administered to the object as a whole or separately. When the components of the pharmaceutical composition are separately administered to an object, the components may be administered to the object simultaneously or sequentially. Preferably, said pharmaceutically acceptable carrier is water, buffered aqueous solution, isotonic saline solution such as PBS (phosphate buffer), glucose, mannitol, dextrose, lactose, starch, magnesium stearate, fiber, magnesium carbonate, 0.3% glycerol, hyaluronic acid, ethanol, or polyalkylene glycol such as polypropylene glycol, triglyceride and the like. The type of the pharmaceutically acceptable carrier used depends particularly on the administration manner of the composition according to the present disclosure, for example, oral, nasal, intradermal, subcutaneous, intramuscular or intravenous administration. The composition according to the present disclosure may contain, as an additive, a lubricant, a preservative, a stabilizer, a wetting agent, an emulsifier, a salt which affects the osmotic pressure, a buffer, a coloring matter, a flavoring substance, and/or an aromatic substance.

(14) The pharmaceutical compositions according to the present disclosure may be administrated by any suitable way, for example, orally, nasally, intradermally, subcutaneously, intramuscularly or intravenously.

(15) “Administering” means providing a substance to an object in a pharmacologically available manner.

(16) The “pharmaceutically effective amount” and “effective amount” used herein refers to a dose which is sufficient to show its benefit to the object to which it is administered. The actual administered amount, as well as the rate and time course of administration, depend on the condition and severity of the object being treated. The prescription for treatment (e.g., the determination of the dose) is ultimately the responsibility of the general practitioner and other physicians and relied on them to make decisions, usually the disease being treated, the condition of the individual patient, the site of delivery, the method of administration, and the other factors known to the doctors are considered.

(17) The term “object” used herein means animals, including warm-blooded mammals, such as humans and primates; birds; domesticated or farm animals, such as cats, dogs, sheep, goats, cows, horses; and pigs; laboratory animals such as mice, rats and guinea pigs; fish; reptiles; zoo animals and wild animals.

(18) Unless otherwise defined, all technical and scientific terms used herein have the same meaning as understood by those having ordinary skill in the art.

(19) Unless otherwise defined, any component, element, attribute or step disclosed about an embodiment of the method and product may be applied to any of the other methods and products disclosed herein.

(20) Each of the patents, patent applications and cited publications in the present disclosure or the descriptions in the present disclosure are hereby entirely incorporated by reference.

(21) The invention is further defined in the following embodiments. It should be understood that the embodiments are only examples for illustration purpose rather than intending to limit the scope of the present invention. From the discussion and the examples above, those having ordinary skill in the art can determine the essential characteristics of the present invention and make changes and modifications to the present invention in various aspects to adapt it to various usages and conditions without departing from its essential and scope.

EXAMPLES

Example 1: Preparation of Polyethylene Glycol Mesylate

(22) ##STR00008##

(23) 50 ml of dichloromethane, 7.18 ml of triethylamine, 206 mg of p-dimethylaminopyridine and 10.0 g of CH.sub.3—(OCH.sub.2CH.sub.2).sub.6—OH (abbreviated as mPEG6-OH) (Jiaxing Biomatrik, M006110503) were successively added to a reaction flask. A solution of 3.28 ml of methanesulfonyl chloride in 50 ml of dichloromethane was added dropwise with stirring at 0° C. After the completion of the dropwise addition, the reaction was stirred at room temperature overnight. The reaction solution was transferred to a separatory funnel and washed with dilute hydrochloric acid and brine successively. The organic layer was separated, dried with 30 g of anhydrous sodium sulfate. After the desiccant was filtered out, the organic layer was concentrated by rotary evaporation to obtain 13.6 g of crude product. The crude product was purified by chromatography to obtain 13.0 g of mPEG6-OMs (CH.sub.3—(OCH.sub.2CH.sub.2).sub.6—OSO.sub.2CH.sub.3). 1H-NMR (400 MHz, CDCl3), δ ppm 4.35-4.41 (2H, m), 3.74-3.80 (2H, m), 3.60-3.70 (18H, m), 3.52-3.58 (2H, m), 3.38 (3H, s), 3.08 (3H, s).

(24) Using a similar procedure, with different polyethylene glycols as starting materials, a variety of polyethylene glycol mesylate as listed in the table below could be prepared.

(25) TABLE-US-00001 TABLE 1 Polyethylene glycol mesylate used in the present disclosure Abbreviation Molecular Structure PEG3-(OMs)2 CH.sub.3SO.sub.2—(OCH.sub.2CH.sub.2).sub.3—OSO.sub.2CH.sub.3 PEG4-(OMs)2 CH.sub.3SO.sub.2—(OCH.sub.2CH.sub.2).sub.4—OSO.sub.2CH.sub.3 PEG6-(OMs)2 CH.sub.3SO.sub.2—(OCH.sub.2CH.sub.2).sub.6—OSO.sub.2CH.sub.3 PEG8-(OMs)2 CH.sub.3SO.sub.2—(OCH.sub.2CH.sub.2).sub.8—OSO.sub.2CH.sub.3 PEG10-(OMs)2 CH.sub.3SO.sub.2—(OCH.sub.2CH.sub.2).sub.10—OSO.sub.2CH.sub.3 mPEG6-OMs CH.sub.3—(OCH.sub.2CH.sub.2).sub.6—OSO.sub.2CH.sub.3 mPEG8-OMs CH.sub.3—(OCH.sub.2CH.sub.2).sub.8—OSO.sub.2CH.sub.3

(26) The specific preparation process is as follows:

(27) PEG3-(OMs)2: 50 ml of dichloromethane, 17.0 ml of triethylamine, 244 mg of p-dimethyaminopyridine and 6.00 g of H—(OCH.sub.2CH.sub.2).sub.3—OH (abbreviated as H-PEG3-OH) (Sinopharm Reagent, 20130125) were successively added to a reaction flask. A solution of 7.8 ml of methanesulfonyl chloride in 20 ml of dichloromethane was added dropwise with stirring at 5 to 10° C. After the completion of the dropwise addition, the reaction was stirred at room temperature overnight. The reaction solution was transferred to a separatory funnel and washed with dilute hydrochloric acid and brine successively. The organic layer was separated, dried with 30 g of anhydrous sodium sulfate. After the desiccant was filtered out, the organic layer was concentrated by rotary evaporation to obtain 12.6 g of crude product. The crude product was purified by chromatography to obtain 11.2 g of pure product. 1H-NMR (400 MHz, CDCl3), δ ppm 4.34-4.41 (4H, m), 3.74-3.80 (4H, m), 3.68 (4H, s), 3.07 (6H, s).

(28) PEG4-(OMs)2: 50 ml of dichloromethane, 17.0 ml of triethylamine, 244 mg of p-dimethylaminopyridine and 7.76 g of H—(OCH.sub.2CH.sub.2).sub.4—OH (abbreviated as H-PEG4-OH) (Aladdin Reagent, J1218031) were successively added to a reaction flask. A solution of 7.8 ml of methanesulfonyl chloride in 50 ml of dichloromethane was added dropwise with stirring at 0° C. After the completion of the dropwise addition, the reaction was stirred at room temperature overnight. The reaction solution was transferred to a separatory funnel and washed with dilute hydrochloric acid and brine successively. The organic layer was separated, dried with 30 g of anhydrous sodium sulfate. After the desiccant was filtered out, the organic layer was concentrated by rotary evaporation to obtain 14.7 g of crude product. The crude product was purified by chromatography to obtain 13.90 g of pure product. 1H-NMR (400 MHz, CDCl3), δ ppm 4.34-4.41 (4H, m), 3.73-3.81 (4H, m), 3.60-3.71 (8H, m), 3.07 (6H, s).

(29) PEG6-(OMs)2: 50 ml of dichloromethane, 10.625 ml of triethylamine, 152.5 mg of p-dimethylaminopyridine and 7.05 g of H—(OCH.sub.2CH.sub.2).sub.6—OH (abbreviated as H-PEG6-OH) (Jiaxing Biomatrik, DH06141230) were successively added to a reaction flask. A solution of 4.875 ml of methanesulfonyl chloride in 40 ml of dichloromethane was added dropwise with stirring at 5-8° C. After the completion of the dropwise addition, the reaction was stirred at room temperature overnight. The reaction solution was transferred to a separatory funnel and washed with dilute hydrochloric acid and brine successively. The organic layer was separated, dried with 30 g of anhydrous sodium sulfate. After the desiccant was filtered out, the organic layer was concentrated by rotary evaporation to obtain 12.5 g of crude product. The crude product was purified by chromatography to obtain 10.5 g of pure product. 1H-NMR (400 MHz, CDCl3), δ ppm 4.34-4.41 (4H, m), 3.73-3.81 (4H, m), 3.60-3.71 (16H, m), 3.08 (6H, s).

(30) PEG8-(OMs)2: 50 ml of dichloromethane, 5.74 ml of triethylamine, 83 mg of p-dimethylaminopyridine and 5.0 g of H—(OCH.sub.2CH.sub.2).sub.8—OH (abbreviated as H-PEG8-OH) (Jiaxing Biomatrik, DH08150114) were successively added to a reaction flask. A solution of 2.64 ml of methanesulfonyl chloride in 50 ml of dichloromethane was added dropwise with stirring at 0° C. After the completion of the dropwise addition, the reaction was stirred at room temperature overnight. The reaction solution was transferred to a separatory funnel and washed with dilute hydrochloric acid and brine successively. The organic layer was separated, dried with 30 g of anhydrous sodium sulfate. After the desiccant was filtered out, the organic layer was concentrated by rotary evaporation to obtain 8.17 g of crude product. The crude product was purified by chromatography to obtain 6.7 g of pure product. 1H-NMR (400 MHz, CDCl3), δ ppm, 4.34-4.41 (4H, m), 3.73-3.81 (4H, m), 3.60-3.71 (24H, m), 3.08 (6H, s).

(31) PEG10-(OMs)2: 50 ml of dichloromethane, 4.63 ml of triethylamine, 66.5 mg of p-dimethylaminopyridine and 5.0 g of H—(OCH.sub.2CH.sub.2).sub.10—OH (abbreviated as H-PEG10-OH) (Jiaxing Biomatrik, DH10140509) were successively added to a reaction flask. A solution of 2.13 ml of methanesulfonyl chloride in 50 ml of dichloromethane was added dropwise with stirring at 0° C. After the completion of the dropwise addition, the reaction was stirred at room temperature overnight. The reaction solution was transferred to a separatory funnel and washed with dilute hydrochloric acid and brine successively. The organic layer was separated, dried with 30 g of anhydrous sodium sulfate. After the desiccant was filtered out, the organic layer was concentrated by rotary evaporation to obtain 8.0 g of crude product. The crude product was purified by chromatography to obtain 5.5 g of pure product. 1H-NMR (400 MHz, CDCl3), δ ppm, 4.34-4.41 (4H, m), 3.73-3.81 (4H, m), 3.60-3.71 (32H, m), 3.08 (6H, s).

(32) mPEG8-OMs: 50 ml of dichloromethane, 2.763 ml of triethylamine, 80 mg of p-dimethylaminopyridine and 5.0 g of CH.sub.3—(OCH.sub.2CH.sub.2).sub.8—OH (abbreviated as mPEG8-OH) (Jiaxing Biomatrik, M008131031) were successively added to a reaction flask. A solution of 1.27 ml of methanesulfonyl chloride in 50 ml of dichloromethane was added dropwise with stirring at 0° C. After the completion of the dropwise addition, the reaction was stirred at room temperature overnight. The reaction solution was transferred to a separatory funnel and washed with dilute hydrochloric acid and brine successively. The organic layer was separated, dried with 30 g of anhydrous sodium sulfate. After the desiccant was filtered out, the organic layer was concentrated by rotary evaporation to obtain 7.0 g of crude product. The crude product was purified by chromatography to obtain 5.44 g of purified product. 1H-NMR (400 MHz, CDCl3), δ ppm 4.35-4.41 (2H, m), 3.74-3.80 (2H, m), 3.60-3.70 (26H, m), 3.52-3.58 (2H, m), 3.38 (3H, s), 3.08 (3H, s).

Example 2: Preparation of 3-MOM-Naltrexone and 3-MOM-Naloxone

(33) ##STR00009##

(34) 200 ml of dichloromethane, 41.7 g of diisopropylethylamine and 19.83 g of naltrexone hydrochloride (Jinan Haohua Industrial Co., Ltd., FR00007) were successively added to a reaction flask. A solution of 17.7 g of chlorotnethyl methyl ether in 200 ml of dichlorornethane was added dropwise with stirring at 0° C. After the dropwise addition was completed, the reaction was stirred overnight. The reaction solution was transferred to a separatory funnel and washed with an alkali solution (400 ml of water +40 g of anhydrous sodium carbonate +10.0 g of sodium hydroxide +30 g of sodium chloride, well mixed). The organic layer was separated, dried with 30 g of anhydrous sodium sulfate. After the desiccant was filtered out, the organic layer was concentrated by rotary evaporation to obtain 21.3 g of product 3-MOM-naltrexone. 1H-NMR. (400 MHz, CDCl3), δ ppm 6.85-6.92 (1H, d, J=8.0 Hz), 6.58-6.65 (1H, d, J=8.4 Hz), 5.20-5.30 (2H, m), 4.68 (1H, s), 3.52 (3H, s), 3.15-3.22 (1H, d, J=6.0 Hz), 2.97-3.12 (2H, m), 2.66-2.75 (1H, m), 2.54-2.64 (1H, m), 2.37-2.50 (3H, m), 2.26-2.35 (1H, m), 2.10-2.20 (1H, m), 1.83-1.93 (1H, m), 1.54-1.71 (2H, m), 0.81-0.92 (1H, m), 0.52-0.62 (2H, m), 0.11-0.19 (2H, m). LC/MS (ESI) m/z 386.5 [M+H]+.

(35) 50 ml of dichloromethane, 13.51 g of diisopropylethylamine and 5.00 g of naloxone hydrochloride (Jinan Haohua Industrial Co., Ltd., GR00037) were successively added to a reaction flask. A solution of 5.55 g of chloromethyl methyl ether in 50 nil of dichloromethane was added dropwise with stirring at 0° C. After the dropwise addition was completed, the reaction was stirred overnight. The reaction solution was transferred to a separatory funnel and washed with an alkali solution (100 ml of water +10 g of anhydrous sodium carbonate +3.0 g of sodium hydroxide +7 g of sodium chloride, well mixed). The organic layer was separated, dried with 10 g of anhydrous sodium sulfate. After the desiccant was filtered out, the organic layer was concentrated by rotary evaporation to obtain 5.14 g of product 3-MOM-naloxone. 1H-NMR (400 MHz, CDCl3), δ ppm 6.86-6.92 (1H, d, J=8.4 Hz), 6.59-6.66 (1H, d, J=8.4 Hz), 5.76-5.90 (1H, m), 5.15-5.30 (4H, m), 5.02 (1H, s), 4.67 (1H, s), 3.52 (3H, s), 3.14-3.19 (2H, m), 3.06-3.14 (1H, d, J=18.4 Hz), 2.96-3.07 (2H, m), 2.53-2.64 (2H, m), 2.34-2.44 (1H, m), 2.25-2.33 (1H, m), 2.10-2.20 (1H, m), 1.82-1.92 (1H, m), 1.54-1.68 (2H, m). LC/MS (ESI) m/z 372.5[M+H]+.

Example 3: Preparation of 3-MOM-6-α-Naltrexol and 3-MOM-6-α-Naloxol

(36) ##STR00010##

(37) Under the protection of argon, 100 ml of tetrahydrofuran and 16.8 g of 3-MOM-naltrexone were added to a reaction flask. 45 ml of a solution of potassium tri-sec-butylborohydride in tetrahydrofuran was added under stirring at 10° C. The reaction was monitored using thin-layer chromatography (TLC). After the reaction was completed, the pH was adjusted to 2 to 4 with dilute hydrochloric acid. The reaction solution was transferred to a separatory funnel and extracted four times with dichloromethane (150, 100, 100, 100 ml). The organic layers were separated, and 10.0 g of sodium hydroxide in 150 ml of aqueous solution was added to the aqueous layer, and the mixture was mixed well and extracted three times with dichloromethane (100 ml×3). The organic layers were combined, dried with 20 g of anhydrous sodium sulfate. After the desiccant was filtered out, the organic layer was concentrated by rotary evaporation to obtain 16.5 g of product 3-MOM-6-α-naltrexol. 1H-NMR (400 MHz, CDCl3), δ ppm 6.78-6.88 (1H, d, J=8.0 Hz), 6.54-6.64 (1H, d, J=8.4 Hz), 5.53-5.62 (1H, d, J=6.4 Hz), 5.29 (1H, s), 5.05 (1H, s), 4.96-5.02 (1H, d, J=6.4 Hz), 4.58-4.68 (1H, d, J=5.2 Hz), 4.12-4.24 (1H, m), 3.48 (3H, s), 2.98-3.12 (3H, m), 2.55-2.70 (2H, m), 2.33-2.42 (2H, d, J=6.8 Hz), 2.15-2.31 (2H, m) 1.48-1.60 (3H, m), 1.28-1.42 (1H, 0.78-0.91 (1H, m), 0.49-0.60 (2H, m), 0.09-0.18 (2H, m). LC/MS (ESI) m/z 388.5 [M+H]+.

(38) Under the protection of argon, 20 ml of tetrahydrofuran and 3.48 g of 3-MOM-naloxone were added to a reaction flask. 13 ml of a solution of potassium tri-sec-butylborohydride in tetrahydrofuran was added under stirring at −15° C. The reaction was monitored using thin-layer chromatography (TLC). After the reaction was completed, the pH was adjusted to 3 to 5 with dilute hydrochloric acid. The reaction solution was transferred to a separatory funnel and extracted four times with dichloromethane (50 ml×4). The organic layers were separated, and 1.0 g of sodium hydroxide in 50 ml of aqueous solution was added to the aqueous layer, and the mixture was mixed well and extracted three times with dichloromethane (50 ml×3). The organic layers were combined, dried with 10 g of anhydrous sodium sulfate. After the desiccant was filtered out, the organic layer was concentrated by rotary evaporation to obtain 2.97 g of product 3-MOM-6-α-naloxol. 1H-NMR (400 MHz, CDCl3), ppm 6.80-6.86 (1H, d, J=8.4 Hz), 6.57-6.63 (1H, d, J=8.4 Hz), 5.73-5.87 (1H, m), 5.53-6.58 (1H, d, J=6.4 Hz), 5.12-5.24 (2H, m), 4.96-5.02 (1H, d, J=6.4 Hz), 4.88 (1H, s), 4.59-4.64 (1H, d, J=5.2 Hz), 4.13-4.21 (1H, m), 3.48 (3H, s), 3.02-3.14 (4H, m), 2.87-2.93 (1H, d, J=6.4 Hz), 2.48-2.66 (2H, m), 2.19-2.25 (2H, d, J=7.6 Hz), 1.82-1.95 (1H, m), 1.44-1.60 (3H, m), 1.27-1.39 (1H, m). LC/MS (ESI) m/z 374.5 [M+H]+.

Example 4: Preparation of 3-MOM-6-β-naltrexol and 3-MOM-6-β-naloxol

(39) ##STR00011##

(40) Under the protection of argon, 30 ml of ethanol, 1.64 g of an alkaline solution of 3-MOM-naloxone and aminoiminomethanesulfinic acid (1.74 g of aminoiminomethanesulfinic acid +0.85 g of sodium hydroxide 30 ml of water) were added to a reaction flask. Reaction was carried out at 80° C. with stirring and the reaction was monitored using TLC. After the reaction was completed, the heating was stopped. 120 ml of brine (10%) was added to the mixture, and the mixture was mixed well and then transferred to a separatory funnel and extracted three times with dichloromethane (50, 25, 25 nil). The organic layers were combined and washed with 10% brine. The organic layers were combined, dried with 10 g of anhydrous sodium sulfate. After the desiccant was filtered out, the organic layer was concentrated by rotary evaporation to obtain 1.29 g of product 3-MOM-6-β-naloxol. 1H-NMR (400 MHz, CDCl3), δ ppm 6.87-6.93 (1H, d, J=8.4 Hz), 6.58-6.64 (1H, d, J=8.4 Hz), 5.73-5.87 (1H, m), 5.10-5.26 (4H, m), 4.96-5.02 (1H, d, J=6.4 Hz), 4.46-4.52 (1H, d, J=5.6 Hz), 3.54-3.64 (1H, m), 3.50 (3H, s), 3.03-3.16 (3H, m), 2.90-2.96 (1H, d, J=5.6 Hz), 2.50-2.66 (2H, m), 2.10-2.28 (2H, m), 1.86-2.02 (1H, m), 1.54-1.68 m), 1.46-1.54 (1H, m), 1.30-1.40 (1H, m). LC/MS (ESI) m/z 374.5 [M+H]+.

(41) Under the protection of argon, 80 ml of ethanol, 4.51 g of an alkali solution of 3-MOM-naltrexone and aminoiminomethanesulfinic acid (4.33 g of aminoiminomethanesulfinic acid +2.06 g of sodium hydroxide +80 ml of water) were added to a reaction flask. Reaction was carried out at 80° C. with stirring and the reaction was monitored using TLC. After the reaction was completed, the heating was stopped, 300 ml of brine (10%) was added, and the mixture was well mixed and transferred to a separatory funnel and extracted three times with dichloromethane (100, 50, 50 ml). The organic layers were combined and washed with 10% brine. The organic layers were combined, dried with 20 g of anhydrous sodium sulfate. After the desiccant was filtered out, the organic layer was concentrated by rotary evaporation to obtain 4.10 g of product 3-MOM-6-β-naltrexol. 1H-NMR (400 MHz, CDCl3), δ ppm 6.87-6.93 (1H, d, J=8.4 Hz), 6.57-6.63 (1H, d, 0.1=8.0 Hz), 5.15-5.26 (2H, m), 5.29 (1H, s), 4.46-4.53 (1H, d, J=5.6 Hz), 3.54-3.64 (1H, m), 3.50 (3H, s), 3.00-3.14 (2H, m), 2.86-2.98 (1H, m), 2.55-2.70 (2H, m), 2.33-2.40 (2H, d, J=6.4 Hz), 2.19-2.30 (1H, m), 2.09-2.20 (1H, m), 1.88-2.04 (1H, m), 1.55-1.70 (2H, m), 1.47-1.54 (1H, m), 1.32-1.43 (1H, m), 0.78-0.91 (1H, m), 0.48-0.60 (2H, m), 0.05-0.18 (2H, m). LC/MS (ESI) m/z 388.6 [M+H]+.

Example 5: Preparation of PEGylated Naltrexol and Naloxol, and Hydrochlorides Thereof

(42) ##STR00012##

(43) Under the protection of argon, 1.00 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 2.02 g of 3-MOM-6-α-naltrexol in 12 ml of tetrahydrofuran and 661 mg of PEG3-(OMs)2 in 12 ml of tetrahydrofuran were successively added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate. After the desiccant was removed, the organic layer was concentrated by rotary evaporation to obtain 2.45 g of residue. Then said residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain 0.55 g of PEG3-(6-α-naltrexol)2-ether (abbreviated as Na1053) product.

(44) The above-mentioned product was dissolved in 20 ml of ethanol, then 2.5 ml of hydrogen chloride ethanol solution (1.0 M) was added. The mixture was mixed well and then concentrated by rotary evaporation to obtain 0.65 g of residue. Said residue was dissolved in 20 ml of water and freeze-dried to obtain 0.45 g of almost white powdery solid, i.e. dihydrochloride salt of Na1053. 1H-NMR (400 MHz, D2O), δ ppm 6.72-6.77 (2H, d, J=8.4 Hz), 6.62-6.68 (2H, d, J=8.4 Hz), 4.81-4.86 (2H, d, J=4.8 Hz), 3.90-4.00 (4H, m), 3.66-3.74 (2H, m), 3.59-3.66 (2H, m), 3.46-3.59 (4H, m), 3.45 (4H, s), 3.25-3.34 (2H, d, J=19.6 Hz), 3.17-3.25 (2H, m), 3.03-3.14 (4H, m), 2.87-2.97 (2H, m), 2.73-2.87 (2H, m), 2.35-2.47 (2H, m), 1.68-1.79 (2H, m), 1.47-1.69 (6H, m), 1.16-1.30 (2H, m), 0.92-1.04 (2H, m), 0.67-0.77 (2H, m), 0.57-0.67 (2H, m), 0.28-0.43 (4H, m). LC/MS (EST) m/z 802.1 [M+H]+ and 401.8 [M+2H]++.

(45) Using a similar procedure, with different polyethylene glycols as starting materials (as shown in Table 3), a variety of PEGylated naltrexol and naloxol, and hydrochlorides thereof as listed in table 2 below could be prepared.

(46) TABLE-US-00002 TABLE 2 Conjugates of the Present Disclosure Molecular Chemical Abbreviation Structure Formula Hydrochloride Form Nal053 PEG3-(6-α- C.sub.46H.sub.60N.sub.2O.sub.10 C.sub.46H.sub.60N.sub.2O.sub.10•2HCl naltrexol)2-ether Nal021 PEG4-(6-α- C.sub.48H.sub.64N.sub.2O.sub.11 C.sub.48H.sub.64N.sub.2O.sub.11•2HCl naltrexol)2-ether Nal025 PEG6-(6-α- C.sub.52H.sub.72N.sub.2O.sub.13 C.sub.52H.sub.72N.sub.2O.sub.13•2HCl naltrexol)2-ether Nal029 PEG8-(6-α- C.sub.56H.sub.80N.sub.2O.sub.15 C.sub.56H.sub.80N.sub.2O.sub.15•2HCl naltrexol)2-ether Nal033 PEG10-(6-α- C.sub.60H.sub.88N.sub.2O.sub.17 C.sub.60H.sub.88N.sub.2O.sub.17•2HCl naltrexol)2-ether Nal022 PEG4-(6-β- C.sub.48H.sub.64N.sub.2O.sub.11 C.sub.48H.sub.64N.sub.2O.sub.11•2HCl naltrexol)2-ether Nal026 PEG6-(6-β- C.sub.52H.sub.72N.sub.2O.sub.13 C.sub.52H.sub.72N.sub.2O.sub.13•2HCl naltrexol)2-ether Nal030 PEG8-(6-β- C.sub.56H.sub.80N.sub.2O.sub.15 C.sub.56H.sub.80N.sub.2O.sub.15•2HCl naltrexol)2-ether Nal037 mPEG6-(6-α- C.sub.33H.sub.51NO.sub.10 C.sub.33H.sub.51NO.sub.10•HCl naltrexol)-ether Nal041 mPEG8-(6-α- C.sub.37H.sub.59NO.sub.12 C.sub.37H.sub.59NO.sub.12•HCl naltrexol)-ether Nal038 mPEG6-(6-β- C.sub.33H.sub.51NO.sub.10 C.sub.33H.sub.51NO.sub.10•HCl naltrexol)-ether Nal045 mPEG6-(6-α- C.sub.32H.sub.49NO.sub.10 C.sub.32H.sub.49NO.sub.10•HCl naloxol)-ether Nal049 mPEG8-(6-α- C.sub.36H.sub.57NO.sub.12 C.sub.36H.sub.57NO.sub.12•HCl naloxol)-ether Nal046 mPEG6-(6-β- C.sub.32H.sub.49NO.sub.10 C.sub.32H.sub.49NO.sub.10•HCl naloxol)-ether

(47) TABLE-US-00003 TABLE 3 Raw Materials for Preparation of the Conjugate of the Present Disclosure Raw Material 1 3-MOM-6- 3-MOM-6- 3-MOM-6- 3-MOM-6- Raw Material 2 α-naltrexol β-naltrexol α-naloxol β-naloxol PEG3-(OMs)2 Nal053 PEG4-(OMs)2 Nal021 Nal022 PEG6-(OMs)2 Nal025 Nal026 PEG8-(OMs)2 Nal029 Nal030 PEG10-(OMs)2 Nal033 mPEG6-OMs Nal037 Nal038 Nal045 Nal046 mPEG8-OMs Nal041 Nal049

(48) Na1021: Under the protection of argon, 1.10 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 3.0 g of 3-MOM-6-α-naltrexol in 12 ml of tetrahydrofuran and 1.15 g of PEG4-(OMs)2 in 12 ml of tetrahydrofuran were subsequently added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate. After the desiccant was removed, the organic layer was concentrated by rotary evaporation to obtain a residue. Then said residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain product PEG4-(6-α-naltrexol)2-ether. The above-mentioned product was dissolved in 20 ml of ethanol, then 3.0 ml of hydrogen chloride ethanol solution (1.0 M) was added, and the mixture was well mixed and then concentrated by rotary evaporation to obtain 1.43 g of residue. The resulting residue was dissolved in 20 ml of water and freeze-dried to obtain 0.97 g of almost white powdery solid, i.e. dihydrochloride salt of Na1021. 1H-NMR (400 MHz, D2O), δ ppm 6.72-6.77 (2H, d, J=8.0 Hz), 6.62-6.68 (2H, d, J=8.0 Hz), 4.81-4.86 (2H, d, J=4.8 Hz), 3.90-4.00 (4H, m), 3.66-3.74 (2H, m), 3.59-3.66 (2H, m), 3.45-3.59 (12H, m), 3.24-3.34 (2H, d, J=19.6 Hz), 3.16-3.24 (2H, m), 3.03-3.14 (4H, m), 2.86-2.97 (2H, m), 2.72-2.84 (2H, m), 2.35-2.47 (2H, m), 1.68-1.79 (2H, m), 1.47-1.68 (6H, m), 1.16-1.30 (2H, m), 0.91-1.04 (2H, m), 0.67-0.77 (2H, m), 0.57- 0.67 (2H, m), 0.28-0.43 (4H, m). LC/MS (ESI) m/z 846.0 [M+H]+ and 423.8 [M+2H]++.

(49) Na1025: Under the protection of argon, 0.73 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 2.92 g of 3-MOM-6-α-naltrexol in 12 ml of tetrahydrofuran and 1.38 g of PEG6-(OMs)2 in 12 ml of tetrahydrofuran were subsequently added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate. After the desiccant was removed, the organic layer was concentrated by rotary evaporation to obtain 4.18 g of residue. Then said residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain product PEG6-(6-α-naltrexol)2-ether. The above-mentioned product was dissolved in 20 ml of ethanol, then 3.0 ml of hydrogen chloride—ethanol solution (1.0 M) was added, and the mixture was well mixed and then concentrated by rotary evaporation to obtain 1.1 g of residue. The resulting residue was dissolved in 20 ml of water and freeze-dried to obtain 0.71 g of almost white powdery solid, i.e. dihydrochloride salt of Na1025. 1H-NMR (400 MHz, D2O), δ ppm 6.72-6.77 (2H, d, J=8.0 Hz), 6.62-6.68 (2H, d, J=8.0 Hz), 4.81-4.86 (2H, d, J=4.8 Hz), 3.90-4.00 (4H, m), 3.66-3.74 (2H, m), 3.59-3.66 (2H, m), 3.45-3.59 (20H, m), 3.24-3.34 (2H, d, 20.0 Hz)), 3.16-3.24 (2H, m), 3.03-3.14 (4H, m), 2.86-2.97 (2H, m), 2.72-2.84 (2H, m), 2.35-2.47 (2H, m), 1.68-1.79 (2H, m), 1.47-1.68 (6H, m), 1.16-1.30 (2H, m), 0.91-1.04 (2H, m), 0.67-0.77 (2H, m), 0.57-0.67 (2H, m), 0.28-0.43 (4H, m), LC/MS (ESI) m/z 934.1 [M+H]+ and 467.8 [M+2H]++.

(50) Na1029: Under the protection of argon, 0.92 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 2.99 g of 3-MOM-6-α-naltrexol in 12 ml of tetrahydrofuran and 1.74 g of PEG8-(OMs)2 in 12 ml of tetrahydrofuran were subsequently added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate. After the desiccant was removed, the organic layer was concentrated by rotary evaporation to obtain 4.07 g of residue. Then said residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain product PEG8-(6-α-naltrexol)2-ether. The above-mentioned product was dissolved in 20 ml of ethanol, then 2.5 ml of hydrogen chloride ethanol solution (1.0 M) was added, and the mixture was well mixed and then concentrated by rotary evaporation to obtain 1.0 g of residue. The resulting residue was dissolved in 20 ml of water and freeze-dried to obtain 0.84 g of almost white powdery solid, i.e. dihydrochloride salt of Na1029. 1H-NMR (400 MHz, D2O), δ ppm 6.72-6.77 (2H, d, J=8.0 Hz), 6.62-6.68 (2H, d, J=8.0 Hz), 4.81-4.88 (2H, d, J=4.8 Hz), 3.90-4.02 (4H, m), 3.66-3.74 (2H, m), 3.44-3.66 (30H, m), 3.24-3.34 (2H, d, J=20.0 Hz), 3.16-3.24 (2H, m), 3.03-3.14 (4H, m), 2.86-2.97 (2H, m), 2.72-2.84 (2H, m), 2.35-2.47 (2H), 1.68-1.79 (2H, m), 0.47-1.68 (6H, m), 1.16-1.30 (2H, m), 0.91-1.04 (2H, m), 0.67-0.77 (2H, m), 0.57-0.67 (2H, m), 0.28-0.43 (4H, m) LC/MS (ESI) m/z 1022.2 [M+H]+ and 511.9 [M+2H]++.

(51) Na1033: Under the protection of argon, 0.92 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 3.00 g of 3-MOM-6-α-naltrexol in 12 ml of tetrahydrofuran and 2.00 g of PEG10-(OMs)2 in 12 ml of tetrahydrofuran were subsequently added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate. After the desiccant was removed, the organic layer was concentrated by rotary evaporation to obtain 4.18 g of residue. Then said residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain product PEG10-(6-α-naltrexol)2-ether. The above-mentioned product was dissolved in 20 ml of ethanol, then 2.5 ml of hydrogen chloride—ethanol solution (1.0 M) was added, and the mixture was well mixed then and concentrated by rotary evaporation to obtain 1.3 g of residue. The resulting residue was dissolved in 20 ml of water and freeze-dried to obtain 1.00 g of almost white powdery solid, i.e. dihydrochloride salt of Na1033. 1 H-NMR (400 MHz, D2O), δ ppm 6.72-6.77 (2H, d, J=8.0 Hz), 6.62-6.68 (2H, d, J=8.0 Hz), 4.81-4.88 (2H, d, J=4.8 Hz), 3.90-4.02 (4H, m), 3.67-3.74 (2H, m), 3.44-3.67 (38H, m), 3.24-3.34 (2H, d, J=20.0 Hz), 3.16-3.24 (2H, m), 3.03-3.14 (4H, m), 2.86-2.97 (2H, m), 2.72-2.84 (2H, m), 2.35-2.47 (2H, m), 1.68-1.79 (2H, m), 1.47-1.68 (6H, m), 1.16-1.30 (2H, m), 0.91-1.04 (2H, m), 0.67-0.77 (2H, m), 0.57-0.67 (2H, m), 0.28-0.43 (4H, m). LC/MS (ESI) m/z 1110.2 [M+H]+ and 555.9 [M+2H]++.

(52) Na1022: Under the protection of argon, 1.05 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 2.70 g of 3-MOM-6-β-naltrexol in 12 ml of tetrahydrofuran and 1.25 g of PEG4-(OMs)2 in 12 ml of tetrahydrofuran were subsequently added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate. After the desiccant was removed, the organic layer was concentrated by rotary evaporation to obtain 3.05 g of residue. Then the resulting residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain product PEG4-(6-β-naltrexol)2-ether. The above-mentioned product was dissolved in 20 ml of ethanol, then 2.5 ml of hydrogen chloride—ethanol solution (1.0 M) was added, and the mixture was well mixed and then concentrated by rotary evaporation to obtain 0.96 g of residue. The resulting residue was dissolved in 20 ml of water and freeze-dried to obtain 0.56 g of almost white powdery solid, i.e. dihydrochloride salt of Na1022. 1H-NMR (400 MHz, D2O), δ ppm 6.69-6.74 (2H, d, J=8.0 Hz), 6.63-6.69 (2H, d, 0.1=8.0 Hz), 4.49-4.56 (2H, d, J=6.4 Hz), 3.90-3.96 (2H, d, J=5.6 Hz), 3.64-3.71 (4H, m), 3.48-3.63 (12H, m), 3.18-3.32 (6H, m), 2.99-3.11 (4H, m), 2.78-2.88 (2H, m), 2.54-2.68 (2H, m), 2.28-2.41 (2H, m), 1.73-1.85 (2H, m), 1.59-1.70 (4H, m), 1.46-1.59 (2H, m), 1.30-1.44 (2H, m), 0.88-1.02 (2H, m), 0.64-0.75 (2H, m), 0.54-0.64 (2H, m), 0.25-0.41 (4H, m). LC/MS (ESI) m/z 845.8 [M+H]+ and 423.9 [M+2H]++.

(53) Na1026: Under the protection of argon, 1.10 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 2.80 g of 3-MOM-6-β-naltrexol in 12 ml of tetrahydrofuran and 1.36 g of PEG6-(OMs)2 in 12 ml of tetrahydrofuran were subsequently added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary, evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate. After the desiccant was removed, the organic layer was concentrated by rotary evaporation to obtain 3.60 g of residue. Then the resulting residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain product PEG6-(6-β-naltrexol)2-ether. The above-mentioned product was dissolved in 20 ml of ethanol, then 2.5 ml of hydrogen chloride ethanol solution (1.0 M) was added, and the mixture was well mixed and then concentrated by rotary evaporation to obtain 1.05 g of residue. The resulting residue was dissolved in 20 ml of water and freeze-dried to obtain 0.61 g of almost white powdery solid, i.e. dihydrochloride salt of Na1026. 1H-NMR (400 MHz, D2O), δ ppm 6.73-6.78 (2H, d, J=8.4 Hz), 6.67-6.72 (2H, d, J=8.0 Hz), 4.52-4.58 (2H, d, J=6.4 Hz), 3.92-3.99 (2H, d, J=6.0 Hz), 3.66-3.74 (4H, m), 3.50-3.65 (20H, m), 3.20-3.34 (6H, m), 3.02-3.15 (4H, m), 2.81-2.91 (2H, m), 2.57-2.71 (2H, m), 2.31-2.45 (2H, m), 1.76-1.88 (2H, m), 1.61-1.73 (4H, m), 1.51-1.62 (2H, m), 1.34-1.48 (2H, m), 0.91-1.04 (2H, m), 0.67-0.77 (2H, m), 0.57-0.67 (2H, m), 0.28-0.43 (4H, m). LC/MS (ESI) m/z 934.1 [M+H]+ and 467.8 [M+2H]++.

(54) Na1030: Under the protection of argon, 1.10 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 2.8 g of 3-MOM-6-β-naltrexol in 12 ml of tetrahydrofuran and 1.45 g of PEG8-(OMs)2 in 12 ml of tetrahydrofuran were subsequently added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate. After the desiccant was removed, the organic layer was concentrated by rotary evaporation to obtain 3.80 g of residue. Then the resulting residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain product PEG8-(6-β-naltrexol)2-ether. The above-mentioned product was dissolved in 20 ml of ethanol, then 2.5 ml of hydrogen chloride—ethanol solution (1.0 M) was added, and the mixture was well mixed and then concentrated by rotary evaporation to obtain 1.0 g of residue. The resulting residue was dissolved in 20 ml of water and freeze-dried to obtain 0.51 g of almost white powdery solid, i.e. dihydrochloride salt of Na1030. 1H-NMR (400 MHz, D2O), δ ppm 6.71-6.77 (2H, d, J=8.0 Hz), 6.65-6.71 (2H, d, J=8.0 Hz), 4.50-4.56 (2H, d, J=6.4 Hz)), 3.91-3.97 (2H, d, J=6.0 Hz), 3.64-3.71 (4H, m), 3.48-3.63 (28H, m), 3.18-3.32 (6H, m), 2.99-3.11 (4H, m), 2.78-2.88 (2H, m), 2.54-2.68 (2H, m), 2.28-2.41 (2H, m), 1.73-1.85 (2H, m), 1.59-1.70 (4H, m), 1.48-1.59 (2H, m), 1.32-1.44 (2H, m), 0.88-1.02 (2H, m), 0.64-0.75 (2H, m), 0.54-0.64 (2H, m), 0.25-0.41 (4H, m). LC/MS (ESI) m/z 512.0 [M+2H]++.

(55) Na1037: Under the protection of argon, 0.37 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 0.92 g of 3-MOM-6-α-naltrexol in 12 ml of tetrahydrofuran and 0.823 g of mPEG6-OMs in 12 ml of tetrahydrofuran were subsequently added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate. After the desiccant was removed, the organic layer was concentrated by rotary evaporation to obtain 1.64 g of residue. Then the resulting residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain product mPEG6-(6-α-naltrexol)-ether. The above-mentioned product was dissolved in 20 ml of ethanol, then 3.5 ml of hydrogen chloride ethanol solution (1.0 M) was added, and the mixture was well mixed and then concentrated by rotary evaporation to obtain 0.65 g of residue. Said residue was dissolved in 20 ml of water and freeze-dried to obtain 0.42 g of almost white powdery solid, i.e. hydrochloride salt of Na1037. 1H-NMR (400 MHz, D2O), δ ppm 6.72-6.78 (1H, d, J=8.0 Hz), 6.62-6.69 (1H, d, J=8.4 Hz), 4.82-4.88 (1H, d, J=4.8 Hz), 3.91-4.01 (2H, m), 3.67-3.75 (1H, m), 3.43-3.67 (23H, m), 3.16-3.34 (5H, m), 3.03-3.15 (2H, m), 2.86-2.97 (1H, m), 2.72-2.85 (1H, m), 2.35-2.48 (1H, m), 1.70-1.82 (1H, m), 1.48-1.70 (3H, m), 1.20- 1.35 (1H, m), 0.91-1.04 (1H, m), 0.67-0.77 (1H, m), 0.57-0.67 (1H, m), 0.28-0.43 (2H, m). LC/MS (ESI) m/z 623.1 [M+H]+.

(56) Na1041: Under the protection of argon, 0.44 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 1.00 g of 3-MOM-6-α-naltrexol in 12 ml of tetrahydrofuran and 1.05 g of mPEG8-OMs in 12 ml of tetrahydrofuran were subsequently added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate, After the desiccant was removed, the organic layer was concentrated by rotary evaporation to obtain 1.76 g of residue. Then the resulting residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain product mPEG8-(6-α-naltrexol)-ether. The above-mentioned product was dissolved in 20 ml of ethanol, then 3.0 ml of hydrogen chloride—ethanol solution (1.0 M) was added, and the mixture was well mixed and then concentrated by rotary evaporation to obtain 0.95 g of residue. The resulting residue was dissolved in 20 ml of water and freeze-dried to obtain 0.67 g of almost white powdery solid, i.e. hydrochloride salt of Na1041. 1H-NMR (400 MHz, D2O), δ ppm 6.72-6.78 (1H, d, J=8.0 Hz), 6.62-6.69 (1H, d, J=8.4 Hz), 4.82-4.88 (1H, d, J=4.8 Hz), 3.91-4.01 (2H, m), 3.67-3.75 (1E, m), 3.43-3.67 (31H, m), 3.16-3.34 (51-1, in), 3.03-3.15 (2H, m), 2.86-2.97 (1H, m), 2.72-2.85 (1H, m), 2.35-2.48 (1H, m), 1.70-1.82 (1H, m), 1.48- 1.70 (3H, m), 1.20-1.35 (1H, m), 0.91-1.04 (1H, m) 0.67-0.77 (1H, m), 0.57-0.67 (1H, m), 0.28-0.43 (2H, m). LC/MS (ESI) m/z 711.1 [M+H]+.

(57) Na1038: Under the protection of argon, 0.55 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 1.20 g of 3-MOM-6-β-naltrexol in 12 ml of tetrahydrofuran and 1.05 g of mPEG6-OMs in 12 ml of tetrahydrofuran were subsequently added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary, evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate. After the desiccant was removed and concentrated by rotary evaporation to obtain 1.90 g of residue. Then the resulting residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain product mPEG6-(6-β-naltrexol)-ether. The above-mentioned product was dissolved in 20 ml of ethanol, then 2.5 ml of hydrogen chloride—ethanol solution (1.0 M) was added, and the mixture was well mixed and then concentrated by rotary evaporation to obtain 1.0 g of residue. Said residue was dissolved in 20 ml of water and freeze-dried to obtain 0.71 g of almost white powdery solid, i.e. hydrochloride salt of Na1038. 1H-NMR (400 MHz, D2O), δ ppm 6.75-6.80 (1H, d, J=8.0 Hz), 6.68-6.75 (1H, d, J=8.4 Hz), 4.52-4.60 (1H, d, J=6.4 Hz), 3.92-4.01 (1H, d, J=6.0 Hz), 3.68-3.76 (2H, t), 3.54-3.67 (20H, m), 3.47-3.54 (2H, m), 3.20-3.36 (6H, m), 3.02-3.17 (2H, m), 2.81-2.91 (1H, m), 2.59-2.73 (1H, m), 2.32-2.46 (1H, m), 1.78-1.89 (1H, m), 1.52-1.74 (3H, m), 1.36-1.50 (1H, m), 0.91-1.04 (1H, m), 0.67-0.77 (1H, m), 0.57-0.67 (1H, m) 0.28-0.43 (2H, m) LC/MS (ESI) m/z 623.1 [M+H]+.

(58) Na1045: Under the protection of argon, 0.60 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 1.40 g of 3-MOM-6-α-naloxol in 12 ml of tetrahydrofuran and 1.38 g of mPEG6-OMs in 12 ml of tetrahydrofuran were subsequently added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate. After the desiccant was removed and concentrated by rotary evaporation to obtain 2.47 g of residue. Then the resulting residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain product mPEG6-(6-α-naloxol)-ether. The above-mentioned product was dissolved in 20 ml of ethanol; then 2.5 ml of hydrogen chloride—ethanol solution (1.0 M) was added, and the mixture was well mixed and then concentrated by rotary evaporation to obtain 1.5 g of residue. The resulting residue was dissolved in 20 ml of water and freeze-dried to obtain 1.11 g of almost white powdery solid, i.e. hydrochloride salt of Na1045. 1H-NMR (400 MHz, D2O), δ ppm 6.72-6.79 (1H, d, J=8.4 Hz), 6.62-6.70 (1H, d, J=8.0 Hz), 5.72-5.88 (1H, m), 5.48-5.60 (2H, t), 4.81-4.88 (1H, d, J=5.2 Hz), 3.89-3.99 (1H, m), 3.40-3.85 (27H, m), 3.27-3.37 (1H, d, J=20.0 Hz)), 3.28 (3H, s), 3.10-3.21 (1H, m) 2.95-3.08 (1H, m), 2.76-2.90 (1H, m), 2.34-2.48 (1H, m), 1.62-1.80 (2H m), 1.45-1.62 (2H, m), 1.21-1.35 (1H, m). LC/MS (ESI) m/z 609.1 [M+H]+.

(59) Na1049: Under the protection of argon, 0.60 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 1.50 g of 3-MOM-6-α-naloxol in 12 ml of tetrahydrofuran and 1.81 g of mPEG8-OMs in 12 ml of tetrahydrofuran were subsequently added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate. After the desiccant was removed, the organic layer was concentrated by rotary evaporation to obtain 2.93 g of residue. Then the resulting residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain product mPEG8-(6-α-naloxol)-ether. The above-mentioned product was dissolved in 20 ml of ethanol, the pH was adjusted to 1 to 2 with hydrogen chloride—ethanol solution (1.0 M), and the mixture was well mixed and then concentrated by rotary evaporation to obtain 1.6 g of residue. Said residue was dissolved in 20 ml of water and freeze-dried to obtain 1.33 g of almost white powdery solid, i.e. hydrochloride salt of Na1049. 1H-NMR (400 MHz, D2O), δ ppm 6.72-6.79 (1H, d, J=8.4 Hz), 6.62-6.70 (1H, d, J=8.4 Hz), 5.72-5.88 (1H, m), 5.48-5.60 (2H, t), 4.81-4.88 (1H, d, J=5.2 Hz), 3.89-3.99 (1H, m), 3.40-3.85 (35H, m), 3.27-3.37 (1H, d, J=20.0 Hz)), 3.28 (3H, s), 3.10-3.21 (1H, m), 2.95-3.08 (1H, m), 2.76-2.90 (1H, m), 2.34-2.48 (1H, m), 1.62-1.80 (2H, m), 1.45-1.62 (2H, m), 1.21-1.35 (1H, m). LC/MS (ESI) m/z 697.1 [M+H]+.

(60) Na1046: Under the protection of argon, 0.55 g of sodium hydride and 20 ml of dimethylformamide were added to a reaction flask and stirred to be a suspension. Then, 1.26 g of 3-MOM-6-β-naloxol in 12 ml of tetrahydrofuran and 1.24 g of mPEG6-OMs in 12 ml of tetrahydrofuran were subsequently added. The temperature was raised to 60° C., reaction was carried out with stirring and monitored using TLC. After the reaction was completed, excess sodium hydride was quenched with water. The reaction solution was concentrated by rotary evaporation, the resulting residue was extracted with dichloromethane and water system, and the organic layer was separated. The organic layer was washed five times with a 5% aqueous sodium carbonate solution, separated and dried with anhydrous sodium sulfate. After the desiccant was removed and concentrated by rotary evaporation to obtain 2.08 g of residue. Then the resulting residue was treated with 15 ml of hydrochloric acid (4.0 M) and stirred at room temperature overnight. After the reaction was completed, the pH of the reaction solution was adjusted to 3 to 5 with 4 M aqueous ammonia. Finally, chromatography purification was carried out to obtain product mPEG6-(6-β-naloxol)-ether. The above-mentioned product was dissolved in 20 ml of ethanol, then 2.5 ml of hydrogen chloride—ethanol solution (1.0 M) was added, and the mixture was well mixed and then concentrated by rotary evaporation to obtain 1.1 g of residue. Said residue was dissolved in 20 ml of water and freeze-dried to obtain 0.85 g of almost white powdery solid, i.e. hydrochloride salt of Na1046. 1H-NMR (400 MHz, D2O), δ ppm 6.75-6.81 (1H, d, 0.1=8.0 Hz), 6.69-6.75 (1H, d, J=8.4 Hz), 5.73-5.88 (1H, m), 5.49-5.59 (2H, m), 4.52-4.59 (1H, d, J=6.8 Hz), 3.45-3.85 (27H, m), 3.23-3.37 (5H, m), 3.10-3.20 (1H, m), 2.96-3.08 (1H, m), 2.62-2.76 (1H, m), 2.32-2.45 (1H, m), 1.76-1.88 (1H, m), 1.54-1.72 (3H, m), 1.33-1.47 (1H, m) LC/MS (ESI) m/z 609.2 [M+H]+.

Example 6: In Vitro Activity Test

(61) CHO-K1/Gα15/OPRM1 cells (Gen Script (Nanjing) Co., Ltd.) stably expressing OPRM1 (Mμ opioid receptor gene) receptor were cultured in 10-cm dishes at 37° C./5% CO.sub.2 incubator. When the cell reached 80% to 85% confluence, cells were digested and collected. Cell suspension was inoculated into a 384-well microplate at a density of 15,000 cells per well, and cultured in a 37° C./5% CO.sub.2 incubator for at least 18 hours before experiment.

(62) Before the test, the conjugates to be tested was diluted with HBSS buffer (containing 20 mM HEPES) and prepared as 5× solution with a concentration five times of the test solution. The final concentrations of the conjugate were 0.128 nM, 0.64 nM, 3.2 nM, 1.6 nM, 80 nM, 400 nM, 2000 mM and 10000 nM, each in duplicate. The concentration of the corresponding control DMSO is 0.1%.

(63) The protocol for preparing agonist detection: cells were inoculated into 384-well microplates, 20 μl of cell suspension per well (containing 15,000 cells); the plate was placed in 37° C./5% CO.sub.2 incubator to continue the culture; cells were taken out after 18 hours, and dye was added, 20 μl per well; the plate was incubated in 37° C./5% CO.sub.2 incubator for 1 hour and equilibrated at room temperature for 15 minutes; and 10 μl of 5× test solution of agonist was added to each well for RFU value detection.

(64) The protocol for preparing inhibitor detection: cells were inoculated into 384-well microplates, 20 μl of cell suspension per well (containing 15,000 cells); the plate was placed in 37° C./5% CO.sub.2 incubator to continue the culture; cells were taken out after 18 hours; for the inhibitor detection, 20 μl of dye was added to the plate, and then 10 μl of prepared conjugate solution was added; the plate was incubated in 37° C./5% CO.sub.2 for 1 hour and equilibrate at room temperature for 15 minutes; 12.5 μl of 5× EC80 positive agonist was added for RFU value detection.

(65) The 384-well microplate containing the conjugate solution (5× test concentration), cell plate and pipette tip box were placed in FLIPR (Molecule Devices), and the agonist detection program was run. The overall detection time of the instrument was 120 seconds. 10 μl of agonist was automatically added to the plate at the 21.sup.st second. The 384-well microplate containing 5× EC80 positive agonist, cell plate and pipette tip box were placed in FLIPRTETRA (Molecule Devices), and the inhibitor detection program was run. The overall detection time of the instrument was 120 seconds. 12.5 μl of positive agonist was automatically added to the plate at the 21.sup.st second. The effects of 13 conjugates on the OPRM1 receptor were shown in Table 4 below

(66) TABLE-US-00004 TABLE 4 IC50 of the conjugates of the present disclosure Inhibition rate (%) of highest concentration Highest (mean ± standard Name IC50(M) concentration deviation) (N = 2) Naltrexone 4.15 × 10−8 10000 nM 101.35 ± 0.65 hydrochloride Naloxone 9.75 × 10−8 10000 nM 100.08 ± 0.40 hydrochloride Nal053 2.50 × 10−7 10000 nM 101.85 ± 0.97 Nal021 2.29 × 10−7 10000 nM 100.31 ± 0.56 Nal025 1.85 × 10−7 10000 nM 100.63 ± 0.48 Nal026 1.83 × 10−7 10000 nM 100.52 ± 0.26 Nal029 1.61 × 10−7 10000 nM  98.00 ± 3.09 Nal033 1.97 × 10−7 10000 nM  99.12 ± 1.00 Nal037 2.25 × 10−7 10000 nM 100.58 ± 0.34 Nal041 2.10 × 10−7 10000 nM  95.82 ± 1.86 Nal045 1.32 × 10−6 10000 nM  90.44 ± 3.22 Nal046 3.84 × 10−6 10000 nM  79.55 ± 5.73 Nal049 1.05 × 10−6 10000 nM  94.31 ± 5.39

(67) The data in the above table shows that the conjugates with two naltrexone and PEG (i.e., Na1053, Na1021, Na1025, Na1026, Na1029, and Na1033) exhibit a lower IC50 than the naloxone-PEG conjugate or the mononaltrexone-PEG conjugate; when the number of the repeating units of polyethylene glycol is a specific value (e.g., 6, 8, 10), the conjugates of the present disclosure exhibits even lower IC50, which is really unexpected.

Example 7: Experimental Study Regarding the Blood-Brain Barrier

(68) The experimental study on whether the conjugate test group can pass through the blood-brain barrier was completed by using the mouse hot plate analgesia experiment. Morphine has an analgesic effect, and if the conjugate to be tested can antagonize the analgesic effect of morphine, then it can be concluded that the conjugate can pass through the blood-brain barrier.

(69) Six conjugates Na1021, Na1029, Na1045, Na1037, Na1041, and Na1049 were subjected to mouse hot plate analgesia experiments for four times, respectively. In each experiment, mice (JOINN Laboratories (Suzhou)) were randomly divided into four groups (eight per group): one blank control group, one morphine control group, and two other groups which were administered with morphine by gavage first, and then low dose and high dose of the conjugates were subcutaneously injected respectively. The mice were put on a hot plate (55±0.1) ° C. before administration, and the time was recorded immediately. The duration from the beginning on the hot plate to the first hind paw licking was recorded as the reaction time of heat-pain sensation (i.e., pain threshold, the unit is second (s)). The pain threshold was set as 100% analgesia if there was no hind paw licking in 60 seconds. It was recorded as 60 seconds when the time exceeded 60 seconds to prevent burn of foot. The mice were again put on a (55±0.1) ° C. hot plate 60 minutes after the administration of drug and the reaction time of heat-pain sensation was recorded. The experimental result was shown in the table below. It can be seen that the difference between the pain threshold before and after administration in the blank control group is not significant (P>0.05), while the pain threshold before and after administration in the morphine control group are significantly different (P<0.01), and the differences between the pain thresholds before and after administration in high-dose groups of Na1037 and Na1041 are not significant (P>0.05). The pain threshold before and after administration in each other drug-administered groups is significantly different (P<0.05). Therefore, it can be concluded that the conjugates Na1037 and Na1041 can pass through the blood-brain barrier so as to antagonize the analgesic effect of morphine. Na1021, Na1029, Na1045, or Na1049 cannot pass through the blood-brain barrier. The value of the pain thresholds before and after administration was compared and the percentage of increase in pain threshold was calculated according to the following formula to determine the effect of drug on antagonizing morphine analgesia.
Percentage of increase in pain threshold=[(average heat pain threshold after administration−average pain threshold before administration)/average pain threshold before administration]×100%

(70) TABLE-US-00005 TABLE 5 Experimental Result of Hot Plate Analgesia Experiment with Nal021 and Nal029 Average Pain Average Pain Threshold Threshold before after Administration Administration Group Method of Treatment (s) Mean ± SD (s) Mean ± SD 1 Blank Control  25.00 ± 10.54 31.13 ± 13.85  2 Morphine Control 19.25 ± 5.57 53.25 ± 10.96** 3 NAL021(4.36 μmol/kg) + 20.63 ± 7.07 56.25 ± 10.61** Morphine 4 NAL021 (21.8 μmol/kg) + 18.63 ± 4.60 59.75 ± 0.71**  Morphine 5 NAL029 (3.66 μmol/kg) + 20.00 ± 5.71 60.00 ± 0**    Morphine 6 NAL029 (18.28 μmol/kg) + 22.13 ± 7.26 56.5 ± 9.90** Morphine Note: * P < 0.05, **P < 0.01

(71) TABLE-US-00006 TABLE 6 Experimental Result of Hot Plate Analgesia Experiment with Nal045 Average Pain Average Pain Threshold Threshold before after Administration Administration Group Method of Treatment (s) Mean ± SD (s) Mean ± SD 1 Blank Control 21.38 ± 5.13 25.00 ± 5.81   2 Morphine Control 20.38 ± 3.93 52.00 ± 9.67**  5 NAL045 (3.10 μmol/kg) + 20.38 ± 4.53 52.13 ± 14.78** Morphine 6 NAL045 (15.52 μmol/kg) + 24.00 ± 8.38 51.00 ± 11.43** Morphine Note: * P < 0.05, **P < 0.01

(72) TABLE-US-00007 TABLE 7 Experimental Result of Hot Plate Analgesia Experiment with Nal037 Average Pain Average Pain Threshold Threshold before after Administration Administration Group Method of Treatment (s) Mean ± SD (s) Mean ± SD 1 Blank Control 21.13 ± 4.26 20.75 ± 7.25 2 Morphine Control 20.88 ± 4.91   38.5 ± 15.58** 5 Nal037 (4 μmol/kg) + 19.38 ± 5.21  31.50 ± 11.83* Morphine 6 Nal037 (20 μmol/kg) +  20.5 ± 3.93 20.88 ± 5.87 Morphine Note: *P < 0.05, **P < 0.01

(73) TABLE-US-00008 TABLE 8 Experimental Result of Hot Plate Analgesia Experiment with Nal041 and Nal049 Average Pain Average Pain Threshold Threshold before after Administration Administration Group Method of Treatment (s) Mean ± SD (s) Mean ± SD 1 Blank Control 17.50 ± 3.59 21.87 ± 5.84   2 Morphine Control 15.12 ± 6.38 39.00 ± 17.25** 3 Nal041 (4 μmol/kg) + 14.25 ± 1.75  38.5 ± 14.74** Morphine 4 Nal041 (20 μmol/kg) + 16.88 ± 6.79 22.00 ± 5.12   Morphine 5 Nal049 (4 μmol/kg) + 14.00 ± 7.09 31.63 ± 10.04** Morphine 6 Nal049 (20 μmol/kg) +  13.5 ± 5.07 25.88 ± 7.64**  Morphine Note: * P < 0.05, **P < 0.01

(74) TABLE-US-00009 TABLE 9 Percentage of Increase in Pain Threshold for Each Compound Group with Different dose of Percentage of Increase in Serial number Each Compound Pain Threshold 1 NAL021 (4.36 μmol/kg) 172.66% 2 NAL021 (21.8 μmol/kg) 220.72% 3 NAL029 (3.66 μmol/kg) 200.00% 4 NAL029 (18.28 μmol/kg) 155.31% 5 NAL045 (3.10 μmol/kg) 155.79% 6 NAL045 (15.52 μmol/kg) 112.50% 7 NAL037 (4 μmol/kg) 62.54% 8 NAL037 (20 μmol/kg) 1.85% 9 NAL041 (4 μmol/kg) 170.18% 10 NAL041 (20 μmol/kg) 30.33% 11 NAL049 (4 μmol/kg) 125.93% 12 NAL049 (20 μmol/kg) 91.70%

Example 8: Experimental Study on Anti-Constipation

(75) The anti-constipation study experiment was completed by using the test of antagonizing the antidiarrheal effect of loperamide hydrochloride. The specific experiment process was: the mice were divided into groups randomly, eight mice each group, wherein one group was administered with 0.5 mL of castor oil by gavage, one group was administered with 2 mg/kg loperamide hydrochloride and 0.5 mL of castor oil by gavage, and conjugate test groups were respectively administered with Na1021 (300 nmol/kg), Na1029 (300 nmol/kg), Na1033 (300 nmol/kg) and Na1045 (600 nmol/kg) by gavage first, and then with 2 mg/kg loperamide and 0.5 mL castor oil by gavage. The cumulative number of diarrhea times in the mice was observed in six hours after the administration, and the loose stools point was counted to determine diarrhea conditions. The result shows that the total number of diarrhea times in the castor oil control group is nearly two times as high as that compared with the loperamide hydrochloride castor oil control group six hours after the administration, indicating that the diarrhea model was effective. The total number of diarrhea times of Na1029 is relatively high six hours after the administration, and the anti-diarrhea effect of antagonizing loperamide hydrochloride is stronger, indicating that the anti-constipation is better. The specific data is shown in Table 10.

(76) TABLE-US-00010 TABLE 10 Experimental Result for each Conjugate to Antagonize the Antidiarrheal Effect of Loperamide Hydrochloride Total Number of Diarrhea Times Six Hours after Group Method of Treatment Administration 1 Castor Oil Control 16.625 ± 5.95  2 Loperamide Hydrochloride + 10.88 ± 7.14 Castor Oil 4 Nal029 (300 nmol/kg) + 20.88 ± 5.82 Loperamide Hydrochloride + Castor Oil 5 Nal033 (300 nmol/kg) + 15.25 ± 4.13 Loperamide Hydrochloride + Castor Oil 6 Nal045 (600 nmol/kg) +  9.75 ± 6.96 Loperamide Hydrochloride + Castor Oil

Example 9: Pharmacodynamical Experimental Study on Anti-Constipation of Conjugate Na1029

(77) The mice were divided into groups randomly, eight mice each group, wherein one group was administered with 0.5 mL of castor oil by gavage, one group was administered with 2 mg/kg loperamide and 0.5 mL castor oil by gavage, other groups were respectively administered with different doses of conjugate Na1029 first, then with 2 mg/kg loperamide hydrochloride and 0.5 mL castor oil by gavage.

(78) It can be seen from the experimental result that compared with the model control group, conjugate Na1029 can significantly antagonize the antidiarrheal effect of loperamide hydrochloride, and the pharmacodynamics is better than that of naltrexone with same dose and have a dose-dependent manner. The EC50 is 0.086 mg/kg approximately. The specific result is shown in Table 11 and FIG. 1.

(79) TABLE-US-00011 TABLE 11 Experimental Result for Conjugate Nal029 to Antagonize Antidiarrheal Effect of Loperamide Hydrochloride Total Number of Diarrhea Tinies Six Hours after Group Method of Treatment Administration 1 Castor Oil Control 11.13 ± 5.54 2 Loperamide Hydrochloride +  5.75 ± 3.69 Castor Oil 3 Nal029 (500 μg/kg) + 18.25 ± 3.99 Loperamide Hydrochloride + Castor Oil 4 Nal029 (100 μg/kg) + 12.13 ± 5.67 Loperamide Hydrochloride + Castor Oil 5 Nal029 (30 μg/kg) +  6.50 ± 5.88 Loperamide Hydrochloride + Castor Oil 6 Nal029 (10 μg/kg) +  5.25 ± 7.42 Loperamide Hydrochloride + Castor Oil 7 Naltrexone (500 μg/kg) + 14.00 ± 5.76 Loperamide Hydrochloride + Castor Oil

(80) The above description is only preferred embodiments, which is only used as examples instead of a limitation to the combination of the features required to practice the present invention. The provided headings are not meant to limit the various embodiments of the present invention.

(81) All publications and patents mentioned in this application are hereby incorporated by reference. Various modifications and variations of the described methods and compositions of the present disclosure are apparent to those having ordinary skill in the art without departing from the scope and spirit of the present invention. Although the present invention is described by way of specific preferred embodiments, it should be understood that the claimed invention should not be inappropriately limited to these specific embodiments. In fact, those various variations of the described modes for practicing the present invention that are obvious to those people having ordinary skill in the art are intended to be included within the protective scope of the appended claims.