A preparation method of deuterium-labeled ractopamine hydrochloride-d5

Abstract

The invention belongs to the technical field of food detection, and in particular relates to a preparation method of deuterium-labeled ractopamine hydrochloride-d5. Deuterium-labeled ractopamine-d5 is obtained by using deuterated phenol as a raw material through a series of reactions including substitution, phenolic hydroxyl protection, reductive amination, deuterated aluminum hydride lithium reduction, palladium carbon reduction, deprotection and salt formation; the reaction process has good atom economy, simple synthesis operation, good reproducibility and stabilizer, and can be used as an internal standard in the field of food quality inspection, and has important practical application value.

Description

Technical Field

[0001] The invention belongs to the technical field of food detection, and in particular relates to a preparation method of deuterium-labeled ractopamine-d5 hydrochloride. Background Art

[0002] Ractopamine is a pharmaceutical raw material, a cardiotonic drug that can be used to treat congestive heart failure. It can also be used to treat muscular dystrophy, grow muscle, reduce fat accumulation, and is beneficial to the growth of fetuses and newborns. The US FDA approved it in 2010 for use in animal nutrition reformulation and is widely used in animal husbandry and breeding. In animal husbandry, it can be used as a "clenbuterol" additive in feed. Its main function is to reduce the fat content ratio in ketone bodies and increase the lean meat output rate. It is the most common type of clenbuterol, and its meat residue toxicity is much lower than other animal feed additives with the same function. The United States claims that legal use at the permissible residue level it has determined will not cause poisoning or short-term harm to humans. However, current experimental data cannot determine whether it will have other side effects on the human body, and it is also unclear whether long-term intake of residual ractopamine by the human body will cause health problems, so it remains controversial. At present, the main detection methods for ractopamine residues in food in China include high performance liquid chromatography, gas chromatography-mass spectrometry, liquid chromatography-mass spectrometry, radioimmunoassay, enzyme-linked immunosorbent assay, etc. However, these methods have problems such as cumbersome pretreatment and large matrix effect, which have a great impact on the measurement results. Isotope dilution mass spectrometry (IDMS) uses stable isotope labeled compounds as internal standard reagents, which well combines the separation ability of chromatography and the qualitative ability of mass spectrometry. By comparing the ratio of ions with the standard ratio of the corresponding mass number, the purpose of accurate quantification can be achieved, and the matrix effect and recovery rate difference caused by the sample pretreatment process can be effectively eliminated, thereby improving the detection accuracy. Therefore, this method is an analytical method with high accuracy and precision. In China, the isotope-labeled ractopamine hydrochloride-d5 standard used in this method is completely dependent on imported products, and factors such as price and delivery period seriously restrict the application of IDMS technology in related detection fields in my country.

[0003] A Chinese patent with publication number CN117263814A discloses a method for synthesizing deuterium-labeled ractopamine, wherein deuterium-labeled 4-acetoxystyrene is obtained by using deuterium water as a source of stable isotope deuterium through a hydrogen-deuterium exchange reaction; then undergoing an epoxidation reaction; and a ring-opening reaction of 4-(3-aminobutyl)phenol; and finally deacetylation to obtain deuterium-labeled ractopamine; the method has simple steps, mild reaction conditions, low cost, and high purity and isotope abundance. A Chinese patent with publication number CN113061094A discloses a method for preparing ractopamine hydrochloride-D6, wherein compound I is used as a starting material, and isotope-labeled ractopamine hydrochloride-D6 is synthesized through HD exchange, reductive amination, deprotection, and salt formation reactions; the method uses cheap labeling raw materials, greatly saves costs, and the experimental process is controllable, easy to operate, and the obtained product has high purity, good reproducibility and stability, and can be prepared on a large scale.

[0004] However, there are no reports on the synthesis of naturally abundant and stable isotope-labeled ractopamine hydrochloride-d5 at home and abroad. The author designed this experiment to synthesize ractopamine hydrochloride-d5. It fundamentally solves the current situation that restricts my country's detection field from relying on imported products. Therefore, the synthesis of ractopamine isotope-labeled compounds is of great significance. Summary of the invention

[0005] In order to solve the technical problems existing in the prior art, the present invention provides a method for preparing deuterium-labeled ractopamine hydrochloride-d5, aiming to propose a method for preparing deuterium-labeled ractopamine hydrochloride-d5 with reasonable design, strong operability, easy product purification, high purity and high yield.

[0006] In order to achieve the above object, the present invention provides the following technical solutions:

[0007] A method for preparing deuterium-labeled ractopamine hydrochloride-d5 comprises the following steps:

[0008] S1. Deuterated phenol (structural formula: Figure 1 1 in the figure) is dissolved in the first solvent, bromine is added dropwise at 0°C for substitution reaction, and stirred under an inert atmosphere until the reaction is complete to obtain deuterated p-bromophenol (structural formula as Figure 1 As shown in 2); wherein the feed ratio of deuterated phenol to bromine is 1:1-1.05;

[0009] S2. After dissolving deuterated p-bromophenol and an alkaline reagent in a second solvent, tert-butyldimethylsilyl chloride is added in batches to protect the phenolic hydroxyl group, and the reaction is carried out overnight under an inert atmosphere to obtain (4-bromophenoxy-2,3,5,6-d4)(tert-butyl)dimethylsilane (structural formula: Figure 13); wherein the alkaline reagent includes any one of imidazole, potassium carbonate, triethylamine, and N,N-diisopropylethylamine; the second solvent includes any one of dichloromethane, 1,2-dichloroethane, and tetrahydrofuran;

[0010] S3. Dissolve (4-bromophenoxy-2,3,5,6-d4)(tert-butyl)dimethylsilane in a third solvent, add n-butyl lithium dropwise at -78°C, react at -78°C for 1-3h under an inert atmosphere, add N,N-dimethylformamide dropwise, and stir at -78°C until the reaction is complete to obtain 4-((tert-butyldimethylsilyl)oxy)benzaldehyde-2,3,5,6-d4 (structural formula: Figure 1 4); wherein the feed ratio of (4-bromophenoxy-2,3,5,6-d4)(tert-butyl)dimethylsilane, n-butyl lithium, and N,N-dimethylformamide is 1:1.3-1.6:2-4;

[0011] S4. Dissolve 4-((tert-butyldimethylsilyl)oxy)benzaldehyde-2,3,5,6-d4 and 1-triphenylphosphine-2-propanone in a fourth solvent, toluene, and heat to 120°C under an inert atmosphere and stir overnight until the reaction is complete to obtain (E)-4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-3-en-2-one (structural formula: Figure 1 5); wherein the feed ratio of 4-((tert-butyldimethylsilyl)oxy)benzaldehyde-2,3,5,6-d4 and 1-triphenylphosphine-2-acetone is 1:1.5-2;

[0012] S5. Dissolve (E)-4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-3-en-2-one in the fifth solvent, add the heterogeneous catalyst palladium carbon in batches and stir evenly, introduce reducing gas hydrogen and stir overnight at room temperature until the reaction is complete, and reduce to obtain 4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-one (structural formula as shown in FIG. Figure 1 6); wherein the fifth solvent comprises any one of ethyl acetate, methanol, ethanol, and tetrahydrofuran;

[0013] S6. Add 4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-one, p-methoxybenzylamine and acetic acid to the sixth solvent, stir at room temperature for 1-3 hours, then add sodium triacetoxyborohydride under ice bath conditions, stir at room temperature overnight until the reaction is complete, and obtain 4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4-N-(4-methoxybenzyl)butan-2-amine (structural formula: Figure 17); wherein the feed ratio of 4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-one, p-methoxybenzylamine, and sodium triacetoxyborohydride is 1:1.1-1.5:2-3; the sixth solvent includes any one of dichloromethane and 1,2-dichloroethane;

[0014] S7. Dissolve 4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4-N-(4-methoxybenzyl)butan-2-amine and potassium carbonate in the seventh solvent, then add 2-bromo-1-(4-hydroxyphenyl)ethan-1-one, and stir overnight at 30°C until the reaction is complete to obtain 2-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl(4-methoxybenzyl)amino)-1-(4-hydroxyphenyl)ethan-1-one (structural formula: Figure 1 8); wherein the feed ratio of 4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4-N-(4-methoxybenzyl)butan-2-amine, 2-bromo-1-(4-hydroxyphenyl)ethan-1-one, and potassium carbonate is 1:1.2-1.5:3;

[0015] S8. Dissolve 2-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl(4-methoxybenzyl)amino)-1-(4-hydroxyphenyl)ethan-1-one in the eighth solvent, add deuterated lithium aluminum hydride in batches at 0°C, stir at room temperature for 45 minutes until the reaction is complete, and reduce with deuterated lithium aluminum hydride to obtain 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl(4-methoxybenzyl)amino)-1-hydroxyethyl-1-d)phenol (structural formula: Figure 1 9); wherein the feed ratio of 2-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl(4-methoxybenzyl)amino)-1-(4-hydroxyphenyl)ethan-1-one and deuterated lithium aluminum hydride is 1:1.2;

[0016] S9. Dissolve 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl(4-methoxybenzyl)amino)-1-hydroxyethyl-1-d)phenol in the ninth solvent, add the heterogeneous catalyst palladium carbon in batches, stir for 4 hours under deuterium atmosphere until the reaction is complete, and hydrogenate with the heterogeneous catalyst palladium carbon to obtain 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl)amino)-1-hydroxyethyl-1-d)phenol (structural formula as shown in FIG. Figure 110 in the figure); wherein the ninth solvent comprises one or a mixture of ethyl acetate, methanol, ethanol, and tetrahydrofuran; preferably, the ninth solvent is a mixed solvent of ethyl acetate and methanol, and the volume ratio of ethyl acetate to methanol is 6:1;

[0017] S10, dissolving 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butyl-2-yl)amino)-1-hydroxyethyl-1-d)phenol and tetrabutylammonium fluoride in the tenth solvent, stirring at room temperature overnight until the reaction is complete, and deprotecting to obtain ractopamine-d5 (structural formula: Figure 1 11 in the figure); wherein the feed ratio of 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butyl-2-yl)amino)-1-hydroxyethyl-1-d)phenol and tetrabutylammonium fluoride is 1:1.1-2;

[0018] S11, after dissolving ractopamine-d5 in the eleventh solvent, hydrochloric acid was added dropwise to adjust the pH of the solution to 2-3, stirred for 0.5-2h, and freeze-dried to obtain the target product ractopamine hydrochloride-d5 (structural formula as Figure 1 12 in the figure); wherein the eleventh solvent comprises any one of dioxane hydrochloride and hydrochloric acid solution.

[0019] The present invention has the following beneficial effects:

[0020] The key intermediate 2-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-2-yl(4-methoxybenzyl)amino)-1-(4-hydroxyphenyl)ethan-1-one is synthesized and then reduced with a substantially equivalent amount of a deuterated reagent to obtain compound 9 (4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-2-yl(4-methoxybenzyl)amino)-1-hydroxyethyl-1-d)phenol) in high yield. The method has good atom economy, simple synthesis operation, good reproducibility and stability. The final stable isotope labeled product has a chemical purity of more than 98.3% and an isotope abundance of more than 98%. It can be used as an internal standard in the field of food quality inspection and has important practical application value. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 Schematic diagram of the synthesis route of deuterium-labeled ractopamine hydrochloride-d5 of the present invention;

[0022] Figure 2 This is the detection spectrum of ractopamine hydrochloride-d5 prepared by the present invention. DETAILED DESCRIPTION

[0023] The following will be combined with the drawings in the embodiments of the present application to clearly and completely describe the technical solutions in the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, not all of the embodiments. Based on the embodiments in the present application, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of this application.

[0024] like Figure 1 As shown, the preparation method of deuterium-labeled ractopamine hydrochloride-d5 in the present invention mainly comprises the following steps:

[0025] S1. Deuterated phenol (structural formula: Figure 1 1 in the figure) is dissolved in the first solvent, bromine is added dropwise at 0°C, and stirred under an inert atmosphere until the reaction is complete to obtain deuterated p-bromophenol (structural formula as Figure 1 2);

[0026] S2. After dissolving deuterated p-bromophenol and an alkaline reagent in a second solvent, tert-butyldimethylsilyl chloride is added in batches, and the mixture is reacted overnight under an inert atmosphere to obtain (4-bromophenoxy-2,3,5,6-d4)(tert-butyl)dimethylsilane (structural formula: Figure 1 3 in the figure);

[0027] S3. Dissolve (4-bromophenoxy-2,3,5,6-d4)(tert-butyl)dimethylsilane in a third solvent, add n-butyl lithium dropwise at -78°C, react at -78°C for 1-3h under an inert atmosphere, add N,N-dimethylformamide dropwise, and stir at -78°C until the reaction is complete to obtain 4-((tert-butyldimethylsilyl)oxy)benzaldehyde-2,3,5,6-d4 (structural formula: Figure 1 4 in );

[0028] S4. Dissolve 4-((tert-butyldimethylsilyl)oxy)benzaldehyde-2,3,5,6-d4 and 1-triphenylphosphine-2-propanone in a fourth solvent, heat to 120°C and stir overnight under an inert atmosphere until the reaction is complete to obtain (E)-4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-3-en-2-one (structural formula: Figure 1 5 in );

[0029] S5. Dissolve (E)-4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-3-en-2-one in the fifth solvent, add palladium carbon in batches and stir evenly, introduce hydrogen and stir overnight at room temperature until the reaction is complete to obtain 4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-one (structural formula: Figure 1 6 in );

[0030] S6. Add 4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-one, p-methoxybenzylamine and catalyst acetic acid to the sixth solvent, stir at room temperature for 1-3 hours, then add sodium triacetoxyborohydride under ice bath conditions, stir at room temperature overnight until the reaction is complete, to obtain 4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4-N-(4-methoxybenzyl)butan-2-amine (structural formula: Figure 1 7 in );

[0031] S7. Dissolve 4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4-N-(4-methoxybenzyl)butan-2-amine and potassium carbonate in the seventh solvent, then add 2-bromo-1-(4-hydroxyphenyl)ethan-1-one, and stir overnight at 30°C until the reaction is complete to obtain 2-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl(4-methoxybenzyl)amino)-1-(4-hydroxyphenyl)ethan-1-one (structural formula: Figure 1 8 in );

[0032] S8. Dissolve 2-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl(4-methoxybenzyl)amino)-1-(4-hydroxyphenyl)ethan-1-one in the eighth solvent, add deuterated lithium aluminum hydride in batches at 0°C, and stir at room temperature for 45 minutes until the reaction is complete to obtain 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl(4-methoxybenzyl)amino)-1-hydroxyethyl-1-d)phenol (structural formula as shown in FIG. Figure 1 9 in );

[0033] S9. Dissolve 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl(4-methoxybenzyl)amino)-1-hydroxyethyl-1-d)phenol in the ninth solvent, add palladium carbon in batches, and stir for 4 hours under a deuterium atmosphere until the reaction is complete to obtain 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl)amino)-1-hydroxyethyl-1-d)phenol (structural formula as shown in FIG. Figure 1 10 in FIG);

[0034] S10, dissolving 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butyl-2-yl)amino)-1-hydroxyethyl-1-d)phenol and tetrabutylammonium fluoride in the tenth solvent, stirring at room temperature overnight until the reaction is complete, to obtain ractopamine-d5 (structural formula: Figure 111 in the figure);

[0035] S11, after dissolving ractopamine-d5 in the eleventh solvent, hydrochloric acid was added dropwise to adjust the pH of the solution to 2-3, stirred for 0.5-2h, and freeze-dried to obtain the target product ractopamine hydrochloride-d5 (structural formula as Figure 1 12 in the figure). Embodiment 1

[0036] Synthesis of deuterated p-bromophenol: Dissolve 800 mg (8.1 mmol) of deuterated phenol in ethyl acetate (10 mL), and add 1.43 g (8.8 mmol) of bromine at 0°C. Stir for 2 h under nitrogen protection. After the reaction is completed, pour the reaction solution into water, extract with ethyl acetate, combine the organic phases, dry with anhydrous sodium sulfate, filter, and remove the solvent by distillation under reduced pressure. The crude product is used directly in the next step without purification. Embodiment 2

[0037] Synthesis of bromophenoxy-2,3,5,6-d4) (tert-butyl) dimethylsilane: At room temperature, 1.8 g (10.2 mmol) of crude deuterated p-bromophenol and 1.4 g (20.4 mmol) of imidazole were dissolved in dichloromethane (20 mL), and 1.85 g (12.2 mmol) of tert-butyldimethylchlorosilane was added in batches. Stir for 16 h under nitrogen protection. After the reaction, the reaction solution was poured into water, extracted with dichloromethane, the organic phases were combined, dried with anhydrous sodium sulfate, filtered, and the solvent was removed by reduced pressure distillation. The crude product was purified by Flash column chromatography to obtain 2.4 g (4-bromophenoxy-2,3,5,6-d4) (tert-butyl) dimethylsilane with a yield of 100.0%. Embodiment 3

[0038] Synthesis of 4-((tert-butyldimethylsilyl)oxy)benzaldehyde-2,3,5,6-d4: Dissolve 2.4g (8.2mmol) (4-bromophenoxy-2,3,5,6-d4) (tert-butyl)dimethylsilane in tetrahydrofuran (30mL), and add 7.8mL (12.4mmol) n-butyllithium (1.6M in hexane) dropwise at -78°C. Stir at -78°C for 1h under nitrogen protection, add 1.21g (16.5mmol) N,N-dimethylformamide dropwise, and stir at -78°C for 3h. After the reaction is completed, pour the reaction solution into an aqueous ammonium chloride solution, extract with ethyl acetate, combine the organic phases, dry with anhydrous sodium sulfate, filter, and distill under reduced pressure to remove the solvent. The crude product was purified by flash column chromatography to obtain 1.6 g of 4-((tert-butyldimethylsilyl)oxy)benzaldehyde-2,3,5,6-d4, with a yield of 85.6%; ESI-MS (m / z): 241 [M+H] + . Embodiment 4

[0039] Synthesis of (E)-4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-3-en-2-one: 1.4 g (5.8 mmol) of 4-((tert-butyldimethylsilyl)oxy)benzaldehyde-2,3,5,6-d4 and 2.8 g (8.75 mmol) of 1-triphenylphosphine-2-propanone were dissolved in toluene (20 mL), and the mixture was heated to 120 °C and stirred for 16 h under nitrogen protection. After the reaction, the solvent was removed by distillation under reduced pressure. The crude product was purified by flash column chromatography to obtain 1.2 g of (E)-4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-3-en-2-one with a yield of 72.3%; ESI-MS (m / z): 281 [M+H] + Embodiment 5

[0040] Synthesis of 4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-one: 1.2 g (15 mmol) (E)-4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-3-en-2-one was dissolved in ethyl acetate (15 mL) at room temperature, and 130 mg of palladium carbon was added in batches. After the addition was completed, the mixture was reacted at room temperature under hydrogen for 15 h. After the reaction was completed, the mixture was filtered and the solvent was removed by distillation under reduced pressure. The crude product was purified by flash column chromatography to obtain 870 mg of 4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-one with a yield of 71.9%; ESI-MS (m / z): 300 [M+NH4] + ,ESI-MS(m / z):305[M+Na] + . Embodiment 6

[0041] Synthesis of 4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4-N-(4-methoxybenzyl)butan-2-amine: 870 mg (3.1 mmol) of 4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-one, 510 mg (3.7 mmol) of p-methoxybenzylamine and catalyst acetic acid were dissolved in 1,2-dichloroethane (15 mL) at room temperature and reacted for 1 h at room temperature. 1.97 g (9.3 mmol) of sodium triacetoxyborohydride was added in batches under ice bath. After addition, the reaction was continued at room temperature for 16 h. After the reaction was completed, the reaction solution was poured into water, extracted with dichloroethane, the organic phases were combined, dried over anhydrous sodium sulfate, filtered and distilled under reduced pressure to remove the solvent. The crude product was purified by flash column chromatography to obtain 1 g of 4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4-N-(4-methoxybenzyl)butan-2-amine, with a yield of 80.6%; ESI-MS (m / z): 404 [M+H] + . Embodiment 7

[0042] Synthesis of 2-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl(4-methoxybenzyl)amino)-1-(4-hydroxyphenyl)ethan-1-one: 1.0 g (2.5 mmol) of 4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4-N-(4-methoxybenzyl)butan-2-amine and 1.1 g (7.4 mmol) of potassium carbonate were dissolved in acetonitrile (15 mL) at room temperature, and 640 mg (3.0 mmol) of 2-bromo-1-(4-hydroxyphenyl)ethan-1-one was added. After the addition was completed, the reaction mixture was reacted at 30°C for 16 hours. After the reaction was completed, the reaction solution was poured into an aqueous solution, extracted with dichloromethane, the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the solvent was removed by distillation under reduced pressure. The crude product was purified by flash column chromatography to give 670 mg of 2-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl(4-methoxybenzyl)amino)-1-(4-hydroxyphenyl)ethan-1-one, yield 50.4%; ESI-MS (m / z): 538 [M+H] + . Embodiment 8

[0043] Synthesis of 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-2-yl(4-methoxybenzyl)amino)-1-hydroxyethyl-1-d)phenol: 670 mg (1.2 mmol) of 2-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-2-yl(4-methoxybenzyl)amino)-1-(4-hydroxyphenyl)ethan-1-one was dissolved in anhydrous tetrahydrofuran (10 mL) at room temperature and stirred evenly. 58 mg (1.4 mmol) of lithium deuterated aluminum hydride was added in batches at 0°C and stirred at room temperature for 45 min. After the reaction was completed, the reaction solution was poured into an aqueous solution, extracted with ethyl acetate, the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and the solvent was removed by distillation under reduced pressure. The crude product was purified by flash column chromatography to give 510 mg of 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl(4-methoxybenzyl)amino)-1-hydroxyethyl-1-d)phenol, with a yield of 75.7%; ESI-MS (m / z): 541 [M+H] + . Embodiment 9

[0044] Synthesis of 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-2-yl)amino)-1-hydroxyethyl-1-d)phenol: 510 mg (0.9 mmol) of 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-2-yl(4-methoxybenzyl)amino)-1-hydroxyethyl-1-d)phenol was dissolved in ethyl acetate / methanol (6 mL / 1 mL) at room temperature, stirred evenly, palladium carbon was added in batches, and the reaction was stirred for 4 h under deuterium gas. After the reaction was completed, the reaction solution was directly filtered and the solvent was removed by vacuum distillation. The crude product was purified by flash column chromatography to obtain 310 mg of 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl)amino)-1-hydroxyethyl-1-d)phenol, with a yield of 78.1%; ESI-MS (m / z): 421 [M+H] + . Embodiment 10

[0045] Synthesis of 4-(3-((2-hydroxy-2-(4-hydroxyphenyl)ethyl-2-d)amino)butyl)benzene-2,3,5,6-d4-ol: 310 mg (0.7 mmol) of 4-((4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butyl-2-yl)amino)-1-hydroxyethyl-1-d)phenol and 212 mg (0.8 mmol) of tetrabutylammonium fluoride were dissolved in tetrahydrofuran (8 mL) at room temperature and stirred for 16 h. After the reaction, the reaction solution was distilled under reduced pressure to remove the solvent to obtain a crude product. The crude product was purified by flash column chromatography and reverse phase preparative chromatography to obtain 111 mg of the product ractopamine-d5, with a yield of 49.1%; ESI-MS (m / z): 307 [M+H] + . Embodiment 11

[0046] Synthesis of ractopamine hydrochloride-d5: 111 mg (0.7 mmol) of ractopamine-d5 was dissolved in water (4 mL) at room temperature, 2M HCl (aq.) was added dropwise, pH = 2-3, stirred for 0.5 h, and directly lyophilized to obtain 120 mg of the final product ractopamine hydrochloride-d5, with a yield of 96.8%. ESI-MS detection showed an isotopic abundance of 98.02%. 1 HNMR (400MHz, D2O) δ7.31-7.18 (m, 2H), 6.93-6.83 (m, 2H), 3.32-3.22 (m, 1H), 3.17 (d, J = 7.6Hz, 2H), 2.76-2.65 (m, 1H), 2.58-2.48 (m, 1H), 2.07-1.93 (m, 1H), 1.88-1.74 (m, 1H), 1.33 (t, J = 6.4, 5.2Hz, 3H); test spectrum as shown Figure 2 shown.

[0047] It should be noted that, in this article, relational terms such as first and second, etc. are only used to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Moreover, the terms "include", "comprise" or any other variants thereof are intended to cover non-exclusive inclusion, so that a process, method, article or device including a series of elements includes not only those elements, but also other elements not explicitly listed, or also includes elements inherent to such process, method, article or device.

[0048] Although the embodiments of the present application have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions and variations may be made to the embodiments without departing from the principles and spirit of the present application, and that the scope of the present application is defined by the appended claims and their equivalents.

Claims

1. A method for preparing deuterium-labeled ractopamine hydrochloride-d5, characterized in that: The following steps are involved: S1, dissolving deuterated phenol in a first solvent, adding bromine, and obtaining deuterated p-bromophenol through a substitution reaction; S2, dissolving deuterated p-bromophenol and an alkaline reagent in a second solvent, adding tert-butyldimethylsilyl chloride, and protecting the phenolic hydroxyl group under an inert atmosphere to obtain (4-bromophenoxy-2,3,5,6-d4) tert-butyldimethylsilane; S3, dissolving (4-bromophenoxy-2,3,5,6-d4) tert-butyldimethylsilane in a third solvent, adding n-butyl lithium dropwise, and reacting with N,N-dimethylformamide under an inert atmosphere to obtain 4-((tert-butyldimethylsilyl)oxy)benzaldehyde-2,3,5,6-d4; S4, dissolving 4-((tert-butyldimethylsilyl)oxy)benzaldehyde-2,3,5,6-d4 and 1-triphenylphosphine-2-propanone in a fourth solvent, toluene, and reacting the mixture under an inert atmosphere to obtain (E)-4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-3-ene-2-one; S5, dissolving (E)-4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)but-3-en-2-one in a fifth solvent, adding a heterogeneous catalyst, and reacting under a reducing atmosphere to obtain 4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-one; S6, adding 4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-one, p-methoxybenzylamine and acetic acid to the sixth solvent, adding sodium triacetoxyborohydride after mixing, and obtaining 4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4-N-(4-methoxybenzyl)butan-2-amine after reductive amination; S7, dissolving 4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4-N-(4-methoxybenzyl)butan-2-amine and potassium carbonate in the seventh solvent, and then adding 2-bromo-1-(4-hydroxyphenyl)ethan-1-one to react to obtain 2-((4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl)(4-methoxybenzyl)amino)-1-(4-hydroxyphenyl)ethan-1-one; S8, dissolving 2-((4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl)(4-methoxybenzyl)amino)-1-(4-hydroxyphenyl)ethan-1-one in the eighth solvent, adding deuterated lithium aluminum hydride, and reducing to obtain 4-(2-((4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl)(4-methoxybenzyl)amino)-1-hydroxyethyl-1-d)phenol; S9, dissolving 4-(2-((4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butyl-2-yl)(4-methoxybenzyl)amino)-1-hydroxyethyl-1-d)phenol in the ninth solvent, adding a heterogeneous catalyst, and hydrogenating under a deuterium atmosphere to obtain 4-(2-((4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butyl-2-yl)amino)-1-hydroxyethyl-1-d)phenol; S10, dissolving 4-(2-((4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butyl-2-yl)amino)-1-hydroxyethyl-1-d)phenol and tetrabutylammonium fluoride in a tenth solvent, and then deprotecting to obtain ractopamine-d5; S11. After dissolving ractopamine-d5 in the eleventh solvent, hydrochloric acid is added dropwise to adjust the pH of the solution to 2-3, and freeze-drying is performed to obtain the target product.

2. The method for preparing deuterium-labeled ractopamine hydrochloride-d5 according to claim 1, characterized in that: In step S1, the feed ratio of deuterated phenol to bromine is 1:1-1.

05.

3. The method for preparing deuterium-labeled ractopamine hydrochloride-d5 according to claim 1, characterized in that: In step S2, the alkaline reagent includes any one of imidazole, potassium carbonate, triethylamine, and N,N-diisopropylethylamine; and the second solvent includes any one of dichloromethane, 1,2-dichloroethane, and tetrahydrofuran.

4. The method for preparing deuterium-labeled ractopamine hydrochloride-d5 according to claim 1, characterized in that: The feed ratio of (4-bromophenoxy-2,3,5,6-d4) tert-butyldimethylsilane, n-butyl lithium and N,N-dimethylformamide in step S3 is 1:1.3-1.6:2-4.

5. The method for preparing deuterium-labeled ractopamine hydrochloride-d5 according to claim 1, characterized in that: The fifth solvent in step S5 includes any one of ethyl acetate, methanol, ethanol, and tetrahydrofuran.

6. The method for preparing deuterium-labeled ractopamine hydrochloride-d5 according to claim 1, characterized in that: In step S6, the feed ratio of 4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-one, p-methoxybenzylamine, and sodium triacetoxyborohydride is 1:1.1-1.5:2-3; the sixth solvent includes any one of dichloromethane and 1,2-dichloroethane.

7. The method for preparing deuterium-labeled ractopamine hydrochloride-d5 according to claim 1, characterized in that: In step S7, the feed ratio of 4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4-N-(4-methoxybenzyl)butan-2-amine, 2-bromo-1-(4-hydroxyphenyl)ethan-1-one, and potassium carbonate is 1:1.2-1.5:

3.

8. The method for preparing deuterium-labeled ractopamine hydrochloride-d5 according to claim 1, characterized in that: In step S8, the feed ratio of 2-((4-(4-((tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl)(4-methoxybenzyl)amino)-1-(4-hydroxyphenyl)ethan-1-one and deuterated lithium aluminum hydride is 1:1.

2.

9. The method for preparing deuterium-labeled ractopamine hydrochloride-d5 according to claim 1, characterized in that: The ninth solvent in step S9 includes one or a mixture of ethyl acetate, methanol, ethanol, and tetrahydrofuran.

10. The method for preparing deuterium-labeled ractopamine hydrochloride-d5 according to claim 1, characterized in that: In step S10, the feed ratio of 4-(2-((4-(4-(tert-butyldimethylsilyl)oxy)phenyl-2,3,5,6-d4)butan-2-yl)amino)-1-hydroxyethyl-1-d)phenol and tetrabutylammonium fluoride is 1:1.1-2.

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