Synthetic method of plerixafor impurity

Abstract

The invention relates to a synthetic method of a plerixafor impurity. According to the method disclosed by the invention, a plerixafor impurity A, namely N1-(2-((4-(1, 4, 8, 11-tetraazacyclotetradecane-1-yl) methyl) benzyl) amino) ethyl)-N3-(2-aminoethyl) propane-1, 3-diamine, and a plerixafor impurity B, namely N1-(4-(1, 4, 8, 11-tetraazacyclotetradecane-1-yl) methyl) benzyl)-N1-(2-(3-aminopropyl) amino) ethyl) propane-1, 3-diamine can be prepared. The invention relates to synthesis of a 1, 3-diamine and plerixafor impurity C: N1-(4-(1, 4, 8, 11-tetraazacyclotetradecane-1-yl) methyl) benzyl)-N1, N3-bis (2-aminoethyl) propane-1, 3-diamine and plerixafor impurity. Four brand-new potential impurities of plerixafor are obtained through a simple synthesis method, and the method has important significance on synthesis of high-quality plerixafor.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical technology, and relates to plerixafor, and also relates to a method for preparing plerixafor impurities. Furthermore, the impurities prepared by the present invention are potential ring-opening impurities of plerixafor, which can be used for quality monitoring of plerixafor API or its preparation, for example, as an impurity reference substance when detecting potential ring-opening impurities in plerixafor. Background Art

[0002] Plerixafor is a hematopoietic stem cell mobilizer, which was publicly launched by GENZYME. Its English name is Plerixafor. The commercial product is an injection solution, and its trade name is Mozobil. The English chemical name of Plerixafor is 1,4-Bis((1,4,8,11-tetraazacyclotetradecan-1-yl)methyl)benzene, and its Chinese chemical name is 1,4-bis((1,4,8,11-tetraazacyclotetradecan-1-yl)methyl)benzene or 1,1'-[1,4-phenylene (methylene)]-di-1,4,8,11-tetraazacyclotetradecan, molecular formula: C28H54N8, molecular weight 502.79, and chemical structure formula is:

[0003]

[0004] Plerixafor API is a white to off-white crystalline solid that is hygroscopic with a partition coefficient of <0.1 between 1-octanol and pH 7 aqueous buffer.

[0005] As a hematopoietic stem cell activator and CXCR4 chemokine antagonist, Mozobil, an injection of plerixafor, was approved by the U.S. Food and Drug Administration for autologous transplantation in patients with non-Hodgkin's lymphoma (NHL) and multiple myeloma (MM) in December 2008. Plerixafor can be used in combination with granulocyte colony-stimulating factor (G-CSF, filgrastim) to block the binding of CXCR4 to SDF-1α, drive the release of CD34+ cells into the peripheral blood, and quickly and effectively increase the number of hematopoietic stem cells in the blood circulation of NHL and MM patients.

[0006] The typical synthetic reaction route of plerixafor is as follows:

[0007]

[0008] Generally speaking, in the above reaction route, 1,4,8,11-tetraazacyclotetradecane (compound 2) is used as the starting material, and acylated with ethyl trifluoroacetate to obtain 1,1',1"-(1,4,8,11-tetraazacyclotetradecane-1,4,8-triyl)tris(2,2,2-trifluoroethanone) (compound 3), and then compound 3 is reacted with 1,4-dichlorobenzyl under the alkaline condition of potassium carbonate and catalyzed by potassium iodide to generate hexa-trifluoroacetyl plerixafor (compound 4), and finally hydrolyzed under the alkaline condition of sodium hydroxide to obtain the target product compound 1, i.e. plerixafor. It is reported that the total yield of this reaction route is about 44% based on compound 2, and it is believed that the purity of the product can reach more than 99%.

[0009] In the process of drug synthesis, impurities need to be controlled, which is very important for the quality control of the final product. Therefore, the synthesis research of plerixafor impurities is of great significance. In order to effectively control the quality of plerixafor API, plerixafor impurities need to be studied and reasonably controlled.

[0010] The main impurities of plerixafor are process impurities. However, during the storage of raw materials and preparations, oxidized and open-ring impurities may be generated. The existing technology does not have sufficient research on the oxidized open-ring impurities of plerixafor. The generation pathway of open-ring impurities is that plerixafor undergoes dehydrogenation and oxidation under aerobic conditions to produce oxidized impurities A', B', C', D', which are then hydrolyzed to produce open-ring impurities A, B, C, D. The schematic generation principle of oxidized hydrolysis open-ring impurities of plerixafor is as follows:

[0011]

[0012]

[0013] In the analytical method for monitoring the above four open-ring impurities A, B, C, and D in the raw material or preparation of plerixafor, it is necessary to use these impurity reference substances. However, if these open-ring impurities are prepared by the above oxidation and hydrolysis routes, it is obviously difficult to control, and multiple impurities may be formed, and separation is difficult and the purity is low. In addition, although these open-ring impurities can be purchased through commercial channels, for example, from https: / / www.qcchemical.com / index.php / Index / products?info=Plerixafor, their correct synthesis will still be of technical value for the preparation of plerixafor.

[0014] Therefore, those skilled in the art are eager to provide a method for preparing the ring-opening impurities of plerixafor, especially impurities A, B, C, and D. Summary of the invention

[0015] The object of the present invention is to provide a synthetic method for preparing plerixafor ring-opening impurities, especially plerixafor impurity A, plerixafor impurity B, plerixafor impurity C, and plerixafor impurity D. The inventors have found that the above four ring-opening impurities can be easily obtained by the method of the present invention, and the present invention is completed based on these findings.

[0016] On the one hand, the present invention provides a method for preparing plerixafor ring-opening impurities, namely impurity A, impurity B, impurity C and impurity D, and the reaction processes thereof are respectively as follows:

[0017]

[0018] The above letters representing each structural compound can directly represent the compound in the diagram in the context of the present invention.

[0019] The method according to the first aspect of the present invention comprises the following steps:

[0020] (1) Preparation of impurity A

[0021] ① Impurity A uses F as a raw material to undergo an amidation reaction with ethyl trifluoroacetate to generate G, and then G undergoes an alkylation reaction with the raw material E to generate N-(2-aminoethyl)-2,2,2-trifluoro-N-(3-(2,2,2-trifluoro-N-(2-((4-((4,8,11-tri(2,2,2-trifluoroacetyl)-1,4,8,11-tetraazacyclotetradec-1-yl)methyl)benzyl)amino)ethyl)acetamido)acetamido)propyl)acetamide)acetamide (H), and then hydrolyzes and removes the protecting group trifluoroacetyl to obtain impurity A;

[0022] ② The solvent used in the preparation method of G is selected from methanol, ethanol, and isopropanol. In the present invention, methanol is preferably used as the solvent; the amount of ethyl trifluoroacetate is 2 to 3 equivalents; the reaction temperature is 0 to 30°C;

[0023] ③ The acidifying agent used in the preparation method of H is selected from triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably potassium carbonate;

[0024] ④ In the preparation method of A, the deprotection reaction hydrolysis reagent is selected from sodium hydroxide, potassium carbonate, potassium hydroxide, preferably sodium hydroxide;

[0025] (2) Preparation of impurity B

[0026] ① Impurity B is subjected to acylation reaction with diethyl oxalate using I as raw material, and then to acylation reaction with p-toluenesulfonyl chloride, and then the protecting group is removed under acidic conditions to obtain L; L is subjected to alkylation reaction with raw material E to generate 2,2,2-trifluoro-N-(3-((2-((3-(2,2,2-trifluoroacetylamino)propyl)(4-((4,8,11-tri(2,2,2-trifluoroacetyl)-1,4,8,11-tetraazacyclotetradec-1-yl)methyl)benzyl)amino)ethyl)amino)propyl)acetamide (M), and then the protecting group is removed by hydrolysis to obtain impurity B;

[0027] ② In the preparation method of L, sodium hydroxide, dilute sulfuric acid, or dilute hydrochloric acid can be used for the deprotection reaction. The present invention preferably uses dilute hydrochloric acid as the deprotection reagent;

[0028] ③ In the preparation method of B, the alkylation reaction acidifying agent can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably N,N-diisopropylethylamine as the alkylation reaction acidifying agent; the deprotection reaction hydrolysis reagent can use sodium hydroxide, potassium hydroxide, concentrated sulfuric acid, preferably concentrated sulfuric acid;

[0029] (3) Preparation of impurity C

[0030] ① Impurity C uses F as raw material to undergo alkylation reaction with N,N-dimethylformamide dimethyl acetal, and then undergoes acylation reaction with di-tert-butyl dicarbonate, and then removes the tert-butyloxycarbonyl group under alkaline conditions to obtain P; P undergoes alkylation reaction with raw material E to generate Q, and then hydrolyzes and removes the protecting group to obtain impurity C;

[0031] ② In the preparation method of N, N-dimethylformamide dimethyl acetal, the amount of N, N-dimethylformamide dimethyl acetal is 1 to 1.2 equivalents, and the temperature is 50 to 70°C;

[0032] ③ In the preparation method of C, the alkylation reaction acid agent can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably N,N-diisopropylethylamine as the alkylation reaction acid agent; the deprotection reaction hydrolysis reagent can use sodium hydroxide, potassium hydroxide, concentrated sulfuric acid, hydrochloric acid, potassium carbonate, preferably hydrochloric acid is used to remove the tert-butyloxycarbonyl group, and then potassium carbonate is used to remove the trifluoroacetyl group;

[0033] (3) Preparation of impurity D

[0034] ① Impurity D is reacted with I as a raw material to undergo an acylation reaction with di-tert-butyl dicarbonate to obtain S; S is reacted with the raw material E to undergo an alkylation reaction to produce tert-butyl (2-((3-aminopropyl)(tert-butoxycarbonyl)amino)ethyl)(3-((4-((4-((4,8,11-tris(2,2,2-trifluoroacetyl)-1,4,8,11-tetraazacyclotetradecyl-1-yl)methyl)benzyl)amino)propyl)amino)aminobenzoate (T), and then the protecting group is removed by hydrolysis to obtain impurity D;

[0035] ② In the preparation method of S, the amount of di-tert-butyl dicarbonate used ranges from 2 to 3 equivalents, preferably 2.5 equivalents;

[0036] ③ In the preparation method of impurity D, the alkylation reaction acidifying agent can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably N,N-diisopropylethylamine as the alkylation reaction acidifying agent; the deprotection reaction hydrolysis reagent can use sodium hydroxide, potassium hydroxide, concentrated sulfuric acid, hydrochloric acid, potassium carbonate, preferably using hydrochloric acid to remove the tert-butyloxycarbonyl group, and then using potassium carbonate to remove the trifluoroacetyl group.

[0037] The method according to the first aspect of the present invention is characterized in that:

[0038] 11-(4-chloromethyl)benzyl-1,4,8-tris(trifluoroacetyl)-1,4,8,11-tetraazacyclotetradecane (E) and

[0039] N,N'-(Propane-1,3-diyl)bis(N-(2-aminoethyl)-2,2,2-trifluoroacetamide)(G)

[0040] Alkylation reaction occurs under weak alkaline conditions, and then the protective group trifluoroacetyl is removed to obtain

[0041] N1-(2-((4-(1,4,8,11-tetraazacyclotetradec-1-yl)methyl)benzyl)amino)ethyl)-N3-(2-aminoethyl)propane-1,3-diamine is plerixafor impurity A;

[0042] 11-(4-chloromethyl)benzyl-1,4,8-tris(trifluoroacetyl)-1,4,8,11-tetraazacyclotetradecane (E) and

[0043] N,N'-((Ethane-1,2-diylbis(azadiyl))bis(propane-3,1-diyl))bis(4-methylbenzenesulfonamide)(L)

[0044] Alkylation reaction occurs under weak alkaline conditions, and then the protective group trifluoroacetyl is removed to obtain

[0045] N1-(4-(1,4,8,11-tetraazacyclotetradec-1-yl)methyl)benzyl)-N1-(2-(3-aminopropyl)amino)ethyl)propane-1,3-diamine is plerixafor impurity B;

[0046] 11-(4-chloromethyl)benzyl-1,4,8-tris(trifluoroacetyl)-1,4,8,11-tetraazacyclotetradecane (E) and

[0047] Di-tert-butyl ((propane-1,3-diylbis(azadiyl)))bis(ethane-2,1-diyl)) dicarbamate (P)

[0048] Alkylation reaction occurs under weak alkaline conditions, and then the protecting groups trifluoroacetyl and tert-butyloxycarbonyl are removed to obtain

[0049] N1-(4-(1,4,8,11-tetraazacyclotetradecane-1-yl)methyl)benzyl)-N1,N3-bis(2-aminoethyl)propane-1,3-diamine is plerixafor impurity C;

[0050] 11-(4-chloromethyl)benzyl-1,4,8-tris(trifluoroacetyl)-1,4,8,11-tetraazacyclotetradecane (E) and

[0051] Di-tert-butylethane-1,2-diylbis((3-aminopropyl)carbamate)(S)

[0052] Alkylation reaction occurs under weak alkaline conditions, and then the protecting groups trifluoroacetyl and tert-butyloxycarbonyl are removed to obtain

[0053] N1-(4-(1,4,8,11-tetraazacyclotetradec-1-yl)methyl)benzyl)-N3-2-((3-aminopropyl)amino)ethyl)propane-1,3-diamine is plerixafor impurity D.

[0054] According to the method of the first aspect of the present invention, the preparation method of G is: 1 , N 1 '-(Propane-1,3-diyl)bis(ethane-1,2-diamine) (F) is used as a raw material and reacts with ethyl trifluoroacetate at low temperature under alkaline conditions to undergo an amidation reaction to generate G.

[0055] According to the method of the first aspect of the present invention, when preparing G, the alkylation reaction acidifying agent can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably potassium carbonate as the alkylation reaction acidifying agent; the deprotection reaction hydrolysis reagent can use sodium hydroxide, potassium carbonate, potassium hydroxide, preferably sodium hydroxide.

[0056] According to the method of the first aspect of the present invention, when preparing G, the amidation reaction acidifying agent can use triethylamine, N,N-diisopropylethylamine, pyridine, preferably N,N-diisopropylethylamine as the acidifying agent for the amidation reaction; the reaction temperature is controlled in the range of 0-30°C.

[0057] According to the method of the first aspect of the present invention, the preparation method of L is: 1 ,N 1 '-(Ethane-1,2-diyl)bis(propane-1,3-diamine) (I) is used as a raw material, undergoes an acylation reaction with diethyl oxalate, and then undergoes an acylation reaction with p-toluenesulfonyl chloride, and then the protecting group is removed under acidic conditions to obtain L.

[0058] According to the method of the first aspect of the present invention, when preparing L, the alkylation reaction acidifying agent can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably N,N-diisopropylethylamine as the alkylation reaction acidifying agent; the deprotection reaction hydrolysis reagent can use sodium hydroxide, potassium hydroxide, concentrated sulfuric acid, and concentrated sulfuric acid is preferred in the present invention.

[0059] According to the method of the first aspect of the present invention, when preparing L, the deprotection reaction can use sodium hydroxide, dilute sulfuric acid, or dilute hydrochloric acid, and dilute hydrochloric acid is preferably used as the deprotection agent.

[0060] According to the method of the first aspect of the present invention, the preparation method of P is: using F as a raw material, undergoing an alkylation reaction with N,N-dimethylformamide dimethyl acetal, then undergoing an acylation reaction with di-tert-butyl dicarbonate, and then removing the tert-butyloxycarbonyl group under alkaline conditions to obtain P.

[0061] According to the method of the first aspect of the present invention, when preparing P, the alkylation reaction acidifying agent can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably N,N-diisopropylethylamine as the alkylation reaction acidifying agent; the deprotection reaction hydrolysis reagent can use sodium hydroxide, potassium hydroxide, concentrated sulfuric acid, hydrochloric acid, potassium carbonate, preferably hydrochloric acid is used to remove the tert-butyloxycarbonyl group, and then potassium carbonate is used to remove the trifluoroacetyl group.

[0062] According to the method of the first aspect of the present invention, the preparation method of S is: using I as a raw material, and subjecting it to an acylation reaction with di-tert-butyl dicarbonate to obtain S.

[0063] According to the method of the first aspect of the present invention, when preparing S, the alkylation reaction acidifying agent can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably N,N-diisopropylethylamine as the alkylation reaction acidifying agent; the deprotection reaction hydrolysis reagent can use sodium hydroxide, potassium hydroxide, concentrated sulfuric acid, hydrochloric acid, potassium carbonate, preferably hydrochloric acid is used to remove the tert-butyloxycarbonyl group, and then potassium carbonate is used to remove the trifluoroacetyl group.

[0064] According to the method of the first aspect of the present invention, when preparing S, the amount of di-tert-butyl dicarbonate used ranges from 2 to 3 equivalents, preferably 2.5 equivalents.

[0065] According to the method of the first aspect of the present invention, impurity A is prepared by the following steps:

[0066] (1) Preparation of G: At room temperature, 25 g of methanol was added to a 100 ml reaction bottle, and then 10 g of the compound of formula F and 2 g of N,N-diisopropylethylamine were added to the reaction bottle in sequence. Stirring and cooling were started, and the temperature was lowered to 0-10°C (5°C). 22.2 g of ethyl trifluoroacetate was added dropwise to the reaction system while controlling the temperature at 0-10°C (5°C). After the addition was completed, the temperature was raised to 25°C, and stirred at 25°C for 4 h. Then, 25 g of acetonitrile was added to the reaction system, and the mixture was concentrated under reduced pressure at 45°C. 25 g of acetonitrile was added to the reaction system again, and the mixture was concentrated under reduced pressure again to obtain a light yellow viscous oily substance, which is the compound of formula G;

[0067] (2) Preparation of H: At room temperature, add 120 g of acetonitrile to a reaction flask containing a compound of formula G, start stirring, add 35 g of a compound of formula E, 4.2 g of potassium carbonate, and 5.1 g of potassium iodide to the reaction flask in sequence, raise the temperature to 80° C. and allow the reaction to proceed for 5 to 7 hours (actually 6 hours), then cool to 20 to 30° C. (25° C., filter, take the filtrate, and concentrate to obtain a viscous oil; add 50 g of anhydrous ethanol, raise the temperature to 70 to 75° C., and keep stirring for 30 minutes. The oily substance is completely dissolved, and then cooled to 20-30°C (25°C) to obtain an ethanol solution, which is added dropwise to 100 ml of water, and the temperature is controlled at 20-30°C (25°C) to stir and crystallize for 1 hour, filtered, and dried to obtain an off-white solid; 230 g of ethyl acetate is added to the yellow solid, stirred and heated to reflux, kept warm and stirred for 30 minutes, slowly cooled to 30±2°C, kept warm and stirred for 10 hours, filtered, and dried to obtain an off-white solid as a compound of formula H;

[0068] (3) Preparation of A: At room temperature, 200 g of isopropanol / acetone mixture was added to a 500 ml reaction bottle, stirring was started, 10 g of sodium hydroxide and 20 g of water were added in sequence, stirred to dissolve, 38.3 g of compound H was added to the reaction bottle, and under nitrogen protection, the reaction solution was heated to 65°C and kept warm for 4 hours, and the solvent was removed by concentration under reduced pressure, 60 g of methanol was added, stirred to dissolve, 400 g of purified water was added dropwise, the temperature was lowered to 0°C with stirring, and the mixture was kept warm for 5 hours to allow precipitation, which was filtered and dried with air at 50°C to obtain an off-white solid, which was the open-loop impurity A.

[0069] According to the method of the present invention, the isopropanol / acetone mixed solution used in the preparation of A is a mixed solution with a volume ratio of isopropanol to acetone of 2 to 8:1, especially a mixed solution with a volume ratio of isopropanol to acetone of 3 to 6:1, and especially a mixed solution with a volume ratio of isopropanol to acetone of 3 to 5:1.

[0070] The method of the present invention exhibits excellent effects. For example, when preparing impurity A, it is unexpectedly found that the use of a specific reaction solvent not only results in high product purity but also high yield. BRIEF DESCRIPTION OF THE DRAWINGS

[0071] Figure 1 : HPLC spectrum of impurity A of plerixafor.

[0072] Figure 2 : Mass spectrum of impurity A of plerixafor.

[0073] Figure 3 : H NMR spectrum of impurity A of plerixafor.

[0074] Figure 4 : HPLC spectrum of impurity A of plerixafor.

[0075] Figure 5 : HPLC spectrum of impurity B of plerixafor.

[0076] Figure 6 : Mass spectrum of impurity B of plerixafor.

[0077] Figure 7 : H NMR spectrum of plerixafor impurity B.

[0078] Figure 8 : HPLC spectrum of plerixafor impurity C.

[0079] Fig. 9 : Mass spectrum of impurity C of plerixafor.

[0080] Fig.10 : H NMR spectrum of plerixafor impurity C.

[0081] Fig.11 : HPLC spectrum of impurity D of plerixafor.

[0082] Fig.12 : Mass spectrum of impurity D of plerixafor.

[0083] Fig.13 : H NMR spectrum of impurity D of plerixafor. DETAILED DESCRIPTION

[0084] The present invention is further illustrated by the following examples, but the present invention is not limited thereto.

[0085] The compound of formula E used as the starting material in the context of the present invention can be easily prepared by known methods, for example, referring to the method described in Diana Brickute's document (Diana Brickute, et al, Development and Evaluation of an 18F-Radiolabeled Monocyclam Derivative for Imaging CXCR4 Expression, Mol. Pharmaceutics 2019, 16, 2106-2117), and replacing α, α′-dibromo-p-xylene with α, α′-dichloro-p-xylene to obtain the compound of formula E (C24H28N4O3F9Cl, Mr626.9).

[0086] The compound of formula F (N, N'-bis(2-aminoethyl)-1,3-propylenediamine, CAS 4741-99-5, Mr 160.3) and the compound of formula I (N, N'-bis(3-aminopropyl)-1,2-ethylenediamine, CAS 10563-26-5, Mr 174.3) used as starting materials in the context of the present invention can be easily purchased from commercial sources, for example, from Qiyue Biological or Kaiyin Chemical. Other conventional reagents are also easily purchased from commercial sources.

[0087] Example 1: Preparation of impurity A

[0088] This example basically prepares the ring-opening impurity A of plerixafor according to the following process route:

[0089]

[0090] (1) Preparation of G:

[0091] At room temperature, 25 g of methanol was added to a 100 ml reaction bottle, and then 10 g (62.4 mmol) of the compound of formula F and 2 g of N, N-diisopropylethylamine (DIPEA) were added to the reaction bottle in sequence. Stirring and refrigeration were started, and the temperature was lowered to 0-10°C (5°C). 22.2 g of ethyl trifluoroacetate was added dropwise to the reaction system at a temperature of 0-10°C (5°C). After the addition was completed, the temperature was raised to 25°C, and the mixture was stirred at 25°C for 4 h. Then 25 g of acetonitrile was added to the reaction system, and the mixture was concentrated under reduced pressure at 45°C. 25 g of acetonitrile was added to the reaction system again, and the mixture was concentrated under reduced pressure again to obtain a light yellow viscous oil, which is a compound of formula G.

[0092] (2) Preparation of H:

[0093] At room temperature, 120 g of acetonitrile was added to the reaction bottle containing the compound of formula G, stirring was started, 35 g (55.8 mmol) of the compound of formula E, 4.2 g of potassium carbonate, and 5.1 g of potassium iodide were added to the reaction bottle in sequence, the temperature was raised to 80° C. to react for 5 to 7 hours (actually 6 hours), the temperature was lowered to 20 to 30° C. (25° C., filtered, the filtrate was collected, and concentrated to obtain a viscous oil;

[0094] Add 50g of anhydrous ethanol, raise the temperature to 70-75°C, keep warm and stir for 30 minutes until the oil is completely dissolved, then cool to 20-30°C (25°C) to obtain an ethanol solution, add it dropwise to 100ml of water, control the temperature at 20-30°C (25°C) to stir and crystallize for 1 hour, filter, and dry to obtain an off-yellow solid; add 230g of ethyl acetate to the off-yellow solid, stir and heat to reflux, keep warm and stir for 30 minutes, slowly cool to 30±2°C, keep warm and stir for 10h, filter, and dry to obtain an off-white solid as a compound of formula H 38.3g (40.6mmol, C35H45O5N8F15, Mr942.8), (calculated from E) yield 72.8%.

[0095] (3) Preparation A:

[0096] At room temperature, 200 g of a mixed solution of isopropanol / acetone (4 / 1, v / v) was added to a 500 ml reaction flask, and stirring was started. 10 g of sodium hydroxide and 20 g of water were added in sequence, and the mixture was stirred to dissolve. 38.3 g (40.62 mmol) of the compound of formula H was added to the reaction flask. Under nitrogen protection, the reaction solution was heated to 65°C and kept warm for 4 hours. The solvent was removed by concentration under reduced pressure, 60 g of methanol was added, and the mixture was stirred to dissolve. 400 g of purified water was added dropwise, and the mixture was cooled to 0°C with stirring. The mixture was kept warm for 5 hours to precipitate. The mixture was filtered and dried with air at 50°C to obtain 16.5 g of an off-white solid, which was an open-loop impurity A (35.66 mmol, C25H50N8, Mr462.7) with a yield of 87.8%.

[0097] The structural identification data of open-ring impurity A are as follows: H-NMR (300 MHz, D2O) δ2.04~2.17 (m, 6H, 3CH2), 3.12~3.47 (m, 28H, 14CH2), 4.19 (s, 2H, CH2), 4.30 (s, 2H, CH2), 7.47~7.55 (m, 4H, 4ArH); ESI-MS (m / z): 463.5 [M+H]+.

[0098] The HPLC spectrum of the plerixafor ring-opening impurity A obtained in Example 1 is shown in Figure 1 , showing that its normalized purity can reach more than 99% (99.83%), and the mass spectrum and nuclear magnetic resonance hydrogen spectrum of the open-ring impurity A are shown in Figure 2 and Figure 3 These results indicate that the method of Example 1 can obtain high-purity open-ring impurity A in a fairly high yield.

[0099] The method for determining the HPLC normalized purity of various materials, especially four open-ring impurities, of the present invention is as follows:

[0100] Test solution: Take appropriate amount of various materials, especially four open-ring impurities, dissolve them in solvent and dilute quantitatively to make a test solution containing about 1 mg per ml; mobile phase: 0.005 mol / L sodium heptane sulfonate solution with pH adjusted to 2 with perchloric acid as mobile phase A, acetonitrile as mobile phase B; use octadecylsilane bonded silica gel as filler chromatographic column (Agilent ZORBAX SB-C18, 4.6×250 mm, 5 μm), use mobile phase A and mobile phase B for gradient elution according to the elution program shown in the following table:

[0101] Time / min Mobile phase A / % Mobile phase B / % 0 70 30 28 50 50 42 30 70 42.01 70 30 55 70 30

[0102] The flow rate was 1.0 ml per minute, the detection wavelength was 210 nm, the column temperature was 30 °C, and the injection volume was 20 μl;

[0103] Inject the sample solution to be tested into the liquid chromatograph, record the chromatogram, and calculate the purity by area normalization. In the determination, the gradient elution program can be appropriately adjusted as needed to meet the determination requirements of different materials, such as Figure 8 The mobile phase used was isocratic elution with mobile phase A / mobile phase B = 60 / 40.

[0104] In some preliminary exploratory tests conducted by the inventors, it was found that when preparing the open-ring impurity A in step (3) of Example 1, when isopropanol was used as a solvent, excellent purity could be obtained but the yield was low, while when an isopropanol / acetone mixed solvent was used, not only excellent purity could be obtained but also the yield was significantly improved, while these effects could not be obtained without using this solvent combination. These exploratory tests were provided by the experimental operation of step (3) of reference example 1 in the following example 1a, and the compound of formula H used was prepared by the methods of steps (1) and (2).

[0105] Example 1a-1: At room temperature, add 200 g of isopropanol to a 500 ml reaction bottle, start stirring, add 10 g of sodium hydroxide and 20 g of water in sequence, stir to dissolve, add 38.3 g (40.62 mmol) of compound H to the reaction bottle, and under nitrogen protection, heat the reaction solution to 65°C and keep it warm for 4 hours. Remove the solvent by concentration under reduced pressure, add 60 g of methanol, stir to dissolve, add 400 g of purified water dropwise, stir and cool to 0°C, keep it warm for 5 hours to precipitate, filter, and dry at 50°C to obtain 13.1 g of an off-white solid, which is an open-ring impurity A (Mr462.7), with a yield of 69.7% and a HPLC normalized purity of 99.24%.

[0106] Example 1a-2: At room temperature, 220 g of isopropanol was added to a 500 ml reaction bottle, stirring was started, 10.5 g of sodium hydroxide and 22 g of water were added in sequence, the mixture was stirred to dissolve, 38.3 g (40.62 mmol) of the compound of formula H was added to the reaction bottle, the reaction solution was heated to 65°C under nitrogen protection and kept warm for 3.5 hours, the solvent was removed by concentration under reduced pressure, 60 g of methanol was added, the mixture was stirred to dissolve, 400 g of purified water was added dropwise, the mixture was cooled to 0°C with stirring, the mixture was kept warm for 5 hours to allow precipitation, the mixture was filtered, and the mixture was dried at 50°C to obtain 12.8 g of an off-white solid, which was an open-ring impurity A (Mr462.7), with a yield of 68.1% and a HPLC normalized purity of 99.36%.

[0107] Example 1a-3: At room temperature, 180 g of isopropanol was added to a 500 ml reaction bottle, stirring was started, 9.5 g of sodium hydroxide and 18 g of water were added in sequence, the mixture was stirred to dissolve, 38.3 g (40.62 mmol) of the compound of formula H was added to the reaction bottle, and the reaction solution was heated to 65°C under nitrogen protection and kept warm for 5 hours. The solvent was removed by concentration under reduced pressure, 60 g of methanol was added, stirred to dissolve, 400 g of purified water was added dropwise, the mixture was cooled to 0°C with stirring, the mixture was kept warm for 5 hours to allow precipitation, filtered, and dried at 50°C to obtain 13.3 g of an off-white solid, which was an open-ring impurity A (Mr462.7), with a yield of 70.8% and a HPLC normalized purity of 99.68%.

[0108] Example 1a-4: Referring to Example 1a-1, the only difference is that the isopropanol is replaced by an equal amount of ethanol, and 38.3 g (40.62 mmol) of the compound of formula H is added to obtain 13.6 g of the open-ring impurity A (Mr462.7), with a yield of 72.4% and a HPLC normalized purity of 95.18%; Referring to Example 1a-1, the only difference is that the isopropanol is replaced by an equal amount of ethanol / acetone (4 / 1, v / v) mixed solution, and 38.3 g (40.62 mmol) of the compound of formula H is added to obtain 14.5 g of the open-ring impurity A (Mr462.7), with a yield of 77.1% and a HPLC normalized purity of 94.53%. Its typical HPLC graph is as follows Figure 4 shown.

[0109] Example 1a-5: The same method as Example 1a-1 was used, except that the isopropanol was replaced by an equal amount of methanol, and 38.3 g (40.62 mmol) of the compound of formula H was added to obtain 14.0 g of the open-ring impurity A (Mr 462.7), with a yield of 74.5% and a HPLC normalized purity of 93.73%; The same method as Example 1a-1 was used, except that the isopropanol was replaced by an equal amount of methanol / acetone (4 / 1, v / v) mixture, and 38.3 g (40.62 mmol) of the compound of formula H was added to obtain 14.9 g of the open-ring impurity A (Mr 462.7), with a yield of 79.3% and a HPLC normalized purity of 94.25%.

[0110] Example 1a-6: The same method as Example 1a-1 was used, except that the isopropanol was replaced by an equal amount of propylene glycol. 38.3 g (40.62 mmol) of the compound of formula H was added to obtain 12.1 g of the open-ring impurity A (Mr 462.7), with a yield of 64.4% and a HPLC normalized purity of 91.42%. The same method as Example 1a-1 was used, except that the isopropanol was replaced by an equal amount of propylene glycol / acetone (4 / 1, v / v) mixture. 38.3 g (40.62 mmol) of the compound of formula H was added to obtain 12.7 g of the open-ring impurity A (Mr 462.7), with a yield of 67.6% and a HPLC normalized purity of 90.63%.

[0111] Example 1a-7: The same method as Example 1 was used, except that the isopropanol / acetone mixture was replaced by an equal amount of isopropanol / acetone (3 / 1, v / v) mixture, and 38.3 g (40.62 mmol) of the compound of formula H was added to obtain 16.9 g of the open-ring impurity A (Mr 462.7), with a yield of 89.9% and a HPLC normalized purity of 99.68%. The same method as Example 1 was used, except that the isopropanol / acetone mixture was replaced by an equal amount of isopropanol / acetone (2 / 1, v / v) mixture, and the compound of formula H was added. The reaction mixture was added with 38.3 g (40.62 mmol) of the compound to obtain 15.8 g of the open-ring impurity A (Mr 462.7), with a yield of 84.1% and a HPLC normalized purity of 99.46%. The reaction mixture was carried out with reference to Example 1, except that the isopropanol / acetone mixture was replaced with an equal amount of isopropanol / acetone (1 / 1, v / v) mixture. The compound of formula H was added with 38.3 g (40.62 mmol) of the compound to obtain 15.2 g of the open-ring impurity A (Mr 462.7), with a yield of 80.9% and a HPLC normalized purity of 99. 53%; refer to Example 1, except that the isopropanol / acetone mixture was replaced by an equal amount of isopropanol / acetone (5 / 1, v / v) mixture, 38.3 g (40.62 mmol) of the compound of formula H was added to obtain 16.7 g of the open-ring impurity A (Mr 462.7), with a yield of 88.9% and a HPLC normalized purity of 99.61%; refer to Example 1, except that the isopropanol / acetone mixture was replaced by an equal amount of isopropanol / acetone (6 / 1, v / v) mixture, 38.3 g (40.62 mmol) of the compound of formula H was added to obtain 16.7 g of the open-ring impurity A (Mr 462.7), with a yield of 88.9% and a HPLC normalized purity of 99.61%. .3g (40.62mmol) was added to obtain 15.5g of open-ring impurity A (Mr462.7), with a yield of 82.5% and a HPLC normalized purity of 99.31%; refer to Example 1, except that the isopropanol / acetone mixture was replaced with an equal amount of isopropanol / acetone (8 / 1, v / v) mixture, and 38.3g (40.62mmol) of compound H was added to obtain 14.3g of open-ring impurity A (Mr462.7), with a yield of 76.1% and a HPLC normalized purity of 99.43%. These results show that when isopropanol is replaced with other alcohols, not only the yield is significantly reduced, but also the purity is reduced or significantly reduced; and when a mixture of isopropanol: acetone = 3 to 5:1 is used, the yield and purity of the final product are significantly higher. Using isopropanol as a solvent helps to obtain a high-purity product, and adding an appropriate amount of acetone can significantly increase the yield of the product.

[0112] Example 2: Preparation of impurity B

[0113] This example basically prepares the ring-opening impurity B of plerixafor according to the following process route:

[0114]

[0115] (1) Preparation of 1,4-bis(3-aminopropyl)piperazine-2,3-dione (J)

[0116] To a 250 ml three-necked flask, add 10 g of the compound of formula I, 10.06 g of diethyl oxalate and 100 ml of ethanol, stir and heat to 80 ° C to react for 20 h. The raw materials are basically reacted. Stop the reaction, cool to room temperature, and concentrate to obtain a light yellow solid. Recrystallize with 150 ml (acetone: ethanol = 20:1) solution to obtain 11.05 g of a white solid as the compound of formula J.

[0117] (2) Preparation of N,N'-((2,3-dioxopiperazine-1,4-diyl)bis(propane-3,1-diyl))bis(2,2,2-trifluoroacetamide) (K)

[0118] To a 100 ml three-necked flask, add 10 g of compound J, 11.08 g of triethylamine and 40 ml of dichloromethane, stir and cool to 0°C, add a solution consisting of 20.88 g of p-toluenesulfonyl chloride and 80 ml of dichloromethane, raise the temperature to 10°C and react for 15 hours. The raw materials are basically reacted, concentrated, and purified by column, the eluent (dichloromethane: methanol = 80:1 ~ 30:1), the eluent is concentrated to obtain 18.16 g of white solid as compound K.

[0119] (3) Preparation of L

[0120] Add 15g of compound K, 100ml of water and 200ml of hydrochloric acid to a 500ml three-necked flask, stir and heat to 90°C to react for 48h. When the raw materials are basically reacted, stop the reaction, cool to -20°C, add 5M sodium hydroxide solution to adjust the pH to about 14, extract with DCM, concentrate, and purify by column, the eluent (dichloromethane: methanol = 50:1 ~ 20:1), concentrate the eluent to obtain an oily substance, add 45ml of hydrochloric acid, concentrate to obtain 10.55g of light yellow solid as compound L.

[0121] (4) Preparation of M

[0122] Add 25 ml of acetonitrile, 12.34 g of the compound of formula E, 10 g of the compound of formula L, 4.02 g of N,N-diisopropylethylamine and 1.72 g of potassium iodide to a 100 ml reaction bottle, keep at 80±5°C and stir for 15±0.5 h, cool down, filter and add 20 ml of acetonitrile to rinse, remove the solvent from the filtrate under reduced pressure; add 200 ml of dichloromethane, stir for 1 h, add 100 ml of water, stir and separate, remove the solvent from the organic phase under reduced pressure; add 12 0ml ethyl acetate, keep warm at 50±5℃ and stir for 30±5min, cool to 25±5℃ and crystallize for 24±1h, filter with suction, and add 30ml ethyl acetate to rinse, add 120ml ethyl acetate to the wet product, keep warm at 50±5℃ and stir for 30±5min, cool to 25±5℃ and crystallize for 24±1h, filter with suction, and add 30ml ethyl acetate to rinse, and dry the wet product in vacuo at 50±5℃ to obtain 15.05g of off-white solid which is compound of formula M.

[0123] (5) Preparation B

[0124] Add 81g of 90% sulfuric acid to the reaction bottle, start stirring, add 15g of Formula M compound, heat to 110℃ for reaction for 48h, cool to 0℃, drop 150ml of anhydrous ethanol for crystallization, filter, air dry the filter cake at 55℃, add the dried solid to the reaction bottle, add 120ml of 1mol / L sodium hydroxide solution, start stirring, dissolve. Extract with DCM several times, combine the extracts and dry with anhydrous sodium sulfate. Concentrate the collected DCM layer under reduced pressure, and air dry at 55℃ to obtain 4.66g of white solid, which is the open-loop impurity B.

[0125] Its structural identification data are as follows: H-NMR (300 MHz, D2O) δ2.07~2.26 (m, 8H, 4CH2), 3.06~3.15 ​​(m, 4H, 2CH2), 3.24~3.43 (m, 12H, 6CH2), 3.60~3.65 (m, 12H, 6CH2), 4.44 (s, 2H, CH2), 4.59 (s, 2H, CH2), 7.67~7.69 (m, 4H, 4ArH); ESI-MS (m / z): 477.4 [M+H]+.

[0126] The HPLC spectrum of the plerixafor ring-opening impurity B obtained in Example 2 is shown in Figure 5 , its normalized purity is 96.26%, and the mass spectrum and H NMR spectrum of the open-ring impurity B are shown in Figure 6 and Figure 7 .

[0127] Example 3: Preparation of Impurity C

[0128] This example basically prepares the ring-opening impurity C of plerixafor according to the following process route:

[0129]

[0130] (1) Preparation of P

[0131] Add 10 g of compound F, 7.43 g of N,N-dimethylformamide dimethyl acetal and 70 ml of chloroform into a 100 ml three-necked flask, stir and heat to 60 °C for 24 h. When the raw materials are basically reacted, stop the reaction, cool to room temperature, and concentrate to obtain 9.59 g of oily substance which is compound N.

[0132] Add 9.5 g of the compound of formula N and 200 ml of water into a 1000 ml three-necked flask, stir and cool to 0°C, add a solution consisting of 24.21 g of di-tert-butyl dicarbonate and 200 ml of ethanol, heat to 15°C and react for 20 hours. The raw materials are basically reacted and concentrated to obtain 19.50 g of an oily substance which is the compound of formula O.

[0133] Add 200 ml of ethanol to the above compound of formula O, stir to dissolve, add a solution consisting of 4.20 g of sodium hydroxide and 200 ml of water, heat to 80°C and react for 22 hours. When the raw materials are basically reacted, stop the reaction, cool to room temperature, concentrate, and purify by column, the eluent (dichloromethane: methanol = 20:1~10:1), concentrate the eluent to obtain 10.05 g of oily substance which is compound of formula P.

[0134] (2) Preparation of Q

[0135] Add 25 ml of acetonitrile, 16.52 g of the compound of formula E, 10 g of the compound of formula P, 5.38 g of N,N-diisopropylethylamine and 2.3 g of potassium iodide to a 100 ml reaction bottle, keep at 80±5°C and stir for 15±0.5 h, cool down, filter and add 20 ml of acetonitrile to rinse, remove the solvent from the filtrate under reduced pressure; add 200 ml of dichloromethane, stir for 1±0.1 h, add 100 ml of water, stir and separate, remove the solvent from the organic phase under reduced pressure; add 120 ml ethyl acetate, keep warm at 50±5℃ and stir for 30±5min, cool to 25±5℃ and crystallize for 24±1h, filter with suction, add 30ml ethyl acetate to rinse, filter, add 120ml ethyl acetate to the wet product, keep warm at 50±5℃ and stir for 30±5min, cool to 25±5℃ and crystallize for 24±1h, filter with suction, add 30ml ethyl acetate to rinse, filter, and dry the wet product in vacuo at 50±5℃ to obtain 18.05g of white solid which is compound of formula Q.

[0136] (3) Preparation of C

[0137] Add 18.05g of compound Q, 100ml of methanol and 20ml of hydrochloric acid to a 500ml three-necked flask, stir and heat to 25℃ to react for 20h. The raw materials are basically reacted, stop the reaction, and concentrate to obtain an oily substance. Add 100ml of methanol, 10.50g of potassium carbonate, and 10ml of water to the oily substance, heat to 70±5℃, keep stirring for 4±0.5h, cool to 15±5℃, add hydrochloric acid solution dropwise to adjust the pH to 2-3, remove the solvent from the reaction solution under reduced pressure, cool to 15±5℃, add 3M sodium hydroxide solution dropwise to adjust the pH to 14, cool the reaction solution to 10±2℃, stir and crystallize for 5h, filter, and dry the filter cake at 55℃ in vacuum to obtain 4.32g of white solid, which is the open-loop impurity C.

[0138] Its structural identification data are as follows: H-NMR (400MHz, D2O) δ2.09~2.11 (m, 4H, 2CH2), 2.19~2.27 (m, 2H, CH2), 3.13~3.54 (m, 28H, 14CH2), 4.29 (s, 2H, CH2), 4.50 (s, 2H, CH2), 7.56~7.61 (m, 4H, 4ArH); ESI-MS (m / z): 463.5 [M+H] + .

[0139] The HPLC spectrum of the plerixafor ring-opening impurity C obtained in Example 3 is shown in Figure 8 , its normalized purity is 97.23%, and the mass spectrum and H NMR spectrum of the open-ring impurity C are shown in Fig. 9 and Fig.10 .

[0140] Example 4: Preparation of Impurity D

[0141] This example basically prepares the ring-opening impurity D of plerixafor according to the following process route:

[0142]

[0143] (1) Preparation of S

[0144] Add 10g of the compound of formula R and 200ml of dichloromethane into a 500ml three-necked flask, stir evenly, cool to 0°C, add 25.04g of di-tert-butyl dicarbonate (abbreviated as (BOC)2O) dropwise, then heat to 10°C and react for 4h. The raw materials are basically reacted. Concentrate the reaction solution and purify it through a column. The eluent (petroleum ether: ethyl acetate = 1:1 to 1:2) is concentrated to obtain 12.08g of a colorless oily substance which is a compound of formula S.

[0145] (2) Preparation of T

[0146] Add 25 ml of acetonitrile, 15.9 g of the compound of formula E, 10 g of the compound of formula S, 5.18 g of N,N-diisopropylethylamine and 2.22 g of potassium iodide to a 100 ml reaction bottle, keep at 80±5°C and stir for 20±0.5 h, cool down, filter and add 20 ml of acetonitrile to rinse, remove the solvent from the filtrate under reduced pressure; add 200 ml of dichloromethane, stir for 1±0.1 h, add 100 ml of water, stir and separate, remove the solvent from the organic phase under reduced pressure; add 120 ml ethyl acetate, keep warm at 50±5℃ and stir for 30±5min, cool to 25±5℃ and crystallize for 24±1h, filter with suction, add 30ml ethyl acetate to rinse, filter, add 120ml ethyl acetate to the wet product, keep warm at 50±5℃ and stir for 30±5min, cool to 25±5℃ and crystallize for 24±1h, filter with suction, add 30ml ethyl acetate to rinse, filter, and dry the wet product in vacuo at 50±5℃ to obtain 15.83g of off-white solid which is compound of formula T.

[0147] (3) Preparation of D

[0148] Add 15.83g of the white solid from the previous step, 100ml of methanol and 20ml of hydrochloric acid to a 500ml three-necked flask, stir and heat to 25℃ for 20h, the raw materials are basically reacted, stop the reaction, concentrate, and obtain an oily substance. Add 100ml of methanol, 9.07g of potassium carbonate, and 10ml of water to the oily substance, heat to 70±5℃, keep stirring for 5h, cool to 15±5℃, add hydrochloric acid solution dropwise to adjust the pH to 2-3, remove the solvent from the reaction solution under reduced pressure, cool to 15±5℃, add 3M sodium hydroxide solution dropwise to adjust the pH to 14, cool the reaction solution to 10±2℃, stir and crystallize for 5h, filter, and dry the filter cake at 55℃ in vacuum to obtain 3.59g of white solid, which is the open-loop impurity D.

[0149] Its structural identification data are as follows: H-NMR (300 MHz, D2O) δ2.10~2.24 (m, 8H, 4CH2), 3.15 (t, 2H, CH2), 3.25~3.31 (m, 6H, 3CH2), 3.45~3.54 (m, 12H, 6CH2), 3.68~3.79 (m, 8H, 4CH2), 4.37 (s, 2H, CH2), 4.57 (s, 2H, CH2), 7.63~7.70 (m, 4H, 4ArH); ESI-MS (m / z): 477.4 [M+H]+.

[0150] The HPLC spectrum of the plerixafor ring-opening impurity D obtained in Example 4 is shown in Fig.11 , its normalized purity is 95.87%, and the mass spectrum and H NMR spectrum of the open-ring impurity D are shown in Fig.12 and Fig.13 .

[0151] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A method for preparing plerixafor ring-opening impurities, namely impurity A, impurity B, impurity C and impurity D, wherein the reaction processes are as follows: The specific steps include: (1) Preparation of impurity A ① Impurity A uses F as a raw material to undergo an amidation reaction with ethyl trifluoroacetate to generate G, and then G undergoes an alkylation reaction with the raw material E to generate N-(2-aminoethyl)-2,2,2-trifluoro-N-(3-(2,2,2-trifluoro-N-(2-((4-((4,8,11-tri(2,2,2-trifluoroacetyl)-1,4,8,11-tetraazacyclotetradec-1-yl)methyl)benzyl)amino)ethyl)acetamido)acetamido)propyl)acetamide)acetamide (H), and then hydrolyzes and removes the protecting group trifluoroacetyl to obtain impurity A; ② The solvent used in the preparation method of G is selected from methanol, ethanol, and isopropanol. In the present invention, methanol is preferably used as the solvent; the amount of ethyl trifluoroacetate is 2 to 3 equivalents; the reaction temperature is 0 to 30°C; ③ The acidifying agent used in the preparation method of H is selected from triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably potassium carbonate; ④ In the preparation method of A, the deprotection reaction hydrolysis reagent is selected from sodium hydroxide, potassium carbonate, potassium hydroxide, preferably sodium hydroxide; (2) Preparation of impurity B ① Impurity B is subjected to acylation reaction with diethyl oxalate using I as raw material, and then to acylation reaction with p-toluenesulfonyl chloride, and then the protecting group is removed under acidic conditions to obtain L; L is subjected to alkylation reaction with raw material E to generate 2,2,2-trifluoro-N-(3-((2-((3-(2,2,2-trifluoroacetylamino)propyl)(4-((4,8,11-tri(2,2,2-trifluoroacetyl)-1,4,8,11-tetraazacyclotetradec-1-yl)methyl)benzyl)amino)ethyl)amino)propyl)acetamide (M), and then the protecting group is removed by hydrolysis to obtain impurity B; ② In the preparation method of L, sodium hydroxide, dilute sulfuric acid, or dilute hydrochloric acid can be used for the deprotection reaction. The present invention preferably uses dilute hydrochloric acid as the deprotection reagent; ③ In the preparation method of B, the alkylation reaction acidifying agent can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably N,N-diisopropylethylamine as the alkylation reaction acidifying agent; the deprotection reaction hydrolysis reagent can use sodium hydroxide, potassium hydroxide, concentrated sulfuric acid, preferably concentrated sulfuric acid; (3) Preparation of impurity C ① Impurity C uses F as raw material to undergo alkylation reaction with N,N-dimethylformamide dimethyl acetal, and then undergoes acylation reaction with di-tert-butyl dicarbonate, and then removes the tert-butyloxycarbonyl group under alkaline conditions to obtain P; P undergoes alkylation reaction with raw material E to generate Q, and then hydrolyzes and removes the protecting group to obtain impurity C; ② In the preparation method of N, N-dimethylformamide dimethyl acetal, the amount of N, N-dimethylformamide dimethyl acetal is 1 to 1.2 equivalents, and the temperature is 50 to 70°C; ③ In the preparation method of C, the alkylation reaction acid agent can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably N,N-diisopropylethylamine as the alkylation reaction acid agent; the deprotection reaction hydrolysis reagent can use sodium hydroxide, potassium hydroxide, concentrated sulfuric acid, hydrochloric acid, potassium carbonate, preferably hydrochloric acid is used to remove the tert-butyloxycarbonyl group, and then potassium carbonate is used to remove the trifluoroacetyl group; (3) Preparation of impurity D ① Impurity D is reacted with I as a raw material to undergo an acylation reaction with di-tert-butyl dicarbonate to obtain S; S is reacted with the raw material E to undergo an alkylation reaction to produce tert-butyl (2-((3-aminopropyl)(tert-butoxycarbonyl)amino)ethyl)(3-((4-((4-((4,8,11-tris(2,2,2-trifluoroacetyl)-1,4,8,11-tetraazacyclotetradecyl-1-yl)methyl)benzyl)amino)propyl)amino)aminobenzoate (T), and then the protecting group is removed by hydrolysis to obtain impurity D; ② In the preparation method of S, the amount of di-tert-butyl dicarbonate used ranges from 2 to 3 equivalents, preferably 2.5 equivalents; ③ In the preparation method of impurity D, the alkylation reaction acidifying agent can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably N,N-diisopropylethylamine as the alkylation reaction acidifying agent; the deprotection reaction hydrolysis reagent can use sodium hydroxide, potassium hydroxide, concentrated sulfuric acid, hydrochloric acid, potassium carbonate, preferably using hydrochloric acid to remove the tert-butyloxycarbonyl group, and then using potassium carbonate to remove the trifluoroacetyl group.

2. The method according to claim 1, characterized in that: 11-(4-chloromethyl)benzyl-1,4,8-tris(trifluoroacetyl)-1,4,8,11-tetraazacyclotetradecane (E) and N,N'-(Propane-1,3-diyl)bis(N-(2-aminoethyl)-2,2,2-trifluoroacetamide)(G) Alkylation reaction occurs under weak alkaline conditions, and then the protective group trifluoroacetyl is removed to obtain N1-(2-((4-(1,4,8,11-tetraazacyclotetradec-1-yl)methyl)benzyl)amino)ethyl)-N3-(2-aminoethyl)propane-1,3-diamine is plerixafor impurity A; 11-(4-chloromethyl)benzyl-1,4,8-tris(trifluoroacetyl)-1,4,8,11-tetraazacyclotetradecane (E) and N,N'-((ethane-1,2-diylbis(azadiyl))bis(propane-3,1-diyl))bis(4-methylbenzenesulfonamide)(L) Alkylation reaction occurs under weak alkaline conditions, and then the protective group trifluoroacetyl is removed to obtain N1-(4-(1,4,8,11-tetraazacyclotetradec-1-yl)methyl)benzyl)-N1-(2-(3-aminopropyl)amino)ethyl)propane-1,3-diamine is plerixafor impurity B; 11-(4-chloromethyl)benzyl-1,4,8-tris(trifluoroacetyl)-1,4,8,11-tetraazacyclotetradecane (E) and Di-tert-butyl ((propane-1,3-diylbis(azadiyl)))bis(ethane-2,1-diyl)) dicarbamate (P) Alkylation reaction occurs under weak alkaline conditions, and then the protecting groups trifluoroacetyl and tert-butyloxycarbonyl are removed to obtain N1-(4-(1,4,8,11-tetraazacyclotetradecane-1-yl)methyl)benzyl)-N1,N3-bis(2-aminoethyl)propane-1,3-diamine is plerixafor impurity C; 11-(4-chloromethyl)benzyl-1,4,8-tris(trifluoroacetyl)-1,4,8,11-tetraazacyclotetradecane (E) and Di-tert-butylethane-1,2-diylbis((3-aminopropyl)carbamate)(S) Alkylation reaction occurs under weak alkaline conditions, and then the protecting groups trifluoroacetyl and tert-butyloxycarbonyl are removed to obtain N1-(4-(1,4,8,11-tetraazacyclotetradec-1-yl)methyl)benzyl)-N3-2-((3-aminopropyl)amino)ethyl)propane-1,3-diamine is plerixafor impurity D.

3. The method according to claim 1, wherein G is prepared by: 1 , N 1 '-(Propane-1,3-diyl)bis(ethane-1,2-diamine) (F) is used as a raw material, and reacts with ethyl trifluoroacetate under alkaline conditions at low temperature to undergo an amidation reaction to generate G; for example, the alkylation reaction acidifying agent can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably potassium carbonate as the alkylation reaction acidifying agent; the deprotection reaction hydrolysis reagent can use sodium hydroxide, potassium carbonate, potassium hydroxide, preferably sodium hydroxide; for example, the amidation reaction acidifying agent can use triethylamine, N,N-diisopropylethylamine, pyridine, preferably N,N-diisopropylethylamine as the amidation reaction acidifying agent; the reaction temperature control range is 0-30°C.

4. The method according to claim 1, wherein L is prepared by: 1 ,N 1 '-(ethane-1,2-diyl)bis(propane-1,3-diamine) (I) is used as a raw material, undergoes an acylation reaction with diethyl oxalate, and then undergoes an acylation reaction with p-toluenesulfonyl chloride, and then removes the protecting group under acidic conditions to obtain L; for example, the alkylation reaction acylating agent can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably N,N-diisopropylethylamine as the acylating agent for the alkylation reaction; the deprotection reaction hydrolysis reagent can use sodium hydroxide, potassium hydroxide, concentrated sulfuric acid; for example, the deprotection reaction can use sodium hydroxide, dilute sulfuric acid, dilute hydrochloric acid, preferably dilute hydrochloric acid as the deprotection reagent.

5. The method according to claim 1, wherein the preparation method of P is: using F as a raw material, undergoing an alkylation reaction with N,N-dimethylformamide dimethyl acetal, then undergoing an acylation reaction with di-tert-butyl dicarbonate, and then removing the tert-butyloxycarbonyl group under alkaline conditions to obtain P.

6. The method according to claim 5, wherein the acidifying agent for the alkylation reaction can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably N,N-diisopropylethylamine as the acidifying agent for the alkylation reaction; the hydrolysis reagent for the deprotection reaction can use sodium hydroxide, potassium hydroxide, concentrated sulfuric acid, hydrochloric acid, potassium carbonate, preferably hydrochloric acid is used to remove the tert-butyloxycarbonyl group, and then potassium carbonate is used to remove the trifluoroacetyl group.

7. according to the method for claim 1, wherein the preparation method of S is: take I as raw material, and acylation reaction with di-tert-butyl dicarbonate produces S.

8. The method according to claim 7, wherein the acidifying agent for the alkylation reaction can use triethylamine, potassium carbonate, N,N-diisopropylethylamine, sodium carbonate, preferably N,N-diisopropylethylamine as the acidifying agent for the alkylation reaction; the hydrolysis reagent for the deprotection reaction can use sodium hydroxide, potassium hydroxide, concentrated sulfuric acid, hydrochloric acid, potassium carbonate, preferably hydrochloric acid is used to remove the tert-butyloxycarbonyl group, and then potassium carbonate is used to remove the trifluoroacetyl group.

9. The method according to claim 7, wherein the amount of di-tert-butyl dicarbonate used ranges from 2 to 3 equivalents, preferably 2.5 equivalents.

10. The method according to claim 1, wherein impurity A is prepared by the following steps: (1) Preparation of G: At room temperature, add 25g of methanol to a 100ml reaction bottle, then add 10g of the compound of formula F and 2g of N,N-diisopropylethylamine to the reaction bottle in sequence, start stirring and refrigeration, cool to 0-10°C (5°C), and add 22.2g of ethyl trifluoroacetate dropwise to the reaction system at a temperature of 0-10°C (5°C). After the addition is complete, heat to 25°C, stir at 25°C for 4h, then add 25g of acetonitrile to the reaction system, concentrate under reduced pressure at 45°C, add 25g of acetonitrile to the reaction system again, and concentrate under reduced pressure again to obtain a light yellow viscous oily substance, which is the compound of formula G; (2) Preparation of H: At room temperature, add 120g of acetonitrile to the reaction bottle containing the compound of formula G, start stirring, add 35g of the compound of formula E, 4.2g of potassium carbonate, and 5.1g of potassium iodide to the reaction bottle in sequence, heat to 80°C to react for 5 to 7 hours (actually 6 hours), cool to 20 to 30°C (25°C, filter, take the filtrate, concentrate to obtain a viscous oil; add 50g of anhydrous ethanol, heat to 70 to 75°C, keep warm and stir for 30 minutes until the oil is All dissolved, then cooled to 20-30°C (25°C) to obtain an ethanol solution, which was added dropwise to 100 ml of water, stirred and crystallized at 20-30°C (25°C) for 1 hour, filtered, and dried to obtain an off-white solid; 230 g of ethyl acetate was added to the yellow solid, stirred and heated to reflux, kept warm and stirred for 30 minutes, slowly cooled to 30±2°C, kept warm and stirred for 10 hours, filtered, and dried to obtain an off-white solid as a compound of formula H; (3) Preparation A: At room temperature, 200 g of isopropanol / acetone mixture was added to a 500 ml reaction bottle, stirring was started, 10 g of sodium hydroxide and 20 g of water were added in sequence, and the mixture was stirred to dissolve. 38.3 g of compound H was added to the reaction bottle, and the reaction solution was heated to 65°C and kept warm for 4 hours under nitrogen protection. The solvent was removed by concentration under reduced pressure, 60 g of methanol was added, and the mixture was stirred to dissolve. 400 g of purified water was added dropwise, the mixture was cooled to 0°C with stirring, and the mixture was kept warm for 5 hours to precipitate. The mixture was filtered and dried with air at 50°C to obtain an off-white solid, which was an open-loop impurity A.

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