CN114853618A

CN114853618A - Preparation method of cationic lipid material DODMA

Info

Publication number: CN114853618A
Application number: CN202210307849.8A
Authority: CN
Inventors: 于文渊; 吉民; 刘来富; 周光密
Original assignee: Jiangsu Dongnan Nano Material Co ltd
Current assignee: Jiangsu Dongnan Nano Material Co ltd
Priority date: 2022-03-25
Filing date: 2022-03-25
Publication date: 2022-08-05

Abstract

The invention discloses a preparation method of cationic lipid material DODMA, which comprises the following steps: oleyl alcohol and 3-dimethylamino-1, 2-propylene glycol are used as raw materials to carry out etherification reaction, and then DODMA is obtained through filtration, concentration and purification. The method has the advantages of simple reaction steps, simple process, fewer byproducts, high yield, greatly reduced production cost and easy industrial production.

Description

Preparation method of cationic lipid material DODMA

Technical Field

The invention belongs to the field of pharmaceutical chemicals, and particularly relates to a preparation method of cationic lipid material DODMA.

Background

DODMA, chemical name 1, 2-dioleyl-3-dimethylamino-propane, English common name DODMA, CAS number 104162-47-2, is an important artificially synthesized lipid material. With the development of nanotechnology, nano biomaterials show good development prospects in gene therapy, cationic lipids comprise a group of amphiphiles showing positive charges, the charges interact with DNA/RNA (deoxyribonucleic acid/ribonucleic acid) with negative charges, so that a compound containing concentrated gene substances is formed, and cationic liposomes compounded with gene materials are expected to be used as non-viral vectors of gene therapy, including DODMA. Lipid nanoparticles with DODMA may have a neutral or low potential at physiological pH values. These extraordinary structures have profound potential for use in gene therapy. The existing route has the defects of multi-step reaction, generally adopting column chromatography for purification, low efficiency and the like.

Disclosure of Invention

The invention aims to provide a preparation method for preparing DODMA according to a green chemical synthesis concept, which avoids the use of highly toxic substances, has simple process, mild conditions, few side reactions and less harm to people and environment, and is suitable for industrial scale-up production.

In order to achieve the purpose, the main technical scheme provided by the invention is as follows:

a preparation method of DODMA (DoDMA) comprises the steps of taking oleyl alcohol (a compound shown in a formula 1) and 3-dimethylamino-1, 2-propanediol (a compound shown in a formula 2) as raw materials, adding an etherification reagent, carrying out etherification reaction in a reaction solvent, filtering and concentrating to obtain a crude product of DODMA (a compound shown in a formula 3).

Preferably, the etherification reagent is selected from one or more of concentrated sulfuric acid, diisopropyl azodicarboxylate (DIAD) and diethyl azodicarboxylate (DEAD), more preferably diisopropyl azodicarboxylate (DIAD).

Preferably, the reaction solvent is one or more selected from tetrahydrofuran, dioxane, ethyl acetate, toluene, xylene or N, N-dimethylformamide, and more preferably is toluene.

The preparation method further comprises the step of purifying the DODMA crude product:

(1) pulping the DODMA crude product at room temperature by using a solvent 1 for liquid separation to obtain a product phase;

(2) adding a solvent 2 into the product phase, stirring and dissolving, cooling and crystallizing, filtering and drying to obtain DODMA;

the solvent 1 and the solvent 2 are the same or different and are selected from one or more of methanol, ethanol, acetone, ethyl acetate, n-hexane and methyl tert-butyl ether.

Preferably, the solvent 1 is n-hexane, and the solvent 2 is methyl tert-butyl ether.

Preferably, the volume mass ratio of the solvent 1 to the crude product is 4-1 mL: 1g, the volume mass ratio of the solvent 2 to the crude product is 4-1 mL: 1g of the total weight of the composition. More preferably, the volume mass ratio of the solvent 1 to the crude product is 4 mL: 1 g. The volume mass ratio of the solvent 2 to the crude product is 2 mL: 1g of the total weight of the composition.

Preferably, the solvent 2 is used in the step (2) for heating and dissolving, and the temperature is 20-60 ℃, preferably 30 ℃; the crystallization temperature is-10 to 0 ℃, and is preferably-10 ℃.

The invention also aims to provide the DODMA prepared by the method, and the TLC purity of the DODMA is more than or equal to 98.0%.

The invention has the following advantages and effects: 1) the use of highly toxic raw materials is avoided, and the harm to the environment and people is greatly reduced; 2) mild reaction conditions, less side reactions, high yield and greatly reduced production cost. 3) The product is separated and purified by washing and recrystallization, thus avoiding the use of a large amount of solvent for column chromatography and having less pollution to the environment.

Drawings

FIG. 1 is a MS spectrum of DODMA obtained in example two.

FIG. 2 shows the DODMA obtained in example two ¹ H-NMR nuclear magnetic hydrogen spectrum.

FIG. 3 shows the TLC results of DODMA obtained in example two.

Detailed Description

The present invention is further illustrated by the following examples, which are illustrative of the present invention and are not to be construed as limiting the invention thereto.

EXAMPLE-preparation of crude DODMA

The method comprises the following steps:

to a clean and dry 5000mL three-necked flask, oleyl alcohol (Mw: 268.48, 268.5g, 1.0mol, 1.0eq), 3-dimethylamino-1, 2-propanediol (Mw: 119.16, 143.0g, 1.2mol, 1.2eq), triphenylphosphine oxide (Mw: 262.29, 314.7g, 1.2mol, 1.2eq), and toluene (2700mL) were added, and the mixture was stirred by magnetic force. Diisopropyl azodicarboxylate (DIAD) (Mw: 202.21, 242.7g, 1.2mol, 1.2eq) was added dropwise to the reaction system at-10 to 0 ℃. After the dropwise addition, reacting for 2 hours at the temperature of-10-0 ℃. Filtering, concentrating the filtrate at 50 ℃ to dryness to obtain 612.6g of crude DODMA product with the yield of 98.8%.

Method two

To a clean and dry 5000mL three-necked flask, oleyl alcohol (Mw: 268.48, 268.5g, 1.0mol, 1.0eq), 3-dimethylamino-1, 2-propanediol (Mw: 119.16, 143.0g, 1.2mol, 1.2eq), triphenylphosphine oxide (Mw: 262.29, 314.7g, 1.2mol, 1.2eq), and toluene (2700mL) were added, and the mixture was stirred by magnetic force. Diethyl azodicarboxylate (DEAD) (Mw: 174.15, 209.0g, 1.2mol, 1.2eq) was added dropwise to the reaction system at-10 to 0 ℃. After the dropwise addition, reacting for 2 hours at the temperature of-10-0 ℃. Filtering, concentrating the filtrate at 50 deg.C to dryness to obtain crude product of DODMA 533.8g with yield of 86.1%.

Method III

Adding oleyl alcohol (Mw: 268.48, 268.5g, 1.0mol, 1.0eq), 3-dimethylamino-1, 2-propanediol (Mw: 119.16, 143.0g, 1.2mol, 1.2eq), concentrated sulfuric acid (8.1g, 3%), toluene (2700mL) into a clean and dry 5000mL three-neck flask, uniformly stirring by magnetic force, putting up a water distribution device, carrying out reflux reaction for 6h, distributing water, adding sodium bicarbonate solid to neutralize sulfuric acid until the pH is neutral, filtering, adding sodium sulfate anhydrous into filtrate for drying for 1h, filtering, and concentrating the filtrate at 50 ℃ to dryness to obtain the DODMA crude product 513.3g with the yield of 82.7%.

Determining the first method as the optimal reaction condition, wherein the etherification reagent is diisopropyl azodicarboxylate.

EXAMPLE purification of DiDODMA

Adding a crude DODMA (733.5g) and n-hexane (2942mL) into a clean and dry 3000mL three-necked bottle, stirring and pulping for 1h at room temperature, separating, keeping a product phase, adding the product phase and methyl tert-butyl ether (1467mL) into a reaction bottle, stirring and dissolving at the temperature of 30 ℃, cooling to-10 ℃ under stirring for crystallization for 6h, filtering, and drying a filter cake at the temperature of 25 ℃ under vacuum for 24h to obtain 510.6g of a DODMA finished product, wherein the yield is 69.6%, the purity is not less than 99.0% by TLC (thin layer chromatography) detection, and a developing agent (dichloromethane: methanol: 12:1, 10% phosphomolybdic acid ethanol solution, and high-temperature color development at 170 ℃). The MS spectrum is shown in figure 1, ¹ the H-NMR nuclear magnetic hydrogen spectrum is shown in FIG. 2, and the TLC result is shown in FIG. 3.

And (3) under certain other conditions, examining the influence of the volume mass ratio of the solvent 1 and the DODMA crude product in the step (2) on the purity and yield of the product. The results are shown in Table 1.

TABLE 1

Volume mass ratio of solvent 1 to crude product	Yield (%)	Purity (%)
			4mL:1g	69.6％	99.0％
3mL:1g	72.3％	97.5％
			2mL:1g	76.1％	96.0％
1mL:1g	80.2％	94.4％

And (3) under certain other conditions, examining the influence of the volume mass ratio of the solvent 2 and the DODMA crude product in the step (2) on the purity and yield of the product. The results are shown in table 2:

TABLE 2

Volume-to-mass ratio of solvent 2 to crude product	Yield (%)	Purity (%)
			4mL:1g	43.1％	98.8％
3mL:1g	55.3％	98.5％
			2mL:1g	69.6％	99.0％
1mL:1g	74.2％	96.4％

Determining the volume mass ratio of the solvent 1 to the DODMA crude product to be 4 mL: 1g, the volume mass ratio of the solvent 2 to the crude product of DODMA is 2 mL: 1g, is the optimal purification condition.

It should be noted that the above-mentioned embodiments are only for illustrating the technical concept and features of the present invention, and the purpose of the present invention is to enable those skilled in the art to understand the content of the present invention and implement the present invention, and not to limit the protection scope of the present invention. All equivalent changes and modifications made according to the spirit of the present invention should be covered within the protection scope of the present invention.

Claims

1. A preparation method of DODMA is characterized in that oleyl alcohol and 3-dimethylamino-1, 2-propylene glycol are used as raw materials, an etherification reagent is added, etherification reaction is carried out in a reaction solvent, and a crude product of DODMA is obtained after filtration and concentration.

2. The process according to claim 1, wherein the etherification reagent is one or more selected from concentrated sulfuric acid, diisopropyl azodicarboxylate, and diethyl azodicarboxylate.

3. The method according to claim 1, wherein the reaction solvent in step (1) is one or more selected from tetrahydrofuran, dioxane, ethyl acetate, toluene, xylene, and N, N-dimethylformamide.

4. The process according to claim 1, wherein the etherification reagent is diisopropyl azodicarboxylate and the reaction solvent is toluene.

5. The process according to any one of claims 1 to 4, characterized by further comprising a step of purifying the crude DODMA product:

6. The purification process according to claim 5, characterized in that the solvent 1 is n-hexane and the solvent 2 is methyl tert-butyl ether.

7. The purification process according to claim 5, wherein the volume-to-mass ratio of the solvent 1 to the crude product is 4-1 mL: 1g, the volume mass ratio of the solvent 2 to the crude product is 4-1 mL: 1g of the total weight of the composition.

8. The purification process according to claim 7, wherein the volume mass ratio of the solvent 1 to the crude product is 4 ml: 1g, the volume mass ratio of the solvent 2 to the crude product is 2 mL: 1g of the total weight of the composition.

9. The method according to claim 5, wherein the solvent 2 is used in the step (2) for heating and dissolving, and the temperature is 20 to 60 ℃ and the crystallization temperature is-10 to 0 ℃.

10. DODMA, which is characterized by being prepared by the method of any one of claims 1 to 9 and having a DODMA content of 98.0% or more.