Preparation method of oleoyl lysophosphatidyl ethanolamine

Abstract

The invention provides a preparation method of oleoyl lysophosphatidyl ethanolamine, which comprises the following steps: S1, reacting a compound I with bis (diisopropylamino) (2-cyanoethoxy) phosphine, Teoc-ethanolamine and tert-butyl hydroperoxide to generate a compound II; s2, the compound II is subjected to an oxidation reaction under the action of 1, 8-diazabicyclo (5, 4, 0)-7-undecene, and a compound III is generated; and S3, enabling the compound III to react with a hydrogen fluoride pyridine solution to generate oleoyl lysophosphatidyl ethanolamine. According to the preparation method disclosed by the embodiment of the invention, the oleoyl lysophosphatidyl ethanolamine (O-LysoPE) can be obtained by adopting a simple compound I as a starting raw material through substitution reaction and oxidation reaction, the synthesis route is short, the operation is simple and convenient, the yield is high, the total yield can reach 64%, and the preparation method is suitable for industrial production.

Description

Technical Field

[0001] The invention belongs to the technical field of compound preparation, and more specifically, relates to a method for preparing oleoyl lysophosphatidylethanolamine. Background Art

[0002] Liposomes originated from the accidental discovery of British scientist Alec Bangham and his colleagues in 1961. They found that when phospholipids are dispersed in an aqueous medium, they spontaneously form closed vesicles. The structure of liposomes is extremely similar to that of cell membranes, making them highly biocompatible and biodegradable. Therefore, liposomes can protect drugs from being degraded by enzymes before reaching the lesion site. At the same time, drugs are "hidden" inside liposomes in the form of physical encapsulation, which can improve drug stability, reduce drug toxicity, increase drug dosage, and achieve better therapeutic effects. The bilayer surface formed by amphiphilic phospholipids can be structurally modified with ligands or other functional groups through physical or chemical means to make liposomes tissue-targeted, prolong the effective residence time of liposomes in the lesion site, and even achieve efficient targeted drug delivery. By changing the charge on the bilayer surface, it can be used for the encapsulation and delivery of DNA and RNA drugs, such as cationic liposomes.

[0003] Lysophosphatidylethanolamine (LPE) is one of the liposomes. It is a biologically active substance in the human body and is produced by the hydrolysis of phospholipid membranes by phospholipase A2. LPE can enter the cell through the LPE transporter on the cell membrane and participate in cell signal transduction. LPE can be metabolized by the human body, degraded into lysophosphatidic acid by phospholipase B, and can also react with oxidants to generate oxidized lysophosphatidylethanolamine (LPEox). The products of LPE metabolism include lysophosphatidylcholine, lysophosphatidylglycerol, etc. The lost LPE can be supplemented through a reasonable diet, corresponding nutrients, etc., which is of certain significance to maintaining good health.

[0004] LPE exists naturally in animal cells or plant cells, and also exists in human serum in small amounts. From a molecular structure point of view, LPE has the hydrophilic and hydrophobic groups of phosphatidylethanolamine. The reduction of hydrophobic groups greatly improves its hydrophilic properties, giving it good emulsification stability, antibacterial and antioxidant capabilities. LPE has important medical application value and is widely used in functional foods and healthcare industries. LPE can be used to fight HIV, its derivatives have anti-tumor effects, and its liposomes can be used as drug carriers.

[0005] The known synthesis method reported in the literature is shown in the following formula:

[0006]

[0007] As shown in the above formula, the synthetic route is to condense acetone glycerol with oleic acid, followed by hydrolysis to obtain the first intermediate, after which the TBS protecting group is added to the hydroxyl group at position 2 of the first intermediate to obtain the second intermediate, and finally the second intermediate is reacted with phosphorus oxychloride and ethanolamine to obtain the final product. However, the entire route has problems such as poor selectivity, many by-products, multiple purification by chromatography columns, and low yield, and is not suitable for mass production. Summary of the invention

[0008] In view of this, the object of the present invention is to provide a method for preparing oleoyl lysophosphatidylethanolamine with a short route, simple operation and high yield.

[0009] In order to solve the above technical problems, the present invention adopts the following technical solutions:

[0010] According to the method for preparing oleoyl lysophosphatidylethanolamine in an embodiment of the present invention, the synthesis route thereof is shown in the following formula (1):

[0011]

[0012] The specific steps include:

[0013] Step S1, reacting compound I with bis(diisopropylamino)(2-cyanoethoxy)phosphine, Teoc-ethanolamine, and tert-butyl peroxide to generate compound II, wherein the structural formula of compound I is shown as I in formula (1), and the structural formula of compound II is shown as II in formula (1);

[0014] Step S2, causing the compound II to undergo an oxidation reaction under the action of 1,8-diazabicyclo(5,4,0)-7-undecene to generate a compound III, wherein the structural formula of the compound III is shown as III in formula (1);

[0015] Step S3, reacting the compound III with a hydrogen fluoride pyridine solution to generate oleoyl lysophosphatidylethanolamine, the structure of which is shown as IV in formula (1).

[0016] In some embodiments of the present invention, the reaction of step S1 is shown in the following formula (2):

[0017]

[0018] The step S1 specifically includes:

[0019] Step S11, allowing the compound I to undergo a first substitution reaction with the bis(diisopropylamino)(2-cyanoethoxy)phosphine in a first solvent under the action of a first base to obtain a first intermediate, wherein the structural formula of the first intermediate is shown as I1 in formula (2);

[0020] Step S12, after the reaction of step S11 is completed, the Teoc-ethanolamine is continuously added into the reaction system to allow the first intermediate to undergo a second substitution reaction with the Teoc-ethanolamine to generate a second intermediate, wherein the structural formula of the second intermediate is shown as I2 in formula (2);

[0021] Step S13, after the reaction of step S12 is completed, continue to add the tert-butyl peroxide into the reaction system to react and generate the compound II.

[0022] Furthermore, the step S1 further includes:

[0023] Step S14, after the reaction of step S13 is completed, the organic phase is evaporated to dryness, and washed with water and methanol in sequence, and the first solvent is evaporated to dryness to obtain the compound II.

[0024] Furthermore, the first solvent is dichloromethane, tetrahydrofuran, or a mixture thereof, the first base is tetrazole, and the molar ratio of the compound I to the first base is 1:(1.1-1.3).

[0025] In some embodiments of the present invention, the oxidation reaction in step S2 is carried out in a second solvent, and the second solvent is n-heptane, toluene, or a mixture thereof.

[0026] Furthermore, in step S2, the reaction temperature is 0 to 30° C., and the reaction time is 2 to 6 hours.

[0027] Furthermore, after the reaction in step S2 is completed, the reaction solution is washed with hydrochloric acid, dried, and evaporated to dryness, and the obtained crude product is directly used in step S3.

[0028] In some embodiments of the present invention, step S3 is carried out in a third solvent, the third solvent is dichloromethane, methanol, or a mixture thereof, the molar ratio of compound III to pyridine hydrofluoric acid is 1:(3-5), the reaction temperature is 20-40°C, and the reaction time is 4-12 hours.

[0029] Furthermore, in the step S3, after the reaction is completed, pyridine is added to adjust the reaction solution to neutrality, the solvent is evaporated and passed through a chromatography column to obtain the oleoyl lysophosphatidylethanolamine.

[0030] The above technical solution of the present invention has at least one of the following beneficial effects:

[0031] According to the preparation method of the embodiment of the present invention, a simple compound I is used as a starting material, and oleoyl lysophosphatidylethanolamine (O-LysoPE) can be obtained through substitution reaction and oxidation reaction. The synthesis route is short, the operation is simple, the yield is high, and the total yield can reach 64%, which is suitable for industrial production. DETAILED DESCRIPTION

[0032] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution of the present invention will be clearly and completely described in combination with the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the described embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field belong to the scope of protection of the present invention.

[0033] First, the preparation method of oleoyl lysophosphatidylethanolamine according to an embodiment of the present invention is described in detail below.

[0034] According to the method for preparing oleoyl lysophosphatidylethanolamine in an embodiment of the present invention, the synthesis route thereof is shown in the following formula (1).

[0035]

[0036] Specifically, the steps include:

[0037] Step S1, reacting compound I with bis(diisopropylamino)(2-cyanoethoxy)phosphine, Teoc-ethanolamine, and tert-butyl peroxide to generate compound II, wherein the structural formula of compound I is shown as I in formula (1), and the structural formula of compound II is shown as II in formula (1);

[0038] Step S2, causing the compound II to undergo an oxidation reaction under the action of 1,8-diazabicyclo(5,4,0)-7-undecene to generate a compound III, wherein the structural formula of the compound III is shown as III in formula (1);

[0039] Step S3, reacting the compound III with a hydrogen fluoride pyridine solution to generate oleoyl lysophosphatidylethanolamine, the structure of which is shown as IV in formula (1).

[0040] That is, according to the preparation method of the present application, compound I is used as the starting material, and after substitution reaction and oxidation reaction, the target product oleoyl lysophosphatidylethanolamine is obtained. The synthetic route is short, the operation is simple, the yield is high, and it is suitable for industrial-scale production.

[0041] Below, each step is explained in detail.

[0042] (i) Step S1, reacting compound I with bis(diisopropylamino)(2-cyanoethoxy)phosphine, Teoc-ethanolamine and tert-butyl peroxide to generate compound II.

[0043] That is, first, compound I is subjected to a substitution reaction with bis(diisopropylamino)(2-cyanoethoxy)phosphine, Teoc-ethanolamine, and tert-butyl peroxide to produce compound II.

[0044] In some embodiments of the present invention, the reaction of step S1 is shown in the following formula (2):

[0045]

[0046] Specifically, step S1 includes:

[0047] Step S11, allowing the compound I to undergo a first substitution reaction with the bis(diisopropylamino)(2-cyanoethoxy)phosphine in a first solvent under the action of a first base to obtain a first intermediate, wherein the structural formula of the first intermediate is shown as I1 in formula (2);

[0048] Step S12, after the reaction of step S11 is completed, the Teoc-ethanolamine is continuously added into the reaction system to allow the first intermediate to undergo a second substitution reaction with the Teoc-ethanolamine to generate a second intermediate, wherein the structural formula of the second intermediate is shown as I2 in formula (2);

[0049] Step S13, after the reaction of step S12 is completed, continue to add the tert-butyl peroxide into the reaction system to react and generate the compound II.

[0050] That is, through the one-pot method, compound I is used as a starting material, and three substitution reactions are carried out with bis(diisopropylamino)(2-cyanoethoxy)phosphine, Teoc-ethanolamine, and tert-butyl peroxide to generate compound II. The one-pot method is simple to operate and has less three wastes. At the same time, by adding the above three reagents one by one, adding the next reagent after the previous substitution reaction is completed to initiate the next substitution reaction, it can avoid the generation of unnecessary by-products, which helps to improve the yield and simplify the post-treatment of compound II.

[0051] Furthermore, the step S1 may also include:

[0052] Step S14, after the reaction of step S13 is completed, the organic phase is evaporated to dryness, and washed with water and methanol in sequence, and the first solvent is evaporated to dryness to obtain the compound II.

[0053] That is to say, the post-treatment of compound II is simple and low-cost. After simple treatment, it can be used in subsequent oxidation reactions, produces less three wastes, and is very suitable for industrial production.

[0054] The first solvent may be, for example, dichloromethane, tetrahydrofuran, or a mixture thereof, preferably dichloromethane, in which each raw material has good solubility, and dichloromethane can be removed by heating and evaporating, which is simple to operate.

[0055] In addition, the first base may be, for example, tetrazole. The molar ratio of compound I to the first base may be, for example, 1:(1.1-1.3).

[0056] (ii) Step S2, subjecting the compound II to an oxidation reaction under the action of 1,8-diazabicyclo(5,4,0)-7-undecene (ie, DBU) to generate compound III.

[0057] In some embodiments of the present invention, the oxidation reaction in step S2 is carried out in a second solvent, and the second solvent is n-heptane, toluene, or a mixture thereof, preferably n-heptane, which can be directly washed and removed after the reaction, and is easy to operate.

[0058] In addition, the reaction temperature is 0-30°C and the reaction time is 2-6 hours. Preferably, the reaction temperature is 5-10°C, and the generation of impurities can be reduced at low temperatures. Preferably, the reaction time is 2 hours, and the oxidation reaction is relatively rapid and efficient.

[0059] Furthermore, after the reaction in step S2 is completed, the reaction solution is washed with hydrochloric acid, dried, and evaporated to dryness, and the obtained crude product is directly used in step S3. That is, after the oxidation reaction, only simple post-treatment is required before the next step.

[0060] (III) Step S3, reacting the compound III with a hydrogen fluoride pyridine solution to generate oleoyl lysophosphatidylethanolamine.

[0061] That is, after obtaining compound III, further oxidation reaction is carried out with hydrogen fluoride pyridine solution to generate the target product oleoyl lysophosphatidylethanolamine.

[0062] In some embodiments of the present invention, step S3 is performed in a third solvent, the third solvent is dichloromethane, methanol, or a mixture thereof, the molar ratio of compound III to pyridine hydrofluoride is 1:3-5, preferably 1:4.3, the reaction temperature is 20-40°C, and the reaction time is 4-12 hours. The third solvent is preferably methanol, which has a high boiling point, and a higher reaction temperature can improve the reaction efficiency and shorten the reaction time.

[0063] Furthermore, in the step S3, after the reaction is completed, pyridine is added to adjust the reaction solution to neutrality, the solvent is evaporated and passed through a chromatography column to obtain the oleoyl lysophosphatidylethanolamine.

[0064] That is to say, according to the preparation method of the present invention, only the last step requires purification by chromatography column, and the post-treatment of all intermediates is relatively simple and easy to operate, with low cost and less three wastes, which is very suitable for industrial-scale production.

[0065] The preparation method of the oleoyl lysophosphatidylethanolamine of the present invention is further described in detail below through specific examples.

[0066] The following examples 1 and 2 correspond to step S1; examples 3 and 4 correspond to step S2; examples 5 and 6 correspond to step S3. It should be noted that the examples of different steps can be combined arbitrarily, that is, example 1, example 4, and example 5 can be combined to form a complete synthetic embodiment, example 2, example 4, and example 6 can also be combined to form a complete synthetic embodiment, and so on, and they are no longer enumerated one by one.

[0067] Example 1 Synthesis of Compound II

[0068] Add 30g of compound I, 200mL of dichloromethane, 21.2g of bis(diisopropylamino)(2-cyanoethoxy)phosphine and 4.9g of tetrazole to a 1L three-necked flask, protect the system with argon, lower it into an ice-water bath and stir, control the temperature to 10-15°C, react for 6 hours, and monitor the progress of the reaction by TLC. After the reaction is complete, add 4.9g of tetrazole and 14.4g of Teoc-ethanolamine, continue to react for 2 hours, monitor the progress of the reaction by TLC, add 9.8g of tert-butyl peroxide after the reaction is complete, continue to react for 2 hours, monitor the progress of the reaction by TLC, and post-process after the reaction is complete. Wash the reaction solution twice with 200mL of water, evaporate the organic phase, add 400mL of n-heptane again, and wash twice with 200mL of methanol. Evaporate the solvent to obtain 41.3g of compound II, with a yield of 82%.

[0069] Example 2 Synthesis of Compound II

[0070] Add 30g of compound I, 200mL of tetrahydrofuran, 21.2g of bis(diisopropylamino)(2-cyanoethoxy)phosphine and 4.9g of tetrazole to a 1L three-necked flask, protect the system with argon, drop into an ice-water bath and stir, control the temperature to 20-30°C, react for 8 hours, and monitor the reaction progress by TLC. After the reaction is complete, add 4.9g of tetrazole and 14.4g of Teoc-ethanolamine, continue to react for 2 hours, monitor the reaction progress by TLC, add 9.8g of tert-butyl peroxide after the reaction is complete, continue to react for 2 hours, monitor the reaction progress by TLC, and post-process after the reaction is complete. Evaporate the organic phase, add 400mL of n-heptane after evaporating the organic phase, wash the reaction solution twice with 200mL of water, and wash twice with 200mL of methanol. Evaporate the solvent to obtain 32.7g of compound II, with a yield of 65%.

[0071] Example 3 Synthesis of Compound III

[0072] 30g of compound II and 200mL of n-heptane as solvent were added to a 1L single-mouth bottle, and the mixture was placed in an ice-water bath with stirring. 7.1g of DBU was added dropwise at 10°C and the reaction was continued for 4 hours. The reaction progress was monitored by TLC. After the reaction was completed, the reaction solution was washed twice with 150mL of 1mol / L hydrochloric acid, and dried and evaporated to obtain 25.5g of compound III with a yield of 91%.

[0073] Example 4 Synthesis of Compound III

[0074] 30g of compound II and 200mL of toluene as solvent were added to a 1L single-mouth bottle, and the mixture was placed in an ice-water bath with stirring. 7.1g of DBU was added dropwise at 20°C and the reaction was continued for 2 hours. The reaction progress was monitored by TLC. After the reaction was completed, the reaction solution was washed twice with 150mL of 1mol / L hydrochloric acid, and dried and evaporated to obtain 23.2g of compound III with a yield of 83%.

[0075] Example 5 Synthesis of Compound IV, Oleoyl Lysophosphatidylethanolamine (O-LysoPE)

[0076] In a 1L single-mouth bottle, add 25g of compound III, 100mL of dichloromethane, and 16.3g of hydrogen fluoride pyridine solution (70%), and react at 30°C for 10 hours. Monitor the progress of the reaction by TLC. After the reaction is completed, add pyridine to adjust the reaction solution to neutrality, evaporate the solvent, and pass through a chromatography column to obtain 13.2g of compound IV, with a yield of 81%.

[0077] The NMR data of the product are shown below:

[0078] 1H NMR (CDCl3, 400MHz): δ = 0.86-0.89 (t, 3H), 1.27-1.30 (m, 20H), 1.6 (s, 2H), 2.0-2.01 (s, 4H) ,2.3-2.34(t,2H),3.2(s,2H),3.9-4.1(m,4H),5.30-5.38(m,2H),5.55(s,1H),8.36(s,2H).

[0079] The results showed that its structure was consistent with that of oleoyllysophosphatidylethanolamine.

[0080] Example 6 Synthesis of Compound IV, Oleoyl Lysophosphatidylethanolamine (O-LysoPE)

[0081] 25 g of compound III, 100 mL of methanol, and 16.3 g of hydrogen fluoride pyridine solution (70%) were added to a 1 L single-mouth bottle and reacted at 40° C. for 6 hours. The reaction progress was monitored by TLC. After the reaction was completed, pyridine was added to adjust the reaction solution to neutrality, and the solvent was evaporated and passed through a chromatography column to obtain 13.9 g of compound IV, with a yield of 86%.

[0082] The NMR data of the product are shown below:

[0083] 1H NMR (CDCl3, 400MHz): δ = 0.86-0.89 (t, 3H), 1.27-1.30 (m, 20H), 1.6 (s, 2H), 2.0-2.01 (s, 4H) ,2.3-2.34(t,2H),3.2(s,2H),3.9-4.1(m,4H),5.30-5.38(m,2H),5.55(s,1H),8.36(s,2H).

[0084] The results showed that its structure was consistent with that of oleoyllysophosphatidylethanolamine.

[0085] In summary, the method for preparing oleoyl lysophosphatidylethanolamine of the present invention has the advantages of short synthesis route, simple operation, low cost and high yield.

[0086] The above is a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for preparing oleoyl lysophosphatidylethanolamine, characterized in that: Its synthetic route is shown in the following formula (1): The specific steps include: Step S1, reacting compound I with bis(diisopropylamino)(2-cyanoethoxy)phosphine, Teoc-ethanolamine, and tert-butyl peroxide to generate compound II, wherein the structural formula of compound I is shown as I in formula (1), and the structural formula of compound II is shown as II in formula (1); Step S2, causing the compound II to undergo an oxidation reaction under the action of 1,8-diazabicyclo(5,4,0)-7-undecene to generate a compound III, wherein the structural formula of the compound III is shown as III in formula (1); Step S3, reacting the compound III with a hydrogen fluoride pyridine solution to generate oleoyl lysophosphatidylethanolamine, the structure of which is shown as IV in formula (1).

2. The preparation method according to claim 1, characterized in that: The reaction of step S1 is shown in the following formula (2): The step S1 specifically includes: Step S11, allowing the compound I to undergo a first substitution reaction with the bis(diisopropylamino)(2-cyanoethoxy)phosphine in a first solvent under the action of a first base to obtain a first intermediate, wherein the structural formula of the first intermediate is shown as I1 in formula (2); Step S12, after the reaction of step S11 is completed, the Teoc-ethanolamine is continuously added into the reaction system to allow the first intermediate to undergo a second substitution reaction with the Teoc-ethanolamine to generate a second intermediate, wherein the structural formula of the second intermediate is shown as I2 in formula (2); Step S13, after the reaction of step S12 is completed, continue to add the tert-butyl peroxide into the reaction system to react and generate the compound II.

3. The preparation method according to claim 2, characterized in that: The step S1 further comprises: Step S14, after the reaction of step S13 is completed, the organic phase is evaporated to dryness, and washed with water and methanol in sequence, and the first solvent is evaporated to dryness to obtain the compound II.

4. The preparation method according to claim 2, characterized in that: The first solvent is dichloromethane, tetrahydrofuran, or a mixture thereof, the first base is tetrazole, and the molar ratio of the compound I to the first base is 1:(1.1-1.3).

5. The preparation method according to claim 1, characterized in that: The oxidation reaction in step S2 is carried out in a second solvent, and the second solvent is n-heptane, toluene, or a mixture thereof.

6. The preparation method according to claim 5, characterized in that: In the step S2, the reaction temperature is 0 to 30° C., and the reaction time is 2 to 6 hours.

7. The preparation method according to claim 6, characterized in that: After the reaction in step S2 is completed, the reaction solution is washed with hydrochloric acid, dried, and evaporated to dryness, and the obtained crude product is directly used in step S3.

8. The preparation method according to claim 1, characterized in that: The step S3 is carried out in a third solvent, the third solvent is dichloromethane, methanol, or a mixture thereof, the molar ratio of the compound III to pyridine hydrofluoric acid is 1:3 to 1:5, the reaction temperature is 20 to 40° C., and the reaction time is 4 to 12 hours.

9. The preparation method according to claim 8, characterized in that: In the step S3, after the reaction is completed, pyridine is added to adjust the reaction solution to neutrality, the solvent is evaporated and the reaction solution is passed through a chromatography column to obtain the oleoyl lysophosphatidylethanolamine.

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