Caffeoyl amino acid methyl ester derivative as well as preparation method and application thereof

Abstract

The invention discloses a caffeoyl amino acid methyl ester derivative, the caffeoyl amino acid methyl ester derivative takes caffeic acid and amino acid methyl ester hydrochloride as raw materials, N-caffeoyl amino acid methyl ester is synthesized through reaction, and the compound can effectively inhibit tyrosinase activity and inhibit apple browning. The invention further discloses a preparation method of the caffeoyl amino acid methyl ester derivative. The preparation method is simple, the synthesis route is short, large-scale preparation is easy, and the price is low.

Description

Technical Field

[0001] The invention belongs to the technical field of organic synthesis, and in particular relates to caffeoyl amino acid methyl ester derivatives, and also relates to a preparation method and application of the caffeoyl amino acid methyl ester derivatives. Background Art

[0002] Caffeic acid, also known as 3,4-dihydroxycinnamic acid, is widely found in natural plants such as fruits, vegetables, coffee, olive oil, beans, artichokes, honeysuckle, etc. As a phenolic acid substance, caffeic acid can effectively capture free radicals and has free radical scavenging and antioxidant functions. In addition, caffeic acid can also be used to prevent bleeding or stop bleeding during surgical operations, as well as stop bleeding in hemorrhagic diseases such as internal medicine and obstetrics and gynecology. Currently, caffeic acid tablets have been marketed as hemostatic drugs.

[0003] In recent years, caffeic acid and its derivatives have received extensive attention in the fields of chemistry, food, pharmacy, cosmetics, etc., and a variety of caffeic acid derivatives have been shown to have a wide range of biological activities. For example, caffeic acid phenethyl ester has antioxidant, anti-inflammatory, anti-tumor, antibacterial, and immunomodulatory biological activities; caffeic acid sulfonamide derivatives have free radical scavenging, anticoagulant and antibacterial activities; caffeoyl p-hydroxyphenol ester has a whitening effect.

[0004] The production of melanin is a common adverse phenomenon in nature, which can cause browning of fruits and vegetables, pigmentation of human skin, tumors, abnormal melanin metabolism, etc. Tyrosinase, also known as polyphenol oxidase and phenol oxidase, is the rate-limiting enzyme in the biosynthesis of melanin, and the use of tyrosinase inhibitors is an important means of melanin production. At present, although two kinds of caffeoyl amino acid methyl esters (N-caffeoyl phenylalanine methyl ester and N-caffeoyl cysteine ​​methyl ester) have been synthesized, and it is found that they have the ability to scavenge free radicals and inhibit arginase. However, due to the diverse amino acid structures and large differences in properties, other caffeoyl amino acid methyl ester derivatives have not been reported, and the inhibitory effect of this type of substance on tyrosinase and the anti-browning effect of fruits and vegetables have not been studied. Summary of the invention

[0005] The invention aims to provide caffeoyl amino acid methyl ester derivatives, which have easy-to-obtain raw material sources and can be used as tyrosinase inhibitors, anti-enzymatic browning agents for fruits and vegetables, and therapeutic drugs for skin diseases with excessive melanin deposition.

[0006] Another object of the present invention is to provide a method for preparing the above-mentioned caffeoyl amino acid methyl ester derivatives, which has a simple synthesis route and a short reaction cycle;

[0007] The third object of the present invention is to provide new applications of the above-mentioned caffeoyl amino acid methyl ester derivatives.

[0008] The first technical solution adopted by the present invention is that the caffeoyl amino acid methyl ester derivatives have the following general structural formula: Among them, R is

[0009] Any of .

[0010] The second technical solution adopted by the present invention is a method for preparing caffeoyl amino acid methyl ester derivatives, and the specific steps are as follows:

[0011] Step 1, dehydrating and deoxygenating the round-bottom flask used for the reaction;

[0012] Step 2, under ice bath conditions, add caffeic acid into a round-bottom flask, and add solvent A to dissolve, after the caffeic acid is completely dissolved, add amino acid methyl ester hydrochloride, after the amino acid methyl ester hydrochloride is dissolved, add an organic base dropwise to obtain a mixed solution A;

[0013] Step 3, the mixed solution A is mixed thoroughly, the condensing agent is added to the mixed solution A, stirred at room temperature for 2h-24h, the reaction is monitored by thin layer chromatography, and after the reaction is complete, 1mL of water is slowly added dropwise to quench the reaction to obtain a mixed solution B;

[0014] Step 4, post-treating the mixed solution B, distilling the mixed solution B under reduced pressure to remove the organic solvent, adding ethyl acetate to fully dissolve, then washing with 1 mol / L hydrochloric acid solution, water, 5% sodium bicarbonate aqueous solution and saturated brine in sequence, drying the obtained organic phase with anhydrous sodium sulfate and filtering, collecting the filtrate, and concentrating the filtrate under reduced pressure to obtain a crude product;

[0015] Step 5: purify the crude product to obtain a pure product.

[0016] The present invention is also characterized in that:

[0017] The amino acid methyl ester hydrochloride is any one of phenylalanine methyl ester hydrochloride, valine methyl ester hydrochloride, alanine methyl ester hydrochloride, glycine methyl ester hydrochloride, tryptophan methyl ester hydrochloride, threonine methyl ester hydrochloride, leucine methyl ester hydrochloride, isoleucine methyl ester hydrochloride, tyrosine methyl ester hydrochloride, N-tert-butyloxycarbonyllysine methyl ester hydrochloride, aspartic acid dimethyl ester hydrochloride, glutamic acid dimethyl ester hydrochloride, methionine methyl ester hydrochloride, and serine methyl ester hydrochloride.

[0018] The solvent A is any one of N,N-dimethylformamide, tetrahydrofuran and dichloromethane; the organic base is any one of triethylamine, pyridine and 4-dimethylaminopyridine; and the condensing agent is any one of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride, Carter condensing agent and dicyclohexylcarbodiimide.

[0019] In step 2, the molar ratio of caffeic acid to amino acid methyl ester hydrochloride is 1:1 to 1.5.

[0020] The added amount of solvent A is 10 ml per millimole of caffeic acid; the molar ratio of caffeic acid to organic base is 1:1.5-3; and the molar ratio of caffeic acid to condensing agent is 1:1-1.5.

[0021] The third technical scheme adopted by the present invention is to disclose the new use of the caffeoyl amino acid methyl ester derivatives prepared by the above method, which can be used as tyrosinase inhibitors, as anti-enzymatic browning agents for fruits and vegetables, and as therapeutic drugs for skin diseases with excessive melanin deposition.

[0022] The invention has the beneficial effects that the preparation method of the caffeoyl amino acid methyl ester derivatives disclosed in the invention is simple, the synthetic route is short, it is easy to prepare in large quantities, and the price is low; and the caffeoyl amino acid compounds have good inhibitory activity on tyrosinase, and can be used as an agent for preventing browning of fruits and vegetables and a therapeutic drug for skin diseases with excessive melanin deposition. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 The caffeic acid amino acid methyl ester derivatives and the preparation method and application example 1 of the present invention are the compound a1 prepared 1 H NMR spectra;

[0024] Figure 2 The caffeic acid amino acid methyl ester derivatives and the preparation method and application example 2 of the present invention are the compound a2 prepared 1 H NMR spectra;

[0025] Figure 3 The caffeoyl amino acid methyl ester derivatives and their preparation method and application example 15 of the present invention are the compound a15 prepared 1 H NMR spectra;

[0026] Figure 4 The caffeoyl amino acid methyl ester derivatives and the preparation method and application example 4 of the present invention are the compound a4 prepared 13 C NMR spectra;

[0027] Figure 5 The caffeic acid amino acid methyl ester derivatives and the preparation method and application example 8 of the present invention are the compound a8 prepared 13 C NMR spectra;

[0028] Figure 6 The caffeoyl amino acid methyl ester derivatives and their preparation method and application example 14 of the present invention are the compound a14 prepared 13 C NMR spectrum. DETAILED DESCRIPTION

[0029] The present invention is described in detail below with reference to the accompanying drawings and specific embodiments.

[0030] Embodiment 1:

[0031] Synthesis of N-caffeoylphenylalanine methyl ester a1, the structural formula of compound a1 is as follows:

[0032]

[0033] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of tetrahydrofuran. After complete dissolution, 10 mmol of phenylalanine methyl ester hydrochloride was added, and 15 mmol of triethylamine was slowly added dropwise to ensure that the reaction was fully mixed, and then 12 mmol of dicyclohexylcarbodiimide (DCC) was added. The reaction was naturally heated to room temperature and stirred for about 18 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized from methanol to obtain the corresponding N-caffeoylphenylalanine methyl ester compound a1 with a yield of 92%. The prepared compound a1 1 H NMR images Figure 1 shown.

[0034] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a1 are shown below:

[0035] 1 H NMR (400MHz, CD 3 OD)δ7.37(d,J=15.7Hz,1H),7.30-7.17(m,5H),7.00(d,J=1.8Hz,1H),6.89(dd,J=8.2,1.9Hz,1H),6.76(d,J=8.2Hz,1H),6 .40(d,J=15.7Hz,1H),4.78(dd,J=8.6,5.8Hz,1H),3.68(s,3H),3.18(dd,J=13.8,5.8Hz,1H),3.02(dd,J=13.8,8.7Hz,1H); 13 CNMR(101MHz,CD 3 OD)δ173.6,169.0,148.9,146.7,143.1,138.1,130.2,129.5,128.1,127.9,122.3,117.5,116.4,115.1,55.5,52.7,38.5; HRMS(ESI)forC 19 H 19 NO 5Na(M+Na + ):Calcd:364.1155,Found:364.1138.

[0036] Embodiment 2:

[0037] Synthesis of N-caffeoyl valine methyl ester a2, the structural formula of compound a2 is as follows:

[0038]

[0039] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of N,N-dimethylformamide. After complete dissolution, 12 mmol of valine methyl ester hydrochloride was added, and 20 mmol of triethylamine was slowly added dropwise to ensure that the reaction was fully mixed, and then 10 mmol of DCC was added. The reaction was naturally heated to room temperature and stirred for about 24 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized from methanol to obtain the corresponding N-caffeoyl valine methyl ester compound a2 with a yield of 78%. The prepared compound a2 1 H NMR images Figure 2 shown.

[0040] The H NMR spectrum and C NMR spectrum data of compound a2 are as follows:

[0041] 1 H NMR (400MHz, CD 3 OD)δ7.42(d,J=15.6Hz,1H),7.03(s,1H),6.92(d,J=7.8Hz,1H),6.77(d,J=8.0Hz,1H),6.54(d,J=15.6 Hz,1H),4.45(d,J=6.1Hz,1H),3.73(s,3H),2.18(dq,J=13.3,6.7Hz,1H),0.98(dd,J=6.7,3.1Hz,6H); 13 CNMR(101MHz,CD 3 OD)δ173.8,169.4,149.0,146.8,143.1,128.1,122.3,117.6,116.4,115.1,59.4,52.5,31.9,19.5,18.6; HRMS(ESI) for C 15 H 19 NO 5 Na(M+Na + ):Calcd:316.1155,Found:316.1142.

[0042] Embodiment 3:

[0043] Synthesis of N-caffeoyl alanine methyl ester a3, the structural formula of compound a3 is as follows:

[0044]

[0045] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of dichloromethane. After complete dissolution, 15 mmol of alanine methyl ester hydrochloride was added, and 20 mmol of pyridine was slowly added to ensure that the reaction was fully mixed, and then 12 mmol of DCC was added. The reaction was naturally heated to room temperature and stirred for about 12 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized with methanol to obtain the corresponding N-caffeoyl alanine methyl ester compound a3 with a yield of 67%.

[0046] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a3 are shown below:

[0047] 1 H NMR (400MHz, CD 3 OD)δ7.41(d,J=15.5Hz,1H),7.02(s,1H),6.90(d,J=7.5Hz,1H),6.76(d,J=7.7Hz,1H ),6.42(d,J=15.5Hz,1H),4.51(q,J=7.1Hz,1H),3.73(s,3H),1.42(d,J=7.2Hz,3H); 13 CNMR(101MHz,CD 3 OD)δ174.9,169.0,149.5,147.0,143.1,127.9,122.3,117.4,116.5,114.9,52.8,49.6,17.5; HRMS(ESI) for C 13 H 15 NO 5 Na(M+Na + ):Calcd:288.0842,Found:288.0830.

[0048] Embodiment 4:

[0049] Synthesis of N-caffeoylglycine methyl ester a4, the structural formula of compound a4 is shown below:

[0050]

[0051] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of N,N-dimethylformamide. After complete dissolution, 12 mmol of glycine methyl ester hydrochloride was added, and 15 mmol of pyridine was slowly added dropwise to ensure that the reaction was fully mixed, and then 15 mmol of DCC was added. The reaction was naturally heated to room temperature and stirred for about 24 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized from methanol to obtain the corresponding N-caffeoylglycine methyl ester compound a4 with a yield of 75%. The prepared compound a4 1 H NMR images Figure 4 shown.

[0052] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a4 are shown below:

[0053] 1 H NMR (400MHz, CD 3 OD)δ7.42(d,J=15.7Hz,1H),7.03(d,J=1.8Hz,1H),6.91(dd,J=8.2,1.8Hz,1 H),6.77(d,J=8.2Hz,1H),6.43(d,J=15.7Hz,1H),4.05(s,2H),3.72(s,3H); 13 CNMR(101MHz,CD 3 OD)δ171.9,169.6,148.9,146.7,143.0,128.1,122.3,117.5,116.4,115.0,52.6,42.0; HRMS(ESI) for C 12 H 13 NO 5 Na(M+Na + ):Calcd:274.0686,Found:274.0675.

[0054] Embodiment 5:

[0055] Synthesis of N-caffeoyltryptophan methyl ester a5, the structural formula of compound a5 is shown below:

[0056]

[0057] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of tetrahydrofuran. After complete dissolution, 10 mmol of tryptophan methyl ester hydrochloride was added, and 20 mmol of 4-dimethylaminopyridine was slowly added. After ensuring that the reaction was fully mixed, 12 mmol of 1-ethyl-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDCI) was added. The reaction was naturally heated to room temperature and stirred for about 12 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized with methanol to obtain the corresponding N-caffeoyltryptophan methyl ester compound a5 with a yield of 89%.

[0058] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a5 are shown below:

[0059] 1 H NMR (400MHz, CD 3 OD)δ7.53(d,J=7.8Hz,1H),7.38(d,J=15.6Hz,1H),7.32(d,J=8.1Hz,1H),7.11-1.05(m,2H),7.03-6.98(m,2H),6.88(d,J=7.9Hz,1H),6 .75(d,J=8.1Hz,1H),6.41(d,J=15.7Hz,1H),4.87-4.82(m,1H),3.66(s,3H),3.35(dd,J=14.6,7.5Hz,1H),3.23(dd,J=14.6,7.5Hz,1H); 13 CNMR(101MHz,CD 3 OD)δ174.1,169.1,149.5,147.0,143.1,138.0,128.7,127.9,124.4,122.43,122.3 6,119.8,119.1,117.5,116.4,114.9,112.3,110.7,55.1,52.7,28.7; HRMS(ESI)for C 21 H 20 N 2 O 5 Na(M+Na + ):Calcd:403.1264,Found:403.1259.

[0060] Embodiment 6:

[0061] Synthesis of N-caffeoylthreonine methyl ester a6, the structural formula of compound a6 is shown below:

[0062]

[0063] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of N,N-dimethylformamide. After complete dissolution, 12 mmol of threonine methyl ester hydrochloride was added, and 20 mmol of triethylamine was slowly added dropwise to ensure that the reaction was fully mixed, and then 15 mmol of Carter condensation agent (BOP) was added. The reaction was naturally heated to room temperature and stirred for about 24 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized with methanol to obtain the corresponding N-caffeoylthreonine methyl ester compound a6 with a yield of 80%.

[0064] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a6 are shown below:

[0065] 1 H NMR (400MHz, CD 3 OD)δ7.44(d,J=15.2Hz,1H),7.05(s,1H),6.93(d,J=7.5Hz,1H),6.77(d,J=7.5Hz,1H) ,6.57(d,J=14.8Hz,1H),4.60(s,1H),4.34(s,1H),3.75(s,3H),1.21(d,J=5.6Hz,3H); 13 CNMR(101MHz,CD 3 OD)δ172.7,169.6,149.3,146.8,143.3,128.0,122.4,117.5,116.5,115.0,68.5,59.5,52.8,20.3; HRMS(ESI) for C 14 H 17 NO 6 Na(M+Na + ):Calcd:318.0948,Found:318.0935.

[0066] Embodiment 7:

[0067] Synthesis of N-caffeoyl leucine methyl ester a7, the structural formula of compound a7 is shown below:

[0068]

[0069] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of dichloromethane. After complete dissolution, 12 mmol of leucine methyl ester hydrochloride was added, and 20 mmol of triethylamine was slowly added dropwise to ensure that the reaction was fully mixed, and then 10 mmol of EDCI was added. The reaction was naturally heated to room temperature and stirred for about 12 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized with methanol to obtain the corresponding N-caffeoyl leucine methyl ester compound a7 with a yield of 75%.

[0070] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a7 are shown below:

[0071] 1 H NMR (400MHz, CD 3 OD)δ7.41(d,J=15.5Hz,1H),7.03(s,1H),6.91(d,J=6.8Hz,1H),6.77(d,J=6.5Hz,1H),6.44(d, J=15.5Hz,1H),4.61-4.52(m,1H),3.72(s,3H),1.70-1.65(m,3H),0.95(dd,J=12.7,6.1Hz,6H); 13 CNMR(101MHz,CD 3 OD)δ174.8,169.3,149.3,146.9,143.1,127.9,122.3,117.5,116.5,114.9,52.7,52.3,41.5,26.0,23.3,21.8; HRMS(ESI) for C 16 H 21 NO 5 Na(M+Na + ):Calcd:330.1312,Found:330.1298.

[0072] Embodiment 8:

[0073] Synthesis of N-caffeoyl isoleucine methyl ester a7, the structural formula of compound a8 is as follows:

[0074]

[0075] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of tetrahydrofuran. After complete dissolution, 10 mmol of isoleucine methyl ester hydrochloride was added, and 30 mmol of triethylamine was slowly added dropwise to ensure that the reaction was fully mixed, and then 12 mmol of DCC was added. The reaction was naturally heated to room temperature and stirred for about 12 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized from methanol to obtain the corresponding N-caffeoyl isoleucine methyl ester compound a8 with a yield of 90%. The prepared compound a8 1 H NMR images Figure 5 shown.

[0076] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a8 are shown below:

[0077] 1 H NMR (400MHz, CD 3 OD)δ7.42(d,J=15.5Hz,1H),7.04(s,1H),6.91(d,J=7.5Hz,1H),6.77(d,J=7.5Hz,1H),6.53(d,J=1 5.6Hz,1H),4.50(d,J=6.1Hz,1H),3.72(s,3H),1.57-1.46(m,1H),1.36-1.15(m,2H),0.94(m,6H); 13 CNMR(101MHz,CD 3 OD)δ173.7,169.1,148.6,146.4,143.0,128.1,122.2,117.6,116.4,115.1,58.3,52.4,38.3,26.3,15.9,11.7; HRMS(ESI) for C 16 H 21 NO 5 Na(M+Na + ):Calcd:330.1312,Found:330.1301.

[0078] Embodiment 9:

[0079] Synthesis of N-caffeoyltyrosine methyl ester a9, the structural formula of compound a9 is shown below:

[0080]

[0081] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of tetrahydrofuran. After complete dissolution, 10 mmol of tyrosine methyl ester hydrochloride was added, and 15 mmol of 4-dimethylaminopyridine was slowly added to ensure that the reaction was fully mixed, and then 15 mmol of BOP was added. The reaction was naturally heated to room temperature and stirred for about 6 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized with methanol to obtain the corresponding N-caffeoyltyrosine methyl ester compound a9 with a yield of 88%.

[0082] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a9 are shown below:

[0083] 1 H NMR (400MHz, CD 3 OD)δ7.36(d,J=15.6Hz,1H),7.03(d,J=8.2Hz,2H),6.99(s,1H),6.88(d,J=7.9Hz,1H),6.74(d,J=8.7Hz,1H),6.70(d,J=8.3Hz,2 H),6.39(d,J=15.6Hz,1H),4.70(dd,J=7.9,6.2Hz,1H),3.69(s,3H),3.07(dd,J=13.9,5.9Hz,1H),2.93(dd,J=13.8,8.4Hz,1H); 13 CNMR(101MHz,CD 3 OD)δ173.8,169.2,157.4,150.1,147.2,143.2,131.2,128.7,127.5,122.4,117.1,116.5,116.3,114.7,55.8,52.6,37.8; HRMS(ESI) for C 19 H 19 NO 6 Na(M+Na + ):Calcd:380.1105,Found:380.1091.

[0084] Embodiment 10:

[0085] N 6 -tert-butyloxycarbonyl-N 2 -Synthesis of caffeoyl lysine methyl ester, the structural formula of compound a10 is as follows:

[0086]

[0087] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of N,N-dimethylformamide. After complete dissolution, 10 mmol of N-tert-butyloxycarbonyllysine methyl ester hydrochloride was added, and 15 mmol of 4-dimethylaminopyridine was slowly added to ensure that the reaction was fully mixed, and then 15 mmol of EDCI was added. The reaction was naturally heated to room temperature and stirred for about 6 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized from methanol to obtain the corresponding N 6 -Boc-N 2 -Caffeoyl lysine methyl ester a10, yield 85%.

[0088] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a10 are shown below:

[0089] 1 H NMR (400MHz, CD 3 OD)δ7.41(d,J=15.4Hz,1H),7.03(s,1H),6.90(d,J=6.9Hz,1H),6.77(d,J=6.9Hz,1H),6.46(d,J=15.3Hz,1H),4. 49(d,J=2.9Hz,1H),3.72(s,3H),3.03(t,J=6.0Hz,2H),1.90-1.81(m,1H),1.79-1.69(m,1H),1.51-1.38(m,13H); 13 CNMR(101MHz,CD 3 OD)δ174.3,169.3,158.5,149.4,146.9,143.1,127.8,122.3,117.4,116.5,114.9,79.8,53.9,52.7,40.9,32.3,30.5,28.8,24.1; HRMS(ESI)for C 21 H 30 N 2 O 7 Na(M+Na + ):Calcd:445.1945,Found:445.1945.

[0090] Embodiment 11:

[0091] Synthesis of N-caffeoyl aspartic acid dimethyl ester a11, the structural formula of compound a11 is as follows:

[0092]

[0093] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of tetrahydrofuran. After complete dissolution, 12 mmol of dimethyl aspartate hydrochloride was added, and 25 mmol of triethylamine was slowly added dropwise to ensure that the reaction was fully mixed, and then 15 mmol of EDCI was added. The reaction was naturally heated to room temperature and stirred for about 18 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized with methanol to obtain the corresponding N-caffeoyl aspartic acid dimethyl ester a11 with a yield of 86%.

[0094] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a11 are shown below:

[0095] 1 H NMR (400MHz, CD 3 OD)δ7.42(d,J=15.7Hz,1H),7.02(d,J=1.9Hz,1H),6.91(dd,J=8.2,1.9Hz,1H),6.76(d,J=8.2H z,1H),6.43(d,J=15.7Hz,1H),4.93-4.87(m,1H),3.74(s,3H),3.69(s,3H),2.97-2.84(m,2H); 13 CNMR(101MHz,CD 3 OD)δ172.7,172.4,169.0,149.0,146.7,143.3,128.1,122.3,117.5,116.4,115.1,53.1,52.5,50.4,36.9; HRMS(ESI) for C 15 H 17 NO 7 Na(M+Na + ):Calcd:346.0897,Found:346.0888.

[0096] Embodiment 12:

[0097] Synthesis of N-caffeoyl glutamate dimethyl ester a12, the structural formula of compound a12 is as follows:

[0098]

[0099] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of N,N-dimethylformamide. After complete dissolution, 15 mmol of dimethyl glutamate hydrochloride was added, and 20 mmol of triethylamine was slowly added dropwise to ensure that the reaction was fully mixed, and then 15 mmol of EDCI was added. The reaction was naturally heated to room temperature and stirred for about 24 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized with methanol to obtain the corresponding N-caffeoyl glutamate dimethyl ester a12 with a yield of 76%.

[0100] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a12 are shown below:

[0101] 1 H NMR (400MHz, CD 3 OD)δ7.41(d,J=15.7Hz,1H),7.02(s,1H),6.92(d,J=8.2Hz,1H),6.77(d,J=8.1Hz,1H),6.43(d,J=15.7Hz,1 H),4.57(dd,J=8.8,5.4Hz,1H),3.74(s,3H),3.67(s,3H),2.46(t,J=7.4Hz,2H),2.21(m,1H),2.01(m,1H); 13 C NMR (101 MHz, CD 3 OD)δ174.7,173.7,169.3,149.0,146.8,143.2,128.2,122.3,117.6,116.5,115.1,53.2,52.8,52.2,31.0,27.8;

[0102] HRMS(ESI)for C 16 H 19 NO 7 Na(M+Na + ):Calcd:360.1054,Found:360.1046.

[0103] Embodiment 13:

[0104] Synthesis of N-caffeoylmethionine methyl ester a13, the structural formula of compound a13 is as follows:

[0105]

[0106] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of tetrahydrofuran. After complete dissolution, 12 mmol of methionine methyl ester hydrochloride was added, and 30 mmol of triethylamine was slowly added dropwise to ensure that the reaction was fully mixed, and then 12 mmol of benzotriazole-1-acyloxy-tris (dimethylamino) hexafluorophosphate (BOP) was added. The reaction was naturally heated to room temperature and stirred for about 2 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized with methanol to obtain the corresponding N-caffeoylmethionine methyl ester a13 with a yield of 94%.

[0107] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a13 are shown below:

[0108] 1 H NMR (400MHz, CD 3 OD)δ7.43(d,J=15.7Hz,1H),7.03(d,J=1.8Hz,1H),6.91(dd,J=8.2,1.9Hz,1H),6.77(d,J=8.2Hz,1H),6.46(d,J=15.7H z,1H),4.69(dd,J=9.0,4.8Hz,1H),3.72(s,3H),2.63-2.48(m,2H),2.18-2.08(m,1H),2.06(s,3H),2.04-1.94(m,1H); 13 C NMR (101 MHz, CD 3 OD)δ174.0,169.3,148.9,146.7,143.1,128.1,122.3,117.6,116.4,115.1,52.8,49.8,32.1,31.1,15.2; HRMS(ESI) for C 15 H 19 NO 5 SNa(M+Na + ):Calcd:348.0876,Found:348.0874.

[0109] Embodiment 14:

[0110] Synthesis of N-caffeoylserine methyl ester a14, the structural formula of compound a14 is shown below:

[0111]

[0112] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of N,N-dimethylformamide. After complete dissolution, 12 mmol of serine methyl ester hydrochloride was added, and 15 mmol of triethylamine was slowly added dropwise to ensure that the reaction was fully mixed, and then 10 mmol of BOP was added. The reaction was naturally heated to room temperature and stirred for about 2 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized from methanol to obtain the corresponding N-caffeoylserine methyl ester a14 with a yield of 87%. The prepared compound a14 1 H NMR images Figure 6 shown.

[0113] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a14 are shown below:

[0114] 1 H NMR (400MHz, CD 3 OD)δ7.43(d,J=14.5Hz,1H),7.03(s,1H),6.92(d,J=7.8Hz,1H),6.77(d,J=7.9Hz,1H),6.52 (d,J=13.5Hz,1H),4.65(s,1H),3.99-3.90(m,1H),3.90-3.80(m,J=7.6Hz,1H),3.76(s,3H); 13 CNMR(101MHz,CD 3 OD)δ172.4,169.2,149.1,146.8,143.1,128.1,122.3,117.7,116.4,115.0,63.0,56.4,52.8; HRMS(ESI) for C 13 H 15 NO 6 Na(M+Na + ):Calcd:304.0792,Found:304.0785.

[0115] Embodiment 15:

[0116] Synthesis of N-caffeoylcysteine ​​methyl ester a15, the structural formula of compound a15 is shown below:

[0117]

[0118] First, the round-bottom flask used for the reaction was dehydrated and deoxygenated, and then 10 mmol of caffeic acid was added under ice bath conditions and dissolved with 100 mL of tetrahydrofuran. After complete dissolution, 15 mmol of methyl glutamate hydrochloride was added, and 30 mmol of triethylamine was slowly added dropwise to ensure that the reaction was fully mixed, and then 15 mmol of BOP was added. The reaction was naturally heated to room temperature and stirred for about 12 hours. After the reaction was completed, the reaction solution was post-treated to obtain a crude product, which was recrystallized from methanol to obtain the corresponding N-caffeoyl glutamate dimethyl ester a15 with a yield of 95%. The prepared compound a15 1 H NMR images Figure 3 shown.

[0119] The H NMR spectrum, C NMR spectrum and high-resolution mass spectrum data of compound a15 are shown below:

[0120] 1 H NMR (400MHz, CD 3 OD)δ7.44(d,J=15.7Hz,1H),7.05(d,J=1.8Hz,1H),6.93(dd,J=8.2,1.8Hz,1H),6.78(d,J=8.2Hz,1H),6.52(d,J= 15.7Hz,1H),4.75(dd,J=6.4,5.1Hz,1H),3.75(s,3H),2.99(dd,J=14.0,4.9Hz,1H),2.92(dd,J=14.0,6.7Hz,1H); 13 CNMR(101MHz,CD 3 OD)δ172.2,169.1,148.9,146.6,143.3,128.1,122.4,117.5,116.5,115.1,56.3,53.0,26.7; HRMS(ESI) for C 13 H 15 NO 5 SNa(M+Na + ):Calcd:320.0563,Found:320.0551.

[0121] The verification experiment of the use of the product prepared in this application is as follows:

[0122] Experiment 1:

[0123] Determination of the inhibitory rate of N-caffeoyl amino acid methyl ester derivatives on tyrosinase monophenolase.

[0124] The method for determining the inhibition rate of tyrosinase monophenolase was based on the research method of (Luo Tianji, Mei Zikun, Song Jingjiu, et al. Synthesis of isoferulamide derivatives and study on their whitening activity [J]. Surfactant Chemical Industry, 2021, 51(11): 1102-1108). The substrate was L-tyrosine and the determination was performed using an enzyme reader (model SpectraMax i3x): 0.2 M phosphate buffer (PBS) with a pH of 6.8 was used to prepare a 0.05% L-tyrosine solution and a 200 U / mL tyrosinase solution. The sample and the positive control kojic acid and vitamin C (Vc) were dissolved in dimethyl sulfoxide (DMSO) to prepare different concentration gradient solutions. PBS, L-tyrosine and sample solution were added to the 96-well plate in sequence, mixed evenly and incubated at 37°C in the dark for 10 minutes, then tyrosinase was added, and the solution was blown with a pipette to mix it thoroughly, and the absorbance value was measured at 475nm immediately after incubation for 25 minutes, and the measurement was repeated 3 times. The specific reagent ratio table is shown in Table 1. The inhibition rate when the sample concentration was 500 μg / mL was calculated using GraphPad Prism according to Formula 1, and the results are shown in Table 2.

[0125] Table 1 Reagent ratio table

[0126]

[0127] Inhibition rate = [(A a -A b) -(A c -A d ) / (A a -A b )]×100%(1)

[0128] Among them A a , A b , A c , A d is the absorbance value of groups a, b, c, and d.

[0129] Table 2 Tyrosinase monophenolase inhibition rate and IC 50 The results of the value

[0130]

[0131]

[0132] As can be seen from Table 2, different compounds have different inhibitory effects on tyrosinase monophenolase. When the mass concentration is 500 μg / mL, compounds a3, a12, a13, a14, a15, and b1 have good inhibitory activity, and the inhibition rate is greater than 50%. Among them, the inhibition ability of compounds a13 and a15 is significantly higher than that of other compounds, and the inhibition rates are 70.74% and 98.23%, respectively. The IC values ​​of compounds a13 and a15 are 50 The values ​​were 1.919 μM and 1.019 μM, respectively, which were significantly lower than kojic acid (IC 50 =2.528μM) and Vc (IC 50 =2.731 μM), especially compound a15, whose inhibitory ability was significantly higher than that of positive control kojic acid and vitamin C, can be used as a new type of tyrosinase inhibitor.

[0133] Experiment 2:

[0134] Determination of the inhibitory rate of N-caffeoyl amino acid methyl ester derivatives on tyrosinase diphenolase

[0135] The determination of tyrosinase diphenolase inhibition rate was based on the method of (Peng Z, Li Y, Tan L, et al. Anti-tyrosinase, antioxidant and antibacterial activities of gallic acid-benzylidenehydrazine hybrids and their application in preservation of fresh-cut apples and shrimps [J]. Food Chemistry, 2022, 378 (01): 132127) and slightly optimized: DMSO was used to dissolve the sample and the positive control kojic acid and Vc to prepare different concentration gradient solutions. A 2 mM L-dopa solution and a 200 U / mL tyrosinase solution were prepared with 0.2 M PBS buffer at pH = 6.8, and the sample and the positive control kojic acid and Vc were dissolved in DMSO. 20 μL sample solution, 30 μL tyrosinase and 100 μL phosphate buffer were added to the 96-well plate in sequence, mixed well and incubated at 25°C in the dark for 20 min, and then 50 μL L-DOPA was added to start the reaction. After 10 min of reaction, the absorbance value was measured at 475 nm. DMSO was used instead of the sample solution as the blank group, and each sample was measured in parallel 3 times. The inhibition rate when the sample concentration was 500 μg / mL was calculated using GraphPad Prism according to Formula 2, and the results are shown in Table 2.

[0136] Inhibition rate = (A 空白 -A 样品) / A 空白 ×100%(2)

[0137] Among them A 空白 , A 样品 is the absorbance value of the blank group and the sample group.

[0138] It can be concluded from Table 2 that in the experiment of inhibiting tyrosinase diphenolase, when the mass concentration was 500 μg / mL, the tyrosinase inhibitory activity of most compounds was low, but the inhibitory activity of compound a15 with the highest inhibition rate was significantly higher than that of the positive control kojic acid and Vc, indicating that N-caffeoyl amino acid methyl ester derivatives have the ability to inhibit tyrosinase diphenolase, and a15 has the strongest inhibitory ability.

[0139] Experiment 3:

[0140] Anti-browning experiment of N-caffeoyl amino acid methyl ester derivatives on apple slices:

[0141] The anti-browning experiment of apple slices was carried out according to the method of (Zhang Shaoying, Xia Hui. Effect of chitosan phytic acid composite coating on browning and quality of fresh-cut apples [J]. Zhejiang Agricultural Sciences, 2023, 64(08): 1972-1976). Fresh Fuji apples from the local market were selected, the apples were cleaned, cut into 10 mm thick slices along the direction of the apple core, and then immersed in distilled water (blank group), 0.01 g / mL compound (sample group), 0.01 g / mL kojic acid solution (control group), and 0.01 g / mL Vc (control group) solution for 10 min. After taking out, the surface of the apple slices was wiped dry with absorbent paper, placed in a culture dish, and stored in a refrigerator at 4°C. The brightness value L*, red-green value a*, and yellow-blue value b* of the apple slices were measured, and the total color difference ΔE value (Table 3) was calculated according to formula 3, and the appearance was recorded. The measurement was performed once a day and 4 parallel measurements were performed.

[0142]

[0143] Table 3 ΔE values ​​of fresh-cut apple slices treated with different samples

[0144]

[0145]

[0146] As can be seen from Table 3, the ΔE values ​​of apple slices treated with different treatments during storage showed different trends. The ΔE value of the blank group rose rapidly from the first day, and the color of the apple slices quickly darkened; the ΔE values ​​of apple slices treated with kojic acid, Vc and samples changed slowly. In particular, the ΔE value of apple slices treated with compound a15 rose significantly slower than that of the control group and the blank group, especially after the third day, the increase rate of the ΔE value of compound a15 was much lower than that of the other two groups. The above results show that compound a15 can effectively delay the browning of apple slices during short-term storage and has the potential to be used as an anti-browning agent for fresh-cut fruits.

[0147] Experiment 4:

[0148] Anti-browning experiment of N-caffeoyl amino acid methyl ester derivatives on apple juice:

[0149] The anti-browning experiment of apple juice refers to the literature method (Chen Chun, Li Jiayan, Huang Zepeng, et al. Effect of oxidized resveratrol on browning of fresh-cut apples and juice [J]. Food Industry Science and Technology, 2019, 20: 285-289, 295). Peel and core the apples, take about 1000g of pulp and use a kitchen juicer to squeeze the juice. The obtained apple juice is divided into a blank group (distilled water), a sample group (0.05g of compound dissolved in 100ml of apple juice) and a control group (0.05g of kojic acid dissolved in 100ml of apple juice, 0.05g of Vc dissolved in 100ml of apple juice). After adding the sample and mixing thoroughly, centrifuge at 5000r / min and 4℃ for 20min, and take the supernatant and store it in a refrigerator at 4℃. The absorbance of apple juice was measured at 420 nm (Table 4) to indicate its browning degree. The chromaticity (L*, a*, b*) was measured at the same time, the total color difference ΔE value was calculated (Table 5), and its appearance was recorded. The measurement was performed once a day and four times in parallel.

[0150] Table 4 Absorbance of apple juice at 420nm

[0151]

[0152]

[0153] Table 5 ΔE value of apple juice

[0154]

[0155] It can be seen from Table 4 that during the storage period, the browning degree of the compound sample groups was lower than that of the blank group, especially the browning degree of the compound a15 group was significantly lower than that of the other groups, and significantly lower than that of the kojic acid group (p<0.001) and the Vc group (p<0.001), indicating that the N-caffeoyl amino acid methyl ester derivative has an anti-browning effect on apple juice, and the anti-browning effect of a15 is very strong, and it has the potential to be used as an anti-browning agent for juice.

[0156] It can be seen from Table 5 that during the storage period, the ΔE of the compound sample groups was lower than that of the blank group, especially the ΔE of the compound a15 group was significantly lower than that of the other groups, and significantly lower than that of the kojic acid group (p<0.001) and the Vc group (p<0.001), indicating that N-caffeoyl amino acid methyl ester derivatives have anti-browning effects on apple juice, and a15 has a very strong anti-browning effect, and has the potential to be used as an anti-browning agent for juice.

Claims

1. A caffeoyl amino acid methyl ester derivative, characterized in that: Its general structure is as follows: Among them, R is Any of .

2. A method for preparing a caffeoyl amino acid methyl ester derivative, characterized in that: The specific steps are as follows: Step 1, dehydrating and deoxygenating the round-bottom flask used for the reaction; Step 2, under ice bath conditions, add caffeic acid into a round-bottom flask, and add solvent A to dissolve, after the caffeic acid is completely dissolved, add amino acid methyl ester hydrochloride, after the amino acid methyl ester hydrochloride is dissolved, add an organic base dropwise to obtain a mixed solution A; Step 3, the mixed solution A is mixed thoroughly, the condensing agent is added to the mixed solution A, stirred at room temperature for 2h-24h, the reaction is monitored by thin layer chromatography, and after the reaction is complete, 1mL of water is slowly added dropwise to quench the reaction to obtain a mixed solution B; Step 4, post-treating the mixed solution B, distilling the mixed solution B under reduced pressure to remove the organic solvent, adding ethyl acetate to fully dissolve, then washing with 1 mol / L hydrochloric acid solution, water, 5% sodium bicarbonate aqueous solution and saturated brine in sequence, drying the obtained organic phase with anhydrous sodium sulfate and filtering, collecting the filtrate, and concentrating the filtrate under reduced pressure to obtain a crude product; Step 5: purify the crude product to obtain a pure product.

3. The method for preparing the caffeoyl amino acid methyl ester derivatives according to claim 2, characterized in that: The amino acid methyl ester hydrochloride is any one of phenylalanine methyl ester hydrochloride, valine methyl ester hydrochloride, alanine methyl ester hydrochloride, glycine methyl ester hydrochloride, tryptophan methyl ester hydrochloride, threonine methyl ester hydrochloride, leucine methyl ester hydrochloride, isoleucine methyl ester hydrochloride, tyrosine methyl ester hydrochloride, N-tert-butyloxycarbonyllysine methyl ester hydrochloride, aspartic acid dimethyl ester hydrochloride, glutamate dimethyl ester hydrochloride, methionine methyl ester hydrochloride, and serine methyl ester hydrochloride.

4. The method for preparing the caffeoyl amino acid methyl ester derivatives according to claim 2, characterized in that: The solvent A is any one of N,N-dimethylformamide, tetrahydrofuran, and dichloromethane; the organic base is any one of triethylamine, pyridine, and 4-dimethylaminopyridine; and the condensing agent is any one of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride, Carter condensing agent, and benzotriazole-1-acyloxy-tris(dimethylamino)hexafluorophosphate.

5. The method for preparing the caffeoyl amino acid methyl ester derivatives according to claim 2, characterized in that: The molar ratio of caffeic acid to amino acid methyl ester hydrochloride in step 2 is 1:1 to 1.

5.

6. The method for preparing the caffeoyl amino acid methyl ester derivatives according to claim 2, characterized in that: The added amount of the solvent A is 10 ml per millimole of caffeic acid; the molar ratio of caffeic acid to the organic base is 1:1.5-3; and the molar ratio of caffeic acid to the condensing agent is 1:1-1.

5.

7. Application of caffeoyl amino acid methyl ester derivatives as anti-enzymatic browning agents for fruits and vegetables.

8. Application of caffeoyl amino acid methyl ester derivatives as tyrosinase inhibitors.

9. Application of caffeoyl amino acid methyl ester derivatives as therapeutic drugs for skin diseases with excessive melanin deposition.

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