CN107722035A - A kind of qinghaosu bridged piperazine derivatives and its preparation method and the application in medicines resistant to liver cancer is prepared - Google Patents

Abstract

The invention discloses a class of artemisinin-piperazine derivatives, a preparation method thereof and the application of the derivatives in the preparation of anti-liver cancer drugs. The method uses triethylamine as an acid-binding agent to prepare artemisinin-piperazine derivatives through a "one-pot method". The reaction conditions are mild, the operation is simple, and the yield is good. In addition, the prepared derivatives have obvious inhibitory effect on human liver cancer SMMC‑7721 cells in vitro, which is equivalent to the inhibitory effect of vincristine, a first-line anti-hepatic cancer clinical chemotherapy drug. The present invention provides a feasible strategy for the application of artemisinin-piperazine derivatives in the preparation of anti-liver cancer drugs.

Description

A class of artemisinin-piperazine derivatives and its preparation method and its use in the preparation of anti-liver cancer drugs application

technical field

The invention relates to a class of artemisinin-piperazine derivatives, a preparation method thereof and an application in preparation of anti-liver cancer drugs.

Background technique

Artemisinin has good antimalarial activity, but there are few studies on its antitumor activity. Although artemisinin and its derivatives have been used clinically for many years, they still have poor oil solubility and water solubility, poor thermal stability, are easily decomposed by moisture, heat and reducing substances, and have high clinical recurrence rate. Application is limited. Therefore, it is a meaningful work to design and prepare a series of new artemisinin derivatives through chemical modification, and to investigate their antitumor activity.

Piperazine can improve the biological activity of drugs by forming multiple hydrogen bonds or ionic bonds, and can also regulate the solubility and acid-base balance of drugs, promote the pharmacokinetics of drugs, and have obvious effects in antitumor. Therefore, it is often introduced into drug molecules as a type of synergistic group.

Carbamates are a class of important sulfur-containing compounds with unique structures and properties, and occupy an important position in synthetic drugs. It has a wide range of biological activities, such as antibacterial, antiviral, antioxidative, and treatment of chronic alcoholism and heavy metal poisoning. In recent years, more and more studies have found that carbamates have good tumor prevention and antitumor activity, which has aroused the research interest of scholars at home and abroad.

In the present invention, the artemisinin-piperazine derivative is prepared by introducing the carbamate structure into the artemisinin molecule by using piperazine as a linker under mild conditions. The growth of human liver cancer cells (SMMC-7721) has obvious inhibitory effect, so the prospect of artemisinin-piperazine derivatives as potential anti-hepatoma drugs is worthy of attention.

Contents of the invention

In view of the above, the object of the present invention is to provide a class of artemisinin-piperazine derivatives. The preparation method of the derivatives is simple and has high anti-liver cancer activity.

Another object of the present invention is to provide a preparation method of artemisinin-piperazine derivatives.

Another object of the present invention is to provide the application of a class of artemisinin-piperazine derivatives in anti-liver cancer.

The purpose of the present invention is to realize through the following technical solutions:

(1). A class of artemisinin-piperazine derivatives, characterized in that it has the following general formula:

where R is:

(2). The preparation method of artemisinin-piperazine derivatives:

Dissolve artemisinin-piperazine and carbon disulfide in an amount four times that of artemisinin-piperazine in acetonitrile, then add triethylamine in an amount ten times that of artemisinin-piperazine, stir at room temperature for 10 minutes, and then add the amount of Halide acetonitrile solution twice the amount of artemisinin-piperazine, gradually raised to 60°C, reacted for 4-12 hours, TLC tracking, phosphomolybdic acid color development, after the reaction, evaporated to remove the solvent, silica gel column chromatography separation and purification (PE:EA=16:1～6:1) to obtain artemisinin-piperazine derivatives.

The artemisinin-piperazine derivative reaction is expressed as follows with a chemical formula:

Advantage of the present invention and the beneficial effect that produce are:

1. The present invention uses triethylamine as an acid-binding agent to efficiently prepare the artemisinin-piperazine through a series reaction of multiple components under the condition of "one-pot cooking". The reaction conditions are mild, and the experimental operation is simple and feasible. It does not have to be carried out under harsh anhydrous and oxygen-free conditions, has no metal reagents, is environmentally friendly, and has high atom utilization.

2. The artemisinin-piperazine derivatives of the present invention have obvious inhibitory effect on the growth of human liver cancer SMMC-7721 cells, so the artemisinin-piperazine derivatives of the present invention can be applied to the preparation of anti-liver cancer drugs.

detailed description

The present invention is further described in detail by the following examples, but the scope of the present invention is not limited by these examples.

Example 1: Preparation of Artemisinin-Piperazine-Benzyl Aminodithiocarbamate (Compound 1)

At room temperature, successively add (1.2mmol, 0.42g) artemisinin piperazine, (4.8mmol, 0.36g) carbon disulfide, 20mL acetonitrile and (12mmol, 1mL) triethylamine into the round bottom flask, stir for 10 minutes, then add 5mL of acetonitrile solution dissolved with (2.5mmol, 0.43g) benzyl bromide was gradually heated to 60°C and reacted for 12h. After the reaction was followed by TLC, the solvent was evaporated and the silica gel column chromatography [eluent: V (petroleum ether) : V (ethyl acetate) = 16:1] purified to obtain the target compound. White solid, yield 72.6%, mp130.4-130.8℃, ¹ H NMR (400MHz, CDCl ₃ )δ: 7.37(t, J=12.6Hz, 2H), 7.32–7.26(m, 3H), 5.29(d, J=10.7Hz, 1H), 4.56(s, 2H) ,4.35(s,2H),4.07(d,J=9.8Hz,1H),3.89(s,2H),3.07(s,2H),2.79(s,2H),2.58(s,1H),2.34( td,J=13.8,3.5Hz,1H),2.00(d,J=14.4Hz,1H),1.94–1.81(m,1H),1.78–1.65(m,2H),1.55(d,J=12.8Hz ,2H),1.49–1.15(m,5H),1.03(dd,J=18.0,7.7Hz,2H),0.95(d,J=6.1Hz,3H),0.83(d,J=6.5Hz,3H) ; ¹³ C NMR (101MHz, CDCl ₃ ) δ: 196.01, 135.90, 129.36, 128.54, 127.44, 103.98, 91.54, 90.35, 80.19, 77.20, 51.63, 46.92, 45.70, 42.07, 35.35, 320.25, 28.35 24.70, 21.57, 20.25, 13.46; IR(KBr)ν/cm ^-1 : 2981, 2941, 2866, 1600, 1491, 1447, 1378, 1310, 1217, 1161, 1101, 1039, 999, 955, 926, 880, 842, 741, 712, 6 for C ₂₇ H ₃₈ N ₂ NaO ₄ S ₂ 541.2168,found 541.2170[M+Na] ⁺ .

Example 2: Preparation of artemisinin-piperazine-aminodithiocarbamate-(2-methyl)benzyl ester (compound 2)

The preparation method was the same as that in Example 1, and benzyl bromide was replaced by o-methylbenzyl bromide to obtain the target compound as a light yellow solid with a yield of 55.6%, mp87.4-88.4°C, ¹ H NMR (400MHz, CDCl ₃ )δ: 7.34(d, J=7.1Hz, 1H), 7.16(tt, J=6.2, 3.7Hz, 3H), 5.27(s, 1H), 4.51(s, 2H) ,4.35(s,2H),4.06(d,J=10.2Hz,1H),3.89(s,2H),3.06(s,2H),2.78(s,2H),2.57(dqd,J=11.2,7.3 ,4.7Hz,1H),2.44–2.27(m,4H),1.99(ddd,J＝14.6,5.0,2.9Hz,1H),1.86(ddt,J＝13.6,6.7,3.4Hz,1H),1.70( dp,J=10.6,3.4Hz,2H),1.60–1.38(m,5H),1.38–1.16(m,4H),1.08–0.91(m,3H),0.82(d,J=7.1Hz,3H) ; ¹³ C NMR (101MHz, CDCl ₃ ) δ: 196.11, 137.36, 133.17, 130.40, 130.38, 127.85, 126.13, 103.92, 91.49, 90.29, 80.14, 51.58, 50.16, 46.87, 45.65, 40.20, 33, 33, 28.45, 25.93, 24.65, 21.52, 20.22, ^19.28 , 13.42; 982,879,734,550,510; HRMS m/z:calcd for C ₂₈ H ₄₁ N ₂ O ₄ S ₂ 533.2429,found 533.2514[M+H] ⁺ .

Example 3: Preparation of artemisinin-piperazine-aminodithiocarbamate-(4-methyl)benzyl ester (compound 3)

The preparation method is the same as in Example 1, and 4-methylbenzyl bromide is used instead of benzyl bromide to obtain the target compound. White solid, yield 50.3%, mp135.9-136.9℃, ¹ H NMR (400MHz, CDCl ₃ ) δ: 7.27(d, J=7.5Hz, 2H), 7.12(d, J=7.8Hz, 2H), 5.27(s, 1H), 4.52(s, 2H), 4.35 (s,2H),4.06(d,J=10.3Hz,1H),3.91(s,2H),3.05(s,2H),2.78(s,2H),2.57(ddd,J=10.9,7.2,4.3 Hz,1H),2.35–2.29(m,4H),2.00(ddd,J＝14.4,4.9,3.0Hz,1H),1.86(ddt,J＝13.6,6.8,3.6Hz,1H),1.70(ddd, J＝13.0, 7.3, 3.5Hz, 2H), 1.59(s, 2H), 1.55(dt, J＝13.8, 4.4Hz, 1H), 1.49–1.40(m, 1H), 1.38(s, 2H), 1.32 (td,J=13.4,3.6Hz,1H),1.22(td,J=11.3,6.5Hz,1H),1.00(dd,J=12.1,3.7Hz,1H),0.94(d,J=6.2Hz, 3H), 0.82 (d, J=7.1Hz, 3H); ¹³ C NMR (101MHz, CDCl ₃ ) δ: 196.19, 137.20, 132.67, 129.28, 129.26, 104.00, 91.55, 90.36, 80.20, 77.20, 51.65, 46.93, 45.72, 41.91, 37.36, 36.26, 34.21, ^28.52 , 25.97, 24.71, 21.59, 21.13, 20.26, 13.46; 1230,1135,1055,980,879,825,741,487; HRMS m/z: calcd for C ₂₈ H ₄₀ NaN ₂ O ₄ S ₂ 555.2429,found 555.2338[M+Na] ⁺ .

Example 4: Preparation of artemisinin-piperazine-aminodithiocarbamate-(2-chloro)benzyl ester (compound 4)

The preparation method was the same as that in Example 1, and benzyl bromide was replaced by o-chlorobenzyl chloride, and the reaction was heated for 4 hours to obtain the target compound, a pink solid, with a yield of 50.6%, mp90.3-91°C, ¹ H NMR (400MHz, CDCl ₃ ) δ: 7.56 (dd, J = 5.8, 3.6 Hz, 1H), 7.37 (dd, J = 5.7, 3.6 Hz, 1H), 7.21 (dd, J = 5.9, 3.5 Hz, 2H),5.27(s,1H),4.72(s,2H),4.35(s,2H),4.06(d,J=10.2Hz,1H),3.91(s,2H),3.50(s,1H), 3.06(s,2H),2.78(s,2H),2.57(ddd,J=10.9,7.2,4.4Hz,1H),2.34(td,J=13.9,3.9Hz,1H),2.03–1.96(m, 1H),1.91–1.82(m,1H),1.71(ddd,J＝12.4,7.3,3.5Hz,2H),1.56(s,2H),1.48–1.41(m,1H),1.38(s,3H) ,0.94(d,J=6.2Hz,3H),0.82(d,J=7.1Hz,3H); ¹³ C NMR(101MHz,CDCl ₃ )δ:195.76,134.45,134.18,131.48,129.46,128.89,126.87, ^104.01 , 91.52, 90.32, 80.21, 77.20, 51.58, 50.55, 45.66, 39.42, 37.30, 36.19, 34.13, 28.48, 25.86, 24.63, 21.53, 20.18, 13.40; 1885,1472,1404,1275,1231,1208,1185,1130,1053,983,926,879,826,743,550; HRMS m/z:calcd for C ₂₇ H ₃₇ ClNaN ₂ O ₄ S ₂ 575.1776,found 575.1778[M ⁺ Na]

Example 5: Preparation of artemisinin-piperazine-aminodithiocarbamate-(4-chloro)benzyl ester (compound 5)

The preparation method was the same as that in Example 1, and benzyl bromide was replaced by p-chlorobenzyl bromide to obtain the target compound, a white solid, with a yield of 71.5%, mp111.3-112.2°C, ¹ H NMR (400MHz, CDCl ₃ ) δ: 7.35–7.30(m, 2H), 7.28(d, J=2.3Hz, 2H), 5.27(s, 1H), 4.54(s, 2H), 4.34(s, 2H), 4.07(d, J=10.2Hz, 1H), 3.97–3.83(m, 2H), 3.49(d, J=5.0Hz, 1H), 3.06(s, 2H), 2.78(s, 2H), 2.57(ddd, J＝11.0, 7.1, 4.3Hz, 1H), 2.34(td, J＝13.9, 3.9Hz, 1H), 2.00(dt, J＝14.6, 3.8Hz, 1H), 1.90–1.82(m, 1H), 1.71(ddd, J＝12.5, 7.4, 3.5Hz, 2H), 1.58–1.42(m, 3H), 1.38(s, 4H), 1.04–0.97(m, 1H), 0.95(d, J＝ 6.2Hz, 3H), 0.82 (d, J=7.1Hz, 3H); ¹³ C NMR (101MHz, CDCl ₃ ) δ: 195.48, 134.85, 133.20, 130.67, 128.64, 104.00, 91.56, 90.35, 80.20, 77.20, 51.63 , ^{50.90,45.70,41.02,37.36,36.25,34.19,28.51,25.98,24.69,21.58,20.26,13.46} ; ,1159,1104,1052,978,926,879; HRMS m/z:calcd for C ₂₇ H ₃₇ ClNaN ₂ O ₄ S ₂ 575.1780,found 575.1782[M+Na] ⁺ .

Example 6: Preparation of artemisinin-piperazine-aminodithiocarbamate-(4-bromo)benzyl ester (compound 6)

The preparation method was the same as that in Example 1, with p-bromochlorobenzyl instead of bromobenzyl to obtain the target compound, a white solid, with a yield of 55.7%, mp136.9-138°C, ¹ H NMR (400MHz, CDCl ₃ )δ: 7.45–7.40(m,2H),7.30–7.24(m,4H),5.27(s,1H),4.53(s,2H),4.34(s,2H), 4.06(d, J=10.3Hz, 1H), 3.90(d, J=27.5Hz, 2H), 3.06(s, 2H), 2.78(s, 2H), 2.57(ddd, J=10.7, 7.2, 4.3Hz ,1H),2.38–2.29(m,1H),2.00(ddd,J＝14.5,4.9,2.9Hz,1H),1.86(ddt,J＝10.1,7.0,3.4Hz,1H),1.75–1.66(m ,1H),1.59–1.46(m,2H),1.38(s,3H),1.30(dd,J＝13.7,3.7Hz,1H),1.22(td,J＝11.2,6.5Hz,1H),1.07– 0.98(m,1H),0.95(d,J=6.2Hz,3H),0.82(d,J=7.2Hz,3H); ¹³ C NMR(101MHz,CDCl ₃ )δ:195.44,135.42,131.59,131.01, 121.32, 104.00, 91.56, 90.35, 80.20, 77.20, ^51.64 , 46.94, 45.71, 41.04, 37.36, 36.25, 34.20, 28.51, 25.98, 24.70, 21.58, 20.25, 13.46; 2865,1476,1425,1378,1305,1276,1229,1158,1042,1020,979,931,879,826,609; HRMS m/z: calcd for C ₂₇ H ₃₈ BrN ₂ O ₄ S ₂ 597.1378,found 597.1465[M+H] ⁺

Example 7: Preparation of artemisinin-piperazine-carbamodithioate-(4-cyano)benzyl ester (compound 7)

The preparation method is the same as in Example 1, p-cyanobenzyl bromide is substituted for benzyl bromide, the reaction is heated for 4 hours, and purified by silica gel column chromatography [eluent: V (petroleum ether): V (ethyl acetate) = 12: 1] to obtain light Yellow solid, yield 65.8%, mp146.7-147.3℃, ¹ H NMR (400MHz, CDCl ₃ ) δ: 7.62–7.57(m,2H),7.52–7.48(m,2H),5.27(s,1H),4.65(s,2H),4.34(s,2H), 4.07(d, J=10.3Hz, 1H), 3.91(d, J=29.7Hz, 2H), 3.07(s, 2H), 2.79(s, 2H), 2.57(ddd, J=10.9, 7.3, 4.4Hz ,1H),2.34(ddd,J＝14.5,13.3,4.0Hz,1H),2.05–1.96(m,1H),1.86(ddt,J＝13.6,6.7,3.6Hz,1H),1.75–1.67(m ,2H),1.57(d,J＝1.9Hz,1H),1.55–1.46(m,1H),1.38(s,4H),1.34–1.27(m,1H),1.27–1.19(m,1H), 1.02(td, J＝13.1, 12.2, 4.0Hz, 1H), 0.95(d, J＝6.2Hz, 3H), 0.82(d, J＝7.2Hz, 3H); ¹³ C NMR (101MHz, CDCl ₃ )δ :194.67,142.62,132.19,129.98,118.78,111.03,104.01,91.56,90.34,80.20,77.20,51.62,50.87,45.69,40.78,37.35,36.23,34.18,28.50,25.97,24.68,21.57,20.24,13.45；IR (KBr)ν/cm ^-1 :3140,2937,2864,2229,1736,1606,1477,1378,1305,1276,1158,1051,980,926,880,543cm ^-1 ; HRMS m/z: calcd for C ₂₈ H ₃₇ NaN ₃ O ₄ S ₂ 566.2116,found 566.2118[M+Na] ⁺ .

Example 8: Preparation of artemisinin-piperazine-carbamodithioate-(4-nitro)benzyl ester (compound 8)

The preparation method is the same as in Example 1, and 4-nitrobenzyl bromide is used instead of benzyl bromide to obtain the target compound, a yellow solid, with a yield of 67.6%, mp82.2-82.9°C, ¹ H NMR (400MHz, CDCl ₃ ) δ: 8.19–8.13(m,2H), 7.59–7.54(m,2H), 5.27(s,1H), 4.69(s,2H), 4.34(s,2H), 4.07(d,J=10.3Hz,1H),3.92(d,J=31.3Hz,2H),3.07(s,2H),2.80(s,2H),2.57(ddd,J=10.9,7.2,4.3Hz ,1H),2.39–2.29(m,1H),2.00(dt,J＝14.4,4.1Hz,1H),1.90–1.82(m,1H),1.71(ddd,J＝12.7,7.6,3.6Hz,2H ),1.59–1.46(m,2H),1.42(s,1H),1.38(s,3H),1.34–1.18(m,2H),1.07–0.92(m,4H),0.90–0.78(m,3H ); ¹³ C NMR (101MHz, CDCl ₃ ) δ: 196.96, 194.68, 142.62, 132.20, 129.98, 111.04, 104.02, 103.98, 91.56, 90.34, 80.21, 77.20, 51.63, 50.88, 45.63, 37.16, 40.79, , ^{34.20,31.16,28.51,28.32,25.97,24.69,22.28,21.58,20.25,13.96,13.46} ; ,1275,1159,1041,983,879,724cm ^-1 ; HRMS m/z:calcd for C ₂₇ H ₃₇ N ₃ NaO ₆ S ₂ 586.2016,found586.2018[M+Na] ⁺ .

Example 9: Preparation of artemisinin-piperazine-aminodithiocarbamate-2-naphthylmethyl ester (compound 9)

The preparation method is the same as in Example 1, with 2-bromomethylnaphthalene instead of benzyl bromide, purified by silica gel column chromatography [eluent: V (petroleum ether): V (ethyl acetate) = 12:1] to obtain a white solid, Yield 60.3%, mp143.8-144.0℃, ¹ H NMR (400MHz, CDCl ₃ ) δ: 7.86–7.77(m,5H), 7.51–7.42(m,3H), 5.27(s,1H), 4.74(s,2H), 4.37(s,2H), 4.07(d,J=10.2Hz,1H),3.93(s,2H),3.07(s,2H),2.79(s,2H),2.57(tt,J=11.2,6.9Hz,1H),2.38–2.29 (m,1H),1.99(dd,J=14.4,4.2Hz,1H),1.86(ddd,J=13.6,6.6,3.4Hz,1H),1.71(ddd,J=12.7,7.6,3.6Hz,2H ),1.58(s,3H),1.40(d,J＝16.5Hz,4H),1.34–1.18(m,1H),1.07–0.90(m,3H),0.90–0.78(m,3H); ¹³ C NMR(101MHz,CDCl ₃ )δ:195.88,133.41,133.26,132.64,128.27,128.11,127.70,127.59,127.27,126.11,125.87,103.96,91.52,90.33,80.17,77.20,51.61,46.91,45.68,42.28,40.07 , ^{37.32,36.23,34.17,28.49,26.85,25.95,24.67,21.55,20.23,13.44} ; ,1130,1020,982,879,755,474; HRMS m/z:calcd for C ₃₁ H ₄₀ NaN ₂ O ₄ S ₂ 591.2429,found 591.2342[M+Na] ⁺ .

Example 10: Preparation of Artemisinin-Piperazine-Amyl Amyl Dithiocarbamate (Compound 10)

The preparation method was the same as that in Example 1, using pentane chloride instead of benzyl bromide, and heating for 10 hours to obtain the target compound as a light yellow solid with a yield of 58.0%, mp 135.8-136.2°C. ¹ H NMR (400MHz, CDCl ₃ ) δ: 5.25(s, 1H), 4.32(s, 2H), 4.05(d, J=10.2Hz, 1H), 4.00–3.82(m, 2H), 3.27(t, J＝7.4Hz, 2H), 3.04(dt, J＝11.8, 5.2Hz, 2H), 2.76(p, J＝4.9Hz, 2H), 2.56(ddt, J＝10.1, 7.1, 3.6Hz, 1H), 2.32(td, J=13.9,3.9Hz,1H),1.98(ddd,J=14.5,5.0,3.0Hz,1H),1.84(ddt,J=13.6,6.8,3.5Hz,2H),1.69(ddt, J＝14.8, 10.2, 5.4Hz, 5H), 1.53(dt, J＝13.6, 4.3Hz, 1H), 1.47–1.26(m, 8H), 1.21(td, J＝11.2, 6.4Hz, 1H), 1.07 –0.96 (m, 1H), 0.96 – 0.85 (m, 6H), 0.81 (d, J＝7.1Hz, 3H); ¹³ C NMR (101MHz, CDCl ₃ ) δ: 196.89, 103.92, 91.51, 90.29, 80.16, 51.60, 50.11, 46.87, 45.68, 37.31, 37.11, 36.22, 34.17, 31.12, 28.48, 28.28, 25.93, 24.66, 22.24, 21.55, 20.23, 13.93, 13.44; IR(KBr)ν/cm ^-1 :37932, 2937,1755,1423,1388,1277,1233,1160,1131,1022,999,932,881cm ^-1 ; HRMS m/z:calcd for C ₂₅ H ₄₂ N ₂ NaO ₄ S ₂ 521.2477,found 521.2479[M+Na] ⁺ .

Embodiment 11: Preparation of artemisinin-piperazine-octadecyl aminodithiocarbamate (compound 11)

The preparation method was the same as in Example 1, and 1-chlorooctadecane was used instead of benzyl bromide to obtain the target compound as a yellow waxy solid with a yield of 49.3%. ¹ H NMR (400MHz, CDCl ₃ ) δ: 5.25(s, 1H), 4.32(s, 2H), 4.05(d, J=10.2Hz, 1H), 3.27(t, J=7.5Hz, 2H), 3.04 (dt,J=11.0,4.9Hz,2H),2.76(dt,J=11.3,4.8Hz,2H),2.56(ddd,J=11.2,7.2,4.2Hz,1H),2.32(td,J=13.9 ,3.9Hz,1H),1.98(ddd,J＝14.5,4.9,2.9Hz,1H),1.84(ddt,J＝13.5,6.7,3.5Hz,1H),1.74–1.63(m,4H),1.53( dt,J=13.9,4.4Hz,1H),1.30(d,J=49.8Hz,39H),1.01(td,J=13.1,12.3,4.0Hz,1H),0.93(d,J=6.2Hz,3H ),0.89–0.77(m,6H); ¹³ C NMR(101MHz,CDCl ₃ )δ:196.84,103.87,91.47,90.27,80.12,51.58,46.87,45.66,37.28,37.12,36.20,34.16,31.84,29.62, 29.60, 29.58, 29.53, 29.44, 29.29, ^29.15 , 28.97, 28.56, 28.45, 25.90, 24.64, 22.61, 21.52, 20.20, 14.07, 13.41; 1376,1310,1275,1231,1130,1055,983,880,834,721,551; HRMS m/z:calcd for C ₃₈ H ₆₉ N ₂ O ₄ S ₂ 681.4621,found 681.4713[M+H] ⁺ .

Example 12: Preparation of artemisinin-piperazine-carbamodithioate-(benzyloxy)methyl ester (compound 12)

The preparation method is the same as in Example 1, using benzyl bromoacetate instead of benzyl bromide, and purifying by silica gel column chromatography [eluent: V (petroleum ether): V (ethyl acetate) = 6:1] to obtain an orange-yellow solid, the product Rate 35%, mp82.3-83.3℃, ¹ H NMR (400MHz, CDCl ₃ ) δ: 7.40–7.36 (m, 4H), 7.33 (dddd, J=11.4, 4.9, 2.9, 1.8Hz, 1H), 5.28 (s, 1H), 5.19 (s, 2H) ),4.31(s,2H),4.21(s,2H),4.07(d,J=10.2Hz,1H),3.95(d,J=32.2Hz,2H),3.08(dt,J=11.3,5.0Hz ,2H),2.80(dt,J＝11.5,5.2Hz,2H),2.63–2.53(m,1H),2.34(ddd,J＝14.5,13.3,3.9Hz,1H),2.05–1.96(m,1H ), 1.87(ddd, J＝13.6, 6.6, 3.4Hz, 1H), 1.71(ddt, J＝11.2, 6.5, 3.5Hz, 2H), 1.60(s, 1H), 1.55(dt, J＝13.6, 4.3 Hz,1H),1.49–1.41(m,1H),1.38(s,3H),1.35–1.18(m,2H),1.07–0.98(m,1H),0.95(d,J＝6.2Hz,3H) , 0.83 (d, J=7.1Hz, 3H); ¹³ C NMR (101MHz, CDCl ₃ ) δ: 194.32, 168.57, 135.47, 128.52, 128.30, 128.29, 104.01, 91.54, 90.36, 80.20, 67.46, 51.64, 46.91, 45.71, ^38.91 , 37.36, 36.26, 34.21, 28.53, 26.88, 25.97, 24.71, 21.59, 20.26, 13.46; 980,925,878,741,697,550; HRMS m/z:calcd for C ₂₉ H ₄₀ NaN ₂ O ₆ S ₂ 599.2328,found 599.2246[M+Na] ⁺ .

Example 13: Anti-human liver cancer SMMC-7721 cell activity research of artemisinin-piperazine derivatives in vitro

Using MTT (tetramethylazozolium salt colorimetric method), select artemisinin-piperazine derivatives (compounds 1-12) to determine their inhibitory rate on human liver cancer SMMC-7721 cells, and calculate the IC ₅₀ value (μM ). The specific operation steps are as follows:

1. Cell Recovery

Cell culture medium: 1640+10%FBS+1% (Penicillin-Streptomycin Solution)

(1) Take the SMMC7721 cells out of the liquid nitrogen, quickly put them into a 37°C water bath, and gently shake the cryopreservation tube to dissolve the cryopreservation solution;

(2) After dissolving, transfer the cells to centrifuge tubes containing 5 mL of medium, collect the cells by centrifugation, centrifuge at 1000 rpm for 5 min at room temperature, and discard the supernatant;

(3) Suspend the cells in a complete medium containing 10% fetal bovine serum, inoculate into a petri dish, gently blow and beat to mix, and culture at 37° C., 5% CO ₂ , and saturated humidity.

2. Cell passage

Cells were passaged when the density of the cells reached 80%:

(1) Discard the culture medium and wash it once with PBS;

(2) Add 1-2mL 0.25% trypsin to digest the cells, observe under a microscope, digest for 30-60s, you can see that the cells are separated from each other and become round, that is, the digestion is complete;

(3) Quickly discard trypsin, add complete medium, pipette the cells to make a single cell suspension, pass passage at a ratio of 1:3, and expand culture at 37°C, 5% CO ₂ , and saturated humidity.

3. Cell processing and MTT detection

(1) Take the SMMC7721 cells in the logarithmic growth phase and in good growth state, adjust the cell density to ¹ ×104 cells/mL with 1640 medium, insert them into a 96-well plate, and set up a blank group at the same time with 100 μL of cell suspension , cultivate overnight at 37°C (add 100 μL sterile PBS to the wells around the cell wells);

(2) 24 hours after the cells adhere to the wall and grow well, absorb the old culture solution, and add the test solution of different concentrations of compounds 1-12 to each culture well. Set 5 parallel repeat wells for each concentration, set equal volume of dimethyl sulfoxide (DMSO) solvent and blank control wells without drug medium at the same time, continue culturing in an incubator with 5% _CO at 37°C.

After culturing for 24h, 48h, and 72h respectively, discard the supernatant, add 10μL (2mg/mL in PBS) MTT to each well, continue to culture for 4h, suck out the culture supernatant in the well, and add 150μL dimethyl sulfoxide to each well After shaking for 10 minutes to fully dissolve the blue-purple crystals, measure the absorbance value (OD value) of each well sample with a microplate reader at a wavelength of 568 nm, remove the highest value and the lowest value, and take the average value of the remaining values.

Calculation of inhibition rate: the inhibition rate of cell growth was calculated according to the following formula:

Inhibition rate (%)=[1-(OD value of test sample-OD value of blank)/(OD value of negative control-OD value of blank)]×100%

Table 1 The inhibitory rate of artemisinin-piperazine derivatives 1-12 to human liver cancer SMMC-7721 cells

Table 2 Artemisinin-piperazine derivatives 1-12 inhibit human liver cancer SMMC-7721 cells _IC50 value (μM)

[a] Zhao Chenyang, Qiu Rong, Zheng Rongliang. Journal of Lanzhou University (Natural Science Edition), 2000, 36(4), 66-68. (Vincristine: IC ₅₀ ＝63.2±1.8μg/mL＝0.074±0.002μM) ; [b] JJLu, LHMeng, YJCai, et al. Cancer. Biol. Ther. 2008, 7, 1017-1023.

Claims (3)

1. a kind of qinghaosu-bridged piperazine derivatives, it is characterized in that it has below formula：

R is in formula：

2. the preparation method of a kind of qinghaosu-bridged piperazine derivatives as claimed in claim 1：It is characterized in that by qinghaosu-piperazine It is that four times of carbon disulfide measured of qinghaosu-piperazine are dissolved in acetonitrile with dosage, it is ten times of amounts of qinghaosu-piperazine then to add dosage Triethylamine, after 10min is stirred at room temperature, addition dosage is the halides acetonitrile solution of qinghaosu-piperazine doubling dose, is gradually increased to 60 DEG C, 4~12h is reacted, TLC tracking, phosphomolybdic acid colour developing, after reaction terminates, solvent, silica gel column chromatography separating purification (PE is evaporated off:EA =16:1~6:1) qinghaosu-bridged piperazine derivatives, are obtained.

3. application of a kind of qinghaosu-bridged piperazine derivatives in medicines resistant to liver cancer is prepared.