Abietane type diterpene compound, preparation method and application thereof

Abstract

Disclosed are an abietane type diterpene compound, a preparation method and an application thereof, relating to the technical field of anti-tumor compounds, where the compound has a chemical structure as shown in the following formula I: ##STR00001##
Leucosceptrum canum is extracted to prepare the abietane type diterpene compound according to the present application, and the prepared compound is effectively applied in inhibiting human lung cancer cell A549 and human myeloid leukemia cell HL-60.

Claims

1. A preparation method of the abietane type diterpene compound of formula (I): ##STR00003## comprising: S1, taking air-dried Leucosceptrum canum as raw materials, crushing, leaching with organic solvent, and concentrating under a reduced pressure at a low temperature to obtain an extract; S2, mixing the extract with water to obtain a suspension, adding ethyl acetate for extraction, and concentrating under the reduced pressure at the low temperature to obtain an ethyl acetate extracting phase; S3, separating the ethyl acetate extracting phase by silica gel column chromatography, and performing gradient elution with petroleum ether-ethyl acetate as an eluent, where the petroleum ether and the ethyl acetate are in a volume ratio of (9.8-10.2):1, (4.8-5.2):1, and (2.8-3.2):1; collecting the eluent corresponding to the volume ratio of petroleum ether to ethyl acetate of (2.8-3.2):1, followed by thin-layer chromatographic analysis and combining similar fractions to obtain initial components; and S4, separating the initial components by preparative liquid chromatography, and performing isocratic elution by using acetonitrile and water as an eluant to obtain the abietane type diterpene compound.

2. The preparation method of the abietane type diterpene compound according to claim 1, wherein the ethanol in the S1 is in a volume-mass ratio of 10-20 milligrams (mL):1 gram (g) to the air-dried Leucosceptrum canum; and the leaching is carried out under temperature of 65-75 degree Celsius (? C.) for 2-3 times, each time with a duration of 4-6 hours (h).

3. The preparation method of the abietane type diterpene compound according to claim 1, wherein the concentrating under the reduced pressure at the low temperature in both the S1 and the S2 is carried out under a temperature of 40-50? C. with a vacuum degree of 0.07-0.09 Megapascal (MPa).

4. The preparation method of the abietane type diterpene compound according to claim 1, wherein the extract and water in the S2 are mixed in a mass ratio of 1:1-3, the ethyl acetate is in a volume ratio of 0.8-1:1 to the suspension, and the extraction is carried out for 2-4 times.

5. The preparation method of the abietane type diterpene compound according to claim 1, wherein the eluent in the S4 comprises 40-50% by volume of acetonitrile, and the rest is water.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) FIG. 1 shows a process of preparing an abietane type diterpene compound Leucosceptrum A.

DETAILED DESCRIPTION OF THE EMBODIMENTS

(2) The present application is hereinafter described in detail in conjunction with the embodiments and the enclosed drawing.

(3) Unless otherwise specified, the materials used in the present application are commercially available or commonly used in the art, and the methods in the following embodiments are conventional in the art if not specified.

(4) Embodiment 1 Physical and chemical properties test of abietane type diterpene compound Leucosceptrum A

(5) 1.1 Determination of Molecular Structural Formula

(6) The Nuclear Magnetic Resonance (NMR) instrument used is a Bruker AVANCE III 500 NMR instrument manufactured by the Bruker Corporation, and the NMR is performed using the reagent of deuterated CDCl.sub.3 (deuterated chloroform); the hydrogen spectrum and carbon spectrum data of the abietane type diterpene compound Leucosceptrum A is shown in Table 1 below.

(7) TABLE-US-00001 TABLE 1 .sup.1H-NMR (500 megahertz (MHz)) and .sup.13C-NMR (125 MHz) data of Leucosceptrum A S/N ?.sub.H ?.sub.C 1 1.08, m, ? 36.6, CH.sub.2 1.94, m, ? 2 1.71, m, ? 19.4, CH.sub.2 1.51, m, ? 3 2.22, dd (14.0, 6.5), ? 41.3, CH.sub.2 1.67, m, ? 4 33.1, qC 5 1.96, d (14.0) 49.6, CH 6 2.74, m, ? 36.1, CH.sub.2 2.72, m, ? 7 205.8, qC 8 115.9, qC 9 137.8, qC 10 40.2, qC 11 144.3, qC 12 6.6, s 124.4, CH 13 119.2, qC 14 155.7, qC 15 143.1, qC 16 5.33, br s 111.2, CH.sub.2 5.02, br s 17 2.10, s 22.0, CH.sub.3 18 1.24, s 32.9, CH.sub.3 19 1.06, s 21.6, CH.sub.3 20 1.25, s 18.0, CH.sub.3 Note: ? stands for a chemical shift; s represents a singlet peak; br s represents a broad singlet peak; m represents multiplet; and dd stands for double of doubles.

(8) 1.2 Molecular Weight Test

(9) The compound is determined in terms of exact molecular weight by high-resolution mass spectrometry HR-ESI-MS, and HIR-ESI-MS [M-H].sup.? is 313.1804; as determined from the NMR data and mass spectrometry data in Table 1, the abietane type diterpene compound Leucosceptrum A has a molecular formula of C.sub.20H.sub.26O.sub.3, a molecular weight of 314, an unsaturation of 8, and a chemical name of 11,14-dihydroxy-8,11,13,15-abietatetraen-7-one with the structure shown in formula I.

(10) 1.3 Melting Point Test

(11) The melting point is tested by WRS-1C melting point apparatus produced by Shanghai Instrument Physical Optics Instrument Co., Ltd., and it is tested that the melting point of the abietane type diterpene compound Leucosceptrum A is 245-246 degree Celsius (? C.).

Embodiment 2

(12) An abietane type diterpene compound Leucosceptrum A is prepared in the present embodiment; and

(13) according to FIG. 1, the compound Leucosceptrum A is prepared as follows:

(14) S1, air-drying and crushing 5 kilograms (kg) of Leucosceptrum canum, leaching with ethanol with volume percent of 95%, where the ethanol is added in a mass-volume ratio of Leucosceptrum canum to ethanol of 1 g:10 milliliters (mL), and the leaching is carried out under temperature of 70? C. for 3 times, each time for 5 hours (h); filtering off insoluble substances, combining filtrates, and then evaporating to dryness at a low temperature and under reduced pressure at a vacuum degree of 0.08 Megapascal (MPa) and a temperature of 45? C. to obtain extract;

(15) S2, uniformly mixing the obtained extract with water according to a mass ratio of 1:2 to prepare a suspension, extracting with an equal volume of ethyl acetate for 3 times, combining extracted solutions, and evaporating to dryness at a low temperature and under the reduced pressure at a vacuum of 0.08 MPa and a temperature of 45? C. to obtain an ethyl acetate extract phase;

(16) S3, evenly mixing the obtained ethyl acetate extract phase with silica gel of a same mass, then placing in a silica gel column for chromatography, and subjecting to gradient elution with petroleum ether-ethyl acetate as the eluent, where the petroleum ether is in volume ratios of 10:1, 5:1, and 3:1 respectively to the ethyl acetate in each gradient; collecting eluent corresponding to the volume ratio of petroleum ether to ethyl acetate of 3:1, followed by thin-layer chromatographic analysis and combining similar fractions to obtain initial components; and

(17) S4, separating the obtained initial components by preparative liquid chromatography, and performing isocratic elution by using acetonitrile and water in a volume ratio of 45:55 as the eluant to obtain the abietane type diterpene compound Leucosceptrum A of 16.0 milligrams (mg) with anti-tumor effect as shown in the formula I.

Embodiment 3

(18) The present embodiment prepares an abietane type diterpene compound Leucosceptrum A according to the following steps:

(19) S1, air-drying and crushing 5 kg of Leucosceptrum canum, leaching with ethanol with volume percent of 90%, where the ethanol is added in a mass-volume ratio of Leucosceptrum canum to ethanol of 1 g: 20 mL, and the leaching is carried out under temperature of 65? C. for 3 times, each time for 4 h; filtering off insoluble substances, combining filtrates, and then evaporating to dryness at a low temperature and under the reduced pressure at a vacuum degree of 0.07 MPa and a temperature of 50? C. to obtain extract;

(20) S2, uniformly mixing the obtained extract with water according to a mass ratio of 1:1 to prepare a suspension, extracting with equal volume of ethyl acetate for 4 times, combining extracted solutions, and evaporating to dryness at a low temperature and under reduced pressure at a vacuum of 0.07 MPa and a temperature of 50? C. to obtain an ethyl acetate extract phase;

(21) S3, evenly mixing the obtained ethyl acetate extract phase with silica gel of a same mass, then placing in a silica gel column for chromatography, and subjecting to gradient elution with petroleum ether-ethyl acetate as the eluent, where the petroleum ether is in volume ratios of 9.8:1, 4.8:1, and 2.8:1 respectively to the ethyl acetate in each gradient; collecting eluent corresponding to the volume ratio of petroleum ether to ethyl acetate of 2.8:1, followed by thin-layer chromatographic analysis and combining similar fractions to obtain initial components; and

(22) S4, separating the obtained initial components by preparative liquid chromatography, and performing isocratic elution by using acetonitrile and water in a volume ratio of 40:60 as the eluant to obtain the abietane type diterpene compound Leucosceptrum A of 11.4 mg with anti-tumor effect as shown in the formula I.

Embodiment 4

(23) The present embodiment prepares an abietane type diterpene compound Leucosceptrum A according to the following steps:

(24) S1, air-drying and crushing 5 kg of Leucosceptrum canum, leaching with the ethanol with volume percent of 95%, where the ethanol is added in a mass-volume ratio of Leucosceptrum canum to ethanol of 1 g:15 mL, and the leaching is carried out under temperature of 75? C. for 2 times, each time for 4 h; filtering off insoluble substances, combining filtrates, and then evaporating to dryness at a low temperature and under the reduced pressure at a vacuum degree of 0.09 MPa and a temperature of 40? C. to obtain extract;

(25) S2, uniformly mixing the obtained extract with water according to a mass ratio of 1:1 to prepare a suspension, extracting with equal volume of ethyl acetate for 4 times, combining extracted solutions, and evaporating to dryness at a low temperature and under reduced pressure at a vacuum of 0.07 MPa and a temperature of 50? C. to obtain an ethyl acetate extract phase;

(26) S3, evenly mixing the obtained ethyl acetate extract phase with the silica gel of a same mass, then placing in a silica gel column for chromatography, and subjecting to gradient elution with petroleum ether-ethyl acetate as the eluent, where the petroleum ether is in volume ratios of 10.2:1, 5.2:1, and 3.2:1 respectively to the ethyl acetate in each gradient; collecting eluent corresponding to the volume ratio of petroleum ether to ethyl acetate of 3.2:1, followed by thin-layer chromatographic analysis and combining similar fractions to obtain initial components; and

(27) S4, separating the obtained initial components by preparative liquid chromatography, and performing isocratic elution by using acetonitrile and water in a volume ratio of 50:50 as the eluant to obtain the abietane type diterpene compound Leucosceptrum A of 8.1 mg with anti-tumor effects as shown in the formula I.

(28) Embodiment 5 Anti-tumor performance test of abietane type diterpene compound Leucosceptrum A

(29) The abietane type diterpene compound Leucosceptrum A prepared in Embodiment 2 is tested for its anti-tumor ability, with a process as follows:

(30) in vitro tumor cell proliferation inhibition assay: exponential phase tumor cells are taken and the cell suspension concentration is adjusted (50,000-100,000 cells/mL), then the cell suspension is inoculated in 96-well plate with 100 microlitres (u L) per well (M L/well), followed by drug administration (100 ?L/well) 24 h after inoculation; a blank control group, a cell control group and 6 drug groups with concentrations (3.12, 6.25, 12.5, 25, 50, 100 micromoles per litre (u mol/L)) are set up respectively, and the drug in the cell control group is cisplatin; after continuous cultivation for 72 h, 100 ?L 3-(4,5)-dimethylthiahiazo(-z-yl)-3,5-di-phenytetrazoliumromide (MTT) (1 mg/mL MTT dissolved in dulbecco's modified eagle medium (DMEM) culture solution) is added to each well, incubated at 37? C. for 4 h, then 150 ?L acidified isopropyl alcohol (containing 0.04 mol/L HCl) is added to each well, and kept under darkness for 30 minutes (min); then the absorbance is measured at 570 nanometers (nm) by enzyme-labeled instrument, the inhibition rate of the tested drug on the proliferation of tumor cells and the half inhibition concentration (IC.sub.50) of the tested substance on the proliferation of tumor cells (72 h) are calculated respectively; three parallels tests are arranged so as to reduce the influence of experimental errors, and the results are as follows.

(31) 5.1 Human Promyelocytic Leukemia Cell HIL-60

(32) (1) Tables 2, 3 and 4 illustrate the inhibitory effect of cisplatin on HIL-60 cells in the control group.

(33) TABLE-US-00002 TABLE 2 Inhibitory effect of parallel test I of cisplatin on HL-60 cells HL-60 cell quality Cisplatin concentrations (?mol/L) S/N Blank control 3.12 6.25 12.5 25 50 100 1 ?0.001 0.654 0.529 0.471 0.346 0.299 0.043 0.018 2 0.001 0.629 0.635 0.385 0.244 0.208 0.104 0.019 3 0.006 0.619 0.673 0.392 0.346 0.325 0.183 0.027 Average 0.002 0.634 0.6123 0.416 0.312 0.2773 0.11 0.0213 Mean squared 0.0036 0.018 0.0746 0.0478 0.0589 0.0614 0.0702 0.0049 error Survival 96.57% 65.51% 49.05% 43.57% 17.09% 3.06%

(34) According to Graphpad Prism 6.0, IC.sub.50=14.36.

(35) TABLE-US-00003 TABLE 3 Inhibitory effect of parallel test II of cisplatin on HL-60 cells HL-60 cell quality Cisplatin concentrations (?mol/L) S/N Blank control 3.12 6.25 12.5 25 50 100 1 ?0.001 0.515 0.435 0.215 0.185 0.109 0.095 0.033 2 0 0.502 0.465 0.365 0.299 0.121 0.056 0.011 3 ?0.002 0.467 0.399 0.327 0.22 0.106 0.077 0.032 Average ?0.001 0.4947 0.433 0.3023 0.2347 0.112 0.076 0.0253 Mean squared 0.001 0.0248 0.033 0.078 0.0584 0.0079 0.0195 0.0124 error Survival 87.56% 61.20% 47.55% 22.80% 15.53% 5.31%

(36) According to Graphpad Prism 6.0, IC.sub.50=10.69.

(37) TABLE-US-00004 TABLE 4 Inhibitory effect of parallel test III of cisplatin on HL-60 cells HL-60 cell Cisplatin concentrations (?mol/L) S/N Blank quality control 3.12 6.25 12.5 25 50 100 1 ?0.002 0.515 0.442 0.391 0.197 0.095 0.038 0.017 2 0 0.536 0.512 0.453 0.201 0.192 0.096 0.019 3 0 0.528 0.509 0.447 0.295 0.202 0.045 0.015 Average ?7E?04 0.5263 0.4877 0.4303 0.231 0.163 0.0597 0.017 Mean squared 0.0012 0.0106 0.0396 0.0342 0.0555 0.0591 0.0317 0.002 error Survival 92.66% 81.78% 43.96% 31.06% 11.45% 3.35%

(38) According to Graphpad Prism 6.0, IC.sub.50=13.16.

(39) IC.sub.50=12.74?1.87 of cisplatin is obtained according to the above data.

(40) (2) The inhibitory effect of compound Leucosceptrum A on BIL-60 cells is shown in Tables 5, 6 and 7.

(41) TABLE-US-00005 TABLE 5 Inhibitory effect of parallel test I of compound Leucosceptrum A on HL-60 cells HL-60 cell quality Leucosceptrum A concentrations (?mol/L) S/N Blank control 3.12 6.25 12.5 25 50 100 1 0.001 0.556 0.431 0.353 0.164 0.065 0.049 0.035 2 0.002 0.541 0.425 0.366 0.241 0.104 0.054 0.021 3 0 0.536 0.487 0.386 0.258 0.097 0.053 0.035 Average 0.001 0.5443 0.4477 0.3683 0.221 0.0887 0.052 0.0303 Mean squared 0.001 0.0104 0.0342 0.0166 0.0501 0.0208 0.0026 0.0081 error Survival 82.21% 67.61% 40.49% 16.13% 9.39% 5.40%

(42) According to Graphpad Prism 6.0, IC.sub.50=9.58.

(43) TABLE-US-00006 TABLE 6 Inhibitory effect of parallel test II of compound Leucosceptrum A on HL-60 cells HL-60 cell quality Leucosceptrum A concentrations (?mol/L) S/N Blank control 3.12 6.25 12.5 25 50 100 1 0.001 0.541 0.452 0.417 0.177 0.054 0.048 0.043 2 0 0.571 0.417 0.398 0.129 0.055 0.045 0.021 3 0.002 0.488 0.395 0.381 0.239 0.064 0.037 0.032 Average 0.001 0.5333 0.4213 0.3987 0.1817 0.0577 0.0433 0.032 Mean 0.001 0.042 0.0287 0.018 0.0551 0.0055 0.0057 0.011 squared error Survival 78.96% 74.70% 33.94% 10.64% 7.95% 5.82%

(44) According to Graphpad Prism 6.0, IC.sub.50=9.19.

(45) TABLE-US-00007 TABLE 7 Inhibitory effect of parallel test III of compound Leucosceptrum A on HL-60 cells HL-60 cell quality Leucosceptrum A concentrations (?mol/L) S/N Blank control 3.12 6.25 12.5 25 50 100 1 0.001 0.577 0.369 0.352 0.201 0.082 0.053 0.026 2 0.003 0.568 0.365 0.348 0.33 0.068 0.053 0.049 3 0.004 0.564 0.408 0.392 0.312 0.057 0.038 0.042 Average 0.0027 0.5697 0.3807 0.364 0.281 0.069 0.048 0.039 Mean 0.0015 0.0067 0.0238 0.0243 0.0699 0.0125 0.0087 0.0118 squared error Survival 66.67% 63.73% 49.09% 11.79% 8.00% 6.41%

(46) According to Graphpad Prism 6.0, IC.sub.50=8.42.

(47) IC.sub.50=9.06?0.59 of Leucosceptrum A is obtained according to the above data.

(48) 5.2 Human Lung Cancer Cell A549

(49) (1) The inhibitory effect of cisplatin on human lung cancer cell A549 is shown in Tables 8, 9 and 10.

(50) TABLE-US-00008 TABLE 8 Inhibitory effect of parallel test I of cisplatin on A549 cells A549 cell Cisplatin concentrations (?mol/L) S/N Blank quality control 3.12 6.25 12.5 25 50 100 1 0.001 0.661 0.641 0.489 0.231 0.212 0.205 0.203 2 0.002 0.652 0.642 0.446 0.254 0.288 0.207 0.198 3 0 0.731 0.715 0.391 0.278 0.221 0.196 0.173 Average 0.001 0.6813 0.666 0.442 0.2543 0.2403 0.2027 0.1913 Mean 0.001 0.0432 0.0424 0.0491 0.0235 0.0415 0.0059 0.0161 squared error Survival 97.75% 64.82% 37.24% 35.18% 29.64% 27.98%

(51) According to Graphpad Prism 6.0, IC.sub.50=14.62.

(52) TABLE-US-00009 TABLE 9 Inhibitory effect of parallel test II of cisplatin on A549 cells A549 cell quality Cisplatin concentrations (?mol/L) S/N Blank control 3.12 6.25 12.5 25 50 100 1 0 0.612 0.602 0.431 0.301 0.267 0.213 0.186 2 0.001 0.653 0.647 0.48 0.238 0.234 0.202 0.202 3 0.003 0.626 0.613 0.491 0.239 0.208 0.199 0.179 Average 0.0013 0.6303 0.6207 0.4673 0.2593 0.2363 0.2047 0.189 Mean squared 0.0015 0.0208 0.0235 0.0319 0.0361 0.0296 0.0074 0.0118 error Survival 98.46% 74.09% 41.02% 37.36% 32.33% 29.84%

(53) According to Graphpad Prism 6.0, IC.sub.50=17.85.

(54) TABLE-US-00010 TABLE 10 Inhibitory effect of parallel test III of cisplatin on A549 cells A549 cell quality Cisplatin concentrations (?mol/L) S/N Blank control 3.12 6.25 12.5 25 50 100 1 0 0.71 0.693 0.485 0.262 0.243 0.201 0.146 2 ?0.001 0.725 0.686 0.433 0.242 0.217 0.219 0.209 3 0.002 0.665 0.655 0.469 0.292 0.283 0.257 0.252 Average 0.0003 0.7 0.678 0.4623 0.265 30.2477 0.2257 0.2023 Mean squared 0.0015 0.0312 0.0202 0.0266 0.0252 0.0332 0.0286 0.0533 error Survival 96.86% 66.03% 37.88% 35.35% 32.21% 28.87%

(55) According to Graphpad Prism 6.0, IC.sub.50=15.38.

(56) According to the above data, the IC.sub.50 of cisplatin is IC.sub.50=15.95?1.69.

(57) (2) The inhibitory effect of compound Leucosceptrum A on A549 cells is shown in Tables 11, 12 and 13.

(58) TABLE-US-00011 TABLE 11 Inhibitory effect of parallel test I of compound Leucosceptrum A on A549 cells A549 cell quality Leucosceptrum A concentrations (?mol/L) S/N Blank control 3.12 6.25 12.5 25 50 100 1 0 0.843 0.811 0.828 0.635 0.184 0.085 0.078 2 0.002 0.835 0.824 0.791 0.663 0.175 0.074 0.067 3 0.001 0.814 0.81 0.728 0.602 0.189 0.085 0.085 Average 0.001 0.8307 0.815 0.7823 0.6333 0.1827 0.0813 0.0767 Mean squared 0.001 0.015 0.0078 0.0506 0.0305 0.0071 0.0064 0.0091 error Survival 98.11% 94.17% 76.22% 21.90% 9.68% 9.12%

(59) According to Graphpad Prism 6.0, IC.sub.50=17.55.

(60) TABLE-US-00012 TABLE 12 Inhibitory effect of parallel test II of compound Leucosceptrum A on A549 cells A549 cell quality Leucosceptrum A concentrations (?mol/L) S/N Blank control 3.12 6.25 12.5 25 50 100 1 0.003 0.796 0.72 0.711 0.636 0.221 0.077 0.073 2 0.002 0.793 0.753 0.786 0.687 0.182 0.074 0.066 3 0.003 0.865 0.869 0.769 0.683 0.173 0.059 0.058 Average 0.0027 0.818 0.7807 0.7553 0.6687 0.192 0.07 0.0657 Mean squared 0.0006 0.0407 0.0783 0.0393 0.0284 0.0255 0.0096 0.0075 error Survival 95.42% 92.31% 81.68% 23.22% 8.76% 7.73%

(61) According to Graphpad Prism 6.0, IC.sub.50=18.38.

(62) TABLE-US-00013 TABLE 13 Inhibitory effect of parallel test III of compound Leucosceptrum A on A549 cells A549 cell quality Leucosceptrum A concentrations (?mol/L) S/N Blank control 3.12 6.25 12.5 25 50 100 1 0.001 0.826 0.767 0.822 0.6 0.188 0.071 0.067 2 0 0.867 0.845 0.77 0.587 0.191 0.079 0.067 3 0.002 0.866 0.827 0.791 0.583 0.207 0.088 0.073 Average 0.001 0.853 0.813 0.7943 0.59 0.1953 0.0793 0.069 Mean squared 0.001 0.0234 0.0408 0.0262 0.0089 0.0102 0.0085 0.0035 error Survival 95.31% 93.11% 69.13% 22.81% 9.19% 7.98%

(63) According to Graphpad Prism 6.0, IC.sub.50=16.62.

(64) According to the above data, the IC.sub.50 of Leucosceptrum A is IC.sub.50=17.52?0.88.

(65) 5.3 The 72-hour IC.sub.50 (mol/L) of Leucosceptrum A and cisplatin on two cancer cells is shown in Table 14.

(66) TABLE-US-00014 TABLE 14 72-hour IC.sub.50(?mol/L) of Leucosceptrum A and cisplatin on two cancer cells Tumor cells HL-60 cells A-549 cells Cisplatin 12.74 ? 1.87 15.95 ? 1.69 Leucosceptrum A 9.06 ? 0.59 17.52 ? 0.88

(67) It can be seen from the above tables 5-7 and 11-13 that the compound Leucoceptrum A with different concentrations has inhibitory effects on human lung cancer cell A549 and human promyelocytic leukemia cell HL-60, and the IC.sub.50 values of Leucoceptrum A after 72 h reach 17.52?0.88 and 9.06?0.59 ?mol/L, respectively, indicating that Leucoceptum A has a strong inhibitory effect on the growth of the above two tumor cells; specifically, the IC.sub.50 value of Leucoceptum A on HL-60 cells is lower than that of control group with cisplatin, which indicates that Leucoceptum A has a very significant inhibitory effect on HL-60 cells.

(68) The above embodiments are only used to illustrate the technical scheme of the present application and not to limit it. Although the present application is described in detail with reference to the preferred embodiments, it should be understood by those of ordinary skill in the art that the technical scheme of the present application may be modified or replaced by the same without departing from the purpose and scope of the technical scheme of the present application, which shall be covered by the scope of the claims of the present application. The technology, shape and construction parts not described in detail in the present application are all prior art.