CN107793403A - A kind of noval chemical compound with epoxy construction and preparation method thereof and medical usage - Google Patents

Abstract

The invention discloses a kind of noval chemical compound with epoxy construction and preparation method thereof and medical usage, belong to drug field, and in particular to the isolated a kind of noval chemical compound with epoxy construction, preparation method and medical usage from the dry mature fruit of cardamom.The compound is reports first, and extracting and developing can purify to obtain from the dry mature fruit of cardamom, purity is high.In vitro test proves that it can suppress the propagation of stomach cancer cell, both can individually play inhibitory activity, and enhancing inhibitory activity can also be combined with other anticarcinogens, and the compound can be developed to the medicine for being prepared into treatment stomach cancer.

Description

A kind of noval chemical compound with epoxy construction and preparation method thereof and medical usage

Technical field

The invention belongs to drug field, and in particular to isolated one kind has from the dry mature fruit of cardamom Noval chemical compound of epoxy construction and preparation method thereof and medical usage.

Background technology

Plant cardamom is perennial herb.Rhizome is sturdy, brownish red.Leaf two arranges, and blade is long and narrow to drape over one's shoulders needle-like, leaf sheath tool palm fibre Yellow pubescence.Spike is extracted out by basal part of stem.Inflorescence significant elongation, flower arrangement is sparse, corolla white.The long oval of fruit, Pericarp matter is tough, not easy to crack.Seed group's point 3 valves, per 5~9 pieces of valve seed, seed fragrant odour and it is high strong.Main product Vietnam, in this Maraba (Malabar) seashore of orchid card and South India.Receive and adopt when real ripe, remove the carpopodium of residual, dry.Use part For the dry fruit of zingiberaceous plant cardamom.

Medicinal material cardamom is zingiberaceous plant cardamom Elettaria cardamomum (L.) Maton nearly mellow fruit of drying It is real, be Tibetan medicine and Uygur medicine medication, with the name of " adding element " first recorded in《Four doctor's allusion quotations》, now recorded by multinational pharmacopeia.Cardamom It is world-renowned autonomic drug and spices, is referred to as " after spices ", with originating in South India, Sri Lanka and melon horse traction etc. Ground, usage history exceedes bimillennium in Ayurvedic medicine, has the effect of wind-dispelling, stomach invigorating.The traditional Chinese medical science is rarely employed at present, small Cardamom was once recorded in nineteen fifty-three version Chinese Pharmacopoeia with the name of " cardamom ".

The composition that cardamom has been reported is mainly volatile oil, and polysaccharide and protein, thus it is speculated that also contains flavonoid Thing, but have not yet to see the separation and identification of flavones ingredient monomer.Volatile oil is active component important in cardamom, content Up to 7%, the volatile oil that ultrasonic extraction obtains is with α-terpinyl acetate, eucalyptol, linalool, alpha-terpineol, bergamio Based on, with supercritical CO₂Extraction gained composition is similar；Such as extracted with n-hexane, volatile oil compositions are different, be limonene, Eucalyptol, terpinolene, laurene.In addition, also containing farnesol, neryl acetate, firpene, nerolidol, sabinene Deng.

Cardamom pharmacological activity is various, especially most with antitumor activity report, is related to multiple positions such as skin, lung, colon Tumour, also have preferable protective effect to intestines and stomach, in addition, also there is antibacterial, anti-oxidant, anti-inflammatory, analgesia etc. to act on.

The content of the invention

It is an object of the invention to provide one kind isolated in a kind of dry mature fruit from cardamom to have ring The noval chemical compound of oxide structure；

Another object of the present invention is to provide the preparation method of the noval chemical compound；

It is used for the medical usage for preparing treatment gastric cancer medicament it is still another object of the present invention to provide the compound.

The above-mentioned purpose of the present invention is achieved by following technical scheme：

Compound (I) with following structural formula,

The preparation method of described compound (I), includes following operating procedure：(a) the dry mature fruit powder of cardamom It is broken, extracted with 80% alcohol heat reflux, merge extract solution, no alcohol taste is concentrated into, successively with petroleum ether, ethyl acetate and water saturation Extracting n-butyl alcohol, respectively obtain petroleum ether extract, acetic acid ethyl ester extract and n-butyl alcohol extract；(b) second in step (a) Acetoacetic ester extract is cleaned with macroreticular resin, successively with 10% ethanol and 75% ethanol elution, is collected 75% ethanol eluate, is subtracted Pressure is concentrated to give 75% ethanol elution thing medicinal extract；(c) 75% ethanol elution medicinal extract is separated with purification on normal-phase silica gel in step (b), is used successively Volume ratio is 90:1、45:1、20:1、10:1 and 1:1 methylene chloride-methanol gradient elution obtains 5 components；(d) step (c) Middle component 3 is further separated with purification on normal-phase silica gel, is successively 25 with volume ratio:1、20:1 and 10:1 methylene chloride-methanol gradient Afford 3 components；(e) reverse phase silica gel that component 2 is bonded with octadecylsilane in step (d) separates, and uses volume basis Concentration is 70% methanol aqueous solution isocratic elution, collects 8-12 column volume eluent, eluent is concentrated under reduced pressure to give pure Compound (I).

Further, the macroreticular resin is AB-8 type macroporous absorbent resins.

Pharmaceutical composition, wherein the described compound (I) and pharmaceutically acceptable carrier containing therapeutically effective amount.

Application of the described compound (I) in the medicine for preparing treatment stomach cancer.

Application of the described pharmaceutical composition in the medicine for preparing treatment stomach cancer.

It when the compounds of this invention is used as medicine, can directly use, or be used in the form of pharmaceutical composition.

The pharmaceutical composition contains the compounds of this invention (I) of therapeutically effective amount, and remaining is pharmaceutically acceptable, right The nontoxic and inert pharmaceutical acceptable carrier of humans and animals and/or excipient.

Described pharmaceutical acceptable carrier or excipient is one or more selected from solid, semisolid and liquid diluent, filler And pharmaceutical preparation assistant agent.The pharmaceutical composition of the present invention is used in the form of per weight dose.Medicine of the present invention can The patient for needing to treat is applied to by oral or injection form.For it is oral when, tablet, sustained release tablets, control can be made into Release piece, capsule, dripping pill, micropill, supensoid agent, emulsion, powder or granule, oral liquid etc.；During for injecting, sterilizing can be made into Water-based or oily solution, aseptic powder injection, liposome or emulsion etc..

Figure of description

Fig. 1 is compound (I) structural formula；

Fig. 2 is the two-dimentional NOESY spectrums of compound (I).

Embodiment

The essentiality content of the present invention is further illustrated with reference to embodiment, but present invention protection model is not limited with this Enclose.Although being explained in detail with reference to preferred embodiment to the present invention, it will be understood by those within the art that, can be right Technical scheme is modified or equivalent substitution, without departing from the spirit and scope of technical solution of the present invention.

Embodiment 1：Compound (I) separation prepares and structural identification

Medicinal material and reagent source：Ethanol, petroleum ether, ethyl acetate, n-butanol, dichloromethane are pure to analyze, purchased from Shanghai Ling Feng chemical reagent Co., Ltd, methanol, analysis is pure, purchased from Jiangsu Han Bang chemical reagent Co., Ltd.Cardamom is dried to Ripe fruit is purchased from Hui nationality's Chinese Medicinal Materials Markets, place of production Fujian.

Preparation method：(a) dry mature fruit (10kg) of cardamom is crushed to mung bean size, with 80% alcohol heat reflux Extract (25L × 3 time), merge extract solution, no alcohol taste (6L) is concentrated into, successively with petroleum ether (6L × 3 time), ethyl acetate (6L × 3 times) and water saturated n-butanol (6L × 3 time) extraction, concentration, respectively obtain petroleum ether extract, acetic acid ethyl ester extract (305g) and n-butyl alcohol extract；(b) in step (a) acetic acid ethyl ester extract dissolved in purified water to 2L, medical absorbent cotton mistake Filter, filtrate are separated with AB-8 types macroreticular resin, successively with 10% ethanol (10L) and 75% (12L) ethanol elution, collect 75% second Alcohol eluen, be concentrated under reduced pressure to obtain 75% ethanol elution thing medicinal extract (142g)；(c) 75% ethanol elution medicinal extract is used in step (b) 200-300 mesh purification on normal-phase silica gel separates, and is successively 90 with volume ratio:1 (10 column volumes), 45:1 (9 column volumes), 20:1 (8 Column volume), 10:1 (8 column volumes) and 1:The methylene chloride-methanol gradient elution of 1 (5 column volumes) obtains 5 components；(d) Component 3 (35g) is further separated with 200-300 mesh purification on normal-phase silica gel in step (c), is successively 25 with volume ratio:1 (8 cylinders Product), 20:1 (10 column volumes) and 10:The methylene chloride-methanol gradient elution of 1 (5 column volumes) obtains 3 components；(e) step Suddenly the reverse phase silica gel ODS-C18 that component 2 (10g) is bonded with octadecylsilane in (d) is separated, and is 70% with concentration expressed in percentage by volume Methanol aqueous solution isocratic elution, collect 8-12 column volume eluent, eluent is concentrated under reduced pressure to give pure compound (I) (23mg)。

Structural identification：White, needle-shaped crystals, it is soluble in chloroform, ethyl acetate, acetone and methanol；HR-ESIMS shows [M+ Na]⁺For m/z 741.3112, it is C that can obtain molecular formula with reference to nuclear-magnetism feature₃₇H₅₀O₁₄, degree of unsaturation 13.Proton nmr spectra Data δ_H(ppm,CDCl₃,400MHz)：H-1 (5.54, d, J=6.6), H-2 (2.25, m), H-2 (2.41, m), H-5 (2.89, M), H-6 (2.61, m), H-6 (2.78, m), H-9 (3.25, s), H-11 (4.99, d, J=5.6), H-12 (5.29, d, J= 5.6), H-15 (4.53, m), H-16 (2.44, m), H-16 (2.75, m), H-17 (3.56, t, J=9.4), H-18 (0.42, s), H-19 (1.23, s), H-21 (7.29, s), H-22 (6.37, s), H-23 (7.35, s), H-28 (1.59, s), H-29 (4.13, d, J=12.6), H-29 (4.42, d, J=12.6), H-30 (5.24, s), H-30 (5.28, s), OMe-3 (3.66, s), OAc-1 (2.11, s), OAc-2'(2.14, s), H-2'(3.97, d, J=4.8), H-3'(1.65, m) and, H-4'(0.87, s), H-5' (1.26, m), H-5'(1.44, m), H-6'(0.84, s)；Carbon-13 nmr spectra data δ_C(ppm,CDCl₃,100MHz)：72.0 (CH, 1-C), 38.5 (CH₂, 2-C), 173.7 (C, 3-C), 87.1 (C, 4-C), 42.9 (CH, 5-C), 31.6 (CH₂, 6-C), 170.1 (C, 7-C), 144.2 (C, 8-C), 46.2 (CH, 9-C), 44.0 (C, 10-C), 79.4 (CH, 11-C), 79.4 (CH, 12- C), 55.9 (C, 13-C), 101.8 (C, 14-C), 68.4 (CH, 15-C), 37.6 (CH₂, 16-C), 42.9 (CH, 17-C), 13.9 (CH₃, 18-C), 17.5 (CH₃, 19-C), 123.7 (C, 20-C), 139.8 (CH, 21-C), 110.7 (CH, 22-C), 142.8 (CH, 23-C), 19.3 (CH₃, 28-C), 68.1 (CH₂, 29-C), 109.8 (CH₂, 30-C), 52.3 (CH₃, OMe-3), 170.3 (C, OAc-1), 20.8 (CH₃, OAc-1), 172.0 (C, OAc-2'), 21.1 (CH₃, OAc-2'), 172.6 (C, 1'-C), 75.9 (CH, 2'-C), 38.6 (CH, 3'-C), 15.2 (CH₃, 4'-C), 23.6 (CH₂, 5'-C), 11.1 (CH₃, 6'-C)；Carbon atom mark Note is referring to Fig. 1.¹³C NMR and DEPT spectrum show 37 carbon signals, respectively 8 methyl, 6 methylene, 12 methines and 11 seasons Carbon.The substitution of β types furans cyclic group (δ H7.29,7.35,6.37, each 1H, s；δ C139.8,142.8,110.7,123.7), ring External double bond (δ H5.24,5.28, each 1H, s；δ C144.2,109.8), three tertiary methyl groups (δ H0.42,1.23,1.59, respectively 3H, s；δ C13.9,17.5,19.3), and methoxy group (δ a H3.66,3H, s；δ C52.3) show that its parent nucleus is Prieurianin type limonin types, it is similar to the Aphanagranin D structures of document report.Pass through two dimensional NMR Spectrum (HSQC,¹H-¹H COSY and HMBC) it can obtain the connected mode of quaternary carbon structure fragment.It is main compared with Aphanagranin D It is the opening in C-1 and C-11 ether rings to want difference, and the missing of C-15 ketone carbonyls.In HMBC spectrums, H-1 (δ H5.54, D, J= It is 6.6Hz) related to acetyl group (δ C170.3) and ester carbonyl group C-3 (δ C173.7), and phase of the methoxyl group (δ H3.66, s) with C-3 Guan Xing, show acetyl group (δ H2.11, s respectively；δ C20.8,170.3) and ester carbonyl group position.H-17 (δ H3.56, t, J= 9.4Hz) and H-16 (δ H2.44, m；2.75, m) correlation with C-15 (δ C68.4) shows that oxygen-containing methin groups are located at C- 15, C-15 form oxygen ring with C-14.In addition, H-12 (δ H5.29d, J=5.6Hz) and ester carbonyl group C-1'(δ C172.6) and H- 2'(3.97, d, J=4.8Hz) with the carbonyl (δ C172.0) of acetyl group illustrate that 2- acetoxy-3s-methylpentyryl is connected On C-12 side chain.The relative configuration of the compound can be composed (Fig. 2) analysis by two-dimentional ROESY and be determined.Pass through Me-19 and H- 1, Me-19 and H-11, H-11 it is related to H-12's and H-12 and H-17 understand these Hydrogen Protons be beta configuration.In turn, H-5 With H-9, H-5 and Me-28, H-9 and Me-18 and Me-18 and H-15 correlation show H-5, H-9, Me-18, Me-28 with H-15 is α-configuration.Therefore, the structure of the compound 1 and relative structure are established as shown in Figure 1.

Therefore, finally determine that the compound structure is as shown in Figure 1.

Embodiment 2：Compound (I) pharmacological action is tested

First, material and instrument

Human stomach cancer cell line SGC-7901, it is purchased from Sai Er Reagent Companies；Compound (I) is made by oneself, and normalization purity is more than 98%；CTX (endoxan, positive drug), trypsase, MTT, dimethyl sulfoxide (DMSO) (DMSO), agarose, glacial acetic acid are purchased from the U.S. Sigma companies；RPMI-1640 culture mediums, PBS phosphate buffer solutions are purchased from GIBCO companies of the U.S.；Hyclone is purchased from Shandong Yin Xiang great achievements company；Trypan blue is purchased from Beijing Bioisystech Co., Ltd of Zhong Shan Golden Bridge；TUNEL kits are purchased from triumphant base biology section Skill Development Co., Ltd；DAB colour reagents box is purchased from Beijing biotech firm of Zhong Shan Golden Bridge；Formaldehyde is purchased from Fisher companies of the U.S.； Catalase is purchased from the new fine chemistry industry development centre in Tianjin day.

WD-9403C uv analyzers are purchased from German Biometra companies；RKI-1002 types CO2gas incubator is purchased from day This Ikemoto companies；Superclean bench is purchased from Suzhou purification experimental facilities Co., Ltd；Ultracentrifuge is purchased from Beijing Medical treatment device Tool factory；Low speed centrifuge is purchased from Beijing medical apparatus and instruments factory；Wilovert S types inverted microscope is purchased from Japanese Olympus companies； Pressure steam sterilizer is purchased from Shanghai Bo Xun Industrial Co., Ltd.s；Thermostat metal water bath contains the limited public affairs of instrument purchased from Hangzhou is difficult to understand Department；Constant temperature blender with magnetic force is purchased from Tianjin Raul Science and Technology Ltd.；Cell counting count board is purchased from Shanghai precision instrument company；It is ultrapure Hydrophone is purchased from Britain PURELAB ulTRA GENETIC.

2nd, test method

1st, stomach cancer cell SGC-7901 culture

1.1 cell recovery

(1) cell cryopreservation tube is taken out from liquid nitrogen container, is immediately placed in 37 DEG C -42 DEG C, in 75% alcohol, then moves to equality of temperature In water-bath；(2) l-3min cryopreservation tubes are gently rocked, melt frozen stock solution, are moved in super-clean bench rapidly；(3) cell will be contained Frozen stock solution suction sterile centrifugation tube, add appropriate RPIM-1640 culture mediums；(4) piping and druming is put into low speed centrifuge after mixing, 800rpm/min is centrifuged 3 minutes, removes supernatant；(5) the appropriate culture medium of addition is blown even, and cell suspension is inoculated into according to concentration In 10mL culture dishes；(6) be placed in containing 5% carbon dioxide, 37 DEG C of saturated humidities incubator in cultivate.

1.2 passage

(1) when the cell attachment in culture dish about 80%, blown and beaten in super-clean bench with the old culture medium of pipette, extract Cell in culture dish for several times, to blow dead cell off；(2) old culture medium is sucked with pipette, adds appropriate 0.25% pancreatin and disappear Change cell, be placed in 30 DEG C of incubators and digest；(3) culture dish is placed under inverted microscope about after 3min and observed, treat cell week Side is shinny, retraction then illustrates that cell departs from from wall after being rounded；(4) pancreatin is sucked in super-clean bench rapidly.Add appropriate Culture medium blows and beats cell repeatedly, cell is completely disengaged culture dish；(5) cell suspension is seeded to different trainings by concentration respectively Support in ware, supply culture medium；(6) it is replaced in containing 5%CO₂, 37 DEG C of saturated humidities incubator in cultivate.

2nd, cell count

(1) cell counting count board and cover glass are got out, the two is clean with alcohol wipe, and cover glass is covered in tally On；(2) after alcohol volatilization, appropriate cell suspension is suctioned out, is added dropwise at cover glass edge, suspension is full of cover glass and tally Between, it is careful not to overflow cover glass and the glass guide channel of both sides, is counted after standing；(3) 4 big lattice are found under the microscope, often Individual big lattice are divided into 16 small lattice again, count the cell number in 4 big lattice respectively, average.On the left of line ball cytometer and top , disregard right side and lower section；(4) average cell number × 10000 of cell concentration (individual/mL)=each grid；(5) by cell Suspension is diluted to concentration needed for experiment.

3rd, cell viability determines

(1) SGC-7901 stomach cancer cell suspensions 0.9mL to be determined is taken；(2) trypan blue piping and druming is added into cell suspension Mix；(3) at least 200 cells are counted using cell counting count board blind, observed under inverted microscope；(4) Microscopic observation cell Staining conditions, it is dead cell to dye light blue person into the cell, and the person of being unstained is living cells, with hundred of TCS shared by living cells Divide the vigor (%) than reflection cell.

4th, MTT detects inhibitory rate of cell growth

Experiment packet：1. negative control group；2. compound (I) group 1, compound (I) (50 μm of ol/L)；3. compound (I) Group 2, compound (I) (100 μm of ol/L)；4. CTX groups 1, CTX (50 μm of ol/L)；5. CTX groups 2, CTX (100 μm of ol/L)；6. join Share medicine group 1, compound (I) (25 μm of ol/L)+CTX (25 μm of ol/L)；7. drug combination group 2, compound (I) (50 μm of ol/L) +CTX(50μmol/L)。

(1) take the logarithm the SGC-7901 cells in growth period, counted with cell counting count board, regulation cell concentration is 5 × l0⁵/ mL；(2) 96 orifice plates are inoculated in every μ L of hole 100 using sample injector.Pay attention to only being inoculated into 60 holes of centre, periphery 36 during inoculation Hole is filled with PBS, while spreads 3 96 orifice plates；(3) 96 orifice plates completed are placed in 37 DEG C, 5%CO₂In cell culture incubator, treat Taken out after 24h cell attachments；(4) 96 orifice plates are divided into compound (I) group, (0,50,100 μm of ol/L), CTX groups (0,50,100 μ Mol/L), 2 medicine joint groups (0/0,25/25,50/50 μm of ol/L), while set up not celliferous blank control group and (only plus cultivate Base) give different disposal respectively；(5) each concentration of each medicine sets 5 multiple holes, cultivates 24,48 and 72h respectively；(6) respectively in spy Fixed end time point takes out 96 orifice plates, and the μ L of MTT (5g/L) solution 20 are added per hole, puts back to incubator and continues to cultivate 4h, terminates Culture.(7) supernatant in hole carefully is sucked, 150 μ L DMSO is added per hole, plate shaker vibration 10min makes crystallization fully molten Solution, micro- Microscopic observation particle disappear.(8) optical density (OD) value in each hole is determined at ELIASA 490nm wavelength, when calculating each Between the inhibitory rate of cell growth put.Inhibiting rate=[1- (dosing group OD values-blank control group OD values)/(negative control group OD values- Blank control group OD values)] × 100%.

5th, TUNEL methods detection apoptotic index

The preparation of 5.1 cell climbing sheets

(1) processing of cover glass：Cover glass is placed in soaked overnight in the concentrated sulfuric acid, next day is first rinsed for several times with running water, It is placed in again in absolute ethyl alcohol and soaks 4h, then rinsed well with deionized water, is placed on for drying laggard horizontal high voltage sterilization in dry case, Taking-up is directly placed into standby in super-clean bench.6 orifice plates are placed in ultraviolet in super-clean bench and irradiate 30min；(2) cover glass is placed：Six Preparation places cover glass again after putting a small amount of culture medium of position instillation of cover glass in every hole of orifice plate so that cover glass is examined with orifice plate The tension force of culture medium is bonded together, and cover glass hikes up when preventing from adding cell suspension, causes double-layer cell adherent；(3) take the logarithm The SGC-7901 cells in growth period, into cell suspension, it is l × 10 to adjust cell concentration with cell counting count board for digestion piping and druming⁶/mL。 (4) 1mL cell suspensions are separately added into the every hole for being placed with cover glass with sample injector, while spread 3 plates, be placed in 37 DEG C, 5%CO₂Training Support culture 24h in case and treat cell attachment；After (5) 24 hour cells are adherent in super-clean bench by 6 orifice plates be divided into negative control group and Experimental group gives different disposal respectively, and negative control group adds 1mL culture mediums, experimental group be further divided into compound (I) group, CTX groups and 3 subgroups of drug combination group, add 1mL medicines per hole.Compound (I) organizes final concentration of 100 μm of ol/L, CTX groups final concentration of 100 μm ol/L, the final concentration of 2 kinds of medicines of 2 medicine joint groups respectively set 2 multiple holes for 50 μm of every group of ol/L.(6) 37 DEG C are placed in, 5%CO₂'s Continue to cultivate in cell culture incubator.

5.2 TUNEL method operating procedures

(1) 6 orifice plates are taken out during cell climbing sheet dosing culture 24h, following steps is carried out successively according to TUNEL specifications；(2) Carefully inhale the supernatant abandoned in every hole；(3) each hole adds PBS (4 DEG C) 2mL, and plate shaker rinses 5min × 3 time；(4) per hole 1mL precooling cell fixers are added, 4 DEG C of refrigerators fix 30min.(5) inhale and abandon fixer, each hole adds PBS (4 DEG C) 2mL, puts down Plate shaking table rinses 5min × 3 time.(6) immerse in confining liquid, (15 DEG C -25 DEG C) closing 10min of room temperature.(7) each hole adds PBS (4 DEG C) 2mL, plate shaker rinse 5min × 3 time.Blotted around sample with blotting paper.(8) 50 μ LUTdT enzymes are added dropwise in each sample Reaction solution, 37 DEG C, lucifuge moistening reaction 60min.(9) each hole adds PBS (4 DEG C) 2mL, and plate shaker rinses 5min × 3 time. Blotted around sample with blotting paper.(10) 50 μ L Streptavidin-HRP working solutions are added dropwise, 37 DEG C, lucifuge moistening is reacted 30min.(11) each hole adds PBS (4 DEG C) 2mL, and plate shaker rinses 5min × 3 time.(12) 100 μ L DAB work is added dropwise Liquid, color development at room temperature reaction 10min.(13) each hole adds PBS (4 DEG C) 2mL, and plate shaker rinses 5min × 3 time.(14) optics Micro- Microscopic observation is taken pictures:10 high power (× 100) visuals field are randomly selected, each visual field counts 100 cells, calculates tune and dies finger Several average value.Apoptotic index (AI)=(positive cell number/total cell number × 100%).The nucleus of positive cell is in brown Color.

6th, statistical analysis

Analyzed using SPSS 11.5, compare between group and analyzed with Oneway-ANOVA methods, compared two-by-two using LSD-t Method, P<0.05 is that difference is statistically significant.

3rd, result and conclusion

1st, MTT detects growth inhibition ratio of the medicine to SGC-7901 stomach cancer cells

Through 3 repetition MTT, testing result is shown, after single medicine acts on stomach cancer cell 72h, 100 μm of ol/L medicine Medicine group of the group compared with 50 μm of ol/L is higher to the growth inhibition ratio of stomach cancer cell, the statistically significant (P of difference<0.05).50μ Growth inhibition effect and drug combination 25/25 μm ol/L group no significant difference of the mol/L compounds (I) to stomach cancer cell (P>0.05).50 μm of ol/L CTX has to Growth of Gastric inhibitory action and 25/25 μm of ol/L groups ratio of drug combination, difference Statistical significance (P<0.05).50/50 μm of ol/L of drug combination is higher than each high concentration list medicine group to Growth of Gastric inhibiting rate (100 μm of ol/L) is to the growth inhibition ratio of stomach cancer cell, the statistically significant (P of difference<0.001), it is shown in Table 1.

2nd, TUNEL methods detection each group apoptotic index

Compound (I), CTX and drug combination group to stomach cancer cell SGC-7901 play the role of suppress growth, each group with Statistically significant (the P of negative control group comparing difference<0.001) karyon that, negative control group has a small amount of cell is dyed to palm fibre Brown, each single medicine group have increase compared with drug combination group apoptotic cell is compared with negative control group, and drug combination group cell withers Index highest is died, it is more notable to stomach cancer cell inhibition.It is shown in Table 2.

Conclusion shows that compound (I), which acts solely on stomach cancer cell, can suppress the propagation of stomach cancer cell, promote cell Apoptosis, with the increase of concentration, the suppression growth to stomach cancer cell is in enhancing trend.Compound (I) and CTX use in conjunction Promote the apoptosis effect of stomach cancer cell more notable afterwards, the growth inhibition ratio and apoptotic index of cell raise compared with independent medication group. Therefore, compound (I) can individually develop into the medicine for the treatment of stomach cancer, can also be developed with anti-cancer agent in combination such as CTX into multiple Square drug therapy stomach cancer.

The each group different time points of table 1 are to stomach cancer cell inhibiting rate (n=5, mean value ± deviation) * P<0.05, * * P<0.01

Group	n	24h	48h	72h
					(I) 50 μm of ol/L (1) of compound	5	14.87±	16.29±	55.70±6.67
(I) 50 μm of ol/L (2) of compound	5	31.00±	59.42±	74.32±5.84
					CTX50μmol/L(3)	5	24.87±	36.83±	40.58±7.15
CTX100μmol/L(4)	5	42.86±	44.21±	50.31±4.63
					25/25 μm of ol/L of drug combination	5	36.43±	38.76±	58.55±9.53
50/50 μm of ol/L of drug combination	5	49.38±	74.56±	96.12±2.25
					F	/	17.918**	29.646**	47.625**
P(1):(2)	/	0.001	<0.001	<0.001
					(3):(4)	/	<0.001	0.169	0.025
(5):(6)	/	0.005	<0.001	<0.001
					(1):(5)	/	<0.001	<0.001	0.49
(3):(5)	/	0.01	0.713	<0.001
					(2):(6)	/	<0.001	0.008	<0.001
(4):(6)	/	0.13	<0.001	<0.001

Apoptotic index (n=3, mean value ± deviation) * * P of each group cell after the medicine of table 2 effect 24h<0.01

Embodiment 3

The preparation of tablet：Compound (I) is first made by the method for embodiment 1, and utilizes organic acid such as tartaric acid or lemon Acid or formic acid or ethanedioic acid etc., inorganic acid salt as made of hydrochloric acid or sulfuric acid or phosphoric acid, it is 1 by itself and excipient weight ratio:9 Ratio adds excipient, pelletizing press sheet.

Embodiment 4

The preparation of oral liquid：Compound (I) is first made by the method for embodiment 1, and utilizes organic acid such as tartaric acid or lemon Lemon acid or formic acid or ethanedioic acid etc., inorganic acid salt as made of hydrochloric acid or sulfuric acid or phosphoric acid, routinely oral liquid preparation method mouth is made Take liquid.

Embodiment 5

The preparation of capsule or granule：Compound (I) is first made by the method for embodiment 1, and utilizes organic acid such as wine Stone acid or citric acid or formic acid or ethanedioic acid etc., the inorganic acid salt as made of hydrochloric acid or sulfuric acid or phosphoric acid, by itself and excipient weight Amount is than being 1:9 ratio adds excipient, and capsule or granule is made.

Embodiment 6

The preparation of parenteral solution：Compound (I) is first made by the method for embodiment 1, and utilizes organic acid such as tartaric acid or lemon Lemon acid or formic acid or ethanedioic acid etc., inorganic acid salt as made of hydrochloric acid or sulfuric acid or phosphoric acid, routinely plus water for injection, refined filtration, Parenteral solution is made in embedding sterilizing.

Embodiment 7

The preparation of aseptic powder injection：By the method for embodiment 1 first be made compound (I), and using organic acid for example tartaric acid or Citric acid or formic acid or ethanedioic acid etc., the inorganic acid salt as made of hydrochloric acid or sulfuric acid or phosphoric acid, are dissolved in sterile water for injection In, stirring makes molten, is filtered with sterile suction funnel, then sterile refined filtration is sub-packed in ampoule, sterile sealing after frozen drying Obtain powder-injection.

Claims (6)

1. the compound (I) with following structural formula,

2. the preparation method of the compound (I) described in claim 1, it is characterised in that include following operating procedure：(a) cardamom Dry mature fruit crush, with 80% alcohol heat reflux extract, merge extract solution, be concentrated into no alcohol taste, successively with petroleum ether, Ethyl acetate and water saturated extracting n-butyl alcohol, respectively obtain petroleum ether extract, acetic acid ethyl ester extract and extracting n-butyl alcohol Thing；(b) acetic acid ethyl ester extract is cleaned with macroreticular resin in step (a), successively with 10% ethanol and 75% ethanol elution, is collected 75% ethanol eluate, be concentrated under reduced pressure to obtain 75% ethanol elution thing medicinal extract；(c) 75% ethanol elution medicinal extract is used just in step (b) Phase silica gel separates, and is successively 90 with volume ratio:1、45:1、20:1、10:1 and 1:1 methylene chloride-methanol gradient elution obtains 5 Individual component；(d) component 3 is further separated with purification on normal-phase silica gel in step (c), is successively 25 with volume ratio:1、20:1 and 10:1 Methylene chloride-methanol gradient elution obtains 3 components；(e) the anti-phase silicon that component 2 is bonded with octadecylsilane in step (d) Glue separates, and with the methanol aqueous solution isocratic elution that concentration expressed in percentage by volume is 70%, collects 8-12 column volume eluent, eluent It is concentrated under reduced pressure to give pure compound (I).

3. preparation method according to claim 2, it is characterised in that：The macroreticular resin is AB-8 type macroporous absorption trees Fat.

4. pharmaceutical composition, wherein compound (I) described in the claim 1 containing therapeutically effective amount and pharmaceutically acceptable Carrier.

5. application of the compound (I) in the medicine for preparing treatment stomach cancer described in claim 1.

6. application of the pharmaceutical composition in the medicine for preparing treatment stomach cancer described in claim 4.