CN113929706B - Compound with anticancer effect - Google Patents

Compound with anticancer effect Download PDF

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CN113929706B
CN113929706B CN202010606686.4A CN202010606686A CN113929706B CN 113929706 B CN113929706 B CN 113929706B CN 202010606686 A CN202010606686 A CN 202010606686A CN 113929706 B CN113929706 B CN 113929706B
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ethyl acetate
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CN113929706A (en
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肖友利
范震
杰斯·艾美特
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Center for Excellence in Molecular Plant Sciences of CAS
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    • C12P17/18—Preparation of heterocyclic carbon compounds with only O, N, S, Se or Te as ring hetero atoms containing at least two hetero rings condensed among themselves or condensed with a common carbocyclic ring system, e.g. rifamycin
    • C12P17/181—Heterocyclic compounds containing oxygen atoms as the only ring heteroatoms in the condensed system, e.g. Salinomycin, Septamycin
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Abstract

The invention discloses a compound with anticancer effect, the molecular structure of which is shown as formula I, and the compound 1 of the invention can be used for preparing anticancer drugs:

Description

Compound with anticancer effect
Technical Field
The invention belongs to the field of medicines, and relates to a compound with an anticancer effect, in particular to a sufferer alkaloid fluenuffine A, a preparation method and application thereof in preparing anticancer medicines.
Background
One leaf of Securinega suffruticosa (academic name: flueggea suffruticosa (pall.) Baill.) is a plant of genus She De of family Euphorbiaceae, which is widely distributed in China, but not found in northwest China, and distributed in other provinces, and one leaf of Securinega suffruticosa is used in traditional Chinese medicine for treating various nerve-related diseases such as paralysis, neurasthenia, somnolence, etc. The Securinega suffruticosa contains abundant Securinega suffruticosa alkaloids, which are considered as bioactive components of Securinega suffruticosa plants, the main alkaloid component in Securinega suffruticosa is Securinega suffruticosa alkali, which is a medicament used for treating sequela of infantile paralysis, and in recent years, some novel active compounds are isolated from Securinega suffruticosa, and these novel compounds have various activities such as antitumor, antiviral, antibacterial and central nervous system. The research on alkaloids in Securinega suffruticosa can find more bioactive components from Securinega suffruticosa, and the plant resource of the traditional Chinese medicine is fully developed and utilized.
Disclosure of Invention
Some alkaloid components in the sufferer are extracted, separated and identified, and the sufferer alkaloid with a novel structure is obtained by separation, has a molecular structure shown in the following formula 1 and is named fluess affine A:
the biosynthesis pathway of the alkaloid of Securinega suffruticosa was studied to find that an oxidase FsBBE of BBE family isolated from Securinega suffruticosa plants catalyzes the oxidative condensation reaction of L-ascorbic acid (L-AA) or dehydroascorbic acid (dehydroascorbic acid, DHA) with Securinega suffruticosa to produce compound 1:
the physiological activity of the compound 1 is also studied, and the compound can be found to obviously inhibit the proliferation of tumor cells SF-268, MCF-7, hePG-2, A549 and LX-2, thereby suggesting that the compound 1 can be used for resisting cancer. These findings form the basis of the present invention. Specifically, the invention comprises the following technical scheme.
A compound with anticancer effect has a molecular structure shown in formula 1.
Another object of the present invention is to provide a process for preparing the above compound 1, mainly comprising the steps of: extracting the branches and/or leaves of Securinega suffruticosa with an organic solvent.
The above method comprises, for example, the steps of:
A. extracting the branches/leaves of the suffruticosa with ethanol, and concentrating the ethanol phase by distillation to obtain an ethanol extract;
B. re-suspending the ethanol extract with dilute hydrochloric acid aqueous solution with pH of about 6.0, and extracting with ethyl acetate; regulating pH of the water layer with ammonia water to about 8.0, extracting with ethyl acetate, mixing ethyl acetate phases, and concentrating by distillation to obtain total alkaloids;
C. subjecting the total alkaloids to silica gel column chromatography, and collecting the fraction mainly containing compound 1;
D. compound 1 was isolated from the fraction obtained in step C and purified.
In a specific embodiment, the above method comprises the steps of:
A. extracting the dried crushed Securinega suffruticosa branch/leaf with ethanol, for example, repeatedly extracting with 5L ethanol for 4 times for 1.8kg dry weight, and concentrating by distillation to obtain ethanol extract;
B. re-suspending the ethanol extract with dilute hydrochloric acid aqueous solution with pH of about 6.0, and extracting with ethyl acetate with equal volume for three times; regulating pH of the water layer with ammonia water to about 8.0, extracting with equal volume of ethyl acetate for three times, mixing ethyl acetate phases, and concentrating by distillation to obtain total alkaloids;
C. by CH 2 Cl 2 :MeOH separating total alkaloids by silica gel column chromatography with mixed solvent as mobile phase, and collecting the fraction mainly containing compound 1;
D. compound 1 was isolated from the fraction obtained in step C and purified.
As another embodiment, the method of preparing compound 1 comprises the steps of:
a compound 1 is prepared by using L-ascorbic acid or dehydroascorbic acid and Securinega suffruticosa as substrate raw materials through oxidative condensation reaction under the catalysis of oxidase FsBBE of BBE family derived from Securinega suffruticosa.
Preferably, the amino acid sequence of the oxidase FsbBE is SEQ ID NO:1:
MNPLKHSSSTPLVFVLLTVCSCATSVTIPELFFQCLSNTTTTSTSIFNVLYTPRNTSYTSILESRIQNLRFNTTDTPKPLAIVTPLDASHIQATIICARKHNLQIRIRSGGHDYEGLSYVSPLPFVVLDLINLRNITVDVENRVAWVGCGATLGEFYYRIAEKTRTLAFPAGACPTVGVGGHFSGGGYGYLLRKFGLAADNILDASLVDVNGRILDRASMGEDLFWAIRGGGGNSFGVVIAWKVNLVPVPSTLTSFKVSKSLEQNTMIQLLNKWQYVANKLPDELSMFAVVSKKNSTISVKFYSLYVGGIDSLLPLMEERFPELGLKRADCNEMSWIESAVSFAGYASNTSLDVLLNHTNNYEIASGRFKGKSDFVKEPVPEAALEGLLKWLSDKDITNAAIYMVPLGGKMGEITETSIPFPHRAGNLYLLAYYVKWEGQGTEAAQKPLSWIRKGYKYMAPYVSKNPREAHLNDRDLDIGTNNISGNTSYEQASIWGTKYFKNNFDRLVRVKTSVDPSDFFRNEQSVPPLLS(SEQ IDNO::1)。
in a specific operation, the oxidase FsBBE as a biocatalyst may be either a crude enzyme such as Securinega suffruticosa total protein, or a purified oxidase FsBBE, or an oxidase FsBBE expressed by a microorganism such as E.coli engineering bacteria.
Another object of the present invention is to provide the use of compound 1 in the preparation of an anticancer drug.
For example, the cancer is selected from the group consisting of neural cancer, breast cancer, liver cancer, lung cancer.
In the above anticancer drugs, compound 1 is used as the main active pharmaceutical ingredient.
Alternatively, the above anticancer drug may be a pharmaceutical composition comprising, in addition to compound 1 as a pharmaceutically active ingredient, at least one auxiliary therapeutic agent; or compound 1 is administered in combination with at least one adjunctive therapeutic agent.
The adjuvant therapeutic agent may be selected from antitumor drug, immunomodulator, etc.
In another embodiment, the co-administration comprises co-administration, or sequential administration.
The anticancer drug can be made into oral preparation, injection, intravenous injection, suppository, inhalant, or topical preparation.
Experiments show that the compound 1 provided by the invention can obviously inhibit proliferation of tumor cells SF-268, MCF-7, hePG-2, A549 and LX-2, and the compound 1 can be used for resisting cancers. The compound 1 has remarkable growth inhibition activity on tumor cells such as glioma cells SF-268, breast cancer cells MCF-7, liver cancer cells HePG-2, non-small cell lung cancer cells A549 and hepatic stellate cells LX-2, provides candidate compounds for researching and developing new antitumor drugs, and provides scientific basis for developing and utilizing traditional Chinese medicine plant resources.
Drawings
FIG. 1 is a diagram of the structure and two-dimensional nuclear magnetic analysis of Compound 1;
FIG. 2 is an absolute configuration diagram of X-ray determination of Compound 1;
FIG. 3 is a diagram of Compound 1 1 H-NMR spectrum;
FIG. 4 is a diagram of Compound 1 13 C-NMR spectrum;
FIG. 5 is a DEPT 135 spectrum of Compound 1;
FIG. 6 is the HSQC spectrum of Compound 1;
FIG. 7 is a diagram of Compound 1 1 H- 1 H COSY profile;
FIG. 8 is an HMBC spectra of Compound 1;
FIG. 9 is the NOESY spectrum of Compound 1;
fig. 10 is the hresis spectrum of compound 1.
Detailed Description
For analysis and identification of organisms in Securinega suffruticosaExtracting the branches and leaves of Securinega suffruticosa with organic solvent such as ethanol and ethyl acetate to obtain total alkaloids, subjecting the total alkaloids to silica gel column chromatography, and separating by semi-preparative high performance liquid phase to obtain compound 1. Using spectroscopic analysis, including high resolution electrospray ionization mass spectrometry (HRESIMS), 1 H and 13 C-NMR combines with two-dimensional nuclear magnetic resonance, infrared (IR), etc., to determine its molecular structure and its absolute configuration by X-ray diffraction.
Considering that some new compounds isolated from Securinega suffruticosa in recent years have various activities such as antitumor, antiviral, antibacterial or central nervous system, we have focused on whether compound 1 has antitumor physiological activity and examined the growth inhibitory activity of compound 1 on several tumor cell lines using cisplatin as an antitumor agent as a positive control. This comparative test shows the activity of compound 1 against various tumors, thereby potentially developing an anticancer drug.
When used in the preparation of anticancer drugs, compound 1 may be a single component or may be combined with other substances in a mixture.
The medicaments of the present invention may be formulated in conventional dosage forms well known to those skilled in the art, the preparation method chosen depending on the intended route of administration. For example, as orally administered drugs, it is possible to take the form of tablets, capsules, microcapsules, pills, pellets, powders, sustained release formulations, solutions, suspensions; as sterile solutions, suspensions or emulsions for parenteral injection; for topical (e.g. skin) administration as a patch, ointment or cream or for rectal administration as a suppository; the dosage forms of the invention also comprise intraocular dosage forms, such as gels and ointments; the dosage forms of the present invention also include liquid formulations (including pre-filled needles), powders, and the like, suitable for intravenous, intraperitoneal, topical irrigation or lavage, or intracranial administration. In other embodiments, the medicament is in a unit dosage form suitable for single administration of an accurate dose.
When the anticancer drug of the present invention is a solid pharmaceutical composition, the pharmaceutical composition may include a pharmaceutically acceptable carrier. Among the acceptable carriers are adjuvants, excipients, preservatives, absorption delaying agents, fillers, binders, adsorbents, pH buffers, disintegrants, solubilizers, flavoring agents, other carriers, other inert ingredients, or combinations thereof. The specific type and amount of such pharmaceutically acceptable carrier may be selected by the skilled artisan based on common general knowledge in the art, or may be determined by simple experimentation. These pharmaceutically acceptable carriers should not affect the pharmacological activity of compound 1 or cause adverse drug reactions.
When the medicine formulation is injection, the medicine formulation can be prepared into injection solution or freeze-dried powder injection. When the freeze-dried powder injection is prepared, a freeze-drying protective agent can be added, wherein the freeze-drying protective agent can be selected from maltose, trehalose, sucrose, mannitol, lactose, glucose, sorbitol, xylitol, erythritol, threonine or a mixture of more than two of the above.
The present invention may also provide controlled/sustained release pharmaceutical formulations containing the compounds of the present invention as active ingredients wherein the rhythm and rate of release of the active ingredient is controlled and regulated to allow less frequent administration or to improve the pharmacokinetic or toxicity profile of a given active ingredient.
The medicament of the invention is typically a single dosage unit, i.e. a unit dose, whereas in other embodiments multiple unit doses may be present, e.g. the number of dosage units required in a particular regimen or clinical situation.
The invention will be further described with reference to specific examples and drawings. It is to be understood that these examples are illustrative of the present invention and are not intended to limit the scope of the present invention. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
In the examples of the present invention, if no specific explanation is given for the experimental operating temperature, this temperature is usually referred to as room temperature (10-30 ℃).
The amounts, amounts and concentrations of various substances are referred to herein, wherein the percentages refer to weight percentages unless otherwise indicated.
The term "about" or "approximately" when used herein to describe a numerical feature means that the number represented may have a tolerance range of + -10%, + -9%, + -8%, + -7%, + -6% or + -5% or a float range.
Unless otherwise defined, the scope of the terms presented herein includes the terms and any number within the scope.
EXAMPLE 1 isolation and characterization of Compound 1
1.1 extraction and separation of Securinega Suffruticosa alkaloids
Securinega suffruticosa stems and leaves were purchased from south China Yang of Henan, 9 months in 2017 and identified by a national academy of sciences of China, shanghai mountain plant science research center Yan Yuehong researcher.
Grinding Securinega suffruticosa branch and She Ganchong together by 1.8kg, repeatedly extracting with 5L ethanol for 4 times by ultrasonic extraction, concentrating by distillation to obtain ethanol extract, re-suspending ethanol extract with 1L diluted hydrochloric acid aqueous solution with pH of 6.0, extracting with equal volume of ethyl acetate for three times, adjusting pH of water layer with ammonia water to 8.0, extracting with equal volume of ethyl acetate for three times, mixing ethyl acetate phases, concentrating by distillation to obtain total alkaloids 27.0g, and concentrating by distillation to obtain total alkaloids with CH 2 Cl 2 Meoh=100:0, 30:1,20:1,15:1,10:1,7:1,4:1,2:1,1:1,0:1 as mobile phase by silica gel column separation, TLC detection, combining similar fractions to obtain 8 fractions: F1-F8, component F5 was separated again by semi-preparative high performance liquid chromatography on a Luna C18 semi-preparative column (250X 10mm,5 μm) with mobile phase: methanol: 0.1% formic acid water=5:95-25:75, 0-10min;25:75-25:75, 10-15min;25:75-95:5, 15-25min; gradient elution is carried out for 25-30min at a ratio of 95:5, and compound 1 (t) is obtained after separation R =22.5 min). Compound 1 was dissolved in deuterated chloroform for nuclear magnetic identification in the solvent ethyl acetate: methanol=7:1, and slowly evaporated at room temperature to give a single crystal of compound 1, which was subjected to X-ray diffraction to determine its absolute configuration.
1.2 isolation and Structure identification of Compound 1
The compound 1 obtained by separation is light yellow solid, HRESIMS (high resolution electrospray ionization mass spectrum) M/z [ M+H ]] + 390.1183 it shows that its molecular formula is C 19 H 19 NO 8 The unsaturation was 11.IR spectrum showed hydroxy (3452 cm) -1 ) Carbonyl (1745 cm) -1 ) And double bond (2943 cm) -1 、1441cm -1 And 1378cm -1 ) Absorbing. 1 H and 13 C-NMR combined with two-dimensional nuclear magnetic resonance spectroscopy showed (see Table 1) that compound 1 had two lipid or amide carbonyl groups (. Delta.) C 171.4,174.3), 1 sp 2 Quaternary carbon delta C 163.7,3 sp 2 Methine (delta) C/H 148.8/7.05,122.4/6.57,111.3/5.86), sp of 4 lower fields 3 Quaternary carbon (delta) C 112.3,101.5,93.1,80.4), 4 sp 3 Methine (delta) C/H 90.9/5.05,74.4/4.52,56.5/3.92,37.0/2.11), one of which is methine at the lower field, 5 sp 3 Methylene (table 1.3.1). Comparison of the above data with reported alkaloid nuclear magnetic data isolated from Securinega suffruticosa shows that compound 1 contains allosecurine or a heterodimer of securine backbone structures. The detailed two-dimensional nuclear magnetic pattern of compound 1 shows that compound 1 has a planar structure of formula 1, and compound 1 has 8 chiral carbon atoms, of which 4 are quaternary carbons. After several attempts, we fortunately obtained a single crystal that was able to determine the absolute configuration quality of the compound, single crystal diffraction results showed that the compound was in the 2r,3r,7s,9s,2's,3' r,4'r, and 6'S configurations. We named Compound 1 as fluenuffine A.
TABLE 1 Compound 1 1 H spectrum 13 C NMR data
FIG. 1 shows the structure of Compound 1 versus two-dimensional nuclear magnetic analysis; FIG. 2 shows an absolute configuration diagram of X-ray determination of Compound 1; FIG. 3 shows Compound 1 1 H-NMR spectrum; FIG. 4 shows Compound 1 13 C-NMR spectrum; FIG. 5 shows DEPT 135 spectra of Compound 1; FIG. 6 shows the HSQC spectrum of Compound 1; FIG. 7 shows Compound 1 1 H- 1 H COSY profile; FIG. 8 shows HMBC spectra of Compound 1; FIG. 9 shows NOESY spectra of Compound 1; figure 10 shows the hresis spectrum of compound 1.
Example 2 test of anti-tumor Activity of Compound 1
Compound 1 (flueniffine A) was tested for its growth inhibitory activity on tumor cells using the SRB method (Skehan P.et al 1990).
The tumor cell lines are glioma cells SF-268, breast cancer cells MCF-7, liver cancer cells HePG-2, non-small cell lung cancer cells A549 and hepatic stellate cells LX-2.
The experimental method comprises the following steps: dissolving the compound 1 with dimethyl sulfoxide (DMSO) to obtain a mother solution with the concentration of 10mmol/L, and diluting to the required concentration with RPMI-1640 medium, wherein the positive control drug is cisplatin. Taking SF-268, MCF-7, hepG-2, A549 and LX-2 cells in logarithmic growth phase, digesting with pancreatin, staining and counting trypan blue, adjusting cell concentration to 3×10 with fresh RPMI-1640 medium after trypan blue exclusion experiment to detect cell activity greater than 95% 4 Cells were seeded in 96-well plates at a volume of 180. Mu.L of cell suspension per well and 3 blanks Kong Diaoling were set at 37℃in 5% CO 2 Culturing in an incubator for 24 hours. After the cells are attached, 20 mu L of a compound solution with a certain concentration is added into each hole, and 20 mu L of RPMI-1640 medium is added into a negative control, so that cisplatin is used as a positive control. Placing at 37deg.C 5% CO 2 After culturing in an incubator for 72 hours, 50. Mu.L of 50% cold trichloroacetic acid was added to fix the cells, and the cells were left at 4℃for 1 hour, washed with distilled water 5 times, and naturally dried in air. Then 100. Mu.L/well of SRB solution prepared with 1% glacial acetic acid at a concentration of 4mg/mL was added, stained at room temperature for 30min, the supernatant removed, and washed 5 times with 1% glacial acetic acid. Finally, 200 mu L/hole 10mmol/mL Tris solution is added for dissolution, an enzyme label instrument is used for measuring the absorbance value (A) at 570nm, and the inhibition rate of the drug to the cell growth is calculated by the following formula: cell growth inhibition ratio (%) = (1-a) Sample group /A Control group )×100％。
Experimental results: IC of compound 1 and positive control drug cisplatin prepared by the invention on tumor cell strains SF-268, MCF-7, hePG-2, A549 and LX-2 50 The values are shown in Table 2.
TABLE 2 inhibition Activity of Compound 1 on tumor cell growthn＝3)
The results show that: the compound 1 has remarkable growth inhibition activity on various tumor cells, so the invention provides a candidate compound for researching and developing new anti-tumor medicaments and provides scientific basis for developing and utilizing Chinese medicinal plant resources.
Sequence listing
<110> molecular plant science Excellent innovation center of China academy of sciences
<120> a compound having anticancer effect
<130> SHPI2010291
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 532
<212> PRT
<213> Flueggea suffruticosa (Pall.) Baill.
<400> 1
Met Asn Pro Leu Lys His Ser Ser Ser Thr Pro Leu Val Phe Val Leu
1 5 10 15
Leu Thr Val Cys Ser Cys Ala Thr Ser Val Thr Ile Pro Glu Leu Phe
20 25 30
Phe Gln Cys Leu Ser Asn Thr Thr Thr Thr Ser Thr Ser Ile Phe Asn
35 40 45
Val Leu Tyr Thr Pro Arg Asn Thr Ser Tyr Thr Ser Ile Leu Glu Ser
50 55 60
Arg Ile Gln Asn Leu Arg Phe Asn Thr Thr Asp Thr Pro Lys Pro Leu
65 70 75 80
Ala Ile Val Thr Pro Leu Asp Ala Ser His Ile Gln Ala Thr Ile Ile
85 90 95
Cys Ala Arg Lys His Asn Leu Gln Ile Arg Ile Arg Ser Gly Gly His
100 105 110
Asp Tyr Glu Gly Leu Ser Tyr Val Ser Pro Leu Pro Phe Val Val Leu
115 120 125
Asp Leu Ile Asn Leu Arg Asn Ile Thr Val Asp Val Glu Asn Arg Val
130 135 140
Ala Trp Val Gly Cys Gly Ala Thr Leu Gly Glu Phe Tyr Tyr Arg Ile
145 150 155 160
Ala Glu Lys Thr Arg Thr Leu Ala Phe Pro Ala Gly Ala Cys Pro Thr
165 170 175
Val Gly Val Gly Gly His Phe Ser Gly Gly Gly Tyr Gly Tyr Leu Leu
180 185 190
Arg Lys Phe Gly Leu Ala Ala Asp Asn Ile Leu Asp Ala Ser Leu Val
195 200 205
Asp Val Asn Gly Arg Ile Leu Asp Arg Ala Ser Met Gly Glu Asp Leu
210 215 220
Phe Trp Ala Ile Arg Gly Gly Gly Gly Asn Ser Phe Gly Val Val Ile
225 230 235 240
Ala Trp Lys Val Asn Leu Val Pro Val Pro Ser Thr Leu Thr Ser Phe
245 250 255
Lys Val Ser Lys Ser Leu Glu Gln Asn Thr Met Ile Gln Leu Leu Asn
260 265 270
Lys Trp Gln Tyr Val Ala Asn Lys Leu Pro Asp Glu Leu Ser Met Phe
275 280 285
Ala Val Val Ser Lys Lys Asn Ser Thr Ile Ser Val Lys Phe Tyr Ser
290 295 300
Leu Tyr Val Gly Gly Ile Asp Ser Leu Leu Pro Leu Met Glu Glu Arg
305 310 315 320
Phe Pro Glu Leu Gly Leu Lys Arg Ala Asp Cys Asn Glu Met Ser Trp
325 330 335
Ile Glu Ser Ala Val Ser Phe Ala Gly Tyr Ala Ser Asn Thr Ser Leu
340 345 350
Asp Val Leu Leu Asn His Thr Asn Asn Tyr Glu Ile Ala Ser Gly Arg
355 360 365
Phe Lys Gly Lys Ser Asp Phe Val Lys Glu Pro Val Pro Glu Ala Ala
370 375 380
Leu Glu Gly Leu Leu Lys Trp Leu Ser Asp Lys Asp Ile Thr Asn Ala
385 390 395 400
Ala Ile Tyr Met Val Pro Leu Gly Gly Lys Met Gly Glu Ile Thr Glu
405 410 415
Thr Ser Ile Pro Phe Pro His Arg Ala Gly Asn Leu Tyr Leu Leu Ala
420 425 430
Tyr Tyr Val Lys Trp Glu Gly Gln Gly Thr Glu Ala Ala Gln Lys Pro
435 440 445
Leu Ser Trp Ile Arg Lys Gly Tyr Lys Tyr Met Ala Pro Tyr Val Ser
450 455 460
Lys Asn Pro Arg Glu Ala His Leu Asn Asp Arg Asp Leu Asp Ile Gly
465 470 475 480
Thr Asn Asn Ile Ser Gly Asn Thr Ser Tyr Glu Gln Ala Ser Ile Trp
485 490 495
Gly Thr Lys Tyr Phe Lys Asn Asn Phe Asp Arg Leu Val Arg Val Lys
500 505 510
Thr Ser Val Asp Pro Ser Asp Phe Phe Arg Asn Glu Gln Ser Val Pro
515 520 525
Pro Leu Leu Ser
530

Claims (8)

1. A compound with anticancer effect has a molecular structure shown in formula 1:
2. a process for preparing compound 1 as claimed in claim 1, characterized by being extracted from the branches and/or leaves of suffruticosa with an organic solvent, comprising the steps of:
A. extracting the branches/leaves of the suffruticosa with ethanol, and concentrating the ethanol phase by distillation to obtain an ethanol extract;
B. resuspension the ethanol extract with dilute hydrochloric acid aqueous solution of pH6.0, extracting with ethyl acetate; regulating pH of the water layer to 8.0 with ammonia water, extracting with ethyl acetate, mixing ethyl acetate phases, and concentrating by distillation to obtain total alkaloids;
C. subjecting the total alkaloids to silica gel column chromatography, and collecting the fraction mainly containing compound 1;
D. compound 1 was isolated from the fraction obtained in step C and purified.
3. The method according to claim 2, comprising the steps of:
A. extracting the dried Securinega suffruticosa branch/leaf crushed material with ethanol, and concentrating by distillation of ethanol phase to obtain ethanol extract;
B. the ethanol extract was resuspended in dilute aqueous hydrochloric acid at pH6.0 and extracted three times with an equal volume of ethyl acetate; regulating pH of the water layer to 8.0 with ammonia water, extracting with equal volume of ethyl acetate for three times, mixing ethyl acetate phases, and concentrating by distillation to obtain total alkaloids;
C. by CH 2 Cl 2 Separating the total alkaloids by silica gel column chromatography with MeOH mixed solvent as mobile phase, and collecting the fraction mainly containing compound 1;
D. compound 1 was isolated from the fraction obtained in step C and purified.
4. A process for preparing compound 1 of claim 1, comprising the steps of:
takes L-ascorbic acid or dehydroascorbic acid and other Securinega suffruticosa as substrate raw materials, prepares the compound 1 through oxidative condensation reaction under the catalysis of oxidase FsBBE of BBE family from Securinega suffruticosa,
the amino acid sequence of the oxidase FsBBE is SEQ ID NO. 1.
5. Use of compound 1 according to claim 1 for the preparation of an anticancer drug.
6. The use according to claim 1, wherein the cancer is selected from the group consisting of neural cancer, breast cancer, liver cancer, lung cancer.
7. An anticancer drug comprising the compound 1 of claim 1 as an active pharmaceutical ingredient.
8. The anticancer drug according to claim 7, wherein the anticancer dosage form is selected from an oral, injection, intravenous injection, suppository, inhalant, or external preparation.
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Citations (1)

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Publication number Priority date Publication date Assignee Title
CN110831599A (en) * 2017-02-03 2020-02-21 布罗德研究所股份有限公司 Compounds, compositions and methods for cancer treatment

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