CN103421706B - Streptomycete, compound and preparation method thereof and the application in the antibacterial antitumor drug of preparation - Google Patents

Abstract

The invention discloses streptomycete, compound and preparation method thereof and the application in the antibacterial antitumor drug of preparation.This streptomycete Streptomyces? sp.SCSIO? can 10428 produce three kinds of napyradiomycins class new antibiotic compound 4-dehydro-4a-dechloronapyradiomycin? A1 (1), 3-dechloro-3-bromonapyradiomycin? A1 (2) and 3-chloro-6,8-dihydroxy-8-α-lapachone (3).This compounds has stronger antagonism gram positive bacterium effect, can be used in preparing resisting gram-positive bacterial drug; Can be used in preparing antitumor drug simultaneously.Does is therefore the present invention that preparation has antibacterial and napyradiomycins class new antibiotic compound 4-dehydro-4a-dechloronapyradiomycin that is anti-tumor activity? A1 (1), 3-dechloro-3-bromonapyradiomycin? A1 (2) and 3-chloro-6,8-dihydroxy-8-α-lapachone (3) provides a kind of new method, provides desirable candidate compounds for developing antibacterial antitumor drug that is efficient, low toxicity.

Description

Streptomyces, compound and preparation method thereof and application in preparation of antibacterial and antitumor drugs

Technical field:

The invention belongs to the field of industrial microbes, and in particular relates to a strain of Streptomyces sp. SCSIO10428 from marine sources capable of producing multiple napyradiomycins antibiotic compounds, napyradiomycin compounds produced by the bacteria, a preparation method thereof, and an antibacterial and antitumor drug. application.

Background technique:

In recent years, the discovery of new marine actinomycete resources and the screening of new active secondary metabolites have become an important direction of international marine microbial research. Searching for metabolites with novel structure, high efficiency and low toxicity from marine actinomycetes has gradually become a new source of drug discovery.

Invention content:

The first object of the present invention is to provide a strain of Streptomyces sp. SCSIO10428 of marine origin, which was preserved in the China Center for Type Culture Collection (CCTCC) on May 13, 2013, address: Wuhan University, Wuhan, China, its The deposit number is: CCTCCNO.M2013186.

The marine-sourced Streptomyces sp. SCSIO10428 of the present invention is isolated from seabed sediments in Xieyang Island, Beihai, Guangxi. The 16SrRNA was extracted by conventional methods, and its 16SrRNA sequence is shown in SEQ ID NO.1. The 16SrRNA was compared by BLAST, and the phylogenetic tree was analyzed. The phylogenetic tree is shown in Figure 1. Thus, the strain was identified as Streptomyces sp.SCSIO10428, which was deposited in the China Center for Type Culture Collection (CCTCC) on May 13, 2013, address: Wuhan University, Wuhan, China, and its preservation number is: CCTCCNO.M2013186 .

The present invention found that Streptomyces sp.SCSIO10428 of marine origin can produce a variety of napyradiomycins antibiotic compounds, including 4-dehydro-4a-dechloronapyradiomycinA1 (1), 3-dechloro-3-bromonapyradiomycinA1 (2) and 3-chloro-6,8 -dihydroxy-8-alpha-lapachone (3).

The second object of the present invention is to provide three kinds of napyradiomycins antibiotics 4-dehydro-4a-dechloronapyradiomycinA1(1), 3-dechloro-3-bromonapyradiomycinA1(2) and 3-chloro-6,8 with antibacterial and cytotoxic activity -dihydroxy-8-α-lapachone (3), its structure is shown in formula (I):

When R ₁ ＝Cl, R ₂ ＝H, R ₃ ＝NoH, R ₄ ＝geranyl, it is 4-dehydro-4a-dechloronapyradiomycinA1,

When R ₁ ＝Br, R ₂ ＝H ₂ , R ₃ ＝Cl, R ₄ ＝geranyl, it is 3-dechloro-3-bromonapyradiomycinA1,

When R ₁ =Cl, R ₂ =H ₂ , R ₃ =NoH, R ₄ =NoH, it is 3-chloro-6,8-dihydroxy-8-α-lapachone.

The third object of the present invention is to provide a kind of 4-dehydro-4a-dechloronapyradiomycinA1 (1), 3-dechloro-3-bromonapyradiomycinA1 (2) and 3-chloro-6,8-dihydroxy-8-α-lapachone (3 ) preparation method.

4-dehydro-4a-dechloronapyradiomycinA1 (1), 3-dechloro-3-bromonapyradiomycinA1 (2) and 3-chloro-6,8-dihydroxy-8-α-lapachone (3) of the present invention are obtained from Streptomyces sp. prepared and isolated from fermentation cultures of

The present invention preferably prepares 4-dehydro-4a-dechloronapyradiomycinA1(1), 3-dechloro-3-bromonapyradiomycinA1(2) and 3-chloro-6,8-dihydroxy from the fermentation culture of Streptomyces sp. SCSIO10428 by the following method -8-α-lapachone (3), the specific steps are as follows:

(a) preparing a fermentation culture of Streptomyces sp. SCSIO10428;

(b) Separate the fermentation broth and mycelium of the fermentation culture, extract the fermentation broth with ethyl acetate, combine the ethyl acetate extracts, and concentrate to obtain extract A; liquid, after recovering ethanol, the remaining water mixture was extracted with ethyl acetate, and the ethyl acetate extract was concentrated to obtain extract B, and extract A and extract B were combined to obtain a crude extract;

(c) The crude extract is subjected to silica gel column chromatography, isooctane/ethyl acetate v/v, 30/1, 16/1, 8/1, 4/1, 2/1, 1/1, 0/1 ; Chloroform/methanol v/v, 1/1, 0/1, gradient elution, collect isooctane/ethyl acetate volume ratio 8/1 eluted fraction Fr.3, collect isooctane/ethyl acetate volume Fraction Fr.4 eluted with ratio 4/1, collect fraction Fr.5 eluted with isooctane/ethyl acetate volume ratio 2/1;

(d) Fraction Fr.3 was subjected to reverse-phase medium-pressure liquid chromatography, acetonitrile/water system gradient elution, 40/60, 65/35, 80/20, 82/18, 84/16, 86/14, 93/17, 96/14, 100/0, v/v, 9 fractions Fr.3A～Fr.3I were obtained in turn, and the fraction Fr.3I was subjected to normal phase silica gel medium pressure liquid chromatography, n-hexane/ethyl acetate Gradient elution of ester system, 98/2, 96/4, 90/10, v/v, to obtain three fractions Fr. The volume ratio of chloroform/methanol=1/1, and then the second fraction Fr.3I22 was purified to obtain compound 2 (3-dechloro-3-bromonapyradiomycinA1);

(e) Fraction Fr.5 was combined into fraction Fr.4, and the combined fraction Fr.4 was subjected to reverse-phase medium-pressure liquid chromatography, acetonitrile/water system gradient elution, 45/55, 50/50, 60/40, 64/36, 66/34, 68/32, 70/30, 75/25, 80/20, 85/15, 90/10, 95/5, 100/0, v/v, in turn 13 fractions Fr.4A～Fr.4M, the fraction Fr.4I was combined into the fraction Fr.3G, and then the combined Fr.3G was subjected to reversed-phase medium pressure liquid chromatography, acetonitrile/water system gradient elution, 60/40, 70/30, 70/30, 75/25, 80/20, 85/15, 85/15, 85/15, 90/0, v/v, sequentially obtained 9 fractions Fr.3G1～Fr .3G9, wherein the second fraction Fr.3G2 is the monomer compound 3 (3-chloro-6,8-dihydroxy-8-α-lapachone); the fraction Fr.3G7 is purified to obtain compound 1 (4-dehydro -4a-dechloronapyradiomycinA1).

The fermentation culture of the Streptomyces sp. SCSIO10428 prepared in the step (a) is preferably prepared by the following method: insert the activated Streptomycesses Streptomycessp.SCSIO10428 into the seed medium, 28 ° C, 200 rpm, and cultivate for 72 hours to obtain the seed liquid. The seed liquid is inserted into the fermentation medium with an inoculum amount of 6% by volume, 28° C., 200 rpm, and shaken for 7 days to obtain a fermentation culture. The formula of the seed medium and the fermentation medium is as follows: Fraction 100%, including soluble starch 1%, bacteriological peptone 0.2%, yeast extract 0.4%, CaCO ₃ 0.1%, KBr 0.01%, Fe ₂ (SO4) ₃ 4H ₂ O 0.004%, sea salt 3% , the balance is water, pH7.0. Mix evenly according to the above components and contents, and then sterilize at 121°C for 30 minutes.

The present invention finds compound 4-dehydro-4a-dechloronapyradiomycinA1 (1), 3-dechloro-3-bromonapyradiomycinA1 (2) and 3-chloro-6, 8-dihydroxy-8-α-lapachone (3) by experiment Coccus (S.aureusATCC29213), Bacillus subtilis (B.SubtilisSCSIOBS01) and Bacillus thuringensis (B.thuringensisSCSIOBT01) and other Gram-positive bacteria have a strong inhibitory effect (compound 3 except for Staphylococcus aureus S.aureusATCC29213). Among them, the MICs of compound 2 against these three Gram-positive bacteria were as low as 0.5, 1 and 1 μgmL ^-1 , respectively, and had stronger antibacterial activity than the antibiotic ampicillin used in the positive control. At the same time, the results showed that compounds 1-3 had no inhibitory effect on Escherichia coli (Escherichia coli ATCC25922), a Gram-negative bacterium. Thus it can be seen that the compounds 4-dehydro-4a-dechloronapyradiomycinA1(1), 3-dechloro-3-bromonapyradiomycinA1(2) and 3-chloro-6,8-dihydroxy-8-α-lapachone(3) have good inhibitory effects on Gram Positive bacteria activity, is expected to be developed into a new antibacterial drug.

Therefore the fourth object of the present invention is to provide 4-dehydro-4a-dechloronapyradiomycinA1 (1), 3-dechloro-3-bromonapyradiomycinA1 (2) or 3-chloro-6,8-dihydroxy-8-α-lapachone (3) Application in the preparation of anti-gram-positive bacteria drugs.

Further preferred is the use of 4-dehydro-4a-dechloronapyradiomycinA1 (1) or 3-dechloro-3-bromonapyradiomycinA1 (2) in anti-Staphylococcus aureus, Bacillus subtilis or Bacillus thuringiensis drugs. Application of 3-chloro-6,8-dihydroxy-8-α-lapachone (3) in anti-Bacillus subtilis or Bacillus thuringiensis drugs.

The present invention finds through experiments that the compounds 4-dehydro-4a-dechloronapyradiomycinA1(1), 3-dechloro-3-bromonapyradiomycinA1(2) are effective on human breast cancer cell MCF7, human glioma cell SF268, human non-small cell lung cancer cell NCI Both -H460 and human liver cancer tumor cell HepG2 have moderate cytotoxic activity (IC ₅₀ value is between 10 and 20 μM); Cell SF268 has moderate cytotoxic activity (IC ₅₀ value about 24 μM). It shows that the compounds 4-dehydro-4a-dechloronapyradiomycinA1(1), 3-dechloro-3-bromonapyradiomycinA1(2) and 3-chloro-6,8-dihydroxy-8-α-lapachone(3) all have cytotoxic activity and are expected to Developed as a new anti-tumor drug.

Therefore, the fifth object of the present invention is to provide 4-dehydro-4a-dechloronapyradiomycinA1 (1), 3-dechloro-3-bromonapyradiomycinA1 (2) or 3-chloro-6,8-dihydroxy-8-α-lapachone (3 ) in the preparation of antitumor drugs.

Preferably, the application of compound 4-dehydro-4a-dechloronapyradiomycinA1(1) or 3-dechloro-3-bromonapyradiomycinA1(2) in the preparation of drugs for treating human breast cancer, human glioma, human non-small cell lung cancer or human liver cancer.

Preferably, the application of the compound 3-chloro-6,8-dihydroxy-8-α-lapachone (3) in the preparation of a drug for treating human glioma.

In summary, Streptomyces sp.SCSIO10428 of the present invention can produce napyradiomycins antibiotic compound 4-dehydro-4a-dechloronapyradiomycinA1 (1), 3-dechloro-3-bromonapyradiomycinA1 (2) and 3-chloro-6 with antibacterial activity, 8-dihydroxy-8-alpha-lapachone (3), this class of compounds has stronger antagonism against Gram-positive bacteria, and can be used for the preparation of anti-Gram-positive bacteria drugs; meanwhile, compounds 1 and 2 are effective against human breast cancer Cell MCF7, human glioma cell SF268, human non-small cell lung cancer cell NCI-H460, and human liver cancer tumor cell HepG2 all have moderate cytotoxic activity, and compound 3 has moderate cytotoxic activity against human glioma cell SF268, so It can be used to prepare antitumor drugs. Therefore the present invention is to prepare napyradiomycins class antibiotic compound 4-dehydro-4a-dechloronapyradiomycinA1 (1), 3-dechloro-3-bromonapyradiomycinA1 (2) and 3-chloro-6,8-dihydroxy-8- with antibacterial and antitumor activity The production and preparation of α-lapachone(3) provides a new method, which provides an ideal candidate compound for the development of highly effective and low-toxic antibacterial and antitumor drugs.

Streptomyces sp.SCSIO10428 of the present invention was deposited in China Center for Type Culture Collection (CCTCC) on May 13, 2013, address: Wuhan University, Wuhan, China, and its deposit number is: CCTCCNO.M2013186.

Description of drawings:

Figure 1 is a phylogenetic tree constructed based on 16S rDNA of Streptomyces sp.SCSIO10428;

Figure 2 is the key ¹ H- ¹ HCOSY and HMBC related NMR spectra of compounds 1-3.

detailed description:

The following examples are to further illustrate the present invention, rather than limit the present invention.

Example 1: Isolation and identification of Streptomyces sp. SCSIO10428

The marine-sourced Streptomyces sp. SCSIO10428 of the present invention is isolated from seabed sediments in Xieyang Island, Beihai, Guangxi. The 16SrRNA was extracted by conventional methods, and its 16SrRNA sequence is shown in SEQ ID NO.1. The 16SrRNA was compared by BLAST, and the phylogenetic tree was analyzed. The phylogenetic tree is shown in Figure 1. Thus, the strain was identified as Streptomyces sp.SCSIO10428, which was deposited in the China Center for Type Culture Collection (CCTCC) on May 13, 2013, address: Wuhan University, Wuhan, China, and its preservation number is: CCTCCNO.M2013186 .

Example 2: Isolation and preparation of active metabolites

1. Seed medium (fermentation medium): based on the total mass fraction of 100%, including 1% soluble starch, 0.2% bacteriological peptone, 0.4% yeast extract, 0.1% CaCO ₃ , 0.01% KBr, Fe ₂ ( SO4) ₃ ·4H ₂ O0.004%, sea salt 3%, the balance is water, pH7.0. Mix evenly according to the above components and contents, and then sterilize at 121°C for 30 minutes.

2. Fermentation

2.1. Seed culture: Inoculate activated Streptomyces sp.SCSIO10428 into 250 mL Erlenmeyer flasks each containing 50 mL of seed culture medium, culture at 28° C. and 200 rpm for 72 hours to obtain seed liquid.

2.2. Fermentation culture: the seed liquid was inserted into 20L fermentation medium with an inoculum amount of 6% (volume percentage), 28° C., 200 rpm, and vibrated for 7 days to obtain a fermentation culture.

3. Extraction: the fermentation culture is centrifuged (3500r·min ^-1 , 10min) to separate the fermentation broth from the mycelia, the fermentation broth is extracted 4 times with ethyl acetate, the ethyl acetate extract is combined, concentrated by vacuum distillation to obtain Extract A (8.5g); the mycelium was extracted 3 times with ethanol at room temperature, the extracts were combined, the ethanol was recovered under reduced pressure, and the remaining water mixture was extracted with ethyl acetate, and the ethyl acetate extract was distilled under reduced pressure Extract B (7.5g) was obtained after concentration, and extract A and extract B were combined to obtain a crude extract (16.0g).

4. Extraction, separation and identification of active compounds

The combined crude extract (16.0 g) of extract A and extract B was mixed with 35 g of silica gel (100-200 mesh), and eluted through a silica gel column gradient (isooctane/ethyl acetate, 30/1, 16 /1, 8/1, 4/1, 2/1, 1/1, 0/1; chloroform/methanol, 1/1, 0/1, v/v, 400mL for each gradient elution), to obtain 9 in sequence Fractions (Fr.1～Fr.9). Fraction Fr.3 (3g, fraction eluted with isooctane/ethyl acetate volume ratio 8/1) was subjected to C18 reverse phase medium pressure liquid chromatography (column type 35x3cm, acetonitrile/water system gradient elution, 40/ 60, 65/35, 80/20, 82/18, 84/16, 86/14, 93/17, 96/14, 100/0, v/v, each gradient elution 600mL), in order to get 9 Fraction (Fr.3A～Fr.3I). The fraction Fr.3I (367 mg, fraction eluted with acetonitrile/water volume ratio 100/0) was subjected to normal phase silica gel medium pressure liquid chromatography (prefabricated silica gel column 40-60 μm, 6 nm, 12 g, n-hexane/ethyl acetate system Gradient elution, 98/2, 96/4, 90/10, v/v) to obtain 3 fractions (Fr.3I1～Fr.3I3) in sequence. Fraction Fr.3I2 (258mg, fraction eluted with n-hexane/ethyl acetate volume ratio 96/4) was subjected to gel column chromatography (SephadexLH-20), and the elution system was chloroform/methanol volume ratio=1/1, Then its second fraction Fr.3I22 (107mg) was subjected to semi-preparative high-performance liquid chromatography (AngilentChemstation, reversed-phase column Phenomenex, Luna5μPhenyl-Hexyl, 250x10mm, 5μm, 10nm; solvent A phase, ultrapure water; solvent B phase , acetonitrile; 80% phase B constant gradient elution, 3.0mLmin ^-1 ), to obtain compound 2 (8.3mg, retention time 26.8min). Fraction Fr.5 (fraction eluted with isooctane/ethyl acetate volume ratio 2/1) was combined into fraction Fr.4 (fraction eluted with isooctane/ethyl acetate volume ratio 4/1), combined The obtained fraction Fr.4 (3.7g) was subjected to C18 reverse phase medium pressure liquid chromatography (column type 35x3cm, acetonitrile/water system gradient elution, 45/55, 50/50, 60/40, 64/36, 66 /34, 68/32, 70/30, 75/25, 80/20, 85/15, 90/10, 95/5, 100/0, v/v, each gradient elution 600mL), in order to obtain 13 Fractions (Fr.4A～Fr.4M). Fraction Fr.4I (472.0 mg, fraction eluted with acetonitrile/water volume ratio 80/20) was combined into fraction Fr.3G (1.7 g, fraction eluted with acetonitrile/water volume ratio 93/17), and then the combined The Fr.3G carries out C18 reverse phase medium pressure liquid chromatography (column type 35x3cm, acetonitrile/water system gradient elution, 60/40, 70/30, 70/30, 75/25, 80/20, 85/15 , 85/15, 85/15, 90/0, v/v, each gradient elution 720mL), obtained 9 fractions (Fr.3G1～Fr.3G9) in turn, the second fraction (Fr.3G2, The fraction eluted with acetonitrile/water volume ratio 70/30) is monomer compound 3 (13.0 mg). A part (200.0 mg) of the obtained fraction Fr.3G7 (568.0 mg, acetonitrile/water volume ratio 85/15 eluted fraction) was subjected to semi-preparative high-performance liquid chromatography (Hitachi Model D-2000 Elitestation, reversed-phase column Phenomenex, Gemini5 μ C18, 250x10mm, 5μm, 11nm; solvent phase A, ultrapure water; solvent phase B, acetonitrile; 90% B phase constant gradient elution, 3.0mLmin ^-1 ), to obtain compound 1 (8.0mg, retention time 29.5min).

Structure Identification:

Compound 1: Pale yellow oil, whose NMR data is assigned as shown in Table 1; (c0.30,MeOH); UV(MeOH)λ _max (logε):202(4.61),257(4.39),312(4.09),362(4.15)nm; IRν _max 3294,2920,1701,1616,1373 ,1261,1145,802cm ^-1 ; high-resolution mass spectrum HRESIMSm/zC ₂₅ H ₂₉ ClO ₅ (measured value [M+Na] ⁺ 467.1595, calculated value 467.1596), the degree of unsaturation is 9. The infrared spectrum IR of compound 1 has a broadband absorption at a wavelength of 3294cm ^-1 , indicating that the molecule contains multiple hydroxyl functional groups, and has absorption at a wavelength of 1701cm ^-1 , indicating that the molecule contains a conjugated carbonyl functional group. The ultraviolet absorption spectrum UV has maximum absorption wavelengths at 257, 312 and 362nm respectively, indicating that the compound contains functional groups such as aromatic rings. The following signals are present in the hydrogen spectrum: 1 active hydroxyl group (δ _H 12.57), 3 aromatic ring methine protons (δ _H 7.19, 6.92 and 6.71), two alkene protons (δ _H 5.02 and 4.98), one linked chlorine Atomic methine protons (δ _H 4.39), three methylene protons (δ _H 2.48, 2.00, and 1.94), and 5 aliphatic chain methyl protons (δ _H 1.68, 1.58, 1.53, 1.44, and 1.09) . The following signals exist in the carbon spectrum: 2 carbonyl carbons (δ _C 195.4 and 188.6), 2 phenolic hydroxyl carbons (δ _C 165.5 and 164.0), 10 sp2 hybridized methine carbons or quaternary carbons (chemical shift values in between δ _C 141.7 and 108.4), an oxygen quaternary carbon (δ _C 83.3), a chlorine quaternary carbon (δ _C 59.6), and other 8 sp3 hybridized methylene or methyl carbons (chemical shift values lower than δC 40), the 1H _- 12 carbon spectrum characteristics of compound 1 showed that it was very similar to napyradiomycins antibiotic compounds. Careful comparison of the spectral data of compound 1 with the known compound napyradiomycinA1 [Shiomi, K.; Nakamura, H.; Iinuma, H.; Naganawa, H.; Isshiki, K.; Takeuchi, T.; J.Antibiot.1986,39,494-501.], found that the two are basically the same, except for the following difference: Compound 1 has one molecule of HCl less in the molecular formula, and one more degree of unsaturation. In the NMR spectrum, compound 1 has an extra double bond between the C-4 position and C-4a: in the hydrogen spectrum, compound 1 has an extra ene hydrogen proton at the H-4 position (δ _H 6.92), At the same time, a group of methylene protons are missing (δ _H 2.48 and 2.41); in the carbon spectrum, compound 1 has one more olefinic carbon at the C-4 and C-4a positions (δ _C 136.9, CH and 137.1, C ), while missing a methylene carbon (C-4 position, δ _C 42.9, CH ₂ ) and a quaternary carbon (C-4a position, δ _C 79.2, C). Further, the difference of this structural fragment can also be confirmed from the following HMBC (Fig. 2): from H-4 to C-4a/C-5/C-10a/C-2 and from H-3 to C-4 relevant. Through these spectral data, the planar structure of compound 1 was determined as shown in 1 in formula (II), named 4-dedydro-4a-dechloronapyradiomycinA1. For the stereo configuration of compound 1, although there is no experimental data to directly prove it, we speculate that its absolute configuration may be (3R, 10aR), which is mainly based on the following two points of evidence: (i), the napyradiomycins that have been reported so far Antibiotic-like compounds, the absolute configuration of which is (3R, 10aR); (ii), from the perspective of the biosynthetic pathway of this type of compound, compound 1 is obtained by removing a molecule of HCl from napyradiomycinA1, so the C-3 position is still The same absolute configuration is retained.

Compound 2: Pale yellow oil, whose NMR data is assigned as shown in Table 1; (c0.30,MeOH); UV(MeOH)λ _max (logε):202(4.62),253(4.35),297(4.19),364(4.06)nm; IRν _max 3348,2939,1701,1612,1450 ,1369,1253,1072,1018,732cm ^-1 ; high-resolution mass spectrum HRESIMSm/zC ₂₅ H ₃₁ ClBrO ₅ (measured value [M+H] ⁺ 525.1047, calculated value 525.1038), the degree of unsaturation is 8. Comparison of Spectral Data of Compound 2 with the Known Compound napyradiomycinA1 [Shiomi, K.; Nakamura, H.; Iinuma, H.; Naganawa, H.; Isshiki, K.; Takeuchi, T.; .Antibiot.1986,39,494-501.], found that the two are basically the same, only the difference of the C-3 halogen substituent: for napyradiomycinA1, δ _C 59.0, δ _H 4.43, dd, J = 4.5 and 11.5Hz (Cl substitution ); while for compound 2, δC 51.0, δH 4.55, dd, _J = 7.5 and 8.5 Hz ( _Br substitution). Therefore, compound 2 is deduced to be 3-dechloro-3-bromonapyradiomycinA1, the structure of which is shown as 2 in formula (II). Further, the correlation signal of HMBC from H-3 to C2/C-4/C4a further confirmed this inference (Fig. 2). The determination of the absolute configuration of compound 2 is to compare the optical rotation data of compound 2 and napyradiomycinA1 (compound 2, c0.30, MeOH; napyradiomycinA1, c0.50, EtOH). Since the optical rotation of compound 2 is positive as that of napyradiomycinA1, its absolute configuration should also be the same as that of napyradiomycinA1, namely (3R, 4aR, 10aR).

Compound 3: Orange needle-shaped, its NMR data assignment is shown in Table 1; (c0.14, CHCl ₃ ); UV(MeOH)λ _max (logε):217(4.42),263(4.19),305(3.95),450(3.42)nm; IRν _max 3348,2939,1674,1608, 1280,1095,1022,887,779cm ^-1 ; high-resolution mass spectrum HRESIMSm/zC ₁₅ H ₁₄ ClO ₅ (measured value [M+H] ⁺ 309.0532, calculated value 309.0524), the degree of unsaturation is 7. The ultraviolet absorption spectrum UV has maximum absorption wavelengths at 263, 305 and 450nm respectively, indicating that the compound contains naphthoquinone chromophores. Comparison of spectral data of compound 3 with known compound (R)-3-chloro-6-hydroxy-8-methoxy-α-lapachone [Motohashi, K.; Sue, M.; Furihata, K.; Ito, S.; Seto, H., Terpenoids produced by actinomycetes: napyradiomycins from Streptomyces antimycoticus NT17.J.Nat.Prod.2008,71,595-601.] found that compound 3 only lacked the C-8 methoxyl group (δ _C 56.0, δ _H 3.87) signal, so it was speculated that the compound 3 is 3-chloro-6,8-dihydroxy-α-lapachone, and its structure is specifically shown as 3 in formula (II). Similarly, the structure of compound 3 was also confirmed by two-dimensional nuclear magnetic correlation spectroscopy (HMBC and COZY, etc.) (Figure 2). The absolute configuration of compound 3 was determined to be 3R, and the optical rotation data of 3 and (R)-3-chloro-6-hydroxy-8-methoxy-α-lapachone were also compared (3, c0.14, CHCl ₃ ); (R)-3-chloro-6-hydroxy-8-methoxy-α-lapachone, c0.12, CHCl ₃ ).

1 in formula (II) is 4-dehydro-4a-dechloronapyradiomycinA1; 2 is 3-dechloro-3-bromonapyradiomycinA1; 3 is 3-chloro-6,8-dihydroxy-8-α-lapachone.

Table 1: NMR data assignment of compounds 1-3 ( ¹ H-NMR data were measured at 500 MHz, ¹³ C-NMR data were measured at 125 MHz, samples were dissolved in CDCl ₃ for detection; coupling constants (Hz) are in parentheses).

Embodiment 3: the antibacterial activity assay of compound 1-3

The compounds 4-dehydro-4a-dechloronapyradiomycinA1(1), 3-dechloro-3-bromonapyradiomycinA1(2) and 3-chloro-6,8-dihydroxy-8-α-lapachone(3) were tested against Escherichia coli (EscherichiacoliATCC25922), Inhibitory activity (MIC) of Staphylococcus aureus (S.aureusATCC29213), Bacillus subtilis (B.SubtilisSCSIOBS01) and Bacillus thuringensis (B.thuringensisSCSIOBT01), references for experimental methods (Kumar, S.; Kannabiran, K., Antifungal activity of Streptomyces VITSVK5spp .againstdrugresistantAspergillusclinicalisolatesfrompulmonarytuberculosispatients.J.Mycol.Med.2010,20,101-107.), the results are shown in Table 2:

Table 2: Antibacterial activity of compounds 1-3

Shown by Table 2, compound 1-3 has stronger inhibitory effect on Gram-positive bacteria such as Staphylococcus aureus (S.aureusATCC29213), Bacillus subtilis (B.SubtilisSCSIOB01) and Bacillus thuringensis (B.thuringensisSCSIOBT01) ( Compound 3 is effective against Staphylococcus aureus S. aureusATCC29213 except). Among them, the MICs of compound 2 against these three Gram-positive bacteria were as low as 0.5, 1 and 1 μgmL ^-1 , respectively, and had stronger antibacterial activity than the antibiotic ampicillin used in the positive control. At the same time, the results showed that compounds 1-3 had no inhibitory effect on Escherichia coli (Escherichia coli ATCC25922), a Gram-negative bacterium. Combined data suggest that 4-dehydro-4a-dechloronapyradiomycinA1(1), 3-dechloro-3-bromonapyradiomycinA1(2) and 3-chloro-6,8-dihydroxy-8-α-lapachone(3) can be used to prepare antagonistic Drugs for Gram-positive bacteria.

Example 4: Determination of Cytotoxic Activity of Compound 1-3

The effects of compounds 4-dehydro-4a-dechloronapyradiomycinA1(1), 3-dechloro-3-bromonapyradiomycinA1(2) and 3-chloro-6,8-dihydroxy-8-α-lapachone(3) on human breast cancer cells MCF7, Inhibitory activity (IC ₅₀ ) of human glioma cell SF268, human non-small cell lung cancer cell NCI-H460 and human liver cancer tumor cell HepG2, experimental method reference (Skehan, P.; Storeng, R.; Scudiero, D. ; Monks, A.; McMahon, J.; Vistica, D.; Warren, JT; ), the results are shown in Table 3:

Table 3: Cytotoxic activity of compounds 1-3

Table 3 shows that compounds 1 and 2 have moderate cytotoxic activity on human breast cancer cell MCF7, human glioma cell SF268, human non-small cell lung cancer cell NCI-H460 and human liver cancer tumor cell HepG2 ( _IC50 values between 10～20μM); Compound 3 has no cytotoxic activity on human breast cancer cell MCF7, human non-small cell lung cancer cell NCI-H460 and human liver cancer tumor cell HepG2 (IC ₅₀ value is greater than 40 μM), but it has no cytotoxic activity on human glial Tumor cell SF268 has moderate cytotoxic activity (IC ₅₀ value is about 24 μM). The combined data indicated that 4-dehydro-4a-dechloronapyradiomycinA1(1), 3-dechloro-3-bromonapyradiomycinA1(2) and 3-chloro-6,8-dihydroxy-8-α-lapachone(3) could be used to prepare anti- Drugs for tumors.

Claims (8)

1. Streptomyces sp. SCSIO10428, its preservation number is: CCTCCNO.M2013186. 2. The compound shown in formula (I), 3-dechloro-3-bromonapyradiomycinA1: 3. A preparation method of compound 4-dehydro-4a-dechloronapyradiomycinA1, 3-dechloro-3-bromonapyradiomycinA1 and 3-chloro-6,8-dihydroxy-8-α-lapachone, characterized in that the compound 4- dehydro-4a-dechloronapyradiomycinA1, 3-dechloro-3-bromonapyradiomycinA1 and 3-chloro-6,8-dihydroxy-8-α-lapachone are prepared and isolated from the fermentation culture of Streptomyces sp. (Streptomycessp.) SCSIO10428 according to claim 1 owned; 1 in formula (II) is 4-dehydro-4a-dechloronapyradiomycinA1; 2 is 3-dechloro-3-bromonapyradiomycinA1; 3 is 3-chloro-6,8-dihydroxy-8-α-lapachone. 4. preparation method according to claim 3, is characterized in that, concrete steps are as follows: a) preparing a fermentation culture of Streptomyces sp. SCSIO10428; b) Separating the fermentation broth and mycelia of the fermentation culture, extracting the fermentation broth with ethyl acetate, combining the ethyl acetate extracts, and concentrating to obtain extract A; first extracting the mycelium with ethanol, and combining the extracts After recovering the ethanol, the remaining water mixture is extracted with ethyl acetate, the ethyl acetate extract is concentrated to obtain extract B, and extract A and extract B are combined to obtain a crude extract; c) The crude extract is subjected to silica gel column chromatography, isooctane/ethyl acetate v/v, 30/1, 16/1, 8/1, 4/1, 2/1, 1/1, 0/1; Chloroform/methanol v/v, 1/1, 0/1, gradient elution, collect isooctane/ethyl acetate volume ratio 8/1 eluted fraction Fr.3, collect isooctane/ethyl acetate volume ratio 4/1 eluted fraction Fr.4, collect isooctane/ethyl acetate volume ratio 2/1 eluted fraction Fr.5; d) Fraction Fr.3 was subjected to reverse-phase medium-pressure liquid chromatography, acetonitrile/water system gradient elution, 40/60, 65/35, 80/20, 82/18, 84/16, 86/14, 93 /17, 96/14, 100/0, v/v, 9 fractions Fr.3A～Fr.3I were obtained in turn, and the fraction Fr.3I was subjected to normal phase silica gel medium pressure liquid chromatography, n-hexane/ethyl acetate System gradient elution, 98/2, 96/4, 90/10, v/v, to obtain 3 fractions Fr.3I1～Fr.3I3 in turn, the fraction Fr.3I2 was subjected to gel column chromatography, and the elution system was Chloroform/methanol volume ratio=1/1, and then its second fraction Fr.3I22 is purified to obtain compound 3-dechloro-3-bromonapyradiomycinA1; e) Fraction Fr.5 was combined into fraction Fr.4, and the combined fraction Fr.4 was subjected to reverse-phase medium pressure liquid chromatography, acetonitrile/water system gradient elution, 45/55, 50/50, 60 /40, 64/36, 66/34, 68/32, 70/30, 75/25, 80/20, 85/15, 90/10, 95/5, 100/0, v/v, get 13 in turn Fractions Fr.4A～Fr.4M, the fraction Fr.4I was combined into the fraction Fr.3G, and then the combined Fr.3G was subjected to reversed-phase medium pressure liquid chromatography, acetonitrile/water system gradient elution, 60 /40, 70/30, 70/30, 75/25, 80/20, 85/15, 85/15, 85/15, 90/0, v/v, sequentially obtained 9 fractions Fr.3G1～Fr. 3G9, wherein the second fraction Fr.3G2 is the monomer compound 3-chloro-6,8-dihydroxy-8-α-lapachone; fraction Fr.3G7 is purified to obtain the compound 4-dehydro-4a-dechloronapyradiomycinA1. 5. The application of the Streptomyces sp. (Streptomycessp.) SCSIO10428 described in claim 1 in the preparation of the compound 3-dechloro-3-bromonapyradiomycinA1 described in claim 2. 6. The application of the compound 3-dechloro-3-bromonapyradiomycinA1 described in claim 2 in the preparation of anti-Gram-positive bacteria drugs. 7. The application according to claim 6, characterized in that the compound 3-dechloro-3-bromonapyradiomycinA1 is used in the preparation of drugs against Staphylococcus aureus, Bacillus subtilis or Bacillus thuringiensis. 8. The use of the compound 3-dechloro-3-bromonapyradiomycinA1 according to claim 2 in the preparation of medicines for treating human breast cancer, human glioma, human non-small cell lung cancer or human liver cancer.