Compound with plant growth regulation effect as well as preparation method and application thereof

Abstract

The invention provides a compound with a plant growth regulation effect as well as a preparation method and application thereof. The radix angelicae alkaloids G and H are separated and identified from the traditional Chinese medicine radix angelicae for the first time; the structure is novel, and the parent nucleus structure contains butenolactone structures which are the same as those in plant signal molecule child strigolactone, trimethyl butenolactone and the like; however, the natural contents of the two angelica dahurica alkaloids are extremely low, so that the angelica dahurica alkaloids G and H are synthesized by utilizing a chemical method, and a derivative is obtained. Researches find that the angelica dahurica alkaloid and the derivative thereof have a remarkable promoting effect on germination of dicotyledonous crop seeds and monocotyledonous crop seeds under low concentration, and have an inhibiting effect under high concentration. The angelica dahurica alkaloid and the derivative thereof can be a class of new plant hormone-like molecules, and can play a role in regulating and controlling the growth of plants.

Description

Technical Field

[0001] The present invention belongs to the field of biotechnology, and in particular relates to a compound having a plant growth regulating function, a preparation method and an application thereof. Background Art

[0002] Angelica dahurica is a commonly used Chinese medicine in clinical practice. It has the effects of dispelling cold, dispelling wind and relieving pain, clearing the nasal passages, drying dampness and stopping leukorrhea, and reducing swelling and draining pus. It has a wide range of pharmacological effects, including anti-inflammatory, analgesic, antispasmodic, antibacterial, antioxidant, and anti-tumor. Its main chemical components include volatile oils, coumarins, polysaccharides, amino acids, alkaloids, and trace elements.

[0003] The efficacy of Angelica dahurica alkaloids has been reported before, such as the literature (Wang Jianxin, Ma Cuihua, Inhibitory effect of Angelica dahurica alkaloids on HeLa cells, Chinese Journal of Gerontology, Vol. 34, No. 7, 2014) and the literature (Li Kun, Li Qingwang, Liu Jing, Study on the anti-tumor activity of Angelica dahurica alkaloids on U14 cervical cancer in mice, Journal of Yanshan University, Vol. 36, No. 1, 2012) both disclosed that Angelica dahurica alkaloids have an inhibitory effect on cervical cancer.

[0004] So far, there is no report on the phytohormone activity of Angelica dahurica alkaloids. Summary of the invention

[0005] The purpose of the present invention is to provide three compounds with plant growth regulating effects and preparation methods and applications thereof.

[0006] To achieve the above-mentioned object of the invention, the technical solution of the present invention is as follows:

[0007] The present invention first provides a compound having a plant growth regulating effect, characterized in that the structural formula is as shown in formula (I), (II) or (III):

[0008]

[0009] Among the above three compounds, the compounds with structural formulas shown in formula (I) and (II) are obtained by the present invention for the first time from the Chinese medicine Angelica dahurica, and are named Angelica dahurica alkaloids G and H respectively; their structures are novel, and the mother core structure contains the same butenolide structure as the plant signaling molecules strigolactone, trimethylbutenolide, etc. However, the natural content of these two Angelica dahurica alkaloids is extremely low, so the present invention synthesizes the above two Angelica dahurica alkaloids by chemical methods, and obtains an Angelica dahurica alkaloid derivative, the structural formula of which is shown in formula (III).

[0010] The study found that the above-mentioned angelica alkaloids and their derivatives showed significant promotion effects on the germination of dicotyledonous and monocotyledonous crop seeds at low concentrations, but showed inhibitory effects at high concentrations. This suggests that the above-mentioned angelica alkaloids and their derivatives may be a new class of plant hormone-like molecules that can regulate plant growth.

[0011] Based on this, the present invention also provides the use of the above-mentioned compound with plant growth regulating effect in the preparation of plant growth regulating reagents; the plant growth regulating reagents include seed germination promoting reagents or seed germination inhibiting reagents.

[0012] At the same time, the present invention also provides a seed germination promoting reagent, the active ingredient of which includes at least one of the above three compounds with plant growth regulating effects, and the content does not exceed 100μM; and a seed germination inhibiting reagent, the active ingredient of which includes at least one of the above three compounds with plant growth regulating effects, and the content is not less than 1000μM.

[0013] As mentioned above, angelica alkaloids G and H are isolated and identified from angelica dahurica, and the isolation method comprises the following steps:

[0014] (1) extracting the angelica dahurica by heating and refluxing to obtain an extract, and concentrating the extract to obtain an extract;

[0015] (2) suspending the extract with water, extracting with petroleum ether, ethyl acetate and n-butanol in sequence, and concentrating the n-butanol extract under reduced pressure to obtain the n-butanol extract;

[0016] (3) gradient elution of the n-butanol extract using a macroporous resin and ethanol to obtain a 20% ethanol elution fraction;

[0017] (4) gradient elution of the 20% ethanol elution fraction using a silica gel column and an ethyl acetate-methanol mixed eluent, and obtaining an ethyl acetate-methanol elution fraction under elution with an ethyl acetate-methanol mixed eluent having a volume ratio of 15:1;

[0018] (5) The ethyl acetate-methanol elution fraction was purified by semi-preparative HPLC using an acetonitrile-water mixed eluent to obtain the above-mentioned Angelica dahurica alkaloids G and H.

[0019] Preferably, in step (5), isocratic elution is performed using an acetonitrile-water mixed eluent with a volume ratio of 20:80 and an acetonitrile-water mixed eluent with a volume ratio of 30:70.

[0020] In addition, the present invention also provides a chemical synthesis method of the above-mentioned Angelica dahurica alkaloids or their derivatives, and the chemical synthesis method comprises the following steps:

[0021] (a) dissolving 4-hydroxyisoleucine in a sulfuric acid solution to obtain a 4-hydroxyisoleucine solution; slowly adding sodium nitrite dropwise to the 4-hydroxyisoleucine solution at 0° C. and reacting under an ice bath; after the reaction is completed, extracting, drying, and concentrating in vacuo to obtain compound 2;

[0022] (b) Compound 2 was dissolved in dichloromethane, molecular sieves and silica gel of equal weight were added, pyridinium chlorochromate was dissolved in anhydrous dichloromethane at 0°C and then added dropwise to the reaction system; after the reaction was completed, the mixture was filtered, dried, concentrated and purified to obtain Compound 3;

[0023] (c) dissolving 4-hydroxyisoleucine in a dilute hydrochloric acid solution and reacting at room temperature; after the reaction is completed, vacuum concentrating to obtain compound 4;

[0024] (d) Compound 4 was dissolved in anhydrous ethanol, and compound 3 and activated 4A molecular sieve were added to react at 80° C. After the reaction, the solvent was recovered, extracted, dried, vacuum concentrated, and purified to obtain compound 5 and compound 7;

[0025] Among them, compound 5 is angelica alkaloid G, and compound 7 is an angelica alkaloid derivative;

[0026] (e) Compound 4 is racemized under the action of sodium methoxide to obtain compound 4', compound 3 and activated 4A molecular sieve are added to compound 4', and the reaction is carried out at 80° C. After the reaction is completed, the solvent is recovered, extracted, dried, vacuum concentrated, and purified to obtain compound 6 and compound 7;

[0027] Among them, compound 6 is Angelica dahurica alkaloid H, and compound 7 is an Angelica dahurica alkaloid derivative.

[0028] Compared with the prior art, the technical effects of the present invention are embodied in:

[0029] The present invention separates and identifies angelica alkaloids G and H from the traditional Chinese medicine angelica for the first time; their structure is novel, and the mother nucleus structure contains the same butenolide structure as that of plant signal molecules such as strigolactone and trimethylbutenolide; however, the natural content of these two angelica alkaloids is extremely low, so the present invention synthesizes angelica alkaloids G and H by chemical methods and obtains a derivative. Studies have found that the angelica alkaloids and their derivatives of the present invention show significant promoting effects on the germination of seeds of dicotyledonous crops and monocotyledonous crops at low concentrations, but show inhibitory effects at high concentrations. It is suggested that the above-mentioned angelica alkaloids and their derivatives may be a new type of plant hormone-like molecules that can regulate the growth of plants. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1The structure of the alkaloid G of Angelica dahurica of the present invention;

[0031] Figure 2 The structure of the alkaloid H of Angelica dahurica of the present invention;

[0032] Figure 3 The chemical synthesis route of the angelica alkaloids and their derivatives of the present invention;

[0033] Figure 4 is the structure of the Angelica dahurica alkaloid derivative (Compound 7) of the present invention;

[0034] Figure 5 It is a line graph showing the effects of the Angelica dahurica alkaloids G, H, compound 7 and nordihydroguaiaretic acid of the present invention on the germination of Arabidopsis seeds;

[0035] Wherein, Time represents the drug treatment time, Germination rate represents the germination rate, CK represents the blank control, NDGA represents nordihydroguaiaretic acid, G represents Angelica dahurica alkaloid G, H represents Angelica dahurica alkaloid H, 7 represents compound 7, the same below;

[0036] Figure 6 It is a line graph showing the effects of the angelica alkaloids G, H and compound 7 of the present invention on tobacco seed germination at 1 μM;

[0037] Figure 7 It is a line graph showing the effects of the Angelica dahurica alkaloids G, H and compound 7 of the present invention on tobacco seed germination at 10 μM;

[0038] Figure 8 It is a line graph showing the effect of the angelica alkaloid H of the present invention on soybean seed germination;

[0039] Fig. 9 It is a line graph showing the effect of gibberellin on soybean seed germination;

[0040] Among them, GA 3 It means gibberellin, the same below;

[0041] Fig.10 It is a line graph showing the effect of the angelica alkaloid H of the present invention on the germination of peanut seeds;

[0042] Fig.11 It is a line graph showing the effect of gibberellins on peanut seed germination;

[0043] Fig.12 It is a line graph showing the effects of the angelica alkaloid H and gibberellin on the germination of mung bean seeds of the present invention;

[0044] Fig.13 It is a line graph showing the effect of the alkaloid H of Angelica dahurica on the germination of wheat seeds;

[0045] Fig.14 It is a line graph showing the effects of the Angelica dahurica alkaloid G and compound 7 of the present invention on wheat seed germination;

[0046] Fig.15 This is a line graph showing the effect of gibberellins on wheat seed germination;

[0047] Fig.16 It is a line graph showing the effect of the alkaloid G of Angelica dahurica on the germination of corn seeds;

[0048] Fig.17 It is a line graph showing the effect of the alkaloid H of Angelica dahurica on the germination of corn seeds;

[0049] Fig.18 is a line graph showing the effect of compound 7 of the present invention on corn seed germination;

[0050] Fig.19 This is a line graph showing the effect of gibberellins on corn seed germination;

[0051] Fig. 20 It is a line graph showing the effect of the alkaloid G of Angelica dahurica on the germination of rice seeds;

[0052] Fig.21 It is a line graph showing the effect of the alkaloid H of Angelica dahurica on the germination of rice seeds;

[0053] Fig. 22 is a line graph showing the effect of compound 7 of the present invention on rice seed germination;

[0054] Fig.23 This is a line graph showing the effect of gibberellins on rice seed germination. DETAILED DESCRIPTION

[0055] The technical solution of the present invention is further described in detail below in conjunction with the accompanying drawings and embodiments.

[0056] The instruments and materials used in the embodiments of the present invention include:

[0057] Autopol IV fully automatic polarimeter (Rudolf, USA); HPLC ion trap time-of-flight mass spectrometry system: UFLC SIL-20AC autosampler, CTO-20AC column oven, SPD M20A UV detector, LC-20ADXR pump, IT-TOF-MS equipped with ESI ion source (Shimadzu, Japan); Varian 500 nuclear magnetic resonance instrument (Varian, USA); Shimadzu LC-20AT semi-preparative HPLC (diode array detector, binary high pressure gradient pump, Labsolution workstation, Shimadzu, Japan); Waters 2998 semi-preparative HPLC (Waters, USA); ODS semi-preparative column (SunFire TM C18, 150 mm×10 mm, 5 μm); Sephadex LH-20 packing (Amerasham Biosciences, Sweden); ODS column chromatography packing (Li-Chroprep RP-C18, 40-63 μm, Merck, Germany); D101 macroporous resin, silica gel for column chromatography (200-300 mesh) and GF254 silica gel prefabricated plate for thin layer chromatography were all produced by Qingdao Ocean Chemical Plant.

[0058] The Angelica dahurica used in the experiment was purchased from Bozhou City, Anhui Province in December 2013 and was identified by Professor Tu Pengfei of Peking University as the dried root of Angelica dahurica (Fisch.ex Hoffm.) Benth.et Hook.f. of the Umbelliferae family.

[0059] Example 1 Isolation and Identification of Angelica Dahurica Alkaloids G and H

[0060] 1. Separation of Angelica dahurica alkaloids G and H

[0061] In this example, two alkaloids of Angelica dahurica were isolated from Angelica dahurica. The specific separation and identification steps are as follows:

[0062] (1) extracting the angelica dahurica by heating and refluxing to obtain an extract, and concentrating the extract to obtain an extract;

[0063] Specifically, 29.6 kg of dried Angelica dahurica was added to 80% ethanol (3×180 L, 2 h each time) and heated under reflux for extraction three times, each time for 2 h, and concentrated under reduced pressure to obtain 1.5 kg of extract;

[0064] (2) suspending the extract with water, extracting with petroleum ether, ethyl acetate and n-butanol in sequence, and concentrating the n-butanol extract under reduced pressure to obtain the n-butanol extract;

[0065] Specifically, the extract was suspended with water, and then extracted with petroleum ether (4 times × 4 L), ethyl acetate (4 times × 4 L) and n-butanol (4 times × 4 L) in sequence, and the solvent was recovered under reduced pressure to obtain 805 g of n-butanol extract;

[0066] (3) gradient elution of the n-butanol extract using a macroporous resin and ethanol to obtain a 20% ethanol elution fraction;

[0067] Specifically, the n-butanol extract was gradiently eluted with 100%, 80%, and 20% ethanol using a macroporous resin to obtain a 100% ethanol elution portion Fr.A, an 80% ethanol elution portion Fr.B, and a 20% ethanol elution portion Fr.C, respectively;

[0068] (4) gradient elution of the 20% ethanol elution fraction using a silica gel column and an ethyl acetate-methanol mixed eluent, and obtaining an ethyl acetate-methanol elution fraction under elution with an ethyl acetate-methanol mixed eluent having a volume ratio of 15:1;

[0069] Specifically, the 20% ethanol elution fraction Fr.C was loaded onto a silica gel column with a gradient elution of ethyl acetate-methanol (15:1→1:1) to obtain 20 fractions Fr.C1 to C20, from which Fr.C1 (10.1 g) was taken for subsequent experimental steps;

[0070] (5) The ethyl acetate-methanol elution fraction was purified by semi-preparative HPLC using an acetonitrile-water mixed eluent to obtain two angelica alkaloids;

[0071] Specifically, Fr.C1 (10.1 g) was separated and purified by semi-preparative HPLC (acetonitrile-water, 20:80) to obtain 12 fractions Fr.C1A to C1L. Fr.C1H (10 mg) was separated and purified by semi-preparative HPLC (acetonitrile-water, 30:70) with isocratic elution to obtain Angelica dahurica alkaloid G (3.1 mg, t R =19min), Angelica dahurica alkaloid H (0.8mg, tR=20min).

[0072] 2. Identification of alkaloids G and H from Angelica dahurica

[0073] Among them, Angelica dahurica alkaloid G is a colorless oily liquid; specific rotation [α] 2 D 5 +55(c 0.1,MeOH); UVλ m M a e x OH (logε):220(0.85),266(3.00); IR(KBr)ν max:1746,1677,1329,1232,1174,1045cm -1 ; 1 H NMR (500 MHz, CDCl 3 )δ4.73(q,J=6.0Hz,1H),4.31(d,J=12.0Hz,1H),4.07(dq,J=9.5,6.0Hz,1H),3.63(s,1H),1.9 5-1.89(m,1H),1.87(s,3H),1.39(d,6.0Hz,3H),1.35(d,J=6.5Hz,3H),1.20(d,J=6.5Hz,3H). 13 C NMR (126 MHz, CDCl 3 )δ176.3,170.3,131.5,128.5,79.6,78.5,59.9,46.4,18.8,18.5,13.9,10.5; positive ion mode HRESIMS m / z 240.1231 [M+H]+ (theoretical value is 240.1230).

[0074] The structural formula of Angelica dahurica alkaloid G is Figure 1 shown.

[0075] Angelica dahurica alkaloid H is a colorless oily liquid; specific rotation [α] 2 D 5 –49(c 0.1,MeOH); UVλ H Me m O a x (logε):238(0.99),265(1.35); IR(KBr)ν max :1747,1678,1330,1234,1176,1070,1023cm -1 ; 1 HNMR (500MHz, CDCl 3 )δ4.76(q,J=7.0Hz,1H),4.72(d,J=7.5Hz,1H),4.33(dq,J=6.5Hz,J=2.5Hz,1H),3.66(s,1H),2. 60-2.54(m,1H),1.93(s,3H),1.43(d,J=6.5Hz,3H),1.38(d,J=7.0Hz,3H),1.05(d,J=7.0Hz,3H). 13 C NMR (126 MHz, CDCl 3)δ175.8,170.2,130.9,128.4,81.6,78.6,55.2,41.0,20.1,18.9,13.1,10.5; positive ion mode HRESIMS m / z 240.1234 [M+H]+ (theoretical value is 240.1230).

[0076] The structural formula of Angelica dahurica alkaloid H is Figure 2 shown.

[0077] Example 2 Chemical synthesis and identification of Angelica dahurica alkaloids and their derivatives

[0078] 1. Chemical synthesis of Angelica dahurica alkaloids and their derivatives

[0079] In order to obtain sufficient amount of Angelica dahurica alkaloids for subsequent research, this embodiment provides a chemical synthesis method of Angelica dahurica alkaloids and their derivatives. The synthesis route of the method is as follows: Figure 3 As shown, the following steps are included:

[0080] (a) 1.0 g (7.7 mmol) of 4-hydroxyisoleucine (Compound 1) was dissolved in 1.5 g (15.4 mmol) of sulfuric acid solution (1.0 M) to obtain a 4-hydroxyisoleucine solution; 2.0 mL of sodium nitrite (3.2 g (46.2 mmol)) was slowly added dropwise to the 4-hydroxyisoleucine solution at 0°C, and the mixture was reacted under ice bath for 3 h; after the reaction, ethyl acetate was added, and the mixture was extracted with saturated sodium carbonate solution (10 mL), and Na 2 SO 4 After drying and vacuum concentration, compound 2 was obtained;

[0081] (b) 20.13 g (1.0 mmol) of the compound was dissolved in 10.0 mL of dichloromethane, and an equal amount of molecular sieves (to remove water) and silica gel (to disperse the reaction system) were added. 431.1 mg (2.0 mmol) of pyridinium chlorochromate (PCC) was dissolved in anhydrous dichloromethane at 0°C and then added dropwise to the reaction system. After the reaction was completed, diatomaceous earth was added, ethyl acetate was added for filtration, and the mixture was back-extracted with water three times and then with Na 2 SO 4 The organic layer was dried, concentrated in vacuo, and purified by silica gel column chromatography (petroleum ether: ethyl acetate = 2:1) to obtain compound 3;

[0082] (c) 1.29 g (10.0 mmol) of 4-hydroxyisoleucine was dissolved in 55 mL (12.0 mmol) of 5% dilute hydrochloric acid solution, and the mixture was stirred at room temperature for reaction; after the reaction was completed, the mixture was concentrated in vacuo to obtain compound 4;

[0083] (d) Compound 4 (1.0 mmol) was dissolved in 5 mL of anhydrous ethanol, and compound 3 and activated 4A molecular sieve were added, and the mixture was stirred at 80° C. After the reaction, the ethanol was dried and extracted with ethyl acetate. 2 SO 4 Drying, vacuum concentration, and separation by silica gel column chromatography (petroleum ether: ethyl acetate = 4:1) gave Compound 5 and Compound 7;

[0084] (e) Compound 4 was racemized under the action of sodium methoxide to obtain compound 4', compound 3 and activated 4A molecular sieve were added to compound 4' (1.0 mmol), and the reaction was carried out at 80° C. After the reaction was completed, the solvent was dried by spin drying, ethyl acetate was added for extraction, and Na 2 SO 4 After drying, vacuum concentration and separation by silica gel column chromatography (petroleum ether:ethyl acetate=4:1), compound 6 and compound 7 were obtained.

[0085] 2. Physicochemical constants and NMR data of intermediates, alkaloids of Angelica dahurica and their derivatives

[0086] Compound 2, Compound 3, Compound 5, Compound 6 and Compound 7 were analyzed.

[0087] (1) Compound 2: colorless oily liquid; specific rotation [α] 2 D 5 –74(c 0.1,MeOH); IR(KBr)ν max :3422, 2977, 1773, 1385, 1210, 1134, 1058, 1003cm -1 ; 1 H NMR (400 MHz, CDCl 3 )δ4.46(d,J=7.2Hz,1H),4.38(qd,J=6.44.4Hz,1H),2.97(s,1H),2.32-2.24(m,1H),1.40(d,J=6.4Hz,3H),1.11(d,J=7.2Hz,3H). 13 C NMR (126 MHz, CDCl 3 )δ177.3,82.1,70.0,41.5,19.7,11.1. Positive ion mode HRESIMS m / z 131.0701[M+H] + (Molecular formula C 6 H 10 O 3 , the theoretical value is 131.0703).

[0088] (2) Compound 3: colorless oily liquid; specific rotation [α] 2 D 5 –20(c 0.1,MeOH); IR(KBr)ν max :1685,1401,1145,1053,808cm -1 ; 1 H NMR (500 MHz, CDCl 3 )δ5.60(s,1H),4.84(s,1H),1.91(s,3H),1.43(s,3H). 13 C NMR (126 MHz, CDCl 3 )δ170.2,136.9,132.2,78.2,18.6,9.3. Positive ion mode HRESIMS m / z 129.0543 (molecular formula C 6 H 8 O 3 , the theoretical value is 129.0546).

[0089] (3) Compound 5: colorless oil; 1 H NMR (400 MHz, CDCl 3 )δ4.75(q,J=6.4Hz,1H),4.30(d,J=12.0Hz,1H),4.07(dq,J=9.6,6.0Hz,1H),3.52(s,1H),1.96 -1.86(m,1H),1.90(s,3H),1.42(d,J=6.0Hz,3H),1.37(d,J=6.4Hz,3H),1.23(d,J=6.4Hz,3H). 13 C NMR (101 MHz, CDCl 3 )δ176.3,170.3,131.6,128.5,79.7,78.6,60.0,46.6,18.7,18.7,14.0,10.6.HRESIMS m / z 240.1231[M+H] + (calcd for C 12 H 17 NO 4 ,240.1230). The NMR data of compound 5 are consistent with those of Angelica dahurica alkaloid G.

[0090] (4) Compound 6: colorless oil, 1 H NMR (600 MHz, CDCl 3)δ4.75(q,J=6.6Hz,1H),4.70(d,J=7.2Hz,1H),4.30(dq,J=6.6,2.4Hz,1H),3.74(s,1H),2.48 -2.43(m,1H),1.90(s,3H),1.39(d,J=6.6Hz,3H),1.34(d,J=6.6Hz,3H),1.02(d,J=7.2Hz,3H). 13 C NMR (151 MHz, CDCl 3 )δ175.8,170.1,130.7,128.1,81.5,78.6,55.0,40.8,20.0,18.7,12.9,10.4.HRESIMS m / z 240.1234[M+H] + (calcd for C 12 H 17 NO 4 ,240.1230). The NMR data of compound 6 are consistent with those of Angelica dahurica alkaloid H.

[0091] (5) Compound 7: yellow oily liquid; specific rotation [α] 2 D 5 –15(c 0.1,MeOH);UVλ H Me m O a x (logε):237(0.94),285(1.947); IR(KBr)ν max :1751,1683,1329,1236,1165,1064cm -1 ; 1 HNMR (400MHz, CDCl 3 )δ5.09(s,1H),4.88(q,J=6.8Hz,2H),1.89(s,6H),1.46(dd,J=6.7,1.4Hz,6H). 13 C NMR (101 MHz, CDCl 3 )δ170.3,141.0,125.4,78.7,18.7,11.1. Positive ion mode HRESIMS m / z 238.1066[M+H] + (The theoretical value is 238.1070).

[0092] The structure of compound 7 is shown in Figure 4 , which is the Angelica dahurica alkaloid derivative of the present invention.

[0093] Example 3 Analysis of the biological activity of alkaloids of Angelica dahurica

[0094] 1. Effects of Angelica dahurica alkaloids on seed germination of Arabidopsis thaliana and tobacco

[0095] Take the drug to be tested and nordihydroguaiaretic acid (NDGA) and prepare a solution with a concentration of 10.0mM / mL with DMSO, and filter it with a filter membrane. Weigh 2.15g of MS salt and 15g of sucrose, dissolve them in 950mL of distilled water, adjust the pH to 5.8 with NaOH, and then add distilled water to make it 1L. Dispense into 300mL conical flasks, 180mL per bottle, and add 1.44g of agar to each bottle. Sterilize at 120℃ and transfer to the clean bench in time at 90℃. After the culture medium cools down slightly, add the corresponding volume of the solution, mix it thoroughly, pour the plate, cover it, and put it on the clean bench for use. Take Arabidopsis and tobacco seeds and place them in a 1.5mL EP tube, add 1mL of 75% ethanol to disinfect for 30s (timer timing), absorb the ethanol and add 1mL of sterile water to wash for 1min. After absorbing the water, add 1mL 2% sodium hypochlorite solution to disinfect for 10min. After absorbing the disinfectant, wash with sterile water for 1min each time, repeat 6 times, and put it in water for standby. Sow the disinfected seeds in a culture dish, 30 seeds for each treatment, repeat 3 times, a total of 90 seeds, seal it with two layers of sealing film, observe whether there are water drops on the upper layer of the culture dish, if there are, test it with a flame, otherwise it will affect the observation. Put it in a 22±1℃ climate box (16h light) to observe whether the alkaloids of Angelica dahurica G, H and compound 7 have any effect on the germination of Arabidopsis and tobacco seeds.

[0096] (1) Effects of Angelica dahurica alkaloids G, H and compound 7 on Arabidopsis thaliana seed germination

[0097] The results are shown in Table 1 and Figure 5 shown.

[0098] Table 1 Effects of Angelica dahurica alkaloids G, H and compound 7 on Arabidopsis thaliana seed germination

[0099]

[0100] From Table 1 and Figure 5 It can be seen that at 16h, 21h and 24h, it can be clearly seen that under the action of 10μM Angelica dahurica alkaloids G and H and compound 7, the germination rate of Arabidopsis thaliana is much higher than that of the blank control group CK. Among them, the promotion effect is most obvious at 21h, and the germination rate of the Angelica dahurica alkaloids G and H and compound 7 treatment groups is 53.63%, 37.62% and 49.89% higher than that of the blank control group CK, respectively. After being treated with Angelica dahurica alkaloids G and compound 7, the germination rates of Arabidopsis thaliana seeds at 16h were 40.58±1.45% and 43.84±4.94%, respectively, which are higher than the germination rate of the positive control group NDGA (39.29±7.26%).

[0101] (2) Effects of Angelica dahurica alkaloids G, H and compound 7 on tobacco seed germination

[0102] The effects of Angelica dahurica alkaloids G, H and compound 7 on the germination of tobacco seeds were observed. The results are shown in Tables 2, 3 and Figure 6 , Figure 7 shown.

[0103] Table 2 Effects of Angelica dahurica alkaloids G, H and compound 7 on tobacco seed germination (concentration: 1 μM)

[0104]

[0105] Table 3 Effects of Angelica dahurica alkaloids G, H and compound 7 on tobacco seed germination (concentration: 10 μM)

[0106]

[0107] From Table 2, Table 3 and Figure 6 , Figure 7 It can be seen that at 40h, 44h, 48h, 56h and 68h, under the action of 1μM angelica alkaloid H, the germination rate of tobacco increased by 27.78%, 33.33%, 53.33%, 52.22% and 21.11% compared with the control group; under the action of 10μM angelica alkaloid H, the germination rate of tobacco increased by 32.22%, 45.55%, 55.55%, 56.67% and 23.33% compared with the control group, and the germination rate was much higher than the seed germination rate of the positive control group NDGA. Angelica alkaloid G and compound 7 (1μM, 10μM) can promote the germination of tobacco, and the germination rate is close to the seed germination rate of the positive control group NDGA.

[0108] 2. Effects of Angelica dahurica alkaloids on the germination of peanut, mung bean, soybean, corn, rice and wheat seeds

[0109] Select seeds of uniform size, add 75% ethanol to disinfect, absorb the ethanol and add 500mL sterile water to wash. After absorbing the water, add 1% sodium hypochlorite solution to disinfect for 10 minutes, wash with sterile water for 1 minute each time, repeat 6 times, and dry the disinfected seeds for later use. Soak the seeds at different concentrations (refer to the literature for soaking time). Then dry the soaked seeds and place them evenly and neatly in a 90mm diameter culture dish covered with double-layer filter paper. Each treatment has 50 seeds, repeated 3 times, a total of 150 seeds. Germination at 25℃, light 16h conditions. Add clean water regularly every day from the day of bed placement to ensure that the germination bed is moist. When the length of the seed radicle exceeds half of the seed length, it can be recorded as germination, and the germination time is counted from the sowing date. After the seeds germinate, record the number of germinations every day and measure and calculate the seed germination index.

[0110] (1) Effects of Angelica dahurica alkaloid H on soybean seed germination

[0111] The results are shown in Table 4 and Figure 8 , Fig. 9 .

[0112] Table 4 Effects of Angelica dahurica alkaloid H on soybean seed germination

[0113]

[0114] From Table 4 and Figure 8 , Fig. 9 It can be seen that three days before sowing, angelica alkaloid H (50, 100, 200 μM) had a promoting effect on the germination rate of soybean seeds, and the effect of angelica alkaloid H (100 μM) was better than that of the positive drug GA 3 (100μM), after treatment with Angelica dahurica alkaloid H (100μM), the germination rates of soybeans in the first three days were 21.11±1.11%, 54.44±4.84%, and 71.11±2.94%, respectively, while the positive drug GA 3 The germination rates of soybeans in the group (100 μM) were 7.78±1.11%, 52.22±4.84%, and 63.33±1.92%. Compared with the control group, angelica alkaloid H significantly promoted the growth of soybean radicle, and the effect was better than that of the positive control GA. 3 .

[0115] (2) Effects of Angelica dahurica alkaloid H on peanut seed germination

[0116] The results are shown in Table 5 and Fig.10 , Fig.11 .

[0117] Table 5 Effects of Angelica dahurica alkaloid H on peanut seed germination

[0118]

[0119] From Table 5 and Fig.10 , Fig.11 It can be seen that on the 3rd and 4th days after sowing, Angelica dahurica alkaloid H (50, 100, 200 μM) promoted the germination rate of peanut seeds, but the effect was not obvious.

[0120] (3) Effects of Angelica dahurica alkaloid H on mung bean seed germination

[0121] The results are shown in Table 6 and Fig.12 .

[0122] Table 6 Effects of Angelica dahurica alkaloid H on mung bean seed germination

[0123]

[0124] From Table 6 and Fig.12 It can be seen that on the first day after sowing, Angelica dahurica alkaloid H (50, 200 μM) had an inhibitory effect on the germination rate of mung bean seeds, but had no obvious effect at 100 μM.

[0125] (4) Effects of Angelica dahurica alkaloids G and H and compound 7 on wheat seed germination

[0126] The results are shown in Tables 7, 8, 9 and Fig.13 , Fig.14 , Fig.15 .

[0127] Table 7 Effects of Angelica dahurica alkaloid G and compound 7 on wheat seed germination

[0128]

[0129] Table 8 Effects of Angelica dahurica alkaloid H on wheat seed germination

[0130]

[0131] Table 9GA 3 Effects on wheat seed germination

[0132]

[0133] From Table 7, Table 8, Table 9 and Fig.13 , Fig.14 , Fig.15 It can be seen that Angelica dahurica alkaloids G (10μM) and H (10μM, 50μM) have a weak promoting effect on wheat seed germination, compound 7 has no obvious effect, and Angelica dahurica alkaloids H (1000μM, 500μM, 200μM, 100μM) has a significant inhibitory effect on wheat seed germination.

[0134] (5) Effects of Angelica dahurica alkaloids G and H and compound 7 on corn seed germination

[0135] The results are shown in Tables 10, 11, 12, 13 and Fig.16 , Fig.17 , Fig.18 , Fig.19 .

[0136] Table 10 Effect of Angelica dahurica alkaloid G on corn seed germination

[0137]

[0138] Table 11 Effects of Angelica dahurica alkaloid H on corn seed germination

[0139]

[0140] Table 12 Effect of compound 7 on corn seed germination

[0141]

[0142] Table 13GA 3 Effects on corn seed germination

[0143]

[0144] From Table 10, Table 11, Table 12, Table 13 and Fig.16 , Fig.17 , Fig.18 , Fig.19 It can be seen that on the 1st and 2nd day after sowing, Angelica dahurica alkaloids G, H and compound 7 (0.01μM, 0.1μM, 1μM, 10μM, 100μM) all significantly promoted the germination rate of corn seeds. Angelica dahurica alkaloids G (0.1μM), Angelica dahurica alkaloids H (0.1μM, 1μM), and compound 7 (1μM, 10μM) were the optimal concentrations for their effects.

[0145] On the first day, the germination rate of corn seeds treated with Angelica dahurica alkaloid G (0.1 μM) reached 56.00 ± 2.31%, the germination rate of corn seeds treated with Angelica dahurica alkaloid H (0.1 μM, 1 μM) reached 70.67 ± 1.33%, 62.67 ± 2.67%, and the germination rate of corn seeds treated with compound 7 (1 μM, 10 μM) reached 64.00 ± 4.00%, 68.00 ± 2.31%, while the germination rate of the control group was 20.00 ± 2.31%. It is worth noting that the germination promoting effect of Angelica dahurica alkaloid H (0.1 μM, 1 μM) on corn seeds and GA 3 (10μM) (corn germination rate was 69.33±4.81%).

[0146] On the second day, the germination rate of corn seeds treated with Angelica dahurica alkaloid G (0.1 μM) reached 85.33 ± 2.67%, the germination rate of corn seeds treated with Angelica dahurica alkaloid H (0.1 μM, 1 μM) reached 88.00 ± 2.31%, 93.33 ± 1.33%, and the germination rate of corn seeds treated with compound 7 (1 μM, 10 μM) reached 80.00 ± 2.31%, 78.67 ± 2.67%, while the germination rate of the control group was 53.33 ± 1.33%. In addition, Angelica dahurica alkaloids G, H and compound 7 (1000 μM) can significantly delay the germination of corn seeds.

[0147] (6) Effects of Angelica dahurica alkaloids G and H and compound 7 on rice seed germination

[0148] The results are shown in Tables 14, 15, 16, 17 and Fig. 20 , Fig.21 , Fig. 22 , Fig.23 .

[0149] Table 14 Effect of Angelica dahurica alkaloid G on rice seed germination

[0150]

[0151] Table 15 Effects of Angelica dahurica alkaloid H on rice seed germination

[0152]

[0153] Table 16 Effect of compound 7 on rice seed germination

[0154]

[0155] Table 17GA 3 Effects on rice seed germination

[0156]

[0157] From Table 14, Table 15, Table 16, Table 17 and Fig. 20 , Fig.21 , Fig. 22 , Fig.23 It can be seen that two days before sowing, Angelica dahurica alkaloids G (1 μM), H (1 μM), and compound 7 (0.1 μM) can significantly promote the germination of rice seeds.

[0158] On the first day, the germination rate of rice seeds treated with Angelica dahurica alkaloid G (0.1 μM) reached 36.67±1.76%, the germination rate of rice seeds treated with Angelica dahurica alkaloid H (1 μM) reached 46.67±1.33%, and the germination rate of rice seeds treated with compound 7 (0.1 μM) reached 58.67±2.91%, while the germination rate of the control group was 12±2.31%. Angelica dahurica alkaloid H (1 μM) has a higher effect on promoting rice seed germination than GA 3 (1μM) (germination rate 33.33±1.76); Compound 7 (0.1μM) has a higher effect on promoting rice seed germination than GA 3 (0.1 μM) (germination rate 22.00 ± 2.00).

[0159] On the second day, the germination rate of rice seeds treated with Angelica dahurica alkaloid G (1 μM) reached 77.33 ± 0.67%, the germination rate of seeds treated with Angelica dahurica alkaloid H (1 μM) reached 86 ± 3.46%, and the germination rate of seeds treated with compound 7 (0.1 μM) reached 78.67 ± 1.33%, while the germination rate of seeds in the control group was 42 ± 4.16%. In addition, Angelica dahurica alkaloids G, H and compound 7 (1000 μM) can significantly delay the germination of rice seeds.

[0160] From the above studies, it can be seen that Angelica dahurica alkaloids G, H and compound 7 can significantly promote the germination of Arabidopsis and tobacco seeds at low concentrations; they have a regulatory effect on the germination of dicotyledonous crops (soybean, peanut, soybean) seeds and monocotyledonous crops (rice, corn, wheat) seeds: at extremely low concentrations (0.01μM, 0.1μM, 1μM, 10μM), they have a significant promoting effect on the germination of rice and corn seeds, while at high concentrations (1000μM), they have an inhibitory effect on the germination of rice, corn and wheat seeds, suggesting that Angelica dahurica alkaloids G and H may be a new class of plant hormone-like molecules. This study is expected to discover new plant hormone-like molecules and apply them to the high-yield planting of medicinal plants or key crops, which has important academic value and application prospects.

Claims

1. A compound having a plant growth regulating effect, characterized in that: The structural formula is shown in formula (I), (II) or (III):

2. Use of the compound having plant growth regulating effect as claimed in claim 1 in the preparation of a plant growth regulating agent.

3. The use according to claim 2 is characterized in that The plant growth regulating agent includes a seed germination promoting agent or a seed germination inhibiting agent.

4. The use according to claim 3, characterized in that In the seed germination promoting agent, the content of the compound having plant growth regulating function is less than 100 μM.

5. The use according to claim 3, characterized in that In the seed germination inhibition reagent, the content of the compound having a plant growth regulating effect is not less than 1000 μM.

6. A seed germination promoting agent, characterized in that: The active ingredient includes at least one of the compounds having the structural formulas shown in formula (I), (II) and (III): And the content does not exceed 100μM.

7. A seed germination inhibitor, characterized in that the active ingredient comprises at least one of the compounds having the structural formulas shown in formula (I), (II) and (III): And the content is not less than 1000μM.

8. The method for preparing the compound having plant growth regulating effect as claimed in claim 1, characterized in that: The following steps are involved: (1) extracting the angelica dahurica by heating and refluxing to obtain an extract, and concentrating the extract to obtain an extract; (2) suspending the extract with water, extracting with petroleum ether, ethyl acetate and n-butanol in sequence, and concentrating the n-butanol extract under reduced pressure to obtain the n-butanol extract; (3) gradient elution of the n-butanol extract using a macroporous resin and ethanol to obtain a 20% ethanol elution fraction; (4) gradient elution of the 20% ethanol elution fraction using a silica gel column and an ethyl acetate-methanol mixed eluent, and obtaining an ethyl acetate-methanol elution fraction under elution with an ethyl acetate-methanol mixed eluent having a volume ratio of 15:1; (5) The ethyl acetate-methanol elution fraction was purified by semi-preparative HPLC using an acetonitrile-water mixed eluent to obtain compounds with plant growth regulating effects having structural formulas (I) and (II).

9. The preparation method according to claim 8, characterized in that: In step (5), isocratic elution is performed using an acetonitrile-water mixed eluent with a volume ratio of 20:80 and an acetonitrile-water mixed eluent with a volume ratio of 30:

70.

10. The method for preparing the compound having plant growth regulating effect as claimed in claim 1, characterized in that: The following steps are involved: (a) dissolving 4-hydroxyisoleucine in a sulfuric acid solution to obtain a 4-hydroxyisoleucine solution; slowly adding sodium nitrite dropwise to the 4-hydroxyisoleucine solution at 0° C. and reacting under an ice bath; after the reaction is completed, extracting, drying, and concentrating in vacuo to obtain compound 2; (b) Compound 2 was dissolved in dichloromethane, molecular sieves and silica gel of equal weight were added, pyridinium chlorochromate was dissolved in anhydrous dichloromethane at 0°C and then added dropwise to the reaction system; after the reaction was completed, the mixture was filtered, dried, concentrated and purified to obtain Compound 3; (c) dissolving 4-hydroxyisoleucine in a dilute hydrochloric acid solution and reacting at room temperature; after the reaction is completed, vacuum concentrating to obtain compound 4; (d) dissolving compound 4 in anhydrous ethanol, adding compound 3 and activated 4A molecular sieve, and reacting at 80° C.; after the reaction, recovering the solvent, extracting, drying, vacuum concentrating, and purifying to obtain compounds with plant growth regulating effects of structural formulas (I) and (III); (e) Compound 4 is racemized under the action of sodium methoxide to obtain compound 4', compound 3 and activated 4A molecular sieve are added to compound 4', and the mixture is reacted at 80° C. After the reaction, the solvent is recovered, extracted, dried, vacuum concentrated, and purified to obtain compounds with structural formulas (II) and (III) having plant growth regulating effects.

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