A diterpenoid alkaloid compound from Delphinium yiliense, its preparation method and uses

Abstract

This invention relates to a diterpenoid alkaloid compound from *Delphinium ililiense*, its preparation method, and its uses. The compound is obtained by solvent extraction using *Delphinium ililiense* as raw material, followed by acid-dissolution and alkali precipitation, and then solvent extraction again. Separation is achieved using two, three, or four methods: silica gel column chromatography, semi-preparative thin-layer chromatography, or LH-20 dextran gel chromatography. Four new diterpenoid alkaloid compounds were obtained: compound 1 named *Delphinium ililiense* A; compound 2 named *Delphinium ililiense* B; compound 3 named *Delphinium ililiense* C; and compound 4 named *Delphinium ililiense* D. The analgesic activity of the obtained compounds was determined. Experimental results showed that compounds *Delphinium ililiense* C and D exhibited significant analgesic activity in an acetic acid-induced writhing mouse model and can be used to prepare analgesic drugs.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, specifically to a diterpenoid alkaloid compound from *Delphinium yiliense*, its preparation method, and its uses. Background Technology

[0002] Diterpenoid alkaloids are a class of structurally complex and bioactive nitrogen-containing natural organic compounds, mainly found in plants of the genera *Aconitum* and *Delphinium* in the Ranunculaceae family. Due to their diverse biological activities and medicinal value, especially their significant effects in antiarrhythmic and analgesic activity, they have become a subject of widespread research interest. Currently, four diterpenoid alkaloids have been developed into drugs and used in clinical treatment. To date, more than 1600 natural diterpenoid alkaloids have been reported, mainly divided into C... 18 -、C 19 -、C 20 Diterpenoid alkaloids are classified into four main categories: diterpenoids, bis(diterpenoids), and diterpenoids. Due to their complex and variable structures, and the coexistence of activity and toxicity, the search for novel, highly efficient, and low-toxicity diterpenoid alkaloid compounds remains a current research hotspot and challenge. *Delphinium iliense* Huth is a perennial herb belonging to the genus *Delphinium* in the Ranunculaceae family. It is mainly distributed in western Xinjiang, China, growing in mountain shrublands or rocky slopes at altitudes of approximately 2000 meters. It is also found in Central Asia. This plant is abundant in Xinjiang and is mainly used for clearing heat and detoxifying, treating diarrhea, dysentery, rheumatism, and enteritis.

[0003] The analgesic activity of diterpenoid alkaloids has been extensively studied. Currently, 3-acetyl aconitine, aconitine, and aconitine have been developed into non-addictive analgesics for clinical use. Diterpenoid alkaloids are both the main active ingredients and the primary source of toxicity in plants of the genera *Aconitum* and *Delphinium*. Therefore, finding lead compounds with good analgesic effects and low toxicity is our main research objective. Currently, the extraction, isolation, structural identification, and related pharmacological studies of diterpenoid alkaloids from *Delphinium yiliense* are still in their initial stages both domestically and internationally. Therefore, systematic and in-depth research on the diterpenoid alkaloid components in *Delphinium yiliense*, clarifying the material basis of their analgesic activity, discovering new diterpenoid alkaloid compounds with specific activities, and identifying highly effective and low-toxicity drug lead compounds are of great significance. Summary of the Invention

[0004] The purpose of this invention is to provide a diterpenoid alkaloid compound from *Delphinium ililiense*, its preparation method, and its uses. The compound is obtained by solvent extraction using *Delphinium ililiense* as raw material, followed by acid-dissolution and alkali precipitation, and then solvent extraction again. Separation is achieved using two, three, or four methods selected from silica gel column chromatography, semi-preparative thin-layer chromatography, or LH-20 dextran gel chromatography to obtain four new diterpenoid alkaloid compounds: compound 1 named *Delphinium ililiense* A; compound 2 named *Delphinium ililiense* B; compound 3 named *Delphinium ililiense* C; and compound 4 named *Delphinium ililiense* D. The analgesic activity of the obtained compounds was determined. Experimental results showed that compounds *Delphinium ililiense* C and D exhibited significant analgesic activity in an acetic acid-induced writhing mouse model and can be used to prepare analgesic drugs.

[0005] The present invention discloses a diterpenoid alkaloid compound isolated from *Delphinium ililiense*, the structural formula of which is as follows:

[0006]

[0007] in:

[0008] Compound 1 is named Ili delphinidin A;

[0009] Compound 2 is named Ili delphinidin B;

[0010] Compound 3 is named Ili delphinidin C;

[0011] Compound 4 is named Ili delphinidin D.

[0012] The preparation method of the diterpenoid alkaloids from *Delphinium illuminense* is carried out according to the following steps:

[0013] a. After pulverizing the whole herb of Delphinium yiliense, extract it at room temperature with an ethanol aqueous solution of 10-95% by volume, anhydrous methanol or chloroform. Anhydrous methanol and chloroform are extracted by cold soaking, percolation, heating and reflux or ultrasonic extraction. The solvent is recovered by vacuum concentration to obtain the extract.

[0014] b. Disperse the extract obtained in step a with sulfuric acid or hydrochloric acid with a mass fraction of 1-5%. Adjust the pH of the resulting acidic aqueous layer to 9-12 with anhydrous sodium carbonate, ammonia or sodium hydroxide. Then extract with organic solvents such as chloroform, ethyl acetate or n-butanol. Concentrate under reduced pressure to recover the organic solvents and obtain total alkaloids.

[0015] Alternatively, the crude extract obtained in step a can be dispersed with sulfuric acid or hydrochloric acid at a mass fraction of 1-5%. The resulting acidic aqueous solution is adjusted to pH 10 using ammonia and anhydrous Na2CO3, and then extracted with ethyl acetate. The ethyl acetate is then evaporated under reduced pressure to obtain total alkaloid extract 1. The alkaline aqueous layer is then further alkalized to pH 12 using 10% NaOH, and then extracted with n-butanol. The n-butanol is then evaporated under reduced pressure to obtain total alkaloid extract 2. Total alkaloid extract 1 and total alkaloid extract 2 are combined to obtain the total alkaloid extract.

[0016] c. The total alkaloids obtained in step b are separated by two, three, or four methods, namely silica gel column chromatography, thin-layer chromatography, dextran gel LH-20 column chromatography, or high-performance liquid chromatography, to obtain compound 1 Ilitrizin A, compound 2 Ilitrizin B, compound 3 Ilitrizin C, and compound 4 Ilitrizin D.

[0017] The method for preparing diterpenoid alkaloids from *Delphinium ilignon* of Yili, including the two separation processes in step c:

[0018] The total alkaloids obtained in step c were separated by normal-phase silica gel column chromatography. Gradient elution was performed using dichloromethane and methanol in volume ratios of 100:1, 80:1, 60:1, 40:1, 20:1, 10:1, 5:1, 3:1, 2:1, and 1:1. One fraction was collected every 500 ml. The fractions were analyzed by silica gel thin-layer chromatography. Fractions with the same composition were combined to obtain 8 components AH.

[0019] The obtained fraction A was further separated by normal-phase silica gel column chromatography or reversed-phase silica gel column chromatography, using gradient elution with petroleum ether-acetone + 0.2% diethylamine (v / v) or methanol and water (v / v) at ratios of 20:1, 10:1, 5:1, 2:1, 1:1 and 0:1, and the same fractions were combined after TLC and HPLC analysis to obtain five fractions A-1 to A-5. Fraction A-3 was purified by semi-preparative high-performance liquid chromatography and isocratically eluted with acetonitrile-0.1% formic acid aqueous solution (v / v) at a ratio of 29:71 to obtain compounds iridocycline A and iridocycline B.

[0020] The obtained fraction C was further separated by normal-phase silica gel column chromatography or reversed-phase silica gel column chromatography, using gradient elution with petroleum ether-acetone + 0.2% diethylamine in volume ratios of 20:1, 10:1, 5:1, 2:1, 1:1 and 0:1, or methanol and water in volume ratios of 5:95-100:0. After silica gel thin-layer chromatography and HPLC analysis, fractions of the same type were combined to obtain six fractions C-1 to C-6. Fraction C-3 was purified by semi-preparative high-performance liquid chromatography, using gradient elution with acetonitrile-0.1% formic acid aqueous solution in volume ratios of 20:80-30:70 to obtain compounds iridotrichum citrate C and iridotrichum citrate D.

[0021] The method for preparing diterpenoid alkaloids from *Delphinium ilignon* of Yili, including the three separation processes in step c:

[0022] The total alkaloid extract obtained in step c was separated by normal-phase silica gel column chromatography or reversed-phase silica gel column chromatography. Gradient elution was performed using chloroform-methanol at volume ratios of 40:1, 20:1, 10:1, 5:1 and 1:1 or methanol-water at volume ratios of 5:95-80:20. One fraction was collected every 500 ml and analyzed by silica gel thin-layer chromatography. Fractions with the same composition were combined to obtain 6 fractions AF.

[0023] The obtained fraction A was further separated by reversed-phase silica gel column chromatography or normal-phase silica gel column chromatography, using a gradient elution with methanol-water (v / v) of 5:95-70:30 or petroleum ether-acetone + 0.2% diethylamine (v / v) of 10:1, 5:1, 2:1, 1:1 and 0:1. After silica gel thin-layer chromatography and HPLC analysis, fractions of the same type were combined to obtain five fractions A-1 to A-5. Fraction A-3 was purified by semi-preparative high-performance liquid chromatography, and then eluted with a gradient of acetonitrile-0.1% formic acid aqueous solution (v / v) of 20:80-40:60 to obtain compounds iridotrichumine A and iridotrichumine B.

[0024] The obtained fraction C was further separated by dextran LH-20 gel column chromatography, and eluted isocratically with chloroform-methanol at a volume ratio of 1:1 or 0:1. After analysis by silica gel thin-layer chromatography and HPLC, the same fractions were combined to obtain five fractions C-1 to C-5. C-4 was purified by semi-preparative high-performance liquid chromatography and eluted isocratically with acetonitrile-0.1% formic acid at a volume ratio of 28:72 to obtain compounds iridotrichum c and iridotrichum c D.

[0025] The method for preparing diterpenoid alkaloids from *Delphinium ilignon* of Ili includes the following four separation methods in step c:

[0026] The total alkaloid extract obtained in step c was separated by dextran gel LH-20 column chromatography. Isocratic elution was performed using a chloroform-methanol solvent system with a volume ratio of 1:1 or 0:1. The fractions were analyzed by silica gel thin-layer chromatography. Fractions with the same volume were combined to obtain 9 fractions AI.

[0027] Fraction A was subjected to reversed-phase silica gel column chromatography and eluted with a methanol-water gradient of 5:95-80:20 (v / v). After TLC and HPLC analysis, the same fractions were combined to obtain four fractions A-1 to A-4. Fraction A-3 was purified by semi-preparative high-performance liquid chromatography and eluted with an acetonitrile-0.1% formic acid aqueous solution gradient of 20:80-47:53 (v / v) to obtain the compound iridocycline A.

[0028] The obtained fraction C was subjected to normal-phase silica gel column chromatography with gradient elution using petroleum ether-acetone + 0.2% diethylamine at volume ratios of 20:1, 10:1, 5:1, 2:1, 1:1 and 0:1. After analysis by silica gel thin-layer chromatography and HPLC, fractions of the same type were combined to obtain four fractions C-1 to C-4. Fraction C-2 was purified by preparative thin-layer chromatography using chloroform-methanol-water at a volume ratio of 82:16:2 to obtain compound erythritol B.

[0029] The obtained fraction E was subjected to normal-phase silica gel column chromatography with gradient elution using petroleum ether-acetone + 0.2% diethylamine at volume ratios of 10:1, 5:1, 2:1, 1:1 and 0:1. After silica gel thin-layer chromatography and HPLC analysis, the same fractions were combined to obtain five fractions E-1 to E-5. Fraction E-3 was purified by semi-preparative high-performance liquid chromatography with gradient elution using acetonitrile-0.1% formic acid aqueous solution at volume ratios of 20:80-47:53 to obtain compound erythritol citrate C.

[0030] The obtained fraction I was separated by dextran gel LH-20 column chromatography, and isocratic elution was performed using a chloroform-methanol solvent system with a volume ratio of 1:1 or 0:1. The fractions were analyzed by silica gel thin-layer chromatography, and the same fractions were combined to obtain four fractions I-1 to I-4. Fraction I-4 was purified by semi-preparative high performance liquid chromatography, and isocratic elution was performed using an acetonitrile-0.1% formic acid aqueous solution with a volume ratio of 28:72 to obtain the compound erythritol D.

[0031] The use of diterpenoid alkaloids isolated from *Delphinium yiliense* in the preparation of analgesic drugs.

[0032] This invention discloses a diterpenoid alkaloid compound from *Delphinium ililiense*, its preparation method, and its uses. Two novel diterpenoid alkaloid compounds were obtained and their analgesic activity was determined in mice. The experimental results showed that the novel diterpenoid alkaloids 3 and 4 isolated from *Delphinium ililiense* had significant analgesic effects, and the strength of the analgesic effect showed a dose-response relationship. In conclusion, these results provide experimental evidence for the compound to be a potential clinical analgesic.

[0033] This invention discloses a diterpenoid alkaloid compound from *Delphinium ililiense*, its preparation method, and its uses. The compound is obtained by solvent extraction using *Delphinium ililiense* as raw material, followed by acid dissolution and alkali precipitation, and then solvent extraction again. Separation is achieved using two, three, or four methods: silica gel column chromatography, preparative thin-layer chromatography, or LH-20 dextran gel chromatography. Four new diterpenoid alkaloid compounds were obtained: 1. *Delphinium ililiense* A; 2. *Delphinium ililiense* B; 3. *Delphinium ililiense* C; 4. *Delphinium ililiense* D. Wherein:

[0034] Compound 1, named Ili delphinidin A, is a colorless and amorphous powder. Its molecular weight was determined by HCl (+) MS (m / z 480.2592 [M+H]). + The theoretical value is 480.2592, which determines its molecular formula as C. 25 H 37 NO8; According to 1 H, 13 Its structure was determined by C10 and two-dimensional NMR data; compound 1 is the first C10 compound discovered. 19 - A diterpenoid alkaloid, named Sinchianidine A. 1 H and 13 The C NMR data attribution is shown in Table 1 [600MHz ( 1 H), 150MHz 13 C), Solvent: CD3OD];

[0035] Compound 2 is Ili delphinidin B, a colorless and amorphous powder, which was analyzed by HRESI(+) MS (m / z 466.2437 [M+H]). + (Theoretical value 466.2435) determines its molecular formula as C 24 H 35 NO8; According to 1 H, 13 Its structure was determined by C10 and two-dimensional NMR data; compound 2 is the first C10 compound discovered. 19 - A diterpenoid alkaloid, named Sinchianidine B. 1 H and 13 CNMR data attribution is shown in Table 1 [600MHz ( 1 H), 150MHz 13 C), Solvent: CD3OD];

[0036] Compound 3 is lysine C, a colorless and amorphous powder, which was analyzed by HCl (+) MS (m / z 466.2436 [M+H]). + (Theoretical value 466.2435) determines its molecular formula as C 24 H 35 NO8; According to 1 H, 13 Its structure was determined by C10 and two-dimensional NMR data; compound 3 is the first C10 compound discovered. 19 - A diterpenoid alkaloid, named Sinchianidine C. 1 H and 13 CNMR data attribution is shown in Table 1 [600MHz ( 1 H), 150MHz 13C), Solvent: CD3OD];

[0037] Compound 4 is lysine D, a colorless and amorphous powder, which was analyzed by HCl (+) MS (m / z 466.2437 [M+H]). + (Theoretical value 466.2435) determines its molecular formula as C 24 H 35 NO8; According to 1 H, 13 Its structure was determined by C10 and two-dimensional NMR data; compound 4 is the first C10 compound discovered. 19 - A diterpenoid alkaloid, named Sinchianidine D. 1 H and 13 CNMR data attribution is shown in Table 1 [600MHz ( 1 H), 150MHz 13 C), Solvent: CD3OD.

[0038] Table 1. Compounds 1-4 1 H (600MHz) and 13 C(150MHz) NMR data [δ(ppm), J(Hz)]

[0039]

[0040] Attached Figure Description

[0041] Figure 1 The compound of this invention, Ili delphinidin A 1 H NMR spectrum;

[0042] Figure 2 The compound of this invention, Ili delphinidin A 13 C NMR spectrum;

[0043] Figure 3 The compound of this invention, Ili delphinidin B 1 H NMR spectrum;

[0044] Figure 4 The compound of this invention, Ili delphinidin B 13 C NMR spectrum;

[0045] Figure 5 The compound of this invention, Ili delphinidin C 1 H NMR spectrum;

[0046] Figure 6 The compound of this invention, Ili delphinidin C 13 C NMR spectrum;

[0047] Figure 7 The compound of this invention, Ili delphinidin D 1 H NMR spectrum;

[0048] Figure 8 The compound of this invention, Ili delphinidin D 13 C NMR spectrum. Detailed Implementation

[0049] All reagents used were of analytical grade. Acetonitrile used in high-performance liquid chromatography (HPLC) was of HPLC grade (Merk, USA). Column chromatography silica gel (100-200 mesh, 200-300 mesh): produced by Qingdao Marine Chemical Plant; thin-layer chromatography silica gel was GF. 254 Produced by Yantai Huangwu Silica Gel Development and Testing Plant; Sephadex LH-20 gel: produced by Pharmacia, Sweden. High-performance liquid chromatography (Agilent Technologies, USA): P680 HPLC pump, ASI-100 autosampler, TCC-100 column oven, UVD170U ultraviolet detector (four wavelengths), quaternary eluent, online degasser, Chromeleon chromatography workstation. Preparative high-performance liquid chromatography (Jiangsu Hanbang Technology Co., Ltd.): NP7005C pump, N3000D ultraviolet detector (dual wavelengths), semi-preparative dynamic mixer, EasyChrom-1000 chromatography workstation. Mass spectrometry was performed using a QSTAR Elite mass spectrometer (Applied Biosystems / MDS Sciex); nuclear magnetic resonance (NMR) was performed using a Varian Vnmrs 600 / 400 NMR spectrometer (Varian, USA); electronic balance (METTER AC-100, Sartorious, Germany); rotary evaporator (N-1001D, Shanghai Ailang Instrument Co., Ltd.); high-precision polarimeter (UAutopol VI, Rudolph, USA);

[0050] The specimen of *Delphinium iliense* Huth, collected in July 2017 from the northern slope of the Tianshan Mountains at an altitude of 1850±20 meters in Chahanwusu Township, Zhaosu County, Ili, was identified by Associate Researcher Feng Ying of the Xinjiang Institute of Ecology and Geography, Chinese Academy of Sciences, as a plant belonging to the genus *Delphinium* of the family Ranunculeceae. The specimen is preserved at the Xinjiang Institute of Physics and Chemistry, Chinese Academy of Sciences.

[0051] Example 1 (Two Separation Methods)

[0052] a. Take 10.0 kg of whole plant of Delphinium yiliense and crush it. Soak the plant in 5% Na2CO3 solution at room temperature for 3 hours. Add 50 L of chloroform:methanol mixed solution with a volume ratio of 6:1. Extract by cold soaking. Evaporate the solvent under reduced pressure to obtain crude extract of Delphinium yiliense.

[0053] b. Disperse the crude extract obtained in step a with a 1% sulfuric acid solution. Adjust the pH of the resulting acidic aqueous layer to 10 with anhydrous Na2CO3, then extract with ethyl acetate. Evaporate the ethyl acetate under reduced pressure to obtain total alkaloid extract 1. Further alkalize the alkaline aqueous layer with 10% NaOH to pH 12, then extract with n-butanol. Evaporate the n-butanol under reduced pressure to obtain total alkaloid extract 2. Combine total alkaloid extract 1 and total alkaloid extract 2 to obtain the total alkaloid extract.

[0054] c. The obtained total alkaloid extract was separated by normal-phase silica gel column chromatography. Gradient elution was performed using chloroform and methanol in volume ratios of 100:1, 60:1, 40:1, 20:1, 10:1, 5:1, 3:1, and 1:1. One fraction was collected for every 500 ml. The fractions were analyzed by silica gel thin-layer chromatography (TLC). Fractions with the same composition were combined to obtain 10 fractions AJ.

[0055] The obtained fraction A was further separated by reversed-phase silica gel (ODS) column chromatography, eluted with a methanol-water gradient of 5:95-100:0 (v / v), and after TLC and HPLC analysis, the same fractions were combined to obtain five fractions A-1 to A-5; fraction A-2 was purified by semi-preparative high-performance liquid chromatography, eluted with an acetonitrile-0.1% formic acid-water gradient of 20:80-35:65 (v / v), to obtain the compound iridocycline A;

[0056] The obtained fraction C was further separated by normal-phase silica gel column chromatography, using a gradient elution with petroleum ether-acetone + 0.2% diethylamine at volume ratios of 20:1, 10:1, 5:1, 2:1, 1:1 and 0:1. After analysis by TLC and HPLC, fractions of the same type were combined to obtain five fractions C-1 to C-5. Fraction C-2 was purified by semi-preparative high-performance liquid chromatography, using a gradient elution with acetonitrile-0.1% formic acid-water at volume ratios of 20:80-35:65 to obtain the compound iridocycline B.

[0057] The obtained fraction E was further separated by normal-phase silica gel column chromatography, using a gradient elution with petroleum ether-acetone + 0.2% diethylamine at volume ratios of 20:1, 10:1, 5:1, 2:1, 1:1 and 0:1. After analysis by TLC and HPLC, fractions of the same type were combined to obtain four fractions E-1 to E-4. Fraction E-2 was purified by semi-preparative high-performance liquid chromatography, using a gradient elution with acetonitrile-0.1% formic acid-water at volume ratios of 20:80-40:60 to obtain the compound erythritol citrate C.

[0058] The obtained fraction I was further separated by normal-phase silica gel column chromatography, using a gradient elution with petroleum ether-acetone + 0.2% diethylamine at volume ratios of 10:1, 10:1, 5:1, 2:1, 1:1 and 0:1. After analysis by TLC and HPLC, fractions of the same type were combined to obtain four fractions I-1 to I-4. Fraction I-4 was purified by semi-preparative high-performance liquid chromatography, using a gradient elution with acetonitrile-0.1% formic acid-water at volume ratios of 20:80-40:60 to obtain the compound erythritolide D.

[0059] The compounds Ilitrizin A, Ilitrizin B, Ilitrizin C, and Ilitrizin D obtained in step c were analyzed by thin-layer chromatography using chloroform-methanol-water in a volume ratio of 82:16:2 as the developing solvent to identify four new diterpenoid alkaloid monomers.

[0060] Example 2 (Two types of separation)

[0061] a. Take 10.0 kg of whole plant of Delphinium yiliense and crush it. Extract it at room temperature with 50 L of 80% ethanol by percolation. Evaporate the solvent under reduced pressure to obtain crude extract of Delphinium yiliense.

[0062] b. Disperse the crude extract obtained in step a with 2% sulfuric acid. Adjust the pH of the resulting acidic aqueous layer to 10 with ammonia, then extract with chloroform, and evaporate the chloroform under reduced pressure to obtain total alkaloid extract 1. Further alkalize the alkaline aqueous layer with 10% NaOH to pH 12, then extract with n-butanol, and evaporate the n-butanol under reduced pressure to obtain total alkaloid extract 2. Combine total alkaloid extract 1 and total alkaloid extract 2 to obtain total alkaloid extract.

[0063] c. The obtained total alkaloid extract was separated by Sephadex LH-20 gel column chromatography. Isocratic elution was performed using chloroform and methanol in a volume ratio of 1:1. One fraction was collected for every 20 ml. The fractions were analyzed by silica gel thin layer chromatography (TLC). Fractions with the same composition were combined to obtain 8 components AH.

[0064] The obtained fraction A was further separated by Sephadex LH-20 gel column chromatography with gradient elution using methanol-water at a volume ratio of 5:95-80:20. After analysis by TLC and high performance liquid chromatography (HPLC), fractions of the same type were combined to obtain six fractions A-1 to A-6. Fraction A-2 was further separated and purified by preparative thin-layer chromatography using chloroform-methanol at a volume ratio of 20:1 to obtain the compound erythritol A.

[0065] The obtained fraction C was further separated by Sephadex LH-20 gel column chromatography with isocratic elution in methanol. After TLC and HPLC analysis, the same fractions were combined to obtain four fractions C-1 to C-4. Fraction C-3 was further separated and purified by preparative thin-layer chromatography with petroleum ether-acetone saturated with ammonia water at a volume ratio of 3:1 as the developing solvent to obtain the compound erythritol B.

[0066] The obtained fraction D was further separated by Sephadex LH-20 gel column chromatography with a gradient elution of methanol-water with a volume ratio of 5:95-80:20 to obtain the compound iridocycline C.

[0067] The obtained H fraction was further separated by Sephadex LH-20 gel column chromatography with a gradient elution of methanol-water at a volume ratio of 5:95-80:20. After TLC and HPLC analysis, the same fractions were combined to obtain four fractions H-1 to H-4. Fraction H-3 was further separated and purified by preparative thin-layer chromatography with petroleum ether-acetone saturated with ammonia at a volume ratio of 2:1 as the developing solvent to obtain the compound iridocycline D.

[0068] The compounds Ilitrizin A, Ilitrizin B, Ilitrizin C, and Ilitrizin D obtained in step c were analyzed by high performance liquid chromatography. Gradient elution was performed using an acetonitrile-formic acid aqueous solution with a volume ratio of 10:90-80:20 to identify four new diterpenoid alkaloid monomer compounds.

[0069] Example 3 (Three types of separation)

[0070] a. Take 10.0 kg of whole plant of Delphinium yiliensis, crush it, extract it with methanol using ultrasound, and concentrate it under reduced pressure to recover methanol to obtain total extract.

[0071] b. Disperse the total extract obtained in step a in a 3% hydrochloric acid aqueous solution, filter, adjust the pH of the resulting acidic aqueous solution to 10 with anhydrous Na2CO3, extract with ethyl acetate, evaporate the ethyl acetate under reduced pressure to obtain total alkaloid extract 1; then alkalize the alkaline aqueous layer with 10% NaOH to pH 12, extract with n-butanol, evaporate the n-butanol under reduced pressure to obtain total alkaloid extract 2, combine total alkaloid extract 1 and total alkaloid extract 2 to obtain total alkaloid extract;

[0072] c. The total alkaloid extract obtained in step b was separated by normal-phase silica gel column chromatography. Gradient elution was performed sequentially using chloroform-methanol 60:1, 40:1, 20:1, 10:1, 5:1 and 1:1 as solvent systems. One fraction was collected every 500 ml. The fractions were analyzed by silica gel thin-layer chromatography (TLC). Fractions with the same characteristics were combined to obtain 8 fractions AH.

[0073] Component A was further separated by reversed-phase silica gel (ODS) column chromatography, eluted with a methanol-water gradient of 5:95-80:20 (v / v), and analyzed by silica gel thin-layer chromatography (TLC) and HPLC. The same fractions were combined to obtain five components A-1 to A-5. Component A-2 was purified by semi-preparative high-performance liquid chromatography, eluted with an acetonitrile-0.1% formic acid gradient of 10:90-40:60 (v / v) to obtain compounds iridotrichumone A and iridotrichumone B.

[0074] Fraction C was separated by Sephadex LH-20 gel column chromatography with a gradient elution of methanol-water at a volume ratio of 5:95-80:20. After TLC and HPLC analysis, the same fractions were combined to obtain four fractions C-1 to C-4. C-2 was purified by semi-preparative high performance liquid chromatography with a gradient elution of acetonitrile-0.1% formic acid at a volume ratio of 22:78-35:65 to obtain compound iridocycline C.

[0075] Fraction E was separated by Sephadex LH-20 gel column chromatography with a gradient elution of methanol-water at a volume ratio of 5:95-80:20. After TLC and HPLC analysis, the same fractions were combined to obtain three fractions E-1 to E-3. Fraction E-2 was purified by semi-preparative high performance liquid chromatography with a gradient elution of acetonitrile-0.1% formic acid at a volume ratio of 20:80-35:65 to obtain the compound iridocycline D.

[0076] The compounds Ilitrizin A, Ilitrizin B, Ilitrizin C, and Ilitrizin D obtained in step c were analyzed by thin-layer chromatography using petroleum ether-acetone saturated with ammonia at a volume ratio of 3:1 as the developing solvent to identify four new diterpenoid alkaloid monomers.

[0077] Example 4 (Three types of separation)

[0078] a. Take 10.0 kg of whole plant of Delphinium yiliense and crush it. Then, extract it with 50 L of chloroform using ultrasound at room temperature. Evaporate the solvent under reduced pressure to obtain crude extract of Delphinium yiliense.

[0079] b. Disperse the crude extract obtained in step a with 4% hydrochloric acid. Adjust the pH of the resulting acidic aqueous solution to 10 with ammonia, then extract with ethyl acetate. Evaporate the ethyl acetate under reduced pressure to obtain total alkaloid extract 1. Further alkalize the alkaline aqueous layer with 10% NaOH to pH 12, then extract with n-butanol. Evaporate the n-butanol under reduced pressure to obtain total alkaloid extract 2. Combine total alkaloid extract 1 and total alkaloid extract 2 to obtain total alkaloid extract.

[0080] c. The total alkaloid extract obtained in step b was separated by reversed-phase silica gel (ODS) column chromatography, eluted with a methanol-water gradient of 5:95-100:0, and a fraction was collected every 500 ml. After analysis by silica gel thin-layer chromatography (TLC), the same fractions were combined to obtain 5 fractions AE.

[0081] Fraction A was separated by Sephadex LH-20 gel column chromatography with gradient elution using methanol-water at a volume ratio of 5:95-80:20. After TLC and HPLC analysis, fractions of the same type were combined to obtain four fractions A-1 to A-4. Fraction A-3 was purified by preparative thin-layer chromatography using petroleum ether-acetone saturated with ammonia at a volume ratio of 3:1 as the developing solvent to obtain the compound iridocycline A.

[0082] The obtained fraction B was separated by Sephadex LH-20 gel column chromatography with gradient elution using methanol-water at a volume ratio of 5:95-80:20. After TLC and HPLC analysis, the same fractions were combined to obtain four fractions B-1 to B-4. Fraction B-2 was purified by preparative thin-layer chromatography with chloroform-methanol-water at a volume ratio of 82:16:2 to obtain the compound erythritol B.

[0083] The obtained fraction D was separated by Sephadex LH-20 gel column chromatography, with gradient elution using methanol-water at a volume ratio of 5:95-80:20 to obtain Yili delphinidin C.

[0084] The obtained fraction E was separated by normal-phase silica gel column chromatography, and gradient elution was performed sequentially with chloroform-methanol 40:1, 20:1, 10:1, 5:1 and 1:1 solvent systems to obtain the compound iridocycline D.

[0085] The erythritol A, erythritol B, erythritol C, and erythritol D obtained in step c were analyzed by high performance liquid chromatography. Gradient elution was performed using an acetonitrile-formic acid aqueous solution with a volume ratio of 25:75-80:20 to identify four new diterpenoid alkaloid monomer compounds.

[0086] Example 5 (Four Types of Separation)

[0087] a. Take 10.0 kg of whole plant of Delphinium yiliense and pulverize it. Then, heat and reflux it with 50 L of 95% ethanol at room temperature. Evaporate the solvent under reduced pressure to obtain crude extract of Delphinium yiliense.

[0088] b. Disperse the crude extract obtained in step a with 2% sulfuric acid. Adjust the pH of the resulting acidic aqueous solution to 12 with 5% NaOH. Extract with n-butanol and evaporate the n-butanol under reduced pressure to obtain the total alkaloid extract.

[0089] c. The obtained total alkaloid extract was separated by Sephadex LH-20 gel column chromatography. Isocratic elution was performed using dichloromethane and methanol in a volume ratio of 1:1. One fraction was collected for every 20 ml. The fractions were analyzed by silica gel thin-layer chromatography (TLC). Fractions with the same characteristics were combined to obtain 5 components AE.

[0090] Component A was further separated by reversed-phase silica gel (ODS) column chromatography, eluted with a methanol-water gradient of 5:95-80:20 (v / v), and after TLC and HPLC analysis, the same fractions were combined to obtain four components B-1 to B-4; component B-2 was purified by preparative thin-layer chromatography, developed with chloroform-methanol (v / v) as the developing solvent to obtain compound erythritol A;

[0091] The obtained fraction C was further separated by normal-phase silica gel column chromatography, using a gradient elution with petroleum ether-acetone + 0.2% diethylamine at volume ratios of 20:1, 10:1, 5:1, 2:1, 1:1 and 0:1. After analysis by TLC and HPLC, the same fractions were combined to obtain four fractions C-1 to C-4. Fraction C-2 was purified by semi-preparative high-performance liquid chromatography, using a gradient elution with acetonitrile-0.1% formic acid at volume ratios of 22:78-35:65 to obtain the compound iridocycline B.

[0092] Fraction D was separated by Sephadex LH-20 gel column chromatography, using dichloromethane and methanol in a 1:1 volume ratio as eluents for isocratic elution. One fraction was collected for every 10 ml, and the fractions were analyzed by silica gel thin-layer chromatography (TLC). Fractions with the same volume were combined to obtain six fractions D-1 to D-6. Fraction D-2 was purified by semi-preparative high-performance liquid chromatography, using a gradient elution of acetonitrile-0.1% formic acid in a volume ratio of 22:78-35:65 to obtain the compound iridocycline C.

[0093] The obtained fraction E was further separated by normal-phase silica gel column chromatography, using a gradient elution with petroleum ether-acetone + 0.2% diethylamine at volume ratios of 10:1, 10:1, 5:1, 2:1, 1:1 and 0:1. After analysis by TLC and HPLC, fractions of the same type were combined to obtain four fractions E-1 to E-4. Fraction E-3 was purified by semi-preparative high-performance liquid chromatography, using a gradient elution with acetonitrile-0.1% formic acid at volume ratios of 22:78-35:65 to obtain the compound iridocycline D.

[0094] The compounds Ilitrizin A, Ilitrizin B, Ilitrizin C, and Ilitrizin D obtained in step c were analyzed by thin-layer chromatography using chloroform-methanol-water in a volume ratio of 82:16:2 as the developing solvent to identify four new diterpenoid alkaloid monomers.

[0095] Example 6 (Four Types of Separation)

[0096] a. Take 10.0 kg of whole plant of Delphinium yiliense and crush it. Then, extract it with 50 L of methanol at room temperature using ultrasound. Evaporate the solvent under reduced pressure to obtain crude extract of Delphinium yiliense.

[0097] b. Disperse the crude extract obtained in step a with 3% sulfuric acid. Adjust the pH of the resulting acidic aqueous solution to 11 with 5% NaOH. Extract with n-butanol and evaporate the n-butanol under reduced pressure to obtain the total alkaloid extract.

[0098] c. The total alkaloid extract obtained in step b was separated by normal-phase silica gel column chromatography. Gradient elution was performed sequentially using chloroform-methanol 60:1, 40:1, 20:1, 10:1, 5:1 and 1:1 as solvent systems. One fraction was collected every 500 ml. The fractions were analyzed by silica gel thin-layer chromatography (TLC). Fractions with the same characteristics were combined to obtain 5 fractions AE.

[0099] The obtained fraction A was further separated by reversed-phase silica gel (ODS) column chromatography, eluted with a methanol-water gradient of 5:95-100:0 (v / v), and after TLC and HPLC analysis, the same fractions were combined to obtain four fractions A-1 to A-4; fraction A-3 was purified by semi-preparative high performance liquid chromatography, eluted with an acetonitrile-0.1% formic acid-water gradient of 10:90-40:60 (v / v) to obtain the compound iridocycline A;

[0100] Fraction C was separated by Sephadex LH-20 gel column chromatography with gradient elution using methanol-water at a volume ratio of 5:95-80:20. After TLC and HPLC analysis, the same fractions were combined to obtain four fractions C-1 to C-4. Fraction C-2 was purified by preparative thin-layer chromatography with chloroform-methanol-water at a volume ratio of 82:16:2 to obtain the compound erythritol B.

[0101] The obtained fraction D was further separated by normal phase silica gel column chromatography, using petroleum ether-acetone + 0.2% diethylamine in volume ratios of 10:1, 10:1, 5:1, 2:1, 1:1 and 0:1 as eluents to obtain compound iridocycline C.

[0102] The obtained fraction E was further separated by normal-phase silica gel column chromatography, using a gradient elution with petroleum ether-acetone + 0.2% diethylamine at volume ratios of 10:1, 10:1, 5:1, 2:1, 1:1 and 0:1 to obtain five fractions E-1 to E-4; fraction E-2 was purified by semi-preparative high-performance liquid chromatography, using a gradient elution with acetonitrile-0.1% formic acid-water at volume ratios of 20:80-30:70 to obtain the compound iridocycline D;

[0103] The compounds Ilitrizin A, Ilitrizin B, Ilitrizin C, and Ilitrizin D obtained in step c were analyzed by thin-layer chromatography using chloroform-methanol-water in a volume ratio of 82:16:2 as the developing solvent to identify four new diterpenoid alkaloid monomers.

[0104] Example 7

[0105] The use of the diterpenoid alkaloids isolated from *Delphinium ilignon* in the preparation of analgesic drugs, as described in this invention, is illustrated using ICR mice as an example:

[0106] Screening of analgesic activity of the obtained diterpenoid alkaloids:

[0107] Laboratory animals:

[0108] ICR mice, weighing 18-22g, half male and half female, were provided by the Department of Experimental Animal Science, Xinjiang Medical University. The mice were housed in an SPF-grade animal experimental center with an indoor temperature of 22±2℃ and a relative humidity of 40%-70%, and were allowed free access to water and food.

[0109] Experimental instruments and reagents:

[0110] Dimethyl sulfoxide (DMSO), glacial acetic acid (Aladdin), sodium chloride injection (Xinjiang Huashidan Pharmaceutical Co., Ltd.);

[0111] Experiment content:

[0112] Drug solution preparation: Dissolve the drug in dimethyl sulfoxide (DMSO) or 0.1M / L hydrochloric acid, then dilute with physiological saline. The blank control group serves as the solvent control. Store at room temperature in the dark.

[0113] Preparation of 0.6% acetic acid solution: 30 minutes before model preparation, prepare 10 ml of 10% acetic acid aqueous solution and place it in an ice water box; 25 minutes later, take 360 ​​μl of 10% acetic acid and prepare 6 ml of 0.6% acetic acid solution (enough for 10 mice), place it in an ice water box, and use it immediately after preparation;

[0114] Acetic acid writhing test in mice: Fifty ICR mice, half male and half female, were randomly divided into 5 groups: a blank control group and each test drug group, with 10 mice in each group. Mice in each group were administered the drug via intraperitoneal injection at a volume of 0.1 mL / 10 g, while the blank control group received a blank solvent system. Thirty minutes after drug administration, mice in each group were injected intraperitoneally with 0.1 mL / 10 g of 0.6% glacial acetic acid. The number of writhing episodes within 20 minutes was recorded. The pain inhibition percentage of each drug group was compared with that of the blank control group. Pain inhibition percentage (%) = (number of writhing episodes in the blank control group - number of writhing episodes in the drug group) / number of writhing episodes in the blank control group × 100%. The experimental results are shown in Table 2.

[0115] Table 2. Effects of the neoditerpenoid alkaloids *Lysimachia yiliensis* C and D on the analgesic effect of acetic acid in mice.

[0116]

[0117] To investigate the analgesic effects of the novel diterpenoid alkaloid compounds, iridin C and iridin D, isolated from *Lysimachia iridis*, on acetic acid-induced abdominal pain, this experiment established a pain model by intraperitoneal injection of acetic acid into mice. The effects of compounds iridin C and D on the number of writhing movements and the latency of writhing movements in mice were observed, thereby evaluating their efficacy.

[0118] Experimental results show that the novel diterpenoid alkaloids, iridin C and iridin D, isolated from *Delphinium ilignata*, significantly reduced the number of writhing movements and prolonged the writhing latency in painful mice. Therefore, it can be concluded that the novel diterpenoid alkaloids, iridin C and iridin D, isolated from *Delphinium ilignata*, have significant analgesic effects on acetic acid-induced abdominal pain. The analgesic effect of iridin C is dose-dependent. In conclusion, these results provide experimental evidence for whether this compound can be used as a potential clinical analgesic.

Claims

1. A diterpenoid alkaloid compound isolated from *Delphinium ilignon*, characterized in that... The structural formula of this compound is: in: Compound 3 is named Ili delphinidin C; Compound 4 is named Ili delphinidin D.

2. A method for preparing the diterpenoid alkaloid compound isolated from *Delphinium ilignon* as described in claim 1, characterized in that... Follow these steps: a. Take 10.0 kg of whole plant of Delphinium yiliense and crush it. Soak the medicinal material in 5% Na2CO3 solution for 3 hours at room temperature. Add 50 L of chloroform:methanol mixed solution with a volume ratio of 6:

1. Extract by cold soaking. Evaporate the solvent under reduced pressure to obtain crude extract of Delphinium yiliense. b. Disperse the crude extract obtained in step a with a 1% sulfuric acid solution. Adjust the pH of the resulting acidic aqueous layer to 10 with anhydrous Na2CO3, then extract with ethyl acetate. Evaporate the ethyl acetate under reduced pressure to obtain total alkaloid 1. Further alkalize the alkaline aqueous layer with 10% NaOH to pH 12, then extract with n-butanol. Evaporate the n-butanol under reduced pressure to obtain total alkaloid 2. Combine total alkaloid 1 and total alkaloid 2 to obtain total alkaloids. c. The total alkaloids obtained were separated by normal-phase silica gel column chromatography. Gradient elution was performed using chloroform and methanol in volume ratios of 100:1, 60:1, 40:1, 20:1, 10:1, 5:1, 3:1, and 1:

1. One fraction was collected every 500 ml. The fractions were analyzed by silica gel thin-layer chromatography. Fractions with the same composition were combined to obtain 10 fractions AJ. The obtained fraction E was further separated by normal-phase silica gel column chromatography, using a gradient elution with petroleum ether-acetone + 0.2% diethylamine at volume ratios of 20:1, 10:1, 5:1, 2:1, 1:1 and 0:

1. After analysis by TLC and HPLC, fractions of the same type were combined to obtain four fractions E-1 to E-4. Fraction E-2 was purified by semi-preparative high-performance liquid chromatography, using a gradient elution with acetonitrile-0.1% formic acid-water at volume ratios of 20:80-40:60 to obtain the compound iridocycline C. The obtained fraction I was further separated by normal-phase silica gel column chromatography, using a gradient elution with petroleum ether-acetone + 0.2% diethylamine at volume ratios of 10:1, 10:1, 5:1, 2:1, 1:1 and 0:

1. After analysis by TLC and HPLC, fractions of the same type were combined to obtain four fractions I-1 to I-4. Fraction I-4 was purified by semi-preparative high-performance liquid chromatography, using a gradient elution with acetonitrile-0.1% formic acid-water at volume ratios of 20:80-40:60 to obtain the compound erythritolide D.

3. A method for preparing the diterpenoid alkaloid compound isolated from *Delphinium ilyezoensis* as described in claim 1, characterized in that... Follow these steps: a. Take 10.0 kg of whole plant of Delphinium yiliense and crush it. Extract it at room temperature with 50 L of 80% ethanol by percolation. Evaporate the solvent under reduced pressure to obtain crude extract of Delphinium yiliense. b. Disperse the crude extract obtained in step a with 2% sulfuric acid. Adjust the pH of the resulting acidic aqueous layer to 10 with ammonia, then extract with chloroform. Evaporate the chloroform under reduced pressure to obtain total alkaloid 1. Further alkalize the alkaline aqueous layer with 10% NaOH to pH 12, then extract with n-butanol. Evaporate the n-butanol under reduced pressure to obtain total alkaloid 2. Combine total alkaloid 1 and total alkaloid 2 to obtain total alkaloids. c. The total alkaloids obtained were separated by Sephadex LH-20 gel column chromatography. Isocratic elution was performed using chloroform and methanol in a volume ratio of 1:

1. One fraction was collected for every 20 ml. The fractions were analyzed by silica gel thin layer chromatography (TLC). Fractions with the same composition were combined to obtain 8 components AH. The obtained fraction D was separated by Sephadex LH-20 gel column chromatography, with gradient elution using methanol-water at a volume ratio of 5:95-80:20 to obtain the compound Ilidin C. The obtained H fraction was separated by Sephadex LH-20 gel column chromatography with gradient elution using methanol-water at a volume ratio of 5:95-80:

20. After TLC and HPLC analysis, the same fractions were combined to obtain four fractions H-1 to H-4. Fraction H-3 was further separated and purified by preparative thin-layer chromatography using petroleum ether-acetone saturated with ammonia at a volume ratio of 2:1 as the developing solvent to obtain the compound iridocycline D.

4. A method for preparing the diterpenoid alkaloid compound isolated from *Delphinium ilignon* as described in claim 1, characterized in that... Follow these steps: a. Take 10.0 kg of whole plant of Delphinium yiliensis, crush it, extract it with methanol using ultrasound, and concentrate it under reduced pressure to recover the methanol to obtain the total extract. b. Disperse the total extract obtained in step a in a 3% hydrochloric acid aqueous solution, filter, adjust the pH of the resulting acidic aqueous solution to 10 with anhydrous Na2CO3, extract with ethyl acetate, evaporate the ethyl acetate under reduced pressure to obtain total alkaloid 1; then alkalize the alkaline aqueous layer with 10% NaOH to pH 12, extract with n-butanol, evaporate the n-butanol under reduced pressure to obtain total alkaloid 2, combine total alkaloid 1 and total alkaloid 2 to obtain total alkaloids; c. The total alkaloids obtained in step b were separated by normal-phase silica gel column chromatography. Gradient elution was performed sequentially using chloroform-methanol 60:1, 40:1, 20:1, 10:1, 5:1 and 1:1 solvent systems. One fraction was collected every 500 ml. The fractions were analyzed by silica gel thin-layer chromatography (TLC). Fractions with the same characteristics were combined to obtain 8 fractions of AH. The obtained fraction C was separated by Sephadex LH-20 gel column chromatography with a gradient elution of methanol-water at a volume ratio of 5:95-80:

20. After TLC and HPLC analysis, the same fractions were combined to obtain four fractions C-1 to C-4. C-2 was purified by semi-preparative high performance liquid chromatography with a gradient elution of acetonitrile-0.1% formic acid at a volume ratio of 22:78-35:65 to obtain the compound iridocycline C. The obtained fraction E was separated by Sephadex LH-20 gel column chromatography with a gradient elution of methanol-water at a volume ratio of 5:95-80:

20. After TLC and HPLC analysis, the same fractions were combined to obtain three fractions E-1 to E-3. Fraction E-2 was purified by semi-preparative high performance liquid chromatography with a gradient elution of acetonitrile-0.1% formic acid at a volume ratio of 20:80-35:65 to obtain the compound iridocycline D.

5. A method for preparing the diterpenoid alkaloid compound isolated from *Delphinium ilyezoensis* as described in claim 1, characterized in that... Follow these steps: a. Take 10.0 kg of whole plant of Delphinium yiliensis, crush it, and extract it with 50 L of chloroform at room temperature using ultrasound. Then evaporate the solvent under reduced pressure to obtain crude extract of Delphinium yiliensis. b. Disperse the crude extract obtained in step a with 4% hydrochloric acid. Adjust the pH of the resulting acidic aqueous solution to 10 with ammonia, then extract with ethyl acetate. Evaporate the ethyl acetate under reduced pressure to obtain total alkaloid 1. Further alkalize the alkaline aqueous layer with 10% NaOH to pH 12, then extract with n-butanol. Evaporate the n-butanol under reduced pressure to obtain total alkaloid 2. Combine total alkaloid 1 and total alkaloid 2 to obtain total alkaloids. c. The total alkaloids obtained in step b were separated by reversed-phase silica gel column chromatography, eluted with a methanol-water gradient of 5:95-100:0 (volume ratio), and each fraction was collected in 500 ml. After analysis by silica gel thin-layer chromatography, the same fractions were combined to obtain 5 fractions AE. The obtained fraction D was separated by Sephadex LH-20 gel column chromatography, with gradient elution using methanol-water at a volume ratio of 5:95-80:20 to obtain Yili delphinidin C. The obtained fraction E was separated by normal-phase silica gel column chromatography, and gradient elution was performed sequentially with chloroform-methanol 40:1, 20:1, 10:1, 5:1 and 1:1 solvent systems to obtain the compound erythritol D.

6. A method for preparing the diterpenoid alkaloid compound isolated from *Delphinium ilyezoensis* as described in claim 1, characterized in that... Follow these steps: a. Take 10.0 kg of whole plant of Delphinium yiliense and pulverize it. Then, heat and reflux it with 50 L of 95% ethanol at room temperature. Evaporate the solvent under reduced pressure to obtain crude extract of Delphinium yiliense. b. Disperse the crude extract obtained in step a with 2% sulfuric acid. Adjust the pH of the resulting acidic aqueous solution to 12 with 5% NaOH. Extract with n-butanol and evaporate the n-butanol under reduced pressure to obtain total alkaloids. c. The total alkaloids obtained were separated by Sephadex LH-20 gel column chromatography. Isocratic elution was performed using dichloromethane and methanol in a volume ratio of 1:

1. One fraction was collected for every 20 ml. The fractions were analyzed by silica gel thin-layer chromatography. Fractions with the same characteristics were combined to obtain 5 fractions AE. The obtained fraction D was separated by Sephadex LH-20 gel column chromatography, using dichloromethane and methanol in a 1:1 volume ratio as eluents for isocratic elution. One fraction was collected for every 10 ml, and the fractions were analyzed by silica gel thin-layer chromatography. Fractions with the same volume were combined to obtain 6 fractions D-1 to D-6. Fraction D-2 was purified by semi-preparative high performance liquid chromatography, using a gradient elution of acetonitrile-0.1% formic acid in a volume ratio of 22:78-35:65 to obtain the compound iridocycline C. The obtained fraction E was further separated by normal-phase silica gel column chromatography, using a gradient elution with petroleum ether-acetone + 0.2% diethylamine at volume ratios of 10:1, 10:1, 5:1, 2:1, 1:1 and 0:

1. After analysis by TLC and HPLC, fractions of the same type were combined to obtain four fractions E-1 to E-4. Fraction E-3 was purified by semi-preparative high-performance liquid chromatography, using a gradient elution with acetonitrile-0.1% formic acid at volume ratios of 22:78-35:65 to obtain the compound erythritol chlortetracycline D.

7. A method for preparing the diterpenoid alkaloid compound isolated from *Delphinium ilyezoensis* as described in claim 1, characterized in that... Follow these steps: a. Take 10.0 kg of whole plant of Delphinium yiliense and crush it. Extract it with 50 L of methanol at room temperature using ultrasound. Evaporate the solvent under reduced pressure to obtain crude extract of Delphinium yiliense. b. Disperse the crude extract obtained in step a with 3% sulfuric acid. Adjust the pH of the resulting acidic aqueous solution to 11 with 5% NaOH. Extract with n-butanol and evaporate the n-butanol under reduced pressure to obtain total alkaloids. c. The total alkaloids obtained in step b were separated by normal-phase silica gel column chromatography. Gradient elution was performed sequentially using chloroform-methanol 60:1, 40:1, 20:1, 10:1, 5:1 and 1:1 as solvent systems. One fraction was collected every 500 ml. The fractions were analyzed by silica gel thin-layer chromatography (TLC). Fractions with the same characteristics were combined to obtain 5 fractions AE. The obtained fraction D was further separated by normal phase silica gel column chromatography, using petroleum ether-acetone + 0.2% diethylamine in volume ratios of 10:1, 10:1, 5:1, 2:1, 1:1 and 0:1 as eluents to obtain the compound iridocycline C. The obtained fraction E was further separated by normal-phase silica gel column chromatography, using a gradient elution with petroleum ether-acetone + 0.2% diethylamine in volume ratios of 10:1, 10:1, 5:1, 2:1, 1:1 and 0:1 to obtain five fractions E-1 to E-4. Fraction E-2 was purified by semi-preparative high-performance liquid chromatography, using a gradient elution with acetonitrile-0.1% formic acid-water in volume ratios of 20:80-30:70 to obtain the compound iridocycline D.

8. The use of the diterpenoid alkaloid compound isolated from Delphinium yiliense according to claim 1 in the preparation of analgesic drugs.

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