New antidepressant α-acidic compound, its preparation method and application

Abstract

The application belongs to the technical field of medicines, and specifically discloses a novel tricyclic alpha acid compound and a preparation method and application thereof; the novel tricyclic alpha acid compounds (±)-Humulunes M-O and (-)-Garcinielliptone HE provided by the application have not been reported so far; the compounds have a significant protective effect on HT22 cells induced by corticosterone and SH-SY5Y cells induced by LPS, and can be used for developing a medicine for treating depression.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, and more specifically, relates to a novel class of α-acid compounds, their preparation methods, pharmaceutical compositions, and their application in the preparation of drugs for treating depression. Background Technology

[0002] Hops ( Humulus lupulus Humulus (L.) is a perennial climbing herb belonging to the genus Humulus in the family Cannabisaceae. Its extracts are widely used in beer brewing and the pharmaceutical industry. Alpha-acids are the main active components in hops, possessing various biological activities such as antioxidant, anti-inflammatory, and antibacterial properties. However, existing methods for efficiently separating and purifying alpha-acids from hops still suffer from problems such as cumbersome procedures and low yields. Therefore, developing an efficient and simple extraction and separation method is of great significance. The inventors isolated a class of alpha-acids, (±)-Humulunes MO and (±)-Garcinielliptone HE, from the dried aerial parts of hops and found that (±)-Humulune MO and (±)-Garcinielliptone HE have significant neuroprotective activities, with (+)-Humulune O exhibiting the best activity, which can be used to prepare drugs for treating depression. Summary of the Invention

[0003] In view of the shortcomings of the existing technology, the present invention provides α-acid compounds, their preparation methods, pharmaceutical compositions and applications.

[0004] To achieve the above objectives, the technical solution of the present invention is as follows: The novel tricyclic α-acid compound is characterized by having a structure shown in any one of the following formulas (1)-(8):

[0005] Tautomers of the α-acid compounds or pharmaceutically acceptable salts are also within the scope of protection of this invention.

[0006] In another aspect, the present invention also provides a method for preparing the above-mentioned α-acid compounds, wherein the α-acid compounds are extracted and separated from hops.

[0007] Furthermore, the preparation method of the α-acid compound includes the following steps: S1. Take dried hop spikes with flowers, add solvent to extract, combine the extracts, filter, concentrate under reduced pressure to obtain extract; S2. After suspending the extract in water, extract it with dichloromethane to obtain the extract; S3. The dichloromethane extract was subjected to gradient elution using a petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction AE. S4. Fraction C was subjected to gradient elution using a petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction C13. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13 was 0.44-0.45. S5. Fraction C13 was subjected to gradient elution using petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction C13.17. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17 was 0.25-0.27. S6. Fraction C13.17 was isocratically eluted using a petroleum ether-dichloromethane-methanol solution via Sephadex LH-20 gel column chromatography to obtain fraction C13.17.7. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17.7 was 0.57-0.60. S7. Fraction C13.17.7 was subjected to gradient elution using methanol-water solution via ODS column chromatography to obtain fractions C13.17.7.5, C13.17.7.7, and C13.17.7.8. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17.7.5 was 0.45-0.52, the Rf value corresponding to fraction C13.17.7.7 was 0.60-0.64, and the Rf value corresponding to fraction C13.17.7.8 was 0.66-0.71. S8. Using methanol-water as the mobile phase, HPLC was used to prepare tricyclic α-acid compounds represented by formulas (5), (6), (7) and / or (8) in fraction C13.17.7.5 using a C18 column. S9. Using acetonitrile-water as the mobile phase, HPLC method and C18 column can be used to prepare tricyclic α-acid compounds represented by formula (3) and / or formula (4) in fraction C13.17.7.7. S10. Using acetonitrile-water as the mobile phase, tricyclic α-acid compounds of formula (1) and / or formula (2) can be prepared in fraction C13.17.7.8 by HPLC using a C18 column.

[0008] Further, in step S8, the volume ratio of methanol to water in the methanol-water solution is (62:38) to (68:32). Preferably, the volume ratio of methanol to water in the methanol-water solution is 65:35.

[0009] Further, in step S8, fraction C13.17.7.5 is further separated based on the characteristic ultraviolet absorption (λmax = 226, 298 nm) of tricyclic α-acids observed by HPLC analysis, including tricyclic α-acids with a retention time of 35-37 min as shown in formula (5); tricyclic α-acids with a retention time of 39-42 min as shown in formula (6); tricyclic α-acids with a retention time of 22-24 min as shown in formula (7); and tricyclic α-acids with a retention time of 26-29 min as shown in formula (8).

[0010] Further, in step S9, the volume ratio of acetonitrile to water in the acetonitrile-water solution is (45:55) to (50:50). Preferably, the volume ratio of acetonitrile to water in the acetonitrile-water solution is 48:52.

[0011] Further, in step S9, fraction C13.17.7.7 is further separated based on the characteristic ultraviolet absorption (λmax = 226, 298 nm) of tricyclic α-acids observed by HPLC analysis, including tricyclic α-acids with a retention time of 21-24 min and tricyclic α-acids with a retention time of 26-28 min as shown in formula (3).

[0012] Further, in step S10, the volume ratio of acetonitrile to water in the acetonitrile-water solution is (42:58)-(47:53). Preferably, the volume ratio of acetonitrile to water in the acetonitrile-water solution is 45:55.

[0013] Further, in step S10, fraction C13.17.7.8 is further separated based on the characteristic ultraviolet absorption (λmax = 226, 298 nm) of tricyclic α-acids observed by HPLC analysis, including tricyclic α-acids with a retention time of 30-34 min and tricyclic α-acids with a retention time of 36-38 min as shown in formula (1).

[0014] Further, in step S2, the extract is suspended in 8-15 times its weight of water and then extracted with dichloromethane.

[0015] In this application, the Rf value is the Rf value of the fluorescent spot observed at scanning wavelengths of 254 nm and 365 nm.

[0016] Further, in step S3, based on a total volume of 100, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when washing out fraction C is (58:42)-(62:38); preferably, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution is 60:40.

[0017] Further, in step S4, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when washing out fraction C13 is (37:63)-(43:57); preferably, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution is 40:60.

[0018] Further, in step S5, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when washing out fraction C13.17 is (18:82)-(23:77); preferably, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution is 20:80.

[0019] Further, in step S6, the volume ratio of petroleum ether, dichloromethane and methanol in the petroleum ether-dichloromethane-methanol solution when washing out fraction C13.17.7 is (7:3:1)-(3:7:1); preferably, the volume ratio of petroleum ether, dichloromethane and methanol in the petroleum ether-dichloromethane-methanol solution is 5:5:1.

[0020] Further, in step S7, the volume ratio of methanol to water in the methanol-water solution when washing out fraction C13.17.7.5 is (47:53)-(53:47); preferably, the volume ratio of methanol to water in the methanol-water solution is 50:50. The volume ratio of methanol to water in the methanol-water solution when washing out fraction C13.17.7.7 is (56:44)-(62:38); preferably, the volume ratio of methanol to water in the methanol-water solution is 60:40. The volume ratio of methanol to water in the methanol-water solution when washing out fraction C13.17.7.8 is (66:34)-(72:28); preferably, the volume ratio of methanol to water in the methanol-water solution is 65:35.

[0021] Further, in step S1, the solvent is an 88-98 V% aqueous ethanol solution; the solvent added is 8-10 times the mass of the hops, and the reflux extraction is performed 1-3 times, with each extraction lasting 1-3 hours.

[0022] In another aspect, the present invention provides a pharmaceutical composition containing the above-mentioned novel tricyclic α-acid compound.

[0023] Furthermore, the pharmaceutical composition includes an synergist and a pharmacodynamically acceptable carrier or excipient.

[0024] In other words, pharmaceutical compositions containing the tricyclic α-acid compounds of the present invention as active ingredients and conventional pharmaceutical excipients, adjuvants or carriers are also included in the present invention.

[0025] Furthermore, the synergist is one or more of the following substances: Fluoxetine, paroxetine, fluvoxamine, sertraline, citalopram, escitalopram, venlafaxine, duloxetine, mirtazapine, bupropion, agomelatine, trazodone, reboxetine, imipramine, amitriptyline, clomipramine, doxepin, maprotiline, moclobemide, liver-soothing and mood-regulating capsules, St. John's wort extract, flupentixol melitracen.

[0026] Furthermore, the dosage form of the pharmaceutical composition is tablets, capsules, granules, oral liquids, powders, drops, or micro-pellets.

[0027] In another aspect, the present invention provides the use of the above-mentioned tricyclic α-acid compounds or the above-mentioned compositions in antidepressant drugs.

[0028] For ease of description, this application defines the compounds having formulas (1) to (8) as compounds (±)-Humulunes MO and (±)-Garcinielliptone HE, respectively.

[0029] Compared with the prior art, the present invention has the following advantages: (1) This invention provides novel α-acid compounds (±)-Humulunes MO and (-)-Garcinielliptone HE, which have not been previously reported. Experimental results show that the eight compounds have good neuroprotective effects. In particular, the tricyclic α-acid compound (+)-Humulune O has a significant protective effect against corticosterone-induced HT22 cells, with efficacy comparable to the positive control drug fluoxetine, and can be used to develop drugs for the treatment of depression.

[0030] (2) The preparation method of the novel tricyclic α-acid compounds is simple to operate, has good reproducibility, and has high extraction purity. Attached Figure Description

[0031] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0032] Figure 1 The (+)-Humulune M obtained in Example 1 of this invention1 1H NMR spectrum (400 MHz, CDCl3); Figure 2 The (+)-Humulune M obtained in Example 1 of this invention 13 C NMR spectrum (100 MHz, CDCl3); Figure 3 The HMBC spectrum of (+)-Humulune M obtained in Example 1 of this invention; Figure 4 The (-)-Humulune M obtained in Example 1 of this invention 1 1H NMR spectrum (400 MHz, CDCl3); Figure 5 The (-)-Humulune M obtained in Example 1 of this invention 13 C NMR spectrum (100 MHz, CDCl3); Figure 6 The HMBC spectrum of (-)-Humulune M obtained in Example 1 of this invention; Figure 7 The (+)-Humulune N obtained in Example 2 of this invention 1 1H NMR spectrum (400 MHz, CDCl3); Figure 8 The (+)-Humulune N obtained in Example 2 of this invention 13 C NMR spectrum (100 MHz, CDCl3); Figure 9 The HMBC spectrum of (+)-Humulune N obtained in Example 2 of this invention; Figure 10 The (-)-Humulune N obtained in Example 3 of this invention 1 1H NMR spectrum (400 MHz, CDCl3); Figure 11 The (-)-Humulune N obtained in Example 3 of this invention 13 C NMR spectrum (100 MHz, CDCl3); Figure 12 The HMBC spectrum of (-)-Humulune N obtained in Example 3 of this invention; Figure 13 The (+)-Humulune O obtained in Example 3 of this invention 1 1H NMR spectrum (400 MHz, CDCl3); Figure 14The (+)-Humulune O obtained in Example 3 of this invention 13 C NMR spectrum (100 MHz, CDCl3); Figure 15 The HMBC spectrum of (+)-Humulune O obtained in Example 3 of this invention; Figure 16 The (-)-Humulune O obtained in Example 3 of this invention 1 1H NMR spectrum (400 MHz, CDCl3); Figure 17 The (-)-Humulune O obtained in Example 3 of this invention 13 C NMR spectrum (100 MHz, CDCl3); Figure 18 The HMBC spectrum of (-)-Humulune O obtained in Example 3 of this invention; Figure 19 The (+)-Garcinielliptone HE obtained in Example 3 of this invention 1 H NMR spectrum (400MHz, CDCl3); Figure 20 The (+)-Garcinielliptone HE obtained in Example 3 of this invention 13 C NMR spectrum (100 MHz, CDCl3); Figure 21 The HMBC spectrum of (+)-Garcinielliptone HE obtained in Example 3 of this invention; Figure 22 The (-)-Garcinielliptone HE obtained in Example 3 of this invention 1 H NMR spectrum (400MHz, CDCl3); Figure 23 The (-)-Garcinielliptone HE obtained in Example 3 of this invention 13 C NMR spectrum (100 MHz, CDCl3); Figure 24 The HMBC spectrum of (-)-Garcinielliptone HE obtained in Example 3 of this invention. Detailed Implementation

[0033] The technical solution of the present invention will now be clearly and completely described with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0034] The present invention will be further explained below with reference to specific embodiments.

[0035] It should be understood that the specific embodiments described herein are for illustrative purposes only and are not intended to limit the scope of the invention.

[0036] In the embodiments, unless otherwise specified, all methods used are conventional methods in the art.

[0037] The terms “comprising,” “including,” or any other variations thereof, as used herein, are intended to cover non-exclusive inclusion. For example, a composition, step, method, article, or apparatus that includes the listed elements is not necessarily limited to those elements, but may include other elements not expressly listed or elements inherent to such composition, step, method, article, or apparatus.

[0038] Where specific techniques or conditions are not specified in the examples, they shall be performed in accordance with the techniques or conditions described in the literature in this field, or in accordance with the product instructions. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased through legitimate channels.

[0039] The endpoints and any values ​​of the ranges disclosed herein are not limited to the precise ranges or values, and these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of the various ranges, the endpoint values ​​of the various ranges and individual point values, and individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed herein.

[0040] Furthermore, the technical features involved in the various embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other.

[0041] Example 1 This embodiment discloses a method for preparing a novel tricyclic α-acid compound, comprising the following steps: S1. Take dried hop heads with flowers, add solvent for extraction, combine the extracts, filter, concentrate under reduced pressure to obtain an extract; specifically, the solvent is an 88-98 V% aqueous ethanol solution; the mass of the solvent added is 8-10 times that of the hops, the reflux extraction is performed 1-3 times, and each extraction is performed for 1-3 hours.

[0042] The specific implementation plan is as follows: Weigh 101.4 kg of dried hops with flowers, add 95% ethanol (10 times the weight of the hops) and extract three times, reflux at 75°C for 1 hour each time, combine the extracts and concentrate to obtain an extract (about 9.0 kg).

[0043] S2. After suspending the extract in water, extract it sequentially with petroleum ether and dichloromethane to obtain a dichloromethane extract; further, in step S2, after suspending the extract in 8-15 times its mass of water, extract it with dichloromethane.

[0044] The specific steps are as follows: Add the extract (about 9.0 kg) to 10 times its weight of water (90 L) to suspend it, and then extract it with 1.5 times the volume of dichloromethane. Extract it 3 times to obtain dichloromethane extract (about 3100 g).

[0045] S3. The extract was subjected to gradient elution using a petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction C. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C was 0.55-0.62. Based on a total volume of 100, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when fraction C was eluted was (50:50)-(70:30); more preferably, the volume ratio was 60:40.

[0046] It should be noted that elution with a petroleum ether-ethyl acetate solution at a volume ratio of (50:50) to (70:30) and identification of the fraction with an Rf value of 0.55-0.62 using silica gel thin-layer chromatography resulted in fraction C. The volume ratio of the petroleum ether-ethyl acetate solution required to obtain fraction C was between (50:50) and (70:30), with the highest efficiency achieved when eluting with a petroleum ether-ethyl acetate solution at a volume ratio of 60:40. The following gradient elution principles are essentially the same.

[0047] In this application, the Rf value is the Rf value of the fluorescent spot observed at scanning wavelengths of 254 nm and 365 nm.

[0048] The specific steps are as follows: The dichloromethane extract was subjected to silica gel column chromatography, using a petroleum ether-ethyl acetate gradient elution at volume ratios of 100:0, 80:20, 60:40, 30:70, and 0:100, with each 500 mL volume representing one fraction, yielding 35 fractions in total: 1-5, 6-10, 11-25, 26-29, and 30-35. These fractions were then identified using silica gel thin-layer chromatography, based on the values ​​at 254 nm and 365 nm. Fluorescent spots with Rf values ​​of 0.81-0.83 (fractions 1-5), 0.67-0.78 (fractions 6-10), 0.55-0.62 (fractions 11-25), 0.40-0.52 (fractions 26-29), and 0.32-0.38 (fractions 30-35) observed at nm were combined to obtain 5 fractions, named A, B, C, D, and E respectively; among them, fraction C was obtained by combining fractions 11-25.

[0049] The characteristic ultraviolet absorption (λ) of tricyclic α-acids observed by HPLC analysis max = 226, 298 nm) Select fraction C for the next step of separation.

[0050] S4. Fraction C is eluted by gradient elution using a petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction C13. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13 is 0.44-0.45. The volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when fraction C13 is eluted (i.e., the volume ratio corresponding to fraction C13) is (37:63)-(43:57), preferably 40:60.

[0051] The specific operating steps include: passing fraction C through silica gel column chromatography, using a petroleum ether-ethyl acetate gradient elution with volume ratios of 100:0, 95:5, 90:10, 85:15, 80:20, 75:25, 70:30, 65:35, 60:40, 55:45, 50:50, 45:55, 40:60, 35:65, 30:70, 27:73, 25:75, 22:78, 20:80, 17:83, 15:85, 12:88, 10:90, 5:95, and 0:100 to sequentially obtain fraction 1- 4, 5-8, 9-14, 15-18, 19-23, 24-30, 31-34, 35-41, 42-49, 50-55, 56-64, 65-71, 72-75, 76-80, 81-90, 91-100, 101-105, 106-109, 110-115, 116-119, 120-124, 125-130, 131-135, 136-138, 139-142, a total of 142 fractions, were then identified using silica gel thin-layer chromatography, based on wavelengths of 254 nm and 365 nm. The Rf values ​​of the fluorescent spots observed at nm were 0.84-0.88 (fractions 1-4), 0.80-0.83 (fractions 5-8), 0.76-0.78 (fractions 9-14), 0.72-0.75 (fractions 15-18), 0.68-0.70 (fractions 19-23), 0.64-0.67 (fractions 24-30), and 0.61-0.63 (fractions 19-23). Fractions 31-34), 0.57-0.59 (fraction 35-41), 0.52-0.55 (fraction 42-49), 0.49-0.51 (fraction 50-55), 0.46-0.48 (fraction 56-64), 0.46-0.47 (fraction 65-71), 0.44-0.45 (fraction 72-75), 0.42-0.43 (fraction 7) 6-80), 0.39-0.41 (fraction 81-90), 0.37-0.38 (fraction 91-100), 0.35-0.36 (fraction 101-105), 0.33-0.34 (fraction 106-109), 0.30-0.32 (fraction 110-115), 0.27-0.29 (fraction 116-119), 0.24-0 Fractions 0.26 (120-124), 0.21-0.23 (125-130), 0.20-0.22 (131-135), 0.18-0.19 (136-138), and 0.15-0.17 (139-142) are combined to obtain fractions C1-C25; among them, fractions 72-75 are combined to form fraction C13.

[0052] S5. Fraction C13 was subjected to gradient elution using a petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction C13.17. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17 was 0.25-0.27. The volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution eluted from fraction C13.17 (i.e., the fraction corresponding to C13.17) was (18:82)-(23:77), preferably 20:80.

[0053] The specific procedures include: analyzing the characteristic ultraviolet absorption (λ) of α-acids observed by HPLC. max= 226, 298nm) C13 was selected for further separation. The C13 fraction was subjected to silica gel column chromatography using volume ratios of 100:0, 95:5, 90:10, 85:15, 80:20, 75:25, 70:30, 65:35, 60:40, 55:45, 50:50, 45:55, 40:60, 35:65, 30:70, 25:75, 20:80, 15:85, 10:90, and 0:100 petroleum ether. Gradient elution with ether-ethyl acetate yielded 86 fractions: 1-5, 6-10, 11-15, 16-20, 21-25, 26-31, 32-35, 36-41, 42-46, 47-51, 52-55, 56-60, 61-68, 69-72, 73-77, 78-82, 83-86, 87-92, 93-96, and 97-102. These fractions were identified by silica gel thin-layer chromatography at wavelengths of 254 nm and 365 nm. The Rf values ​​of the fluorescent spots observed at nm were 0.83-0.85 (fractions 1-5), 0.79-0.82 (fractions 6-10), 0.75-0.78 (fractions 11-15), 0.71-0.73 (fractions 16-20), 0.67-0.69 (fractions 21-25), 0.63-0.66 (fractions 26-31), 0.54-0.62 (fractions 32-35), 0.47-0.50 (fractions 36-41), 0.43-0.45 (fractions 42-46), 0.41-0.42 (fractions 47-51), and 0.39-0.4. Fractions 0 (52-55), 0.37-0.38 (56-60), 0.35-0.36 (61-68), 0.33-0.34 (69-72), 0.31-0.32 (73-77), 0.28-0.30 (78-82), 0.25-0.27 (83-86), 0.22-0.24 (87-92), 0.19-0.21 (93-96), and 0.15-0.18 (97-102) were combined to obtain 20 fractions, C13.1-C13.20. Fractions 83-86 were combined to form fraction C13.17.

[0054] S6. Fraction C13.17 was isocratically eluted using Sephadex LH-20 gel column chromatography with petroleum ether-dichloromethane-methanol to obtain fraction C13.17.7. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17.7 was 0.57-0.60. The volume ratio of petroleum ether, dichloromethane and methanol in the petroleum ether-dichloromethane-methanol elution fraction C13.17.7 (i.e., the fraction corresponding to C13.17.7) was (7:3:0.5)-(3:7:1.5), preferably 5:5:1.

[0055] The specific procedures include: analyzing the characteristic ultraviolet absorption (λ) of α-acids observed by HPLC. max Fraction C13.17 (226, 298 nm) was selected for further separation. Fraction C13.17 was subjected to Sephadex LH-20 gel column chromatography with isocratic elution using dichloromethane-methanol at a volume ratio of 5:5. A total of 57 fractions (1-4, 5-8, 9-12, 13-15, 16-18, 19-22, 23-25, 26-29, 30-33, 34-36, 37-40, 41-43, 44-46, 47-50, 51-53, 54-57) were collected sequentially. These fractions were identified by silica gel thin-layer chromatography at wavelengths of 254 nm and 365 nm. The Rf values ​​of the fluorescent spots observed at nm were 0.79-0.82 (fractions 1-4), 0.75-0.78 (fractions 5-8), 0.72-0.74 (fractions 9-12), 0.67-0.70 (fractions 13-15), 0.64-0.66 (fractions 16-18), 0.61-0.63 (fractions 19-22), 0.57-0.60 (fractions 23-25), 0.53-0.55 (fractions 26-29), and 0.49-0. Fractions 52 (30-33), 0.43-0.47 (34-36), 0.40-0.42 (37-40), 0.37-0.39 (41-43), 0.34-0.36 (44-46), 0.31-0.33 (47-50), 0.27-0.30 (51-53), and 0.23-0.26 (54-57) were combined to obtain 16 fractions, C13.17.1 to C13.17.16. Fractions 23-25 ​​were combined to form fraction C13.17.7.

[0056] S7. Fraction C13.17.7 was subjected to gradient elution using methanol-water solution via ODS column chromatography to obtain fraction C13.17.7.8. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17.7.8 was 0.66-0.71. The volume ratio of methanol to water in the methanol-water solution eluted from fraction C13.17.7.8 (i.e., the volume ratio corresponding to fraction C13.17.7.8) was (66:34)-(72:28), preferably 70:30.

[0057] The specific procedures include: analyzing the characteristic ultraviolet absorption (λ) of α-acids observed by HPLC. max= 226, 298 nm) C13.17.7 was selected for further separation. Fraction C13.17.7 was subjected to ODS column chromatography using a methanol-water gradient elution with volume ratios of 30:70, 35:65, 40:60, 45:55, 50:50, 55:45, 60:40, 70:30, 80:20, 90:10, and 100:0. Fifty-five fractions (1-5, 6-10, 11-15, 16-20, 21-25, 26-31, 32-35, 36-41, 42-46, 47-51, and 52-55) were collected sequentially. These fractions were identified by silica gel thin-layer chromatography at wavelengths of 254 nm and 365 nm. The Rf values ​​of the fluorescent spots observed at nm were 0.22-0.24 (fractions 1-5), 0.27-0.32 (fractions 6-10), 0.34-0.37 (fractions 11-15), 0.39-0.43 (fractions 16-20), 0.45-0.52 (fractions 21-25), 0.53-0.58 (fractions 26-31), 0.60-0.64 (fractions 32-35), 0.66-0.71 (fractions 36-41), 0.72-0.74 (fractions 42-46), 0.75-0.78 (fractions 47-51), and 0.79-0.82 (fractions 52-55). These values ​​were combined to obtain 11 fractions, namely C13.17.7.1-C13.17.7.11. Among them, fractions 36-41 are combined to form fraction C13.17.7.8.

[0058] S8. The tricyclic α-acid compound having the structure shown in formula (1) and / or (2) was isolated from fraction C13.17.7.8 by HPLC. The mobile phase used in the HPLC method is acetonitrile-water solution.

[0059] This application uses acetonitrile-water as the mobile phase and employs HPLC method, utilizing C 18 Tricyclic α-acid compounds with the structural formulas shown in formulas (1) and (2) were prepared by chromatographic column in fraction C13.17.7.8.

[0060] Wherein, the volume ratio of acetonitrile to water in the acetonitrile-water solution is (42:58) to (47:53). Preferably, the volume ratio of acetonitrile to water in the acetonitrile-water solution is 45:55.

[0061] The characteristic ultraviolet absorption (λ) of tricyclic α-acids observed by HPLC analysis max= 226, 298 nm) Further separation of fraction C13.17.7.8 was performed, including α-acid compounds with structural formula as shown in formula (1) with a retention time of 30-34 min (purity: 99%); including α-acid compounds with structural formula as shown in formula (2) with a retention time of 36-38 min (purity: 99%).

[0062] The physical properties and detection data of compound 1 obtained in Example 1 are as follows: Pale yellow prismatic crystals; High-resolution mass spectrometry (HR-ESI-MS) m / z 379.2111 ([M + H]) + The calculated value is 379.2115), suggesting the molecular formula of the compound is C. 21 H 30 O6 has an unsaturation degree of 7.

[0063] 1 1H NMR (CDCl3, 400 MHz) spectrum ( Figure 1 The signal indicates a hydroxyl proton in the low-field region. d H 13.48 (1H, s, OH); Two methylene hydrogen proton signals are present in the lower field region, linked to electron-withdrawing groups. d H 4.76 (1H, overlapped, H-13), 4.73 (1H, overlapped, H-8); A single methylene hydrogen proton signal is present in the high-field region. d H 2.21 (1H, m, H-19), 3 sets of methylene proton signals d H 3.00 (2H, m, H-7), 2.77 (1H, dd, J =15.0, 7.0 Hz, H-18a), 2.91 (1H, dd, J = 14.8, 7.1 Hz, H-18b), 3.05 (2H, overlapped, H-12), 4 singlet methyl proton signals d H 1.32 (3H, s, 10-CH3), 1.21 (3H, s, 11-CH3), 1.26 (3H, s, 15-CH3), 1.38 (3H, s, 16-CH3), two bimodal methyl proton signals. d H 0.98(3H, d, J= 6.6 Hz, 20-CH3), 0.96 (3H, d, J = 6.6 Hz, 21-CH3). The above information suggests that this compound may be an α-acid.

[0064] 13 C10 NMR (CDCl3, 100 MHz) spectrum Figure 2 The HSQC spectrum showed a total of 21 carbon signals, among which... d C 204.3 (C-17) represents a single carbonyl carbon signal. d C 159.7 (C-1), 105.7 (C-2), 161.8 (C-3), 99.6 (C-4), 163.7 (C-5), and 102.3 (C-6) represent a group of aromatic carbon signals. d C 92.0 (C-13), 71.7 (C-14), 92.5 (C-8), and 72.1 (C-9) represent the carbon signals associated with oxygen. d C 25.5 (C-19) represents a single methylene carbon signal. d C 26.6 (C-12), 51.7 (C-18), and 27.3 (C-7) represent three methylene carbon signals. d C 24.8 (C-16), 26.4 (C-15), 22.9 (C-20), 22.9 (C-21), 23.9 (C-11), and 26.0 (C-10) represent six methyl carbon signals. Combined with the proton NMR spectrum information, this suggests that the compound contains one benzene ring, two 2-hydroxyisopropyl segments, one isovaleryl segment, and the remaining two unsaturated groups are occupied by two furan rings.

[0065] 1 H NMR, 13 The signal assignments for C NMR are shown in Table 1.

[0066] Table 1. NMR data of compound 1 d (ppm)

[0067] HMBC Map ( Figure 3 In ), H2-7 can be observed ( d H 3.03) and C-3 ( d C161.8), C-2 ( d C 105.7), C-1 ( d C 159.7), C-8 ( d C 92.5), C-9 ( d C The remote correlation signal of 72.1) and H2-12 ( d H 2.97) and C-4 ( d C 99.6), C-3 ( d C 161.8), C-5 ( d C 163.7), C-13 ( d C 92.0), C-18 ( d C The remote correlation signal of 71.7) indicates that two furan rings are connected to the C-4, C-5 and C-2, C-3 positions of the benzene ring, respectively, and that two 2-hydroxyisopropyl segments are connected at C-8 and C-13, respectively; in addition, H-19 ( d H 2.21) and C-17 ( d C 204.3), C-18 ( d C 51.7), C-20 ( d C 22.9), C-21 ( d C 22.9) remote correlation signals and H2-18 ( d H 2.78, 2.91) and C-17 ( d C 204.3), C-6 ( d C 102.3), C-19 ( d C The long-range correlation signal (25.5) confirmed the presence of the isovaleryl fragment and its connection at the C-6 position. Based on this information, the planar structure of compound 1 was determined, classifying it as an α-acid. Comparison of calculated circular dichroism (ECD) data with measured ECD data confirmed the absolute configuration of the compound to be 8. R ,13 R .

[0068] In summary, the structure of the new compound 1 (compound (+)-Humulune M) was determined as follows:

[0069] The physical properties and detection data of compound 2 obtained in Example 1 are as follows: Pale yellow prismatic crystals; High-resolution mass spectrometry (HR-ESI-MS) m / z 379.2111 ([M + H]) + The calculated value is 379.2115, suggesting the molecular formula of the compound is C. 21 H 30 O6 has an unsaturation degree of 7.

[0070] 1 1H NMR (CDCl3, 400 MHz) spectrum ( Figure 4 The signal indicates a hydroxyl proton in the low-field region. d H 13.48 (1H, s, OH); Two methylene hydrogen proton signals are present in the lower field region, linked to electron-withdrawing groups. d H 4.76 (1H, overlapped, H-13), 4.73 (1H, overlapped, H-8); A single methylene hydrogen proton signal is present in the high-field region. d H 2.21 (1H, m, H-19), 3 sets of methylene proton signals d H 3.00 (2H, m, H-7), 2.77 (1H, dd, J =15.0, 7.0 Hz, H-18a), 2.91 (1H, dd, J = 14.8, 7.1 Hz, H-18b), 3.05 (2H, overlapped, H-12), 4 singlet methyl proton signals d H 1.32 (3H, s, 10-CH3), 1.21 (3H, s, 11-CH3). This information suggests that the compound may be an α-acid.

[0071] 13 C10 NMR (CDCl3, 100 MHz) spectrum Figure 5 The HSQC spectrum showed a total of 21 carbon signals, among which... d C 204.3 (C-17) represents a single carbonyl carbon signal. dC 159.7 (C-1), 105.7 (C-2), 161.8 (C-3), 99.6 (C-4), 163.7 (C-5), and 102.3 (C-6) represent a group of aromatic carbon signals. d C 92.0 (C-13), 71.7 (C-14), 92.5 (C-8), and 72.1 (C-9) represent the carbon signals associated with oxygen. d C 25.5 (C-19) represents a single methylene carbon signal. d C 26.6 (C-12), 51.7 (C-18), and 27.3 (C-7) represent three methylene carbon signals. d C 24.8 (C-16), 26.4 (C-15), 22.9 (C-20), 22.9 (C-21), 23.9 (C-11), and 26.0 (C-10) represent six methyl carbon signals. Combined with the proton NMR spectrum information, this suggests that the compound contains one benzene ring, two 2-hydroxyisopropyl segments, one isovaleryl segment, and the remaining two unsaturated groups are occupied by two furan rings.

[0072] 1 H NMR, 13 The signal assignments for C NMR are shown in Table 2.

[0073] Table 2 NMR data of compound 2 d (ppm)

[0074] HMBC Map ( Figure 6 In ), H2-7 can be observed ( d H 3.03) and C-3 ( d C 161.8), C-2 ( d C 105.7), C-1 ( d C 159.7), C-8 ( d C 92.5), C-9 ( d C The remote correlation signal of 72.1) and H2-12 ( d H 2.97) and C-4 ( d C 99.6), C-3 ( d C 161.8), C-5 ( d C 163.7), C-13 ( d C 92.0), C-18 ( d C The remote correlation signal of 71.7) indicates that two furan rings are connected to the C-4, C-5 and C-2, C-3 positions of the benzene ring, respectively, and that two 2-hydroxyisopropyl segments are connected at C-8 and C-13, respectively; in addition, H-19 ( d H 2.21) and C-17 ( d C 204.3), C-18 ( d C 51.7), C-20 ( d C 22.9), C-21 ( d C 22.9) remote correlation signals and H2-18 ( d H 2.78, 2.91) and C-17 ( d C 204.3), C-6 ( d C 102.3), C-19 ( d C The long-range correlation signal (25.5) confirmed the presence of the isovaleryl fragment and its connection at the C-6 position. Based on this information, the planar structure of compound 2 was determined, classifying it as an α-acid. Comparison of calculated circular dichroism (ECD) data with measured ECD data confirmed the absolute configuration of the compound to be 8. S ,13 S .

[0075] In summary, the structure of the new compound 2 (compound (-)-Humulune M) was determined as follows:

[0076] Example 2 This embodiment discloses a method for preparing a novel tricyclic α-acid compound, comprising the following steps: S1. Take dried hop heads with flowers, add solvent for extraction, combine the extracts, filter, concentrate under reduced pressure to obtain an extract; specifically, the solvent is an 88-98 V% aqueous ethanol solution; the mass of the solvent added is 8-10 times that of the hops, the reflux extraction is performed 1-3 times, and each extraction is performed for 1-3 hours.

[0077] The specific implementation plan is as follows: Weigh 101.4 kg of dried hops with flowers, add 95% ethanol (10 times the weight of the hops) and extract three times, reflux at 75°C for 1 hour each time, combine the extracts and concentrate to obtain an extract (about 9.0 kg).

[0078] S2. After suspending the extract in water, extract it sequentially with petroleum ether and dichloromethane to obtain a dichloromethane extract; further, in step S2, after suspending the extract in 8-15 times its mass of water, extract it with dichloromethane.

[0079] The specific steps are as follows: Add the extract (about 9.0 kg) to 10 times its weight of water (90 L) to suspend it, and then extract it with 1.5 times the volume of dichloromethane. Extract it 3 times to obtain dichloromethane extract (about 3100 g).

[0080] S3. The extract was subjected to gradient elution using a petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction C. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C was 0.55-0.62. Based on a total volume of 100, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when fraction C was eluted was (50:50)-(70:30); more preferably, the volume ratio was 60:40.

[0081] It should be noted that elution with a petroleum ether-ethyl acetate solution at a volume ratio of (50:50) to (70:30) and identification of the fraction with an Rf value of 0.55-0.62 using silica gel thin-layer chromatography resulted in fraction C. The volume ratio of the petroleum ether-ethyl acetate solution required to obtain fraction C was between (50:50) and (70:30), with the highest efficiency achieved when eluting with a petroleum ether-ethyl acetate solution at a volume ratio of 60:40. The following gradient elution principles are essentially the same.

[0082] In this application, the Rf value is the Rf value of the fluorescent spot observed at scanning wavelengths of 254 nm and 365 nm.

[0083] The specific steps are as follows: The dichloromethane extract was subjected to silica gel column chromatography, using a petroleum ether-ethyl acetate gradient elution at volume ratios of 100:0, 80:20, 60:40, 30:70, and 0:100, with each 500 mL volume representing one fraction, yielding 35 fractions in total: 1-5, 6-10, 11-25, 26-29, and 30-35. These fractions were then identified using silica gel thin-layer chromatography, based on the values ​​at 254 nm and 365 nm. Fluorescent spots with Rf values ​​of 0.81-0.83 (fractions 1-5), 0.67-0.78 (fractions 6-10), 0.55-0.62 (fractions 11-25), 0.40-0.52 (fractions 26-29), and 0.32-0.38 (fractions 30-35) observed at nm were combined to obtain 5 fractions, named A, B, C, D, and E respectively; among them, fraction C was obtained by combining fractions 11-25.

[0084] The characteristic ultraviolet absorption (λ) of tricyclic α-acids observed by HPLC analysis max = 226, 298 nm) Select fraction C for the next step of separation.

[0085] S4. Fraction C is eluted by gradient elution using a petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction C13. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13 is 0.44-0.45. The volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when fraction C13 is eluted (i.e., the volume ratio corresponding to fraction C13) is (37:63)-(43:57), preferably 40:60.

[0086] The specific operating steps include: passing fraction C through silica gel column chromatography, using a petroleum ether-ethyl acetate gradient elution with volume ratios of 100:0, 95:5, 90:10, 85:15, 80:20, 75:25, 70:30, 65:35, 60:40, 55:45, 50:50, 45:55, 40:60, 35:65, 30:70, 27:73, 25:75, 22:78, 20:80, 17:83, 15:85, 12:88, 10:90, 5:95, and 0:100 to sequentially obtain fraction 1- 4, 5-8, 9-14, 15-18, 19-23, 24-30, 31-34, 35-41, 42-49, 50-55, 56-64, 65-71, 72-75, 76-80, 81-90, 91-100, 101-105, 106-109, 110-115, 116-119, 120-124, 125-130, 131-135, 136-138, 139-142, a total of 142 fractions, were then identified using silica gel thin-layer chromatography, based on wavelengths of 254 nm and 365 nm. The Rf values ​​of the fluorescent spots observed at nm were 0.84-0.88 (fractions 1-4), 0.80-0.83 (fractions 5-8), 0.76-0.78 (fractions 9-14), 0.72-0.75 (fractions 15-18), 0.68-0.70 (fractions 19-23), 0.64-0.67 (fractions 24-30), and 0.61-0.63 (fractions 19-23). Fractions 31-34), 0.57-0.59 (fraction 35-41), 0.52-0.55 (fraction 42-49), 0.49-0.51 (fraction 50-55), 0.46-0.48 (fraction 56-64), 0.46-0.47 (fraction 65-71), 0.44-0.45 (fraction 72-75), 0.42-0.43 (fraction 7) 6-80), 0.39-0.41 (fraction 81-90), 0.37-0.38 (fraction 91-100), 0.35-0.36 (fraction 101-105), 0.33-0.34 (fraction 106-109), 0.30-0.32 (fraction 110-115), 0.27-0.29 (fraction 116-119), 0.24-0 Fractions 0.26 (120-124), 0.21-0.23 (125-130), 0.20-0.22 (131-135), 0.18-0.19 (136-138), and 0.15-0.17 (139-142) are combined to obtain fractions C1-C25; among them, fractions 72-75 are combined to form fraction C13.

[0087] S5. Fraction C13 was subjected to gradient elution using a petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction C13.17. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17 was 0.25-0.27. The volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution eluted from fraction C13.17 (i.e., the fraction corresponding to C13.17) was (18:82)-(23:77), preferably 20:80.

[0088] The specific procedures include: analyzing the characteristic ultraviolet absorption (λ) of α-acids observed by HPLC. max= 226, 298nm) C13 was selected for further separation. The C13 fraction was subjected to silica gel column chromatography using volume ratios of 100:0, 95:5, 90:10, 85:15, 80:20, 75:25, 70:30, 65:35, 60:40, 55:45, 50:50, 45:55, 40:60, 35:65, 30:70, 25:75, 20:80, 15:85, 10:90, and 0:100 petroleum ether. Gradient elution with ether-ethyl acetate yielded 86 fractions: 1-5, 6-10, 11-15, 16-20, 21-25, 26-31, 32-35, 36-41, 42-46, 47-51, 52-55, 56-60, 61-68, 69-72, 73-77, 78-82, 83-86, 87-92, 93-96, and 97-102. These fractions were identified by silica gel thin-layer chromatography at wavelengths of 254 nm and 365 nm. The Rf values ​​of the fluorescent spots observed at nm were 0.83-0.85 (fractions 1-5), 0.79-0.82 (fractions 6-10), 0.75-0.78 (fractions 11-15), 0.71-0.73 (fractions 16-20), 0.67-0.69 (fractions 21-25), 0.63-0.66 (fractions 26-31), 0.54-0.62 (fractions 32-35), 0.47-0.50 (fractions 36-41), 0.43-0.45 (fractions 42-46), 0.41-0.42 (fractions 47-51), and 0.39-0.4. Fractions 0 (52-55), 0.37-0.38 (56-60), 0.35-0.36 (61-68), 0.33-0.34 (69-72), 0.31-0.32 (73-77), 0.28-0.30 (78-82), 0.25-0.27 (83-86), 0.22-0.24 (87-92), 0.19-0.21 (93-96), and 0.15-0.18 (97-102) were combined to obtain 20 fractions, C13.1-C13.20. Fractions 83-86 were combined to form fraction C13.17.

[0089] S6. Fraction C13.17 was isocratically eluted using Sephadex LH-20 gel column chromatography with petroleum ether-dichloromethane-methanol to obtain fraction C13.17.7. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17.7 was 0.57-0.60. The volume ratio of petroleum ether, dichloromethane and methanol in the petroleum ether-dichloromethane-methanol elution fraction C13.17.7 (i.e., the fraction corresponding to C13.17.7) was (7:3:0.5)-(3:7:1.5), preferably 5:5:1.

[0090] The specific procedures include: analyzing the characteristic ultraviolet absorption (λ) of α-acids observed by HPLC. max Fraction C13.17 (226, 298 nm) was selected for further separation. Fraction C13.17 was subjected to Sephadex LH-20 gel column chromatography with isocratic elution using dichloromethane-methanol at a volume ratio of 5:5. A total of 57 fractions (1-4, 5-8, 9-12, 13-15, 16-18, 19-22, 23-25, 26-29, 30-33, 34-36, 37-40, 41-43, 44-46, 47-50, 51-53, 54-57) were collected sequentially. These fractions were identified by silica gel thin-layer chromatography at wavelengths of 254 nm and 365 nm. The Rf values ​​of the fluorescent spots observed at nm were 0.79-0.82 (fractions 1-4), 0.75-0.78 (fractions 5-8), 0.72-0.74 (fractions 9-12), 0.67-0.70 (fractions 13-15), 0.64-0.66 (fractions 16-18), 0.61-0.63 (fractions 19-22), 0.57-0.60 (fractions 23-25), 0.53-0.55 (fractions 26-29), and 0.49-0. Fractions 52 (30-33), 0.43-0.47 (34-36), 0.40-0.42 (37-40), 0.37-0.39 (41-43), 0.34-0.36 (44-46), 0.31-0.33 (47-50), 0.27-0.30 (51-53), and 0.23-0.26 (54-57) were combined to obtain 16 fractions, C13.17.1 to C13.17.16. Fractions 23-25 ​​were combined to form fraction C13.17.7.

[0091] S7. Fraction C13.17.7 was subjected to gradient elution using methanol-water solution via ODS column chromatography to obtain fraction C13.17.7.7. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17.7.7 was 0.60-0.64. The volume ratio of methanol to water in the methanol-water solution eluted from fraction C13.17.7.7 (i.e., the volume ratio corresponding to fraction C13.17.7.7) was (56:44)-(62:38), preferably 60:40.

[0092] The specific procedures include: analyzing the characteristic ultraviolet absorption (λ) of α-acids observed by HPLC. max= 226, 298 nm) C13.17.7 was selected for further separation. Fraction C13.17.7 was subjected to ODS column chromatography using a methanol-water gradient elution with volume ratios of 30:70, 35:65, 40:60, 45:55, 50:50, 55:45, 60:40, 70:30, 80:20, 90:10, and 100:0. Fifty-five fractions (1-5, 6-10, 11-15, 16-20, 21-25, 26-31, 32-35, 36-41, 42-46, 47-51, and 52-55) were collected sequentially. These fractions were identified by silica gel thin-layer chromatography at wavelengths of 254 nm and 365 nm. The Rf values ​​of the fluorescent spots observed at nm were 0.22-0.24 (fractions 1-5), 0.27-0.32 (fractions 6-10), 0.34-0.37 (fractions 11-15), 0.39-0.43 (fractions 16-20), 0.45-0.52 (fractions 21-25), 0.53-0.58 (fractions 26-31), 0.60-0.64 (fractions 32-35), 0.66-0.71 (fractions 36-41), 0.72-0.74 (fractions 42-46), 0.75-0.78 (fractions 47-51), and 0.79-0.82 (fractions 52-55). These values ​​were combined to obtain 11 fractions, namely C13.17.7.1-C13.17.7.11. Among them, fractions 32-35 are combined to form fraction C13.17.7.7.

[0093] S8. The tricyclic α-acid compound having the structure shown in formula (3) and / or (4) was isolated from fraction C13.17.7.7 by HPLC. The mobile phase used in the HPLC method is acetonitrile-water solution.

[0094] This application uses acetonitrile-water as the mobile phase and employs HPLC method, utilizing C 18 Tricyclic α-acid compounds with structural formulas as shown in formulas (3) and (4) were prepared by chromatographic column in fraction C13.17.7.7.

[0095] Wherein, the volume ratio of acetonitrile to water in the acetonitrile-water solution is (45:55) to (50:50). Preferably, the volume ratio of acetonitrile to water in the acetonitrile-water solution is 48:52.

[0096] The characteristic ultraviolet absorption (λ) of tricyclic α-acids observed by HPLC analysis max= 226, 298 nm) Further separation of fraction C13.17.7.7 was performed, including α-acid compounds with structural formula as shown in formula (1) with a retention time of 21-24 min (purity: 99%); including α-acid compounds with structural formula as shown in formula (2) with a retention time of 26-28 min (purity: 99%).

[0097] The physical properties and detection data of compound 3 obtained in Example 2 are as follows: White powder; high-resolution mass spectrometry (HR-ESI-MS) m / z 379.2111 ([M + H]) + The calculated value is 379.2115), suggesting the molecular formula of the compound is C. 21 H 30 O6 has an unsaturation degree of 7.

[0098] 1 1H NMR (CDCl3, 400 MHz) spectrum ( Figure 7 The signal indicates a hydroxyl proton in the low-field region. d H 13.48 (1H, s, OH); Two methylene hydrogen proton signals are present in the lower field region, linked to electron-withdrawing groups. d H 4.76 (1H, overlapped, H-13), 4.73 (1H, overlapped, H-8); A single methylene hydrogen proton signal is present in the high-field region. d H 2.21 (1H, m, H-19), 3 sets of methylene proton signals d H 3.04 (2H, m, H-7), 2.77 (1H, dd, J =15.0, 7.0 Hz, H-18a), 2.91 (1H, dd, J = 14.8, 7.1 Hz, H-18b), 3.07 (1H, overlapped, H-12a), 2.96 (1H, overlapped, H-12b), 4 single-peak methyl proton signals d H 1.32 (3H, s, 10-CH3), 1.21 (3H, s, 11-CH3), 1.26 (3H, s, 15-CH3), 1.38 (3H, s, 16-CH3), two bimodal methyl proton signals. d H 0.98 (3H, d,J = 6.6 Hz, 20-CH3), 0.96 (3H, d, J = 6.6Hz, 21-CH3). The above information suggests that this compound may be an α-acid.

[0099] 13 C10 NMR (CDCl3, 100 MHz) spectrum Figure 8 The HSQC spectrum showed a total of 21 carbon signals, among which... d C 204.3 (C-17) represents a single carbonyl carbon signal. d C 159.7 (C-1), 105.7 (C-2), 161.8 (C-3), 99.6 (C-4), 163.7 (C-5), and 102.3 (C-6) represent a group of aromatic carbon signals. d C 92.0 (C-13), 71.7 (C-14), 92.5 (C-8), and 72.1 (C-9) represent the carbon signals associated with oxygen. d C 25.5 (C-19) represents a single methylene carbon signal. d C 26.6 (C-12), 51.7 (C-18), and 27.3 (C-7) represent three methylene carbon signals. d C 24.8 (C-16), 26.4 (C-15), 22.9 (C-20), 22.9 (C-21), 23.9 (C-11), and 26.0 (C-10) represent six methyl carbon signals. Combined with the proton NMR spectrum information, this suggests that the compound contains one benzene ring, two 2-hydroxyisopropyl segments, one isovaleryl segment, and the remaining two unsaturated groups are occupied by two furan rings.

[0100] 1 H NMR, 13 The signal assignments for C NMR are shown in Table 3.

[0101] Table 3 NMR data of compound 3 d (ppm)

[0102] HMBC Map ( Figure 9 In ), H2-7 can be observed ( d H 3.03) and C-3 ( dC 161.8), C-2 ( d C 105.7), C-1 ( d C 159.7), C-8 ( d C 92.5), C-9 ( d C The remote correlation signal of 72.1) and H2-12 ( d H 2.97) and C-4 ( d C 99.6), C-3 ( d C 161.8), C-5 ( d C 163.7), C-13 ( d C 92.0), C-18 ( d C The remote correlation signal of 71.7) indicates that two furan rings are connected to the C-4, C-5 and C-2, C-3 positions of the benzene ring, respectively, and that two 2-hydroxyisopropyl segments are connected at C-8 and C-13, respectively; in addition, H-19 ( d H 2.21) and C-17 ( d C 204.3), C-18 ( d C 51.7), C-20 ( d C 22.9), C-21 ( d C 22.9) remote correlation signals and H2-18 ( d H 2.78, 2.91) and C-17 ( d C 204.3), C-6 ( d C 102.3), C-19 ( d C The long-range correlation signal (25.5) confirmed the presence of the isovaleryl fragment and its connection at the C-6 position. Based on this information, the planar structure of compound 3 was determined, classifying it as an α-acid. Comparison of calculated circular dichroism (ECD) data with measured ECD data confirmed the absolute configuration of the compound to be 8. R ,13 S .

[0103] In summary, the structure of the new compound 3 (compound (+)-Humulune N) was determined as follows:

[0104] The physical properties and detection data of compound 4 obtained in Example 2 are as follows: White powder; high-resolution mass spectrometry (HR-ESI-MS) m / z 379.2111 ([M + H]) + The calculated value is 379.2115), suggesting the molecular formula of the compound is C. 21 H 30 O6 has an unsaturation degree of 7.

[0105] 1 1H NMR (CDCl3, 400 MHz) spectrum ( Figure 10 The signal indicates a hydroxyl proton in the low-field region. d H 13.48 (1H, s, OH); Two methylene hydrogen proton signals are present in the low-field region, linked to electron-withdrawing groups. d H 4.76 (1H, overlapped, H-13), 4.73 (1H, overlapped, H-8); A single methylene hydrogen proton signal is present in the high-field region. d H 2.21 (1H, m, H-19), 3 sets of methylene proton signals d H 3.04 (2H, m, H-7), 2.77 (1H, dd, J =15.0, 7.0 Hz, H-18a), 2.91 (1H, dd, J = 14.8, 7.1 Hz, H-18b), 3.07 (1H, overlapped, H-12a), 2.96 (1H, overlapped, H-12b), 4 single-peak methyl proton signals d H 1.32 (3H, s, 10-CH3), 1.21 (3H, s, 11-CH3), 1.26 (3H, s, 15-CH3), 1.38 (3H, s, 16-CH3), two bimodal methyl proton signals. d H 0.98 (3H, d, J = 6.6 Hz, 20-CH3), 0.96 (3H, d, J= 6.6Hz, 21-CH3). The above information suggests that this compound may be an α-acid.

[0106] 13 C10 NMR (CDCl3, 100 MHz) spectrum Figure 11 The HSQC spectrum showed a total of 21 carbon signals, among which... d C 204.3 (C-17) represents a single carbonyl carbon signal. d C 159.7 (C-1), 105.7 (C-2), 161.8 (C-3), 99.6 (C-4), 163.7 (C-5), and 102.3 (C-6) represent a group of aromatic carbon signals. d C 91.9 (C-13), 71.7 (C-14), 92.5 (C-8), and 72.1 (C-9) represent the carbon signals associated with oxygen. d C 25.5 (C-19) represents a single methylene carbon signal. d C 26.6 (C-12), 51.7 (C-18), and 27.3 (C-7) represent three methylene carbon signals. d C 24.8 (C-16), 26.4 (C-15), 22.9 (C-20), 22.9 (C-21), 23.9 (C-11), and 26.0 (C-10) represent six methyl carbon signals. Combined with the proton NMR spectrum information, this suggests that the compound contains one benzene ring, two 2-hydroxyisopropyl segments, one isovaleryl segment, and the remaining two unsaturated groups are occupied by two furan rings.

[0107] 1 H NMR, 13 The signal assignments for C NMR are shown in Table 4.

[0108] Table 4 NMR data of compound 4 d (ppm)

[0109] HMBC Map ( Figure 12 In ), H2-7 can be observed ( d H 3.03) and C-3 ( d C 161.8), C-2 ( d C 105.7), C-1 ( d C 159.7), C-8 ( d C 92.5), C-9 ( d C The remote correlation signal of 72.1) and H2-12 ( d H 2.97) and C-4 ( d C 99.6), C-3 ( d C 161.8), C-5 ( d C 163.7), C-13 ( d C 92.0), C-18 ( d C The remote correlation signal of 71.7) indicates that two furan rings are connected to the C-4, C-5 and C-2, C-3 positions of the benzene ring, respectively, and that two 2-hydroxyisopropyl segments are connected at C-8 and C-13, respectively; in addition, H-19 ( d H 2.21) and C-17 ( d C 204.3), C-18 ( d C 51.7), C-20 ( d C 22.9), C-21 ( d C 22.9) remote correlation signals and H2-18 ( d H 2.78, 2.91) and C-17 ( d C 204.3), C-6 ( d C 102.3), C-19 ( d C The long-range correlation signal (25.5) confirmed the presence of the isovaleryl fragment and its connection at the C-6 position. Based on this information, the planar structure of compound 4 was determined, classifying it as an α-acid. Comparison of calculated circular dichroism (ECD) data with measured ECD data confirmed the absolute configuration of the compound to be 8. S ,13 R .

[0110] In summary, the structure of the new compound 4 (compound (-)-Humulune N) was determined as follows: .

[0111] Example 3 This embodiment discloses a method for preparing a novel tricyclic α-acid compound, comprising the following steps: S1. Take dried hop heads with flowers, add solvent for extraction, combine the extracts, filter, concentrate under reduced pressure to obtain an extract; specifically, the solvent is an 88-98 V% aqueous ethanol solution; the mass of the solvent added is 8-10 times that of the hops, the reflux extraction is performed 1-3 times, and each extraction is performed for 1-3 hours.

[0112] The specific implementation plan is as follows: Weigh 101.4 kg of dried hops with flowers, add 95% ethanol (10 times the weight of the hops) and extract three times, reflux at 75°C for 1 hour each time, combine the extracts and concentrate to obtain an extract (about 9.0 kg).

[0113] S2. After suspending the extract in water, extract it sequentially with petroleum ether and dichloromethane to obtain a dichloromethane extract; further, in step S2, after suspending the extract in 8-15 times its mass of water, extract it with dichloromethane.

[0114] The specific steps are as follows: Add the extract (about 9.0 kg) to 10 times its weight of water (90 L) to suspend it, and then extract it with 1.5 times the volume of dichloromethane. Extract it 3 times to obtain dichloromethane extract (about 3100 g).

[0115] S3. The extract was subjected to gradient elution using a petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction C. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C was 0.55-0.62. Based on a total volume of 100, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when fraction C was eluted was (50:50)-(70:30); more preferably, the volume ratio was 60:40.

[0116] It should be noted that elution with a petroleum ether-ethyl acetate solution at a volume ratio of (50:50) to (70:30) and identification of the fraction with an Rf value of 0.55-0.62 using silica gel thin-layer chromatography resulted in fraction C. The volume ratio of the petroleum ether-ethyl acetate solution required to obtain fraction C was between (50:50) and (70:30), with the highest efficiency achieved when eluting with a petroleum ether-ethyl acetate solution at a volume ratio of 60:40. The following gradient elution principles are essentially the same.

[0117] In this application, the Rf value is the Rf value of the fluorescent spot observed at scanning wavelengths of 254 nm and 365 nm.

[0118] The specific steps are as follows: The dichloromethane extract was subjected to silica gel column chromatography, using a petroleum ether-ethyl acetate gradient elution at volume ratios of 100:0, 80:20, 60:40, 30:70, and 0:100, with each 500 mL volume representing one fraction, yielding 35 fractions in total: 1-5, 6-10, 11-25, 26-29, and 30-35. These fractions were then identified using silica gel thin-layer chromatography, based on the values ​​at 254 nm and 365 nm. Fluorescent spots with Rf values ​​of 0.81-0.83 (fractions 1-5), 0.67-0.78 (fractions 6-10), 0.55-0.62 (fractions 11-25), 0.40-0.52 (fractions 26-29), and 0.32-0.38 (fractions 30-35) observed at nm were combined to obtain 5 fractions, named A, B, C, D, and E respectively; among them, fraction C was obtained by combining fractions 11-25.

[0119] The characteristic ultraviolet absorption (λ) of tricyclic α-acids observed by HPLC analysis max (225, 295 nm) Select fraction C for the next step of separation.

[0120] S4. Fraction C is eluted by gradient elution using a petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction C13. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13 is 0.44-0.45. The volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when fraction C13 is eluted (i.e., the volume ratio corresponding to fraction C13) is (37:63)-(43:57), preferably 40:60.

[0121] The specific operating steps include: passing fraction C through silica gel column chromatography, using a petroleum ether-ethyl acetate gradient elution with volume ratios of 100:0, 95:5, 90:10, 85:15, 80:20, 75:25, 70:30, 65:35, 60:40, 55:45, 50:50, 45:55, 40:60, 35:65, 30:70, 27:73, 25:75, 22:78, 20:80, 17:83, 15:85, 12:88, 10:90, 5:95, and 0:100 to sequentially obtain fraction 1- 4, 5-8, 9-14, 15-18, 19-23, 24-30, 31-34, 35-41, 42-49, 50-55, 56-64, 65-71, 72-75, 76-80, 81-90, 91-100, 101-105, 106-109, 110-115, 116-119, 120-124, 125-130, 131-135, 136-138, 139-142, a total of 142 fractions, were then identified using silica gel thin-layer chromatography, based on wavelengths of 254 nm and 365 nm. The Rf values ​​of the fluorescent spots observed at nm were 0.84-0.88 (fractions 1-4), 0.80-0.83 (fractions 5-8), 0.76-0.78 (fractions 9-14), 0.72-0.75 (fractions 15-18), 0.68-0.70 (fractions 19-23), 0.64-0.67 (fractions 24-30), and 0.61-0.63 (fractions 19-23). Fractions 31-34), 0.57-0.59 (fraction 35-41), 0.52-0.55 (fraction 42-49), 0.49-0.51 (fraction 50-55), 0.46-0.48 (fraction 56-64), 0.46-0.47 (fraction 65-71), 0.44-0.45 (fraction 72-75), 0.42-0.43 (fraction 7) 6-80), 0.39-0.41 (fraction 81-90), 0.37-0.38 (fraction 91-100), 0.35-0.36 (fraction 101-105), 0.33-0.34 (fraction 106-109), 0.30-0.32 (fraction 110-115), 0.27-0.29 (fraction 116-119), 0.24-0 Fractions 0.26 (120-124), 0.21-0.23 (125-130), 0.20-0.22 (131-135), 0.18-0.19 (136-138), and 0.15-0.17 (139-142) are combined to obtain fractions C1-C25; among them, fractions 72-75 are combined to form fraction C13.

[0122] S5. Fraction C13 was subjected to gradient elution using a petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction C13.17. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17 was 0.25-0.27. The volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution eluted from fraction C13.17 (i.e., the fraction corresponding to C13.17) was (18:82)-(23:77), preferably 20:80.

[0123] The specific procedures include: analyzing the characteristic ultraviolet absorption (λ) of α-acids observed by HPLC. max= 225, 295nm) C13 was selected for further separation. The C13 fraction was subjected to silica gel column chromatography using volume ratios of 100:0, 95:5, 90:10, 85:15, 80:20, 75:25, 70:30, 65:35, 60:40, 55:45, 50:50, 45:55, 40:60, 35:65, 30:70, 25:75, 20:80, 15:85, 10:90, and 0:100 petroleum ether. Gradient elution with ether-ethyl acetate yielded 86 fractions: 1-5, 6-10, 11-15, 16-20, 21-25, 26-31, 32-35, 36-41, 42-46, 47-51, 52-55, 56-60, 61-68, 69-72, 73-77, 78-82, 83-86, 87-92, 93-96, and 97-102. These fractions were identified by silica gel thin-layer chromatography at wavelengths of 254 nm and 365 nm. The Rf values ​​of the fluorescent spots observed at nm were 0.83-0.85 (fractions 1-5), 0.79-0.82 (fractions 6-10), 0.75-0.78 (fractions 11-15), 0.71-0.73 (fractions 16-20), 0.67-0.69 (fractions 21-25), 0.63-0.66 (fractions 26-31), 0.54-0.62 (fractions 32-35), 0.47-0.50 (fractions 36-41), 0.43-0.45 (fractions 42-46), 0.41-0.42 (fractions 47-51), and 0.39-0.4. Fractions 0 (52-55), 0.37-0.38 (56-60), 0.35-0.36 (61-68), 0.33-0.34 (69-72), 0.31-0.32 (73-77), 0.28-0.30 (78-82), 0.25-0.27 (83-86), 0.22-0.24 (87-92), 0.19-0.21 (93-96), and 0.15-0.18 (97-102) were combined to obtain 20 fractions, C13.1-C13.20. Fractions 83-86 were combined to form fraction C13.17.

[0124] S6. Fraction C13.17 was isocratically eluted using Sephadex LH-20 gel column chromatography with petroleum ether-dichloromethane-methanol to obtain fraction C13.17.7. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17.7 was 0.57-0.60. The volume ratio of petroleum ether, dichloromethane and methanol in the petroleum ether-dichloromethane-methanol elution fraction C13.17.7 (i.e., the fraction corresponding to C13.17.7) was (7:3:0.5)-(3:7:1.5), preferably 5:5:1.

[0125] The specific procedures include: analyzing the characteristic ultraviolet absorption (λ) of α-acids observed by HPLC. max Fraction C13.17 was selected for further separation at wavelengths of 225 nm and 295 nm. Fraction C13.17 was subjected to Sephadex LH-20 gel column chromatography with isocratic elution using dichloromethane-methanol at a volume ratio of 5:5. Fifty-seven fractions (1-4, 5-8, 9-12, 13-15, 16-18, 19-22, 23-25, 26-29, 30-33, 34-36, 37-40, 41-43, 44-46, 47-50, 51-53, and 54-57) were collected sequentially. These fractions were identified by silica gel thin-layer chromatography at wavelengths of 254 nm and 365 nm. The Rf values ​​of the fluorescent spots observed at nm were 0.79-0.82 (fractions 1-4), 0.75-0.78 (fractions 5-8), 0.72-0.74 (fractions 9-12), 0.67-0.70 (fractions 13-15), 0.64-0.66 (fractions 16-18), 0.61-0.63 (fractions 19-22), 0.57-0.60 (fractions 23-25), 0.53-0.55 (fractions 26-29), and 0.49-0. Fractions 52 (30-33), 0.43-0.47 (34-36), 0.40-0.42 (37-40), 0.37-0.39 (41-43), 0.34-0.36 (44-46), 0.31-0.33 (47-50), 0.27-0.30 (51-53), and 0.23-0.26 (54-57) were combined to obtain 16 fractions, C13.17.1 to C13.17.16. Fractions 23-25 ​​were combined to form fraction C13.17.7.

[0126] S7. Using methanol-water solution, fraction C13.17.5 was subjected to gradient elution via ODS column chromatography to obtain fraction C13.17.7.5. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17.7.5 was 0.45-0.52. The volume ratio of methanol to water in the methanol-water solution eluted from fraction C13.17.7.5 (i.e., the volume ratio corresponding to fraction C13.17.7.5) was (47:53)-(53:47), preferably 50:50.

[0127] The specific procedures include: analyzing the characteristic ultraviolet absorption (λ) of α-acids observed by HPLC. max= 225, 295 nm) C13.17.7 was selected for further separation. Fraction C13.17.7 was subjected to ODS column chromatography using a methanol-water gradient elution with volume ratios of 30:70, 35:65, 40:60, 45:55, 50:50, 55:45, 60:40, 70:30, 80:20, 90:10, and 100:0. Fifty-five fractions (1-5, 6-10, 11-15, 16-20, 21-25, 26-31, 32-35, 36-41, 42-46, 47-51, and 52-55) were collected sequentially. These fractions were identified by silica gel thin-layer chromatography at wavelengths of 254 nm and 365 nm. The Rf values ​​of the fluorescent spots observed at nm were 0.22-0.24 (fractions 1-5), 0.27-0.32 (fractions 6-10), 0.34-0.37 (fractions 11-15), 0.39-0.43 (fractions 16-20), 0.45-0.52 (fractions 21-25), 0.53-0.58 (fractions 26-31), 0.60-0.64 (fractions 32-35), 0.66-0.71 (fractions 36-41), 0.72-0.74 (fractions 42-46), 0.75-0.78 (fractions 47-51), and 0.79-0.82 (fractions 52-55). These values ​​were combined to obtain 11 fractions, namely C13.17.7.1-C13.17.7.11. Among them, fractions 21-25 are combined to form fraction C13.17.7.5.

[0128] S8. The tricyclic α-acid compounds having structures as shown in formula (5), formula (6), formula (7) and / or (8) were separated from fraction C13.17.7.5 by HPLC. The mobile phase used in the HPLC method is methanol-water solution.

[0129] This application uses methanol-water as the mobile phase and employs HPLC method, utilizing C 18 Tricyclic α-acid compounds with the structural formulas shown in formulas (5), (6), (7), and (8) were prepared by chromatographic column in fraction C13.17.7.5.

[0130] Wherein, the volume ratio of methanol to water in the methanol-water solution is (62:38) to (68:32). Preferably, the volume ratio of methanol to water in the methanol-water solution is 65:35.

[0131] The characteristic ultraviolet absorption (λ) of tricyclic α-acids observed by HPLC analysis max= 225, 295 nm) Further separation of fraction C13.17.7.5 was performed, including α-acid compounds with the structural formula as shown in formula (5) with a retention time of 35-37 min (purity: 99%); α-acid compounds with the structural formula as shown in formula (6) with a retention time of 39-42 min (purity: 99%); α-acid compounds with the structural formula as shown in formula (7) with a retention time of 22-24 min (purity: 99%); and α-acid compounds with the structural formula as shown in formula (8) with a retention time of 26-29 min (purity: 99%).

[0132] The physical properties and detection data of compound 5 obtained in Example 3 are as follows: Pale yellow oily liquid; High-resolution mass spectrometry (HR-ESI-MS) m / z 365.1951 ([M + H]) + The calculated value is 365.1959, suggesting the molecular formula of the compound is C. 20 H 28 O6 has an unsaturation degree of 7.

[0133] 1 1H NMR (CDCl3, 400 MHz) spectrum ( Figure 13 )show d H 13.50 (1H, s, 1-OH) represents a single phenolic hydroxyl proton signal, while the lower field region shows three methylene hydrogen proton signals linked to electron-withdrawing groups. d H 4.75 (1H, t, J = 8.3Hz, H-8), 4.72 (1H, t, J = 8.0 Hz, H-13), 3.69 (1H, p, J = 6.8 Hz, H-18), the higher field region shows two sets of methylene hydrogen proton signals. d H 3.08 (1H, dd, J = 15.2, 9.5 Hz, H-7a), 2.98 (1H, dd, J =15.2 Hz, 8.2 Hz, H-7b), 3.04 (2H, dd, J = 9.2, 1.3 Hz, H2-12), the high field region shows four single-peak methyl hydrogen proton signals. d H1.37 (3H, s, 15-CH3), 1.26 (3H, s, 16-CH3), 1.33 (3H, s, 10-CH3), 1.22 (3H, s, 11-CH3), two bimodal methyl hydrogen proton signals. d H 1.19 (3H, d, J = 6.8 Hz, 19-CH3), 1.17 (3H, d, J = 6.8 Hz, 20-CH3). The above 1H NMR spectrum information suggests that this compound may be an α-acid.

[0134] 13 C10 NMR (CDCl3, 100 MHz) spectrum Figure 14 The HSQC spectrum showed a total of 20 carbon signals, including one carbonyl carbon signal. d C 208.7 (C-17), a set of benzene ring carbon signals d C 161.8 (C-3), 99.5 (C-4), 163.3 (C-5), 101.2 (C-6), 160.1 (C-1), 105.9 (C-2), four carbon signals associated with oxygen. d C 91.9 (C-8), 71.8 (C-9), 92.5 (C-13), 72.1 (C-14), 1 methylene carbon signal d C 38.6 (C-18), 2 methylene carbon signals d C 26.7 (C-12), 27.3 (C-7), 6 methyl carbon signals d C 26.0 (C-10), 26.3 (C-15), 19.6 (C-19), 18.9 (C-20), 24.0 (C-11), 24.7 (C-16). The above information is combined... 1 The 1H NMR spectrum indicates that, apart from the benzene ring and carbonyl group, the remaining two unsaturations of this compound are occupied by two rings. In addition, the structure also contains two 2-hydroxyisopropyl segments and one isobutyryl segment.

[0135] 1 H NMR, 13 The signal assignments for C NMR are shown in Table 5.

[0136] Table 5 NMR data of compound 5 d(ppm)

[0137] HMBC Map ( Figure 18 In ), H2-7 can be observed ( d H 3.08) and C-3 ( d C 161.8), C-2 ( d C 105.9), C-1 ( d C 160.1), C-8 ( d C 91.9), C-9 ( d C The remote correlation signal of 71.8) and H2-12 ( d H 3.04) and C-4 ( d C 99.5), C-3 ( d C 161.8), C-5 ( d C 163.3), C-13 ( d C 92.5), C-14 ( d C The remote correlation signal of 72.1) indicates that two furan rings are connected to the C-4, C-5 and C-2, C-3 positions of the benzene ring, respectively, and that two 2-hydroxyisopropyl segments are connected at C-8 and C-13, respectively; in addition, H-18 ( d H 3.69) and C-17 ( d C 208.7), C-19 ( d C 19.6), C-20 ( d C 18.9), C-6 ( d C The long-range correlation signal (101.2) confirmed the presence of the isobutyryl fragment and its connection at the C-6 position. Based on this information, the planar structure of compound 5 was determined, classifying it as an α-acid. Comparison of calculated circular dichroism (ECD) data with measured ECD data confirmed the absolute configuration of the compound to be 8. R ,13 S .

[0138] In summary, the structure of the new compound 5 (compound (+)-Humulune O) was determined as follows:

[0139] The physical properties and detection data of compound 6 obtained in Example 3 are as follows: Pale yellow oily liquid; High-resolution mass spectrometry (HR-ESI-MS) m / z 365.1951 ([M + H]) + The calculated value is 365.1959, suggesting the molecular formula of the compound is C. 20 H 28 O6 has an unsaturation degree of 7.

[0140] 1 1H NMR (CDCl3, 400 MHz) spectrum ( Figure 16 )show d H 13.50 (1H, s, 1-OH) represents a single phenolic hydroxyl proton signal, while the lower field region shows three methylene hydrogen proton signals linked to electron-withdrawing groups. d H 4.75 (1H, dd, J = 9.5,6.3 Hz, H-8), 4.72 (1H, dd, J = 9.5, 5.4 Hz, H-13), 3.69 (1H, p, J = 6.7 Hz, H-18), the higher field region shows two sets of methylene hydrogen proton signals. d H 3.08 (1H, dd, J = 15.2, 9.5 Hz, H-7a),2.98 (1H, dd, J = 15.2 Hz, 8.2 Hz, H-7b), 3.04 (2H, dd, J = 9.4, 3.5 Hz, H2-12), the high field region shows four single-peak methyl hydrogen proton signals. d H 1.38 (3H, s, 15-CH3), 1.32 (3H, s, 16-CH3), 1.26 (3H, s, 10-CH3), 1.21 (3H, s, 11-CH3), two bimodal methyl hydrogen proton signals. d H 1.19 (3H, d, J = 6.7 Hz, 19-CH3), 1.17 (3H, d, J= 6.7 Hz, 20-CH3). The above 1H NMR spectrum information suggests that this compound may be an α-acid.

[0141] 13 C10 NMR (CDCl3, 100 MHz) spectrum Figure 17 The HSQC spectrum showed a total of 20 carbon signals, including one carbonyl carbon signal. d C 208.7 (C-17), a set of benzene ring carbon signals d C 161.8 (C-3), 99.6 (C-4), 163.3 (C-5), 101.3 (C-6), 160.1 (C-1), 105.9 (C-2), four carbon signals linked to oxygen. d C 92.0 (C-13), 71.7 (C-14), 92.5 (C-8), 72.1 (C-9), 1 methylene carbon signal d C 38.6 (C-18), 2 methylene carbon signals d C 26.6 (C-12), 27.3 (C-7), 6 methyl carbon signals d C 26.0 (C-15), 26.5 (C-16), 19.6 (C-19), 18.9 (C-20), 23.9 (C-10), 24.7 (C-11). The above information is combined... 1 The 1H NMR spectrum indicates that, apart from the benzene ring and carbonyl group, the remaining two unsaturations of this compound are occupied by two rings. In addition, the structure also contains two 2-hydroxyisopropyl segments and one isobutyryl segment.

[0142] 1 H NMR, 13 The signal assignments for C NMR are shown in Table 6.

[0143] Table 6 NMR data of compound 6 d (ppm)

[0144] HMBC Map ( Figure 18 In ), H2-7 can be observed ( d H 3.09) and C-3 ( d C 161.8), C-2 ( d C105.9), C-1 ( d C 160.1), C-8 ( d C 92.0), C-9 ( d C The remote correlation signal of 71.7) and H2-12 ( d H 3.04) and C-4 ( d C 99.6), C-3 ( d C 161.8), C-5 ( d C 163.3), C-13 ( d C 92.5), C-14 ( d C The remote correlation signal of 72.1) indicates that two furan rings are connected to the C-4, C-5 and C-2, C-3 positions of the benzene ring, respectively, and that two 2-hydroxyisopropyl segments are connected at C-8 and C-13, respectively; in addition, H-18 ( d H 3.69) and C-17 ( d C 208.7), C-19 ( d C 19.6), C-20 ( d C 18.9), C-6 ( d C The long-range correlation signal (101.3) confirmed the presence of the isobutyryl fragment and its connection at the C-6 position. Based on this information, the planar structure of compound 6 was determined, classifying it as an α-acid. Comparison of calculated circular dichroism (ECD) data with measured ECD data confirmed the absolute configuration of the compound to be 8. S ,13 S .

[0145] In summary, the structure of the new compound 6 (compound (-)-Humulune O) was determined as follows:

[0146] The physical properties and detection data of compound 7 obtained in Example 3 are as follows: Pale yellow oily liquid; High-resolution mass spectrometry (HR-ESI-MS) m / z 365.1951 ([M + H]) + The calculated value is 365.1959, suggesting the molecular formula of the compound is C.20 H 28 O6 has an unsaturation degree of 7.

[0147] 1 1H NMR (CDCl3, 400 MHz) spectrum ( Figure 19 )show d H 13.50 (1H, s, 1-OH) represents a single phenolic hydroxyl proton signal, while the lower field region shows three methylene hydrogen proton signals linked to electron-withdrawing groups. d H 4.75 (1H, dd, J = 9.5,6.3 Hz, H-8), 4.72 (1H, dd, J = 9.5, 5.4 Hz, H-13), 3.69 (1H, p, J = 6.7 Hz, H-18), the higher field region shows two sets of methylene hydrogen proton signals. d H 3.08 (1H, dd, J = 15.2, 9.5 Hz, H-7a),2.98 (1H, dd, J = 15.2 Hz, 8.2 Hz, H-7b), 3.04 (2H, dd, J = 9.4, 3.5 Hz, H2-12), the high field region shows four single-peak methyl hydrogen proton signals. d H 1.38 (3H, s, 15-CH3), 1.32 (3H, s, 16-CH3), 1.26 (3H, s, 10-CH3), 1.21 (3H, s, 11-CH3), two bimodal methyl hydrogen proton signals. d H 1.19 (3H, d, J = 6.7 Hz, 19-CH3), 1.17 (3H, d, J = 6.7 Hz, 20-CH3). The above 1H NMR spectrum information suggests that this compound may be an α-acid.

[0148] 13 C10 NMR (CDCl3, 100 MHz) spectrum Figure 20 The HSQC spectrum showed a total of 20 carbon signals, including one carbonyl carbon signal. d C 208.7 (C-17), a set of benzene ring carbon signals dC 161.8 (C-3), 99.6 (C-4), 163.3 (C-5), 101.3 (C-6), 160.1 (C-1), 105.9 (C-2), four carbon signals linked to oxygen. d C 92.0 (C-13), 71.7 (C-14), 92.5 (C-8), 72.1 (C-9), 1 methylene carbon signal d C 38.6 (C-18), 2 methylene carbon signals d C 26.6 (C-12), 27.3 (C-7), 6 methyl carbon signals d C 26.0 (C-15), 26.5 (C-16), 19.6 (C-19), 18.9 (C-20), 23.9 (C-10), 24.7 (C-11). The above information is combined... 1 The 1H NMR spectrum indicates that, apart from the benzene ring and carbonyl group, the remaining two unsaturations of this compound are occupied by two rings. In addition, the structure also contains two 2-hydroxyisopropyl segments and one isobutyryl segment.

[0149] 1 H NMR, 13 The signal assignments for C NMR are shown in Table 7.

[0150] Table 7 NMR data of compound 7 d (ppm)

[0151] HMBC Map ( Figure 21 In ), H2-7 can be observed ( d H 3.09) and C-3 ( d C 161.8), C-2 ( d C 105.9), C-1 ( d C 160.1), C-8 ( d C 92.0), C-9 ( d C The remote correlation signal of 71.7) and H2-12 ( d H 3.04) and C-4 ( d C 99.6), C-3 ( d C 161.8), C-5 ( d C 163.3), C-13 ( d C 92.5), C-14 ( d C The remote correlation signal of 72.1) indicates that two furan rings are connected to the C-4, C-5 and C-2, C-3 positions of the benzene ring, respectively, and that two 2-hydroxyisopropyl segments are connected at C-8 and C-13, respectively; in addition, H-18 ( d H 3.69) and C-17 ( d C 208.7), C-19 ( d C 19.6), C-20 ( d C 18.9), C-6 ( d C The long-range correlation signal (101.3) confirmed the presence of the isobutyryl fragment and its connection at the C-6 position. Based on this information, the planar structure of compound 7 was determined, classifying it as an α-acid. Comparison of calculated circular dichroism (ECD) data with measured ECD data confirmed the absolute configuration of the compound to be 8. R ,13 S .

[0152] In summary, the structure of the new compound 7 (compound (+)-Garcinielliptone HE) was determined as follows:

[0153] The physical properties and detection data of compound 8 obtained in Example 3 are as follows: Pale yellow oily liquid; High-resolution mass spectrometry (HR-ESI-MS) m / z 365.1951 ([M + H]) + The calculated value is 365.1959, suggesting the molecular formula of the compound is C. 20 H 28 O6 has an unsaturation degree of 7.

[0154] 1 1H NMR (CDCl3, 400 MHz) spectrum ( Figure 22 )show d H 13.50 (1H, s, 1-OH) represents a single phenolic hydroxyl proton signal, while the lower field region shows three methylene hydrogen proton signals linked to electron-withdrawing groups. dH 4.75 (1H, dd, J = 9.5,6.3 Hz, H-8), 4.72 (1H, dd, J = 9.5, 5.4 Hz, H-13), 3.69 (1H, p, J = 6.7 Hz, H-18), the higher field region shows two sets of methylene hydrogen proton signals. d H 3.08 (1H, dd, J = 15.2, 9.5 Hz, H-7a),2.98 (1H, dd, J = 15.2 Hz, 8.2 Hz, H-7b), 3.04 (2H, dd, J = 9.4, 3.5 Hz, H2-12), the high field region shows four single-peak methyl hydrogen proton signals. d H 1.38 (3H, s, 15-CH3), 1.32 (3H, s, 16-CH3), 1.26 (3H, s, 10-CH3), 1.21 (3H, s, 11-CH3), two bimodal methyl hydrogen proton signals. d H 1.19 (3H, d, J = 6.7 Hz, 19-CH3), 1.17 (3H, d, J = 6.7 Hz, 20-CH3). The above 1H NMR spectrum information suggests that this compound may be an α-acid.

[0155] 13 C10 NMR (CDCl3, 100 MHz) spectrum Figure 23 The HSQC spectrum showed a total of 20 carbon signals, including one carbonyl carbon signal. d C 208.7 (C-17), a set of benzene ring carbon signals d C 161.8 (C-3), 99.6 (C-4), 163.3 (C-5), 101.3 (C-6), 160.1 (C-1), 105.9 (C-2), four carbon signals linked to oxygen. d C 92.0 (C-13), 71.7 (C-14), 92.5 (C-8), 72.1 (C-9), 1 methylene carbon signal d C38.6 (C-18), 2 methylene carbon signals d C 26.6 (C-12), 27.3 (C-7), 6 methyl carbon signals d C 26.0 (C-15), 26.5 (C-16), 19.6 (C-19), 18.9 (C-20), 23.9 (C-10), 24.7 (C-11). The above information is combined... 1 The 1H NMR spectrum indicates that, apart from the benzene ring and carbonyl group, the remaining two unsaturations of this compound are occupied by two rings. In addition, the structure also contains two 2-hydroxyisopropyl segments and one isobutyryl segment.

[0156] 1 H NMR, 13 The signal assignments for C NMR are shown in Table 8.

[0157] Table 8 NMR data of compound 8 d (ppm)

[0158] HMBC Map ( Figure 24 In ), H2-7 can be observed ( d H 3.09) and C-3 ( d C 161.8), C-2 ( d C 105.9), C-1 ( d C 160.1), C-8 ( d C 92.0), C-9 ( d C The remote correlation signal of 71.7) and H2-12 ( d H 3.04) and C-4 ( d C 99.6), C-3 ( d C 161.8), C-5 ( d C 163.3), C-13 ( d C 92.5), C-14 ( d CThe remote correlation signal of 72.1) indicates that two furan rings are connected to the C-4, C-5 and C-2, C-3 positions of the benzene ring, respectively, and that two 2-hydroxyisopropyl segments are connected at C-8 and C-13, respectively; in addition, H-18 ( d H 3.69) and C-17 ( d C 208.7), C-19 ( d C 19.6), C-20 ( d C 18.9), C-6 ( d C The long-range correlation signal (101.3) confirmed the presence of the isobutyryl fragment and its connection at the C-6 position. Based on this information, the planar structure of compound 8 was determined, classifying it as an α-acid. Comparison of calculated circular dichroism (ECD) data with measured ECD data confirmed the absolute configuration of compound 8. S ,13 R .

[0159] In summary, the structure of the new compound 8 (compound (-)-Garcinielliptone HE) was determined as follows:

[0160] Comparative Example 1 Comparative Example 1 of this invention provides a method for preparing a novel tricyclic α-acid compound. The steps are similar to those in Example 1, except that in step S5, when fraction C13 is subjected to silica gel column chromatography, petroleum ether-ethyl acetate with a volume ratio of petroleum ether to ethyl acetate of less than 18:82 or greater than 23:77 is used for elution. The results show that compounds 1 and 2 are not obtained.

[0161] Comparative Example 2 Comparative Example 2 of this invention provides a method for preparing a novel tricyclic α-acid compound. The steps are similar to those in Example 2, except that in step S6, when fraction C13.17 is subjected to silica gel column chromatography, elution is performed using a petroleum ether-dichloromethane-methanol mixture with a volume ratio of petroleum ether, dichloromethane, and methanol of less than 7:3:1 or greater than 3:7:1. The results show that compounds 3 and 4 are not obtained.

[0162] Comparative Example 3 Comparative Example 3 of this invention provides a method for preparing novel tricyclic α-acid compounds. The steps are similar to those in Example 3, except that in step S8, the volume ratio of methanol to water in the mobile phase used for separating fraction C13.17.7.5 by HPLC is either lower than 62:38 or higher than 68:32. The results show that compounds 5, 6, 7, and 8 could not be prepared.

[0163] To better understand the essence of this invention, the following describes the new applications of the above-mentioned α-acid compounds in the pharmaceutical field in conjunction with pharmacological tests and results.

[0164] Test case This experimental example discloses the antidepressant activity of the above compound (±)-Humulunes O in corticosterone-induced HT22 cells (a commonly used cell model for antidepressants).

[0165] (1) Experimental materials and instruments Mouse hippocampal neuron cells HT22 were purchased from the Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences. DMEM medium, PBS buffer, fetal bovine serum (FBS) and trypsin were all purchased from Gibco, USA. Fluoxetine (Shanghai Yuanye Biotechnology Co., Ltd.); CCK-8 Cell Viability Assay Kit (Elabscience); 96-well plate (Corning Corporation, USA); Carbon dioxide cell incubator (Thermo Fisher Scientific, USA). Full-wavelength microplate reader (Thermo Fisher Scientific, USA); BIOFUGE STRATOS centrifuge (Thermo Fisher Scientific, USA); IX73 inverted electron microscope (Olympus Corporation, Japan); Pipettes (Eppendorf, Germany); Electronic balance (Sartorius GmbH, Germany); Clean benches, centrifuge tubes, straws and other related consumables.

[0166] (2) Establishment and evaluation of a corticosterone-damaged HT22 cell model Remove the labeled HT22 cell cryopreservation tubes from liquid nitrogen and immediately place them in a 37°C water bath to thaw them rapidly within 1 minute. After disinfecting the cryopreservation tubes with alcohol, transfer the cryopreservation solution into a 15 mL sterile centrifuge tube, add the appropriate culture medium, mix well, and centrifuge to remove the supernatant. Repeat the above steps once for washing, then add 10 mL of culture medium, mix well, resuspend the cells, transfer them to a 10 mL culture dish, and incubate in a 37°C, 5% CO2 incubator.

[0167] When the cell density reaches 70%-80%, passage the cells. First, remove the old culture medium, wash twice with PBS, add trypsin containing EDTA to the culture dish and incubate at 37°C for 3 min. Then, add culture medium to stop digestion, centrifuge at 800 rpm for 5 min, remove the digestion solution, add serum-containing culture medium again and repeatedly pipette the cells to form a cell suspension. Finally, according to the number of cells, transfer them to a new culture dish to complete the cell passage.

[0168] HT22 cells were seeded at 1×10⁵ cells / well in 96-well plates with 6 replicates per group. 80 μL / well was added and the cells were cultured for 24 h. Fluoxetine, a positive control drug, was dissolved and diluted to 10 μM. (±)-Humulunes O was dissolved and diluted to 10 μM and added to 10 μL / well to treat the cells for 1 h. Then, 10 μL of 25 μM corticosterone was added and the cells were cultured at 37°C for 48 h. Finally, 10 μL of a CCK-8 assay kit was added, and the absorbance at 450 nm was measured using a microplate reader after 1 h. Cell viability was calculated.

[0169] Cell viability (%) = [(Control A - Sample A) / (Control A - Blank A)] × 100%.

[0170] (3) Experimental results The experimental results (Table 9) show that both compounds (+)-Humulune O and (-)-Humulune O have antidepressant effects; in particular, compared with the model group, compound (+)-Humulune O has a significant protective effect against corticosterone-induced HT22 neuronal damage, and its efficacy is comparable to that of the positive control drug fluoxetine (a first-line chemical drug for the clinical treatment of depression, 10 μM).

[0171] Table 9. Neuroprotective effects of compounds on corticosterone-induced HT22 cell damage ( ±s, n=5)

[0172] Note: Compared with the blank group: ###P <0.001; Compared with the model group: * P <0.05, *** P <0.001 Experimental Example 2 This experimental example discloses the antidepressant activity of compounds (±)-Humulunes MO and (±)-Garcinielliptone HE in LPS-induced SH-SY5Y cells (a commonly used cell model for antidepressants).

[0173] (1) Experimental materials and instruments Human neuroblastoma SH-SY5Y was purchased from the Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences. DMEM medium, PBS buffer, fetal bovine serum (FBS), and trypsin were all purchased from Gibco, USA. Fluoxetine (Shanghai Yuanye Biotechnology Co., Ltd.); CCK-8 Cell Viability Assay Kit (Elabscience); 96-well plate (Corning Corporation, USA); Carbon dioxide cell incubator (Thermo Fisher Scientific, USA). Full-wavelength microplate reader (Thermo Fisher Scientific, USA); BIOFUGE STRATOS centrifuge (Thermo Fisher Scientific, USA); IX73 inverted electron microscope (Olympus Corporation, Japan); Pipettes (Eppendorf, Germany); Electronic balance (Sartorius GmbH, Germany); Clean benches, centrifuge tubes, straws and other related consumables.

[0174] (2) Establishment and evaluation of LPS-damaged SH-SY5Y cell model Remove the labeled SH-SY5Y cell cryovials from liquid nitrogen and immediately place them in a 37°C water bath to thaw them rapidly within 1 minute. After disinfecting the cryovials with alcohol, transfer the cryopreservation solution into a 15 mL sterile centrifuge tube, add the appropriate culture medium, mix well, and centrifuge to remove the supernatant. Repeat the above steps once for washing, then add 10 mL of culture medium, mix well, resuspend the cells, transfer them to a 10 mL culture dish, and incubate in a 37°C, 5% CO2 incubator.

[0175] When the cell density reaches 70%-80%, passage the cells. First, remove the old culture medium, wash twice with PBS, add trypsin containing EDTA to the culture dish and incubate at 37°C for 3 min. Then, add culture medium to stop digestion, centrifuge at 800 rpm for 5 min, remove the digestion solution, add serum-containing culture medium again and repeatedly pipette the cells to form a cell suspension. Finally, according to the number of cells, transfer them to a new culture dish to complete the cell passage.

[0176] SH-SY5Y cells were seeded at 1×10⁵ cells / well in 96-well plates with 6 replicates per group. 80 μL / well was added and the cells were cultured for 24 h. Fluoxetine, a positive control drug, was dissolved and diluted to 10 μM. Compounds (±)-Humulunes MO and (±)-Garcinielliptone HE were dissolved and diluted to 10 μM, and 10 μL / well was added to treat the cells for 1 h. Then, 10 μL of 25 μM LPS was added, and the cells were cultured at 37°C for 48 h. Finally, 10 μL of a CCK-8 assay kit was added, and the absorbance at 450 nm was measured using a microplate reader after 1 h. Cell viability was calculated.

[0177] Cell viability (%) = [(Control A - Sample A) / (Control A - Blank A)] × 100%.

[0178] (3) Experimental results The experimental results (Table 10) showed that, compared with the model group, compounds (±)-Humulunes MO and (±)-Garcinielliptone HE had significant protective effects against LPS-induced SH-SY5Y neuronal damage, demonstrating better antidepressant efficacy and significantly superior to the positive control drug fluoxetine (a first-line chemical drug for the clinical treatment of depression).

[0179] Table 10 Neuroprotective effects of compounds on LPS-induced SH-SY5Y cell damage ( ±s, n=5)

[0180] Note: Compared with the blank group: #### P <0.0001: Compared with the model group *** P <0.001, **** P <0.0001 In summary, the novel α-acid compounds (±)-Humulune MO and (±)-Garcinielliptone HE described in this invention have good antidepressant effects; in particular, (+)-Humulune O exhibits significant antidepressant activity, comparable to the efficacy of fluoxetine, a first-line antidepressant, and can be used as a drug for the treatment of depression.

[0181] Application Example 1 This invention discloses a capsule formulation using (+)-Humulune O as the active pharmaceutical ingredient, the components of which are as follows: (+)-Humulune O18.0 mg Starch 6.0 g Sodium metabisulfite 0.2 g Magnesium stearate 0.2 g Anhydrous ethanol appropriate amount Make 100 pills.

[0182] The specific preparation process is as follows: Take (+)-Humulune O, starch, and sodium metabisulfite and mix them evenly. Add anhydrous ethanol to make a soft material, pass it through a 24-mesh sieve, make it into granules, dry it, add magnesium stearate, mix it evenly, and fill it into capsules.

[0183] Application Example 2 This invention discloses a granule formulation using compound (-)-Humulune O as the active pharmaceutical ingredient, the components of which are as follows: (-)-Humulune O35.0 mg Starch 6.0 g Sodium bisulfite 0.2 g Magnesium stearate 0.2 g Anhydrous ethanol appropriate amount Make 100 bags.

[0184] The specific preparation process is as follows: Take (-)-Humulune O, mix it with starch and sodium bisulfite, add anhydrous ethanol to make a soft material, pass it through a 24-mesh sieve to make granules, dry it, add magnesium stearate, mix it well, and pack it into bags.

[0185] Application Example 3 This invention discloses an oral liquid using compound (+)-Humulune N as a raw material, the components of which are as follows: (+)-Humulune N 25.0 mg 3.0 g sucrose Sodium bisulfite 0.2 g Methylparaben 0.2 g Sodium bicarbonate 0.1 g 1000 mL of water for injection Make 100 pieces.

[0186] The specific preparation process is as follows: After the above components are mixed, they can be dispensed using conventional oral liquid preparation methods.

[0187] Application Example 4 This invention discloses an injectable formulation using compound (-)-Humulune N as the active pharmaceutical ingredient, the components of which are as follows: (-)-Humulune N 45.0 mg Vitamin C 0.2 g Sodium chloride 6.0 g Sodium bicarbonate 0.1 g 1000 mL of water for injection Make 100 pieces.

[0188] The specific preparation process is as follows: After the above components are mixed, 100 vials can be obtained by using conventional injection preparation methods.

[0189] Application Example 5 This invention discloses a tablet using compound (+)-Humulune M and fluoxetine as active pharmaceutical ingredients, the composition of which is as follows: (+)-Humulune M20.0 mg Fluoxetine 0.5 g Hydroxypropyl methylcellulose 18 g 0.4 g talc Lactose 0.2 g Magnesium stearate 0.2 g Anhydrous ethanol appropriate amount Make 100 pieces.

[0190] The specific preparation process is as follows: Mix (+)-Humulune M, fluoxetine, hydroxypropyl methylcellulose, talc, lactose, and magnesium stearate evenly, add anhydrous ethanol to make a soft mass, pass it through a 24-mesh sieve to make granules, dry it, add magnesium stearate, mix well, and compress into tablets.

[0191] Application Example 6 This invention discloses a capsule formulation using compound (-)-Humulune M and sertraline as active pharmaceutical ingredients, the components of which are as follows: (-)-Humulune M18.0 mg Sertraline 2.0 g Starch 2.0 g Sodium metabisulfite 0.2 g Magnesium stearate 0.2 g Anhydrous ethanol appropriate amount Make 100 pills.

[0192] The specific preparation process is as follows: Take (-)-Humulune M, sertraline, starch, and sodium metabisulfite, mix them, add anhydrous ethanol to make a soft material, pass it through a 24-mesh sieve to make granules, dry it, add magnesium stearate, mix it, and fill it into capsules.

[0193] Application Example 7 This invention discloses an injection using compounds (-)-Garcinielliptone HE and escitalopram as active pharmaceutical ingredients, the components of which are as follows: (-)-Garcinielliptone HE30.0 mg Escitalopram 2.0 g Vitamin C 0.2 g Sodium chloride 6.0 g Sodium bicarbonate 0.1 g 1000 mL of water for injection Make 100 pieces.

[0194] The specific preparation process is as follows: After the above components are mixed, 100 vials can be obtained by using conventional injection preparation methods.

[0195] The above embodiments are merely examples of several implementation methods of the present invention, and their descriptions are relatively specific and detailed, but they should not be construed as limiting the scope of the invention patent.

[0196] It should be noted that those skilled in the art can make various modifications and improvements without departing from the inventive concept, and these all fall within the scope of protection of this invention. Therefore, the scope of protection of this patent should be determined by the appended claims.

[0197] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A compound or a pharmaceutically acceptable salt thereof, characterized in that, It has a structure shown in any of the following equations (1) to (7): 。 2. A method for preparing the compound as described in claim 1, characterized in that, The tricyclic α-acid compound was extracted and isolated from hops with flowers and fruit spikes; Includes the following steps: S1. Take dried hop spikes with flowers, add solvent to extract, combine the extracts, filter, concentrate under reduced pressure to obtain extract; S2. After suspending the extract in water, extract it with dichloromethane to obtain the extract; S3. The dichloromethane extract was subjected to gradient elution using a petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction AE. S4. Fraction C was subjected to gradient elution using petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction C13; when identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13 was 0.44-0.

45. S5. Fraction C13 was subjected to gradient elution using petroleum ether-ethyl acetate solution via silica gel column chromatography to obtain fraction C13.

17. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17 was 0.25-0.

27. S6. Fraction C13.17 was isocratically eluted using a petroleum ether-dichloromethane-methanol solution via Sephadex LH-20 gel column chromatography to obtain fraction C13.17.

7. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17.7 was 0.57-0.

60. S7. Fraction C13.17.7 was subjected to gradient elution using methanol-water solution via ODS column chromatography to obtain fractions C13.17.7.5, C13.17.7.7, and C13.17.7.

8. When identified by silica gel thin-layer chromatography, the Rf value corresponding to fraction C13.17.7.5 was 0.45-0.52, the Rf value corresponding to fraction C13.17.7.7 was 0.60-0.64, and the Rf value corresponding to fraction C13.17.7.8 was 0.66-0.

71. S8. Using methanol-water as the mobile phase, the tricyclic α-acid compounds shown in formulas (5), (6), and (7) were prepared by HPLC using a C18 column in fraction C13.17.7.

5. S9. Using acetonitrile-water as the mobile phase, HPLC can be used to prepare tricyclic α-acid compounds represented by formula (3) and / or formula (4) in fraction C13.17.7.7 using a C18 column; S10. Using acetonitrile-water as the mobile phase, tricyclic α-acid compounds of formula (1) and / or formula (2) can be prepared in fraction C13.17.7.8 by HPLC using a C18 column.

3. The preparation method according to claim 2, characterized in that, In step S8, the volume ratio of methanol to water in the methanol-water solution is (62:38) - (68:32).

4. The preparation method according to claim 2, characterized in that, In step S8, the fraction C13.17.7.5 is further separated based on the characteristic ultraviolet absorption of tricyclic α-acids observed by HPLC analysis. The retention times of tricyclic α-acids with the structural formula (5) are 35-37 min; the retention times of tricyclic α-acids with the structural formula (6) are 39-42 min; the retention times of tricyclic α-acids with the structural formula (7) are 22-24 min; and the retention times of tricyclic α-acids with the structural formula (8) are 26-29 min.

5. The preparation method according to claim 2, characterized in that, In step S9, the volume ratio of acetonitrile to water in the acetonitrile-water solution is (45:55) to (50:50).

6. The preparation method according to claim 2, characterized in that, In step S9, the fraction C13.17.7.7 is further separated based on the characteristic ultraviolet absorption of tricyclic α-acids observed by HPLC analysis, including the tricyclic α-acid compounds with the structural formula as shown in formula (3) with a retention time of 21-24 min; and the tricyclic α-acid compounds with the structural formula as shown in formula (4) with a retention time of 26-28 min.

7. The preparation method according to claim 2, characterized in that, In step S10, the volume ratio of acetonitrile to water in the acetonitrile-water solution is (42:58)-(47:53).

8. The preparation method according to claim 2, characterized in that, In step S10, fraction C13.17.7.8 is further separated based on the characteristic ultraviolet absorption of tricyclic α-acids observed by HPLC analysis, including tricyclic α-acids with a retention time of 30-34 min as shown in formula (1) and tricyclic α-acids with a retention time of 36-38 min as shown in formula (2).

9. The preparation method according to claim 2, characterized in that, In step S2, the extract is suspended in 8-15 times its weight of water and then extracted with dichloromethane.

10. The preparation method according to claim 2, characterized in that, In step S3, based on a total volume of 100, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when washing out fraction C is (58:42)-(62:38).

11. The preparation method according to claim 2, characterized in that, In step S3, based on a total volume of 100, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when washing out fraction C is 60:

40.

12. The preparation method according to claim 2, characterized in that, In step S4, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when washing out fraction C13 is (37:63)-(43:57).

13. The preparation method according to claim 2, characterized in that, In step S4, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when washing out fraction C13 is 40:

60.

14. The preparation method according to claim 2, characterized in that, In step S5, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate fraction C13.17 is (18:82)-(23:77).

15. The preparation method according to claim 2, characterized in that, In step S5, the volume ratio of petroleum ether to ethyl acetate in the petroleum ether-ethyl acetate solution when washing out fraction C13.17 is 20:

80.

16. The preparation method according to claim 2, characterized in that, In step S6, the volume ratio of petroleum ether, dichloromethane and methanol in the petroleum ether-dichloromethane-methanol fraction C13.17.7 is (7:3:1)-(3:7:1).

17. The preparation method according to claim 2, characterized in that, In step S6, the volume ratio of petroleum ether, dichloromethane, and methanol in the petroleum ether-dichloromethane-methanol solution when washing out fraction C13.17.7 is 5:5:

1.

18. The preparation method according to claim 2, characterized in that, In step S7, the volume ratio of methanol to water in the methanol-water solution of the washed fraction C13.17.7.5 is (47:53)-(53:47); the volume ratio of methanol to water in the methanol-water solution of the washed fraction C13.17.7.7 is (56:44)-(62:38); and the volume ratio of methanol to water in the methanol-water solution of the washed fraction C13.17.7.8 is (66:34)-(72:28).

19. The preparation method according to claim 2, characterized in that, In step S7, the volume ratio of methanol to water in the methanol-water solution of the washed fraction C13.17.7.5 is 50:50; the volume ratio of methanol to water in the methanol-water solution of the washed fraction C13.17.7.7 is 60:40; and the volume ratio of methanol to water in the methanol-water solution of the washed fraction C13.17.7.8 is 70:

30.

20. The preparation method according to claim 2, characterized in that, In step S1, the solvent is an 88-98% V% aqueous ethanol solution; the solvent added is 8-10 times the mass of hops, and the reflux extraction is performed 1-3 times, with each extraction lasting 1-3 hours.

21. A pharmaceutical composition, characterized in that, It includes the compound of claim 1.

22. The pharmaceutical composition according to claim 21, characterized in that, The dosage form of the pharmaceutical composition is any one of tablets, capsules, granules, oral liquids, syrups, ointments, powders, drops, or microcapsules.

23. Use of the compound of claim 1 or the pharmaceutical composition of any one of claims 21-22 in the preparation of a medicament for treating depression.

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