Preparation method and application of (s,e)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenyl acetate in zedoary turmeric

Abstract

This invention relates to the field of pharmaceutical technology, and particularly to a method for preparing (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate from Curcuma zedoaria and its application. The preparation method includes the following steps: (1) crushing Curcuma zedoaria rhizomes, extracting with ethyl acetate, concentrating the extract to obtain a residual extract, separating the extract by silica gel column chromatography with a 100-200 mesh, and using a gradient of a petroleum ether-ethyl acetate mixed solution with a volume ratio of 1:0-0:1. (1) Elution yielded six fractions Fr.I to VI; (2) Fraction Fr.IV was repeatedly separated by HW-40F gel column chromatography, followed by silica gel column chromatography with a 100-200 mesh, and then eluted with a gradient of petroleum ether-ethyl acetate mixed solutions at volume ratios of 8:2, 2:1, and 1:1 to obtain three fractions Fr.IV2b1 to Fr.IV2b3; (3) Fraction Fr.IV2b1 was separated by preparative high-performance liquid chromatography and then purified by semi-preparative high-performance liquid chromatography to obtain the target compound. This compound has α-glucosidase inhibitory activity and can be used in the preparation of pharmaceutical products with hypoglycemic function.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, and in particular to a method for preparing (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate from Curcuma zedoaria and its application. Background Technology

[0002] Curcuma zedoaria (chiristm.) Rosc. is a plant belonging to the ginger family and the genus Curcuma. It is also known as white turmeric, peng'e mao, pengyao, warm turmeric, and guangmao. This perennial monocotyledonous herb has its rhizome used medicinally. Traditional Chinese medicine believes that Curcuma zedoaria is warm in nature, pungent and bitter in taste, and has the effects of promoting qi circulation, breaking up blood stasis, relieving muscle tension, and alleviating pain. It can be used to treat muscle pain caused by pathogenic factors, regulate food stagnation, amenorrhea, dysmenorrhea, and treat injuries from falls and blows. Modern pharmacological research shows that Curcuma zedoaria has anti-tumor properties and other pharmacological effects, such as antiplatelet aggregation, antibacterial, antiviral, hypoglycemic, analgesic, and anti-inflammatory effects.

[0003] In order to find new compounds with significant hypoglycemic activity from this plant, the applicant conducted phytochemical studies on Curcuma zedoaria, which is native to Lingshan County, Guangxi Zhuang Autonomous Region. As a result, a new diphenylheptane compound (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate was isolated from Curcuma zedoaria. This compound showed potential inhibitory activity against α-glucosidase. Currently, no studies have been reported on the isolation and extraction of this diphenylheptane compound from Curcuma zedoaria. Summary of the Invention

[0004] The purpose of this invention is to provide a method for preparing (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate from Curcuma zedoaria, and its application, in order to address the problems mentioned above.

[0005] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0006] The (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate in Curcuma zedoaria of this invention has the following structural formula:

[0007]

[0008] This invention also provides a method for preparing (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate from Curcuma zedoaria, comprising the following steps:

[0009] (1) The rhizome of Curcuma zedoaria was crushed and extracted with ethyl acetate. The extract was concentrated to obtain a residual extract. The extract was separated by silica gel column chromatography with a 100-200 mesh and eluted with a gradient of petroleum ether-ethyl acetate mixed solution with a volume ratio of 1:0-0:1 to obtain six components Fr.I~VI.

[0010] (2) The fraction Fr.IV was separated three times by HW-40F gel column chromatography, followed by silica gel column chromatography with 100-200 mesh, and then eluted with a gradient of petroleum ether-ethyl acetate mixed solution with volume ratios of 8:2, 2:1 and 1:1 to obtain three fractions Fr.IV2b1~Fr.IV2b3;

[0011] (3) The component Fr.IV2b1 was separated by preparative high performance liquid chromatography and then purified by semi-preparative high performance liquid chromatography to obtain the (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate.

[0012] In this invention, preferably, in step (1), the extraction is carried out at room temperature, and the extraction is performed 3 times, each time for 3 days.

[0013] In this invention, preferably, in step (1), the method for concentrating the extract is vacuum concentration.

[0014] In this invention, preferably, in step (1), the gradient elution is performed by sequentially eluting with a petroleum ether-ethyl acetate mixed solution with volume ratios of 1:0, 9:1, 8:2, 2:1, 1:1, and 0:1.

[0015] In this invention, preferably, in step (3), the preparative high performance liquid chromatography separation conditions are as follows: separation is performed using an eluent of CH3OH:H2O at a volume ratio of 55:45.

[0016] In this invention, preferably, in step (3), the semi-preparative high performance liquid chromatography purification conditions are as follows: separation and purification using an eluent of CH3OH:H2O at a volume ratio of 55:45.

[0017] The (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate extracted from Curcuma zedoaria in this invention has α-glucosidase inhibitory activity and can be used as an α-glucosidase inhibitor in the preparation of pharmaceutical products with hypoglycemic function.

[0018] In summary, due to the adoption of the above technical solution, the present invention has the following beneficial effects:

[0019] 1. This invention is the first to isolate a novel diphenylheptane compound (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenyl acetate from Curcuma zedoaria, which is of positive significance for the further in-depth research and utilization of Curcuma zedoaria.

[0020] 2. The (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate isolated from Curcuma zedoaria in this invention exhibits potential inhibitory activity against α-glucosidase, IC50. 50 The value was 71.14±5.82 μmol / L, which can be used as an α-glucosidase inhibitor in the preparation of pharmaceutical products with hypoglycemic function.

[0021] 3. The separation method of the present invention is reasonable, simple to operate, safe, suitable for industrial production, and has industrialization significance. Attached Figure Description

[0022] Figure 1 This is the structural diagram of compound 1.

[0023] Figure 2 It is key to compound 1 1 H– 1 H COSY(-) and HMBC(→) spectra.

[0024] Figure 3 It is the C-5 adjacent proton in the (S)-MTPA ester and (R)-MTPA ester of compound 1. 1 H-NMR chemical shift difference.

[0025] Figure 4 This is a graph showing the relationship between compound 1 and the concentration of α-glucosidase.

[0026] Figure 5 This is a graph showing the relationship between acarbose and α-glucosidase concentration.

[0027] Figure 6 This is a Lineweaver-Burk plot showing the concentration change of compound 1.

[0028] Figure 7 This is a Lineweaver-Burk plot showing the changes in acarbose concentration. Detailed Implementation

[0029] To more clearly illustrate the present invention, the following specific embodiments will be used to further explain the invention.

[0030] The present invention isolates (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate from Curcuma zedoaria, with the structural formula shown below. Figure 1 As shown.

[0031] The compound is a yellow oil, [α]24.9D-6.80 (c 0.1, MeOH); UV(MeOH)λmax(logε):203(4.46),224(4.07),264(3.96)nm; IR(KBr)ν max 3425,1729,1606,1514,1450,1375,1266,1031cm -1 Combined with ESI-MS: m / z 371 [M+H] + The molecular formula C can be obtained from NMR data. 22 H 26 O5, with an unsaturation degree of 10; the ultraviolet spectrum shows three maximum absorption peaks at wavelengths of 203 nm, 224 nm, and 264 nm; the infrared spectrum shows that this compound contains hydroxyl groups (3425 cm⁻¹). -1 ), carbonyl (1729cm) -1 ) and benzene ring (1514cm) -1 ).

[0032] The method for extracting the above-mentioned (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate from Curcuma zedoaria includes the following steps:

[0033] (1) The rhizome of Curcuma zedoaria was crushed and extracted with ethyl acetate. The extract was concentrated to obtain a residual extract. The extract was separated by silica gel column chromatography with a 100-200 mesh and eluted with a gradient of petroleum ether-ethyl acetate mixed solution to obtain six components Fr.I~VI.

[0034] Among them, the rhizome of Curcuma zedoaria (50 kg) was collected in October 2021 from Lingshan County, Guangxi Zhuang Autonomous Region. It was identified by Associate Professor Huang Deqing of the School of Pharmacy, Guilin Medical University as the rhizome of Curcuma zedoaria (chiristm.) Rosc., a plant of the ginger family. It was extracted at room temperature, three times, for three days each time. The extract was concentrated by vacuum concentration. The elution conditions for silica gel column chromatography were to use a petroleum ether-ethyl acetate mixed solution with a gradient elution of 1:0, 9:1, 8:2, 2:1, 1:1 and 0:1 by volume, so as to obtain six components Fr.I~VI.

[0035] (2) The fraction Fr.IV was repeatedly separated by HW-40F gel column chromatography, followed by silica gel column chromatography with 100-200 mesh, and then eluted with a gradient of petroleum ether-ethyl acetate mixed solution with volume ratios of 8:2, 2:1, and 1:1 to obtain three fractions Fr.IV2b1 to Fr.IV2b3.

[0036] (3) The component Fr.IV2b1 was separated by preparative high performance liquid chromatography and then purified by semi-preparative high performance liquid chromatography to obtain the (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate.

[0037] The separation conditions for preparative high performance liquid chromatography (HPLC) are as follows: using CH3OH:H2O eluent at a volume ratio of 55:45. The purification conditions for semi-preparative HPLC are as follows: using CH3OH:H2O eluent at a volume ratio of 55:45.

[0038] The instruments and reagents used in the embodiments of this invention are as follows: optical rotation was measured by a digital JASCO P-1020; IR measurements were performed on a PerkinElmer Spectrum Two FT-IR spectrometer with KBr compression; UV measurements were performed on a Shimadzu UV2401PC UV spectrometer; ESIMS spectra were determined by liquid chromatography-mass spectrometry (Shimadzu LCMS-8030, Japan); 1D and 2D NMR measurements were performed on a Bruker DRX-400MHz NMR spectrometer; an Agilent preparative HPLC system (Agilent LC1260 Infinity, Agilent Technologies, USA); a microplate reader (Thermo Scientific, USA); silica gel (100-200 mesh, Qingdao Ocean Chemical Plant, China); pre-coated silica gel GF254 for thin-layer chromatography (Qingdao Ocean Chemical Plant, China); a reversed-phase column (Zorbax SB-C18, 5μm, 9.4×250mm, Agilent Technologies, USA); and a Sephadex gel chromatography column. LH-20 (40–70 μm, Amersham Pharmacia Biotech AB, Uppsala, Sweden); petroleum ether, ethyl acetate, methanol (AR, Guangdong Xilong Technology Co., Ltd.); (R)-(-)-MTPA-Cl, (S)-MTPA-Cl (SIGMA, Germany); PBS powder (Beijing Regen Biotechnology Co., Ltd.); α-glucosidase (G0660-750UN, SIGMA, Germany); acarbose (109A032, Beijing Solarbio Technology Co., Ltd.); 4-pNPG (4-p-nitrobenzene-α-D-glucopyranoside) (N0493, Tokyo Chemical Industry Co., Ltd., Tokyo, Japan).

[0039] I. Preparation Examples

[0040] Example 1

[0041] (1) The rhizome of Curcuma zedoaria was crushed and extracted with ethyl acetate at room temperature. The extraction was repeated 3 times for 3 days each time. The extract was concentrated under vacuum to obtain the residual extract. The extract was separated by silica gel column chromatography with a 100-200 mesh. The extract was eluted sequentially with a petroleum ether-ethyl acetate mixture with volume ratios of 1:0, 9:1, 8:2, 2:1, 1:1, and 0:1 to obtain six components Fr.I to VI.

[0042] (2) The fraction Fr.IV was repeatedly separated by HW-40F gel column chromatography, followed by silica gel column chromatography with 100-200 mesh, and then eluted with a gradient of petroleum ether-ethyl acetate mixed solution with volume ratios of 8:2, 2:1, and 1:1 to obtain three fractions Fr.IV2b1 to Fr.IV2b3.

[0043] (3) The component Fr.IV2b1 was separated by preparative high performance liquid chromatography under the following conditions: CH3OH:H2O was used as the eluent at a volume ratio of 55:45; then it was purified by semi-preparative high performance liquid chromatography under the following conditions: CH3OH:H2O was used as the eluent at a volume ratio of 55:45 to obtain compound 1(S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate.

[0044] Compound 1 is a yellow oily substance, [α] 24.9D -6.80 (c 0.1, MeOH); UV(MeOH)λmax(logε): 203 (4.46), 224 (4.07), 264 (3.96) nm; IR(KBr)ν max 3425,1729,1606,1514,1450,1375,1266,1031cm -1 Combined with ESI-MS: m / z 371 [M+H] + The molecular formula C can be obtained from NMR data. 22 H 26 O5, with an unsaturation degree of 10; the ultraviolet spectrum shows three maximum absorption peaks at wavelengths of 203 nm, 224 nm, and 264 nm; the infrared spectrum shows that this compound contains hydroxyl groups (3425 cm⁻¹). -1 ), carbonyl (1729cm) -1 ) and benzene ring (1514cm) -1 ).

[0045] 1H of compound 1 and 13 The C NMR data are shown in Table 1 below.

[0046] Table 1 Compound 1 1 H and 13 C NMR data (δinppm, JinHz).

[0047]

[0048]

[0049] a Data was recorded at 400MHz and DMSO-d6

[0050] b Data was recorded at 100MHz and DMSO.

[0051] from 1 The H-NMR data show that compound 1 has a 1,3,4-trisubstituted aromatic proton signal [δ]. H [6.73 (d, J = 2.0 Hz, 1H), 6.66 (d, J = 8.0 Hz, 1H), 6.56 (dd, J = 8.0, 2.0 Hz, 1H)] and a 1,4-disubstituted aromatic proton signal [δ] H 7.16 (d, J = 8.6 Hz, 2H), 6.70 (d, J = 8.6 Hz, 2H)], and there are also 2 methyl proton signals [δ H 3.72(s,3H), 2.01(s,3H)], 4 methylene proton signals [δ H 2.45 (q, J = 6.9 Hz, 2H), 2.12 (p, J = 6.8 Hz, 2H), 1.79 (q, J = 7.3 Hz, 2H), 1.66 (q, J = 7.5 Hz, 2H)], 1 methylene proton signal [δ H 4.83 (t, J = 6.2 Hz, 1H)], 2 olefin hydrogen signals [δ H 6.25 (d, J = 15.9 Hz, 1H), 5.99 (dt, J = 15.9, 6.8 Hz, 1H)]. 13C-NMR data indicate that the compound has a single ester group carbon signal (δ). C 170.3), 12 aromatic carbon signals (δ C 112.5, 115.4, 120.3, 127.0, 128.3, 132.2, 144.6, 147.4, 156.7), 1 oxygen-carbon signal (δ C 72.6), 4 methylene carbons (δ C 28.4, 30.6, 33.5, 35.6) and two methyl carbon signals (δ C21.0, 55.5). By comparing the NMR data of compound 1 with those of diphenylheptane compounds reported in the literature, it can be seen that compound 1 belongs to the diphenylheptane class of compounds.

[0052] 1 H- 1 H COSY spectrum ( Figure 2 ) Display H-1(δ H 6.25) / H-2(δ H 5.99) / H-3(δ H 2.12) / H-4(δ H 4.66) / H-5(δ H 4.83) / H-6(δ H 1.79) / H-7(δ H 2.45) Related signals; from the HMBC spectrum ( Figure 2 H-7 (δ) can be seen in ) H 2.45) and C-1″(δ C 132.2), C-2″(δ C 112.5), C-6″(δ C 120.3), C-6(δ) C 35.6), C-5(δ) C 72.6) There is a relevant signal, δ H 3.72(s,3H) and C-3″(δ) C 115.3) has a relevant signal, H-2″(δ) H 6.73) and C-7(δ) C 30.6), C-4″(δ C 133.5), C-6″(δ C 120.3) has a relevant signal, H-5″ (δ H 6.66) and C-1″(δ C 132.2), C-3″(δ C 147.4) has a relevant signal, H-1(δ) H 6.25) and C-3(δ) C 28.4), C-2′(δ C 127.0), C-6′(δ C 127.0) has a relevant signal, H-2(δ) H 5.99) and C-1′(δ C 128.3) has a relevant signal, H-6′(δ H 7.16) and C-4′(δ C 156.7), C-1(δ) C129.7) has a relevant signal, and based on the above, the planar structure of compound 1 can be inferred. Δ 1,2 The large coupling constant J = 15.9 between the H-1 and H-2 of the double bond determines that the configuration is E.

[0053] The absolute configuration of C-5 was determined by a modified Mosher method, in which the hydroxyl group on C-5 reacted with Mosher reagents (S)-MTPA-Cl and (R)-(-)-MTPA-Cl, respectively, under pyridine conditions and in a dry environment to generate (S)-MTPA (1-a) and (R)-MTPA esters (1-b). [The following text appears to be incomplete and requires further context: "Collect its..."] 1 The 1H NMR data are as follows:

[0054] (S)-MTPA ester (1-a): Yellow oily substance; 1 H NMR(600MHz,Pyridine-d5)δ6.427026(d,J=15.1Hz,1H,H-1),6.268784(dt,J=15.1,6.8Hz,1H,H-2),3.76958 3(m,1H,H-5),2.716774(m,2H,H2-7),2.239983(m,2H,H2-3),2.0563085(m,2H,H2-6),1.920613(m,2H,H2-4).

[0055] (R)-MTPA ester (1-b): Yellow oily substance. 1 H NMR(600MHz,Pyridine-d5)δ6.436814(d,J=15.2Hz,1H,H-1),6.273929(dd,J=15.2,7.9Hz,1H,H-2),3.76437 2(m,1H,H-5),2.658674(m,2H,H2-7),2.237619(m,2H,H2-3),2.051375(m,2H,H2-6),1.925677(m,2H,H2-4).

[0056] Analyze the proton chemical shift difference around the chiral center C-3 in MTPA esters, such as Figure 3 Its configuration was determined to be S, and its structural formula is as follows. It was named (S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenyl acetate [(S,E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenyl acetate].

[0057]

[0058] II. Assay for α-glucosidase inhibitory activity

[0059] Referring to the method described in the literature XL Zhou, SB Li, MQ Yan, Q. Luo, LS Wang, LL Shen, MMLiao, CH Lu, XY Liu, and CQ Liang, Phytochemistry, 183, 112618 (2021), the α-glucosidase inhibitory activity of compound 1 was determined using acarbose as a positive control. The specific experimental steps are as follows:

[0060] The compound was prepared as a stock solution of 1% DMSO, and then diluted with phosphate buffer (pH 6.8) to produce solutions of different concentrations for experiments. 50 μL of sample solutions or acarbose solutions of different concentrations (1, 10, 50, 100, 200, and 300 μM) were added to 96-well plates, followed by 50 μL of buffer solution and 100 μL of 0.2 U / mL α-glucosidase. The mixture was incubated at 37°C for 15 minutes, then 50 μL of 2 mM 4-PNPG solution was added, and the mixture was incubated again at 37°C. The absorbance was measured at 405 nm.

[0061] The inhibition rate of compound 1 against α-glucosidase was calculated using the following formula.

[0062] α-glucosidase inhibition rate (%) = [Ac - (As - Ab) / Ac] × 100%

[0063] Where Ac is the absorbance without the control group, Ab is the absorbance without the 4-PNPG control group, and As is the absorbance of the sample to be tested.

[0064] Enzyme inhibition kinetics assay: The reversibility of inhibition was investigated for the inhibitory activity of the enzyme against different concentrations (0 and 300 μM) of the compound at enzyme concentrations of 0.2, 0.4, 0.6, and 0.8 U / mL. The reaction rate was tested using four different concentrations of 4-PNPG (0.5, 0.7, 1.0, and 2.0 mmol / L) and three different concentrations of compound 1. Double reciprocal curves of the compound were obtained, and the type of inhibition was evaluated.

[0065] The results showed that compound 1 has potential inhibitory activity against α-glucosidase, IC50. 50 The value was 71.14 ± 5.82 μmol / L, and the IC50 of acarbose was... 50 The value was 120.58 ± 39.21 μmol / L. Kinetic analysis showed that... Figure 4 and Figure 5 In the equation, all straight lines pass through the origin (0,0), and the slope of the lines decreases with increasing sample concentration. Therefore, the inhibitory effect of compound 1 and acarbose on α-glucosidase is reversible. Furthermore, in... Figure 6 As can be seen, compound 1 intersects on the x-axis, and compound 1 inhibits α-glucosidase in a non-competitive manner; Figure 7 In acarbose, the lines intersect on the y-axis, indicating that its inhibitory effect is competitive.

[0066] The above description is a detailed description of the preferred embodiments of the present invention. However, the embodiments are not intended to limit the scope of the patent application of the present invention. All equivalent changes or modifications made under the technical spirit of the present invention should fall within the patent scope covered by the present invention.

Claims

1. (S, E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenyl acetate, characterized in that, The structure is as follows: 。 2. The process for the preparation of (S, E)-4-(5-hydroxy-7-(4-hydroxy-3- methoxyphenyl)hept-1 -en-1 -yl)phenyl acetate according to claim 1, characterized in that, Includes the following steps: (1) The rhizome of Curcuma zedoaria was crushed and extracted with ethyl acetate. The extract was concentrated to obtain the residual extract. The extract was separated by silica gel column chromatography with a 100-200 mesh and eluted with a gradient of petroleum ether-ethyl acetate mixed solution with a volume ratio of 1:0-0:1 to obtain six components Fr.I ~ VI. (2) The fraction Fr.Ⅳ was repeatedly separated by HW-40F gel column chromatography three times, followed by silica gel column chromatography with 100-200 mesh, and then eluted with a gradient of petroleum ether-ethyl acetate mixed solution with volume ratios of 8:2, 2:1 and 1:1 to obtain three fractions Fr.Ⅳ2b1~Fr.Ⅳ2b3; (3) The component Fr.Ⅳ2b1 was separated by preparative high performance liquid chromatography and then purified by semi-preparative high performance liquid chromatography to obtain the (S, E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate; The separation conditions for preparative high performance liquid chromatography (HPLC) are as follows: using CH3OH: H2O eluent at a volume ratio of 55:45; the purification conditions for semi-preparative HPLC are as follows: using CH3OH: H2O eluent at a volume ratio of 55:

45.

3. The preparation method according to claim 2, characterized in that, In step (1), the extraction is carried out at room temperature, and the extraction is performed 3 times, each time for 3 days.

4. The preparation method according to claim 2, characterized in that, In step (1), the extraction solution is concentrated by vacuum concentration.

5. The preparation method according to claim 2, characterized in that, In step (1), the gradient elution is performed by sequentially eluting with a petroleum ether-ethyl acetate mixed solution with volume ratios of 1:0, 9:1, 8:2, 2:1, 1:1, and 0:

1.

6. The use of (S, E)-4-(5-hydroxy-7-(4-hydroxy-3-methoxyphenyl)hept-1-en-1-yl)phenylacetate from Curcuma zedoaria according to claim 1 in the preparation of hypoglycemic pharmaceutical products.

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