Use of arjunolic acid in preparing glycosidase inhibitor

[0017] Example 1 :Preparation of arjunolic acid in dried leaves of Lagerstroemia indica

[0018] 1.1 Instruments and reagents

[0019] Proton Nuclear Magnetic Resonance Spectroscopy ( 1 H-NMR), carbon nuclear magnetic resonance spectroscopy ( 13 C-NMR) and two-dimensional nuclear magnetic resonance spectroscopy (2D NMR) are measured by INOVA superconducting nuclear magnetic resonance spectrometer (VARIAN INOVA-400MHz) (tetramethylsilyl ether is the internal standard); electrospray mass spectrometry (ESI-MS) is determined by Measured by Bruker Esquire 3000+ mass spectrometer, silica gel for column chromatography (100-200, 200-300) and silica gel GF254 (10-40 mesh) for thin layer chromatography were purchased from Qingdao Ocean Chemical Factory; all reagents used were analytical pure , The boiling range of petroleum ether is 60-90°C; the ultraviolet lamp of 254nm and 365nm is used for thin plate (TLC) detection; the developer uses 10% sulfuric acid-ethanol and bromocresol green solution.

[0020] 1.2 Plant source and identification

[0021] The medicinal material used for extraction was Lagerstroemia tomentosa Prezl (Lagerstroemia tomentosa Prezl), which was collected from Yunnan Province in August 2001 and was identified by researcher Peng Hua from the Kunming Institute of Botany, Chinese Academy of Sciences.

[0022] 1.3 Extraction and separation

[0023] The sample (the dried leaves of Lagerstroemia invertebrate, weighing 0.5 kg) was crushed and extracted twice with 95% ethanol at room temperature for 24 hours each time. After cooling, the extracts were combined and concentrated under reduced pressure to obtain 51 g of brown viscous crude extract . The crude extract was dissolved in 2 liters of hot water, petroleum ether was defatted (2 liters/each, 3 times in total), then extracted with ethyl acetate (2 liters/each, 5 times in total), and the solvent was evaporated under reduced pressure. The obtained ethyl acetate extract (12.5 g) was mixed with 10 g of 100-200 mesh silica gel, and then chromatographed on a silica gel column (200-300 mesh, 150 g), and then petroleum ether-acetone (100:0-0: 100) is the eluent for gradient elution, and each 100 ml is a first fraction. The fraction of the same composition is detected by thin-layer chromatography, and the α-glucosidase inhibitory activity is tested after decompression and desolvation. The α-glucosidase inhibitory activity measured by combining the 55-76 fractions was 65.7% (the crude extract was determined at a concentration of 100 μg/ml), and then the solvent was distilled under reduced pressure, and methanol was used as the solvent for multiple recrystallizations , The colorless needle-crystal compound, Arjunolic acid (42 mg) was obtained.

[0024] 1.4 Structural identification of Ajiang's acid

[0025] Arganic acid: colorless needle crystals; melting point 250-252°C (uncorrected); ESI-MS m/z: 487[M-H] -; Proton Nuclear Magnetic Resonance Spectroscopy (CD 3 OD, 400MHz): δ 5.29 (1H), 3.73 (1H, multiplet), 3.54 (1H, double peak, J = 10.0 Hz), 3.39 (1H, double peak, J = 9.6 Hz), 3.35 (2H, single Peak), 3.31 (1H, doublet, J=10.0Hz), 2.89 (1H, multiplet), 1.22 (3H, singlet), 1.14 (3H, singlet), 1.10 (3H, singlet), 0.98 ( 3H, singlet), 0.95 (3H, singlet), 0.86 (3H, singlet), 0.74 (3H, singlet); carbon nuclear magnetic resonance spectrum (Me 2 CO-d 6 , 100MHz): δ 181.84 (C, C-28), 145.37 (C, C-13), 123.40 (CH, C-12), 78.13 (CH, C-3), 69.65 (CH, C-2), 66.23(CH 2 , C-23), 48.92 (CH, C-5), 48.13 (CH, C-9), 47.86 (CH 2 , C-1), 47.21(CH 2 , C-19), 47.61 (C, C-17), 44.10 (C, C-4), 43.00 (C, C-14), 42.70 (CH, C-18), 39.02 (s, C-10) , 34.87(CH 2 , C-21), 33.81 (CH 2 , C-7), 33.55(CH 3, C-29), 33.32 (CH 2 , C-22), 31.60 (C, C-20), 28.76 (CH 2 , C-15), 26.45 (CH 3 , C-27), 24.60 (CH 2 , C-11), 24.02 (CH 3 , C-30), 23.97 (CH 2 , C-16), 19.08 (CH 2 , C-6), 17.76 (CH 3 , C-25), 17.52 (CH 3 , C-26), 13.86(CH 3 , C-24).

[0026] According to the above-mentioned structural identification data, it can be seen that the compound’s 1 H-NMR and 13 The C-NMR spectrum is basically the same as that of the compound ajiangmanic acid [Kundu, AP; Mahato, SBPhytochemistry, 1993, 32, 999; Yu Shao; Bing-Nan Zhou, Long-Ze Lin et al; phytochemistry, 1996, 38(6) , 1487-1492], thereby identifying the structure of the compound as arjunolic acid, namely 2α, 3β, 23-trihydroxy-oleanolic-12-en-28-acid.

[0027] Formula 1)

[0028] Example 2 :Preparation of arjunolic acid in fresh leaves of Lagerstroemia indica

[0029] 2.1 Apparatus and reagents: same as in Example 1.

[0030] 2.2 Plant source and identification: the same as in Example 1.

[0031] 2.3 Extraction and separation

[0032] The samples (fresh leaves weighing 0.5 kg) were chopped immediately with a knife and extracted twice with 95% ethanol at room temperature for 24 hours each time. After cooling, the extracts were combined and concentrated under reduced pressure to obtain 35 grams of brownish black viscous Like crude extract. The crude extract was dissolved in 2 liters of hot water, petroleum ether was defatted (2 liters/each, 3 times in total), then extracted with ethyl acetate (2 liters/each, 5 times in total), and the solvent was evaporated under reduced pressure. The obtained ethyl acetate extract (8.3 g) was mixed with 10 g of 100-200 mesh silica gel, and was chromatographed on a silica gel column (200-300 mesh, 150 g), and then petroleum ether-acetone (100:0-0: 100) is the eluent for gradient elution, and each 100 ml is a first fraction. The fractions of the same composition are detected by thin-layer chromatography, and the α-glucosidase inhibitory activity is tested after decompression. The α-glucosidase inhibitory activity measured by combining the 45-53 fractions was 59.7% (the crude extract was measured at a concentration of 100 μg/ml), and then the solvent was distilled under reduced pressure, and methanol was used as the solvent for multiple repetitions. Crystallized to obtain a colorless needle-crystal compound, ajiangramenic acid (28 mg).

[0033] 2.4 The structure identification of arjunolic acid: the same as in Example 1.

[0034] Example 3 :Preparation of arjunolic acid in the stems, branches and root bark of Lagerstroemia indica

[0035] 3.1 Instruments and reagents: the same as in Example 1.

[0036] 3.2 Plant origin and identification: the same as in Example 1.

[0037] 3.3 Extraction and separation

[0038] The sample (dry weight of stems, branches and root bark 0.5 kg) was crushed and extracted twice with 95% ethanol at room temperature for 24 hours each time. After cooling, the extracts were combined and concentrated under reduced pressure to obtain 43 grams of brown viscous crude extract. Things. The crude extract was dissolved in 2 liters of hot water, petroleum ether was defatted (2 liters/each, 3 times in total), then extracted with ethyl acetate (2 liters/each, 5 times in total), and the solvent was evaporated under reduced pressure. The obtained ethyl acetate extract (9.6 g) was mixed with 10 g of 100-200 mesh silica gel, and was chromatographed on a silica gel column (200-300 mesh, 150 g), and then petroleum ether-acetone (100:0-0: 100) is the eluent for gradient elution, and each 100 ml is a first fraction. The fractions of the same composition are detected by thin-layer chromatography, and the α-glucosidase inhibitory activity is tested after decompression. The α-glucosidase inhibitory activity measured by combining the 50th to 57th fractions was 51.5% (the crude extract was determined at a concentration of 100 μg/ml), and then the solvent was distilled under reduced pressure, and methanol was used as the solvent for multiple recrystallizations , The colorless needle-crystal compound, Arjunolic acid (26 mg) was obtained.

[0039] 3.4 Structural identification of arjunolic acid: the same as in Example 1.