Compound isolated from balsam pear and its preparation method and application
A compound, the technology of bitter melon, is applied in the application field of preparation of hypoglycemic drugs, which can solve the problems of increasing the burden of pancreatic islet β cells, difficult to correct hypoglycemia, and pancreatic islet function failure, and achieve excellent hypoglycemic activity, novel structure, and quality. Effect
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Embodiment 1
[0031] Take dried unripe bitter melon fruits and extract them under reflux with 70% ethanol, extract twice for 2 hours each time, filter, combine the filtrates, and recover the solvent under reduced pressure to obtain ethanol bitter melon extract. After acid hydrolysis (see Example 3-9 for the method), the bitter melon hydrolyzate was obtained. Separate the bitter melon hydrolyzate on 200-300 mesh silica gel, use petroleum ether-ethyl acetate 50:1, 30:1, 15:1 gradient elution, 5 column volumes for each gradient elution, qualitative silica gel thin layer, combine The same fraction, 30:1 elution pooled part to obtain a mixture of compounds Ⅰ, Ⅱ, Ⅲ and Ⅳ. The mixture was prepared by high performance liquid chromatography, and the liquid phase conditions were: temperature: room temperature; UV detection wavelength: 203nm; mobile phase: 92% methanol-water; flow rate: 3.5mL / min; maximum injection volume / time: <1mg , There is good resolution between the 4 chromatographic peaks, so i...
Embodiment 2
[0042] The above compounds inhibit the activity of PTP1B
[0043] The experiment of inhibiting PTP1B enzyme activity was carried out in a 96-well plate. The experimental group was sequentially added with 83 μL enzyme-containing buffer solution, 10 μL test sample solution and 4 μL substrate solution, and the positive control group was sequentially added with 83 μL enzyme-containing buffer solution and 10 μL sodium vanadate solution. and 4 μL of substrate solution. To the reagent control group, 83 μL of enzyme-containing solution, 10 μL of DMSO solution and 4 μL of substrate solution were added, reacted in a constant temperature incubator at 37°C for 30 minutes, and then terminated with 5 μL of NaoH. The ultraviolet absorption intensity of the product at 410 nm was measured by a microplate reader.
[0044] Solution preparation
[0045] ① Storage solution
[0046] Take 0.6055g Tris and 0.0584g EDTA and dilute to 20mL with distilled water, and adjust the pH to 7.5 with concentr...
Embodiment 3
[0072] The hydrochloric acid hydrolysis method of embodiment 3 balsam pear extract
[0073] 3.1 Weigh 10g of bitter gourd extract, ultrasonically dissolve it in 50mL of methanol, slowly add 10mL of concentrated hydrochloric acid dropwise (stirring at the same time) and perform ultrasonic hydrolysis. Ultrasonic conditions: frequency: 50kHz; power: 3KW; time: 30 minutes; hydrolysis at a temperature of 40°C for 12h. Add 1000mL of water to the hydrolyzed reaction liquid, and after standing still, the precipitated part is washed with water repeatedly to neutrality, then stand still, collect the precipitate, filter, and dry to obtain the bitter gourd hydrolyzate.
[0074] 3.2 Weigh 10g of balsam pear extract, dissolve it in 50mL of methanol by ultrasonic, slowly add 10mL of concentrated hydrochloric acid dropwise (while stirring), and perform ultrasonic hydrolysis. Ultrasonic conditions: frequency: 50kHz; power: 3KW; time: 30 minutes; hydrolyzed at a temperature of 40°C for 12 hour...
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