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The application of papaya saponin Ⅰ in the detection of lysosome

Active Publication Date: 2022-02-25
JINAN UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] It was reported earlier that saponin Ⅰ has potential antitumor activity, but so far, there is no report that saponin Ⅰ can target lysosomes

Method used

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  • The application of papaya saponin Ⅰ in the detection of lysosome
  • The application of papaya saponin Ⅰ in the detection of lysosome
  • The application of papaya saponin Ⅰ in the detection of lysosome

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] The synthesis of embodiment 1 Polyphyllin I-Cy3

[0050] 1.1 Experimental steps

[0051] Synthesis steps of Polyphyllin I-Cy3: Weigh 5mg polyphyllin I (Polyphyllin I) and dissolve it in 0.5mL dimethylformamide (DMF, Shanghai Titan Technology Co., Ltd.), add 3.45mg Cy3-COOH (Ruixi Biological ), 1.34mg 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC, Biide Pharmaceuticals) and 71.47μg 4-dimethylaminopyridine (DMAP, Adamas Reagent Co., Ltd. ) Dissolved completely (Paulus saponin Ⅰ: Cy3-COOH: EDC: DMAP = 1: 1.2: 1.2: 0.1 (molar ratio). Stir the reaction at 50°C for 6 h, and pass through a rotary evaporator (type YRE2000E, Yuhua, Gongyi City) Instrument Co., Ltd.) under the condition of 40 ℃ and 120r / min, the solvent was removed by rotary evaporation under reduced pressure, then washed 3 times with acetone, recrystallized with ethanol, and dried in vacuo to obtain the product. Using high performance liquid chromatography (HPLC, Hangzhou Xinqiao Bio Technol...

Embodiment 2

[0057] Example 2 Polyphyllin I-Cy3 targets lysosomes

[0058] 2.1 Experimental steps:

[0059] 2.1.1 Cell culture

[0060] Liver cancer HepG2 cells were purchased from the American Type Culture Collection (ATCC). At 37°C, 5% CO 2 Cultured in DMEM medium (Invitrogen) containing 10% (v / v) fetal bovine serum (Gibco) and 1% (w / v) penicillin / streptomycin (Invitrogen) in an incubator. When the cells are subcultured, first remove the original medium, wash it twice with PBS buffer, add trypsin (Gibco company) to digest the cells, and leave it in the cell incubator for about 1 min. If the cells have fallen off, add the medium to stop the digestion . The cells were blown down gently, centrifuged at 216g for 3min, the supernatant was removed, and the cells were resuspended in fresh DMEM medium containing 10% (v / v) fetal bovine serum and 1% (w / v) penicillin / streptomycin, The passage ratio was 1:4. The cells used for confocal observation were all cultured in confocal culture dishes (...

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Abstract

The invention discloses the application of papaya saponin I in detecting lysosomes. In the present invention, it has been found through experiments that patricia saponin I has the effect of specifically targeting lysosomes. In order to facilitate observation and detection, the present invention further synthesizes a red fluorescent probe with papaya saponin I and a fluorescent dye, which is named as Polyphyllin I-Cy3, its structural formula is as shown in (II). The Polyphyllin I-Cy3 red fluorescent probe synthesized by the present invention can permeate the cell membrane and can be used for lysosome-specific fluorescent staining in living cells. It has good targeting, fast detection speed, good repeatability, convenient use, and no cytotoxicity, etc. advantage.

Description

technical field [0001] The invention belongs to the field of biological detection, and in particular relates to the application of saponin I in the detection of lysosomes. Background technique [0002] Lysosomes (lysosomes) are single-membrane-coated sac-like structures with a diameter of about 0.025 to 0.8 microns, which contain a variety of hydrolytic enzymes. They are organelles with certain morphological structures and important physiological functions in biological cells. exogenous and endogenous macromolecules. Since Belgian scholar Cristian de Duve and others discovered lysosomes in rat liver cells in 1955, people have studied lysosomes in all aspects, and found that lysosomes are related to many diseases such as metabolic diseases, inflammation, tumors and immunity, etc. occurrence is closely related. Therefore, it is of great significance to develop probes that can accurately label and visualize lysosomes. [0003] Cy3 dye is an amino-reactive dye with a molecula...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07J71/00C09K11/06G01N21/64
CPCC07J71/0005C09K11/06G01N21/6428G01N21/6458C09K2211/1029C09K2211/1088G01N2021/6439
Inventor 汪洋张静何庆瑜
Owner JINAN UNIVERSITY
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