Application of Fenghuang Dancong tea extract in preparation of anti-breast-cancer drugs

Abstract

The invention discloses application of Fenghuang Dancong tea extract in preparation of anti-breast-cancer drugs. The Fenghuang Dancong tea extract is acquired by means of water extraction; water-soluble active ingredients in the Fenghuang Dancong tea extract have good antitumor activity; the Fenghuang Dancong tea extract is highly worthy of research for human breast cancer. Raw material for the Fenghuang Dancong tea extract herein is low in price and easy to obtain; the extraction process is simple and convenient and has simple conditions; the Fenghuang Dancong tea is safe to human; the Fenghuang Dancong tea extract is expectedly promotive in the application of Fenghuang Dancong tea and its isogeneic tea drinks in the fields such as anti-breast-cancer medicine, cosmetics, health products and foods; the Fenghuang Dancong tea extract is also applicable to the fields such as foods, health products and biomedicine to resist oxidation and improve body immunity.

Description

technical field

[0001] The invention belongs to the technical field of medicine and food, and particularly relates to the application of Fenghuang Dancong tea extract in preparing anti-breast cancer drugs. Background technique

[0002] Phoenix Dancong tea (C.sinensis cv.Fenghuang Dancong), is one of the green tea varieties, belongs to the oolong tea category, belongs to semi-fermented tea, and is also one of the six major tea varieties in China. In addition, Fenghuang Dancong tea and Lingtou Dancong and Fenghuang Narcissus is one of the main varieties of Guangdong oolong tea. Fenghuang Dancong has a production history of more than 900 years, with a long history and a wide reputation. Chaozhou, the main producing area of ​​Fenghuang Dancong, is one of the three major oolong tea producing areas in my country. Chaozhou Fenghuang Mountain is the origin of the national-level fine tea tree variety "Phoenix Narcissus". Several generations of tea farmers have separated and screened...

Examples

Embodiment 1

[0039] Example 1 Cell Viability Experiment

[0040] A MTT method was used to measure the effects of the water extract of Fenghuang Dancong tea on four human malignant tumor cells (human breast cancer cell MDA-MB231, human colon cancer cell SW480, human cervical cancer cell Hela, human liver cancer cell HepG2) and one The method for the toxicity of normal cells (human hepatocyte L02) (tumor cells and normal cells are purchased from the U.S. Model Culture Collection Bank), mainly includes the following steps:

[0041] (1) Dissolve the tea water extract of Fenghuang Dancong tea in PBS buffer solution, put it on an ultrasonic instrument and sonicate for 1-2 minutes, and prepare a 5 mg / mL stock solution for later use;

[0042] (2) Different tumor cells and normal cells in the logarithmic growth phase were taken at a density of 2×10 4 cells / mL were inoculated in 96-well plates (100 μL / well), and allowed to grow on the wall for 24 hours. Then add 100 μL of drugs with different conc...

Embodiment 2

[0044] Example 2 Flow cytometry detection of the influence of Fenghuang Dancong tea water extract on the distribution of tumor cell cycle

[0045] A method for detecting the influence of Phoenix Dancong tea water extract on the cell cycle distribution of human breast cancer MDA-MB231 by flow cytometry, mainly comprising the following steps:

[0046] (1) Dissolve the tea water extract of Fenghuang Dancong tea in PBS buffer solution, put it on an ultrasonic instrument for 1-2 minutes, and prepare a 5 mg / mL solution for later use;

[0047] (2) MDA-MB231 cell density is 2×10 4 / mL in a 6mL culture dish for 24 hours, after the cells are completely adhered to the wall, add the water extract of Fenghuang Dancong tea leaves with different concentrations prepared by using the stock solution prepared in step (1), incubate for 72 hours, and collect the cells. Fix with pre-cooled 70% ethanol overnight at -20°C. Centrifuge at 1500rpm for 10min, discard the culture medium, turn it upside ...

Embodiment 3D

[0049] Embodiment 3DHE probe detects intracellular reactive oxygen species (ROS) level

[0050] A method using DHE probes to detect the influence of Phoenix Dancong tea water extract on the level of reactive oxygen species (ROS) in human breast cancer MDA-MB231 cells, mainly comprising the following steps:

[0051] (1) Dissolve the tea water extract of Fenghuang Dancong tea in PBS buffer solution, put it on an ultrasonic instrument and sonicate for 1-2 minutes, and prepare a 60 μg / mL solution for later use.

[0052] (2) MDA-MB231 cell density is 2×10 4 / mL in a 96-well plate for 24 hours, add Phoenix Dancong tea water extracts with concentrations of 60 μg / mL, 30 μg / mL, and 15 μg / mL, and incubate for 2 hours, remove the medium in each well, and add 100 μL of 10 μM DHE Probes (reagents purchased from Sigma) were incubated for 30 min. The excitation and emission wavelengths of the fluorescent microplate reader were set to 300nm and 610nm respectively, and the fluorescence absor...

technical field

[0001] The invention belongs to the technical field of medicine and food, and particularly relates to the application of Fenghuang Dancong tea extract in preparing anti-breast cancer drugs. Background technique

[0002] Phoenix Dancong tea (C.sinensis cv.Fenghuang Dancong), is one of the green tea varieties, belongs to the oolong tea category, belongs to semi-fermented tea, and is also one of the six major tea varieties in China. In addition, Fenghuang Dancong tea and Lingtou Dancong and Fenghuang Narcissus is one of the main varieties of Guangdong oolong tea. Fenghuang Dancong has a production history of more than 900 years, with a long history and a wide reputation. Chaozhou, the main producing area of ​​Fenghuang Dancong, is one of the three major oolong tea producing areas in my country. Chaozhou Fenghuang Mountain is the origin of the national-level fine tea tree variety "Phoenix Narcissus". Several generations of tea farmers have separated and screened...

Examples

Embodiment 1

[0039] Example 1 Cell Viability Experiment

[0040] A MTT method was used to measure the effects of the water extract of Fenghuang Dancong tea on four human malignant tumor cells (human breast cancer cell MDA-MB231, human colon cancer cell SW480, human cervical cancer cell Hela, human liver cancer cell HepG2) and one The method for the toxicity of normal cells (human hepatocyte L02) (tumor cells and normal cells are purchased from the U.S. Model Culture Collection Bank), mainly includes the following steps:

[0041] (1) Dissolve the tea water extract of Fenghuang Dancong tea in PBS buffer solution, put it on an ultrasonic instrument and sonicate for 1-2 minutes, and prepare a 5 mg / mL stock solution for later use;

[0042] (2) Different tumor cells and normal cells in the logarithmic growth phase were taken at a density of 2×10 4 cells / mL were inoculated in 96-well plates (100 μL / well), and allowed to grow on the wall for 24 hours. Then add 100 μL of drugs with different conc...

Embodiment 2

[0044] Example 2 Flow cytometry detection of the influence of Fenghuang Dancong tea water extract on the distribution of tumor cell cycle

[0045] A method for detecting the influence of Phoenix Dancong tea water extract on the cell cycle distribution of human breast cancer MDA-MB231 by flow cytometry, mainly comprising the following steps:

[0046] (1) Dissolve the tea water extract of Fenghuang Dancong tea in PBS buffer solution, put it on an ultrasonic instrument for 1-2 minutes, and prepare a 5 mg / mL solution for later use;

[0047] (2) MDA-MB231 cell density is 2×10 4 / mL in a 6mL culture dish for 24 hours, after the cells are completely adhered to the wall, add the water extract of Fenghuang Dancong tea leaves with different concentrations prepared by using the stock solution prepared in step (1), incubate for 72 hours, and collect the cells. Fix with pre-cooled 70% ethanol overnight at -20°C. Centrifuge at 1500rpm for 10min, discard the culture medium, turn it upside ...

Embodiment 3D

[0049] Embodiment 3DHE probe detects intracellular reactive oxygen species (ROS) level

[0050] A method using DHE probes to detect the influence of Phoenix Dancong tea water extract on the level of reactive oxygen species (ROS) in human breast cancer MDA-MB231 cells, mainly comprising the following steps:

[0051] (1) Dissolve the tea water extract of Fenghuang Dancong tea in PBS buffer solution, put it on an ultrasonic instrument and sonicate for 1-2 minutes, and prepare a 60 μg / mL solution for later use.

[0052] (2) MDA-MB231 cell density is 2×10 4 / mL in a 96-well plate for 24 hours, add Phoenix Dancong tea water extracts with concentrations of 60 μg / mL, 30 μg / mL, and 15 μg / mL, and incubate for 2 hours, remove the medium in each well, and add 100 μL of 10 μM DHE Probes (reagents purchased from Sigma) were incubated for 30 min. The excitation and emission wavelengths of the fluorescent microplate reader were set to 300nm and 610nm respectively, and the fluorescence absor...

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