Natural flavonoid from green tea

(+)-Gallocatechin (10-300 μM, 48 h) causes apoptosis and prevents HCT-116 and SW-480 cells from proliferating [1].

Apoptosis analysis [1]
Cell Types: HCT-116 Cell
Tested Concentrations: 0-100 µM
Incubation Duration: 48 h
Experimental Results: Apoptosis rates at concentrations of 60, 80 and 100 µM: 14.1%, 25.0%, 23.5%.

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Cell Proliferation Analysis[1]
Tea polyphenols were dissolved in DMSO and stored at −20 °C before use. Cells were seeded in 96-well plates (1 × 10~4 cells/well). After 24 h, indicated concentrations of drugs were added to the wells. The final concentration of DMSO was 1%. Controls were exposed to culture medium containing 1% DMSO without drugs. Following the indicated incubation period, cell proliferation was evaluated using an MTS assay according to the manufacturer’s instructions. Briefly, the medium was replaced with 100 μL of fresh medium and 20 μL of MTS reagent (CellTiter 96 Aqueous Solution) in each well, and the plate was returned to the incubator for 1–2 h. A 60 μL aliquot of medium from each well was transferred to an ELISA 96-well plate and its absorbance at 490 nm was recorded. Since 1% DMSO did not influence the proliferation of the two cell lines, results were expressed as percent of control (DMSO vehicle set at 100%).

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Cell Cycle Analysis[1]
HCT-116 cells were plated at a density of 2 × 10~5 cells onto 24-well tissue culture plates. After culturing for 1 day, the medium was changed and the tea polyphenols were added. Cells were incubated for 48 h before they were harvested. These cells were fixed gently with 80% ethanol in a freezer for 2 h and were then treated with 0.25% Triton X-100 for 5 min in an ice bath. Cells were resuspended in 150 µL of PBS containing 40 µg/mL propidium iodide (PI) and 0.1 mg/mL RNase. The cells were incubated in a dark room for 20 min at room temperature, and cell cycle analysis was performed using a FACScan flow cytometer and FlowJo 9.0 software. For each measurement, at least 10,000 cells were counted.

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Apoptosis Analysis[1]
HCT-116 cells were seeded in 24-well tissue culture plates (2 × 10~5 cells/well). On the second day, the medium was changed and cells were treated with test compounds. After treatment for 48 h, the cells floating in the medium were collected. The adherent cells were detached with 0.05% trypsin. Then the culture medium containing FBS and floating cells was added to inactivate the trypsin. After being pipetted gently, the cells were centrifuged for 5 min at 1500× g. The supernatant was removed and the cells were stained with annexin V-FITC and PI according to the manufacturer’s instructions. Annexin V-FITC detects translocation of phosphatidylinositol from the inner to the outer cell membrane during early apoptosis, and PI can enter the cell in late apoptosis or necrosis. Untreated cells were used as control for the double staining. The cells were analyzed immediately after staining using a FACScan flow cytometer and FlowJo 9.0 software. For each measurement, at least 20,000 cells were counted.

[1]. Epigallocatechin Gallate (EGCG) is the most effective cancer chemopreventive polyphenol in green tea. Nutrients. 2012 Nov 8;4(11):1679-91.

Gallocatechin is a catechin belonging to the flavanoids, with hydroxyl groups substituted at the 3, 3', 4', 5, 5', and 7 positions (trans isomer). It is isolated from black locust (Acacia mearnsii) as a metabolite. It is a catechin and also a flavanoid-3,3',4',5,5',7-hexanol. (+)-Gallocatechin has been reported in tea (Camellia sinensis), oak (Quercus acutissima), and other organisms with relevant data. See also: anthocyanin alcohols (subclass); Croferlemer (monomer); green tea leaves (partial). Green tea is a popular beverage consumed daily by millions worldwide. Previous studies have shown that some polyphenolic compounds in green tea possess anticancer activity. However, systematic evaluation remains limited. This study determined the cancer chemopreventive potential of 10 representative polyphenols (caffeic acid, CA; gallic acid, GA; catechin, C; epicatechin, EC; gallocatechin, GC; catechin gallate, CG; gallocatechin gallate, GCG; epicatechin gallate, ECG; epigallocatechin, EGC; epigallocatechin gallate, EGCG) and explored their structure-activity relationships. The effects of these 10 polyphenols on the proliferation of HCT-116 and SW-480 human colorectal cancer cells were assessed using the MTS assay. Cell cycle distribution and apoptosis were analyzed by flow cytometry after staining with propidium iodide (PI)/RNase or Annexin V/PI. Among the 10 polyphenols, EGCG exhibited the strongest antiproliferative effect and significantly induced cell cycle arrest in the G1 phase and apoptosis. Studies on the relationship between chemical structure and anticancer activity have revealed that C and EC do not have antiproliferative effects, while GA does have certain antiproliferative effects. When C and EC are esterified with GA to generate CG and ECG, their antiproliferative effects are significantly enhanced. A similar association exists between EGC and EGCG. Gallic acid groups significantly enhance the anticancer potential of catechins. This property can be used for the semi-synthesis of flavonoid derivatives in the future to develop novel anticancer drugs. [1]

C15H14O7

306.2675

306.073

C, 58.83; H, 4.61; O, 36.57

970-73-0

(-)-Epigallocatechin;970-74-1;(+)-Gallocatechin-13C3

65084

White to off-white solid powder

1.7±0.1 g/cm3

685.6±55.0 °C at 760 mmHg

189 - 191 °C

368.5±31.5 °C

0.0±2.2 mmHg at 25°C

1.776

-0.1

6

7

1

22

380

2

C1[C@@H]([C@H](OC2=CC(=CC(=C21)O)O)C3=CC(=C(C(=C3)O)O)O)O

XMOCLSLCDHWDHP-SWLSCSKDSA-N

InChI=1S/C15H14O7/c16-7-3-9(17)8-5-12(20)15(22-13(8)4-7)6-1-10(18)14(21)11(19)2-6/h1-4,12,15-21H,5H2/t12-,15+/m0/s1

(2R,3S)-2-(3,4,5-trihydroxyphenyl)-3,4-dihydro-2H-chromene-3,5,7-triol

(+)-Gallocatechin; Gallocatechin; 970-73-0; Gallocatechol; (+)-gallocatechol; d-Gallocatechin; 1617-55-6; (2R,3S)-gallocatechin;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product requires protection from light (avoid light exposure) during transportation and storage.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~100 mg/mL (~326.51 mM)
H2O : ~3.33 mg/mL (~10.87 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (8.16 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (8.16 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (8.16 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)