green tea extract containing a highly absorbent catechin prevents diet-induced lipid metabolism disorder - highly absorbent
We studied the effect of Benifuuki extract (
A variety of tea leaves containing table alloc subtable ech-3-O-(3-O-methyl)gallate (EGCG3”Me))
High in feedingfat/high-sucrose (HF/HS)diet.
Then compare this tea variety with the extract from Yabukita (
Popular tea varieties that lack methylation tea).
For 6 weeks, whether or not taking tea extract from tea varieties, feed the HF/HS diet, which contains almost the same ingredients except for the methylation of the chechin (i. e. , Yabukita (0. 2% and 1%)or Benifuuki (0. 2% and 1%)
Add Ben mu 0.
2% the plasma TG and NEFAs levels were significantly reduced compared to mice supplemented with Yabukita 0. 2%.
Diet containing benifuki 1% reduces fat tissue weight, liver TG, and expression of liver fat-generating genes.
These results show that benifuki has a larger lipid-
Reducing blood lipid effect than Bush North pine needles.
In combination, these data suggest that catech salts of methylation directly result in strong lipid-
Reduce the activity of the wood.
Male C57/BL6 mice (12 weeks)
From the Charles River (Kanagawa, Japan).
They stay at a temperature. and humidity-
Control room with 12-h-
Light-dark cycle (
Lights from eight o'clock A. M. to eight o'clock P. M).
All mice adapted for a week while feeding AIN93G diet. AIN-
93G and HF/HS diets were obtained from Oriental KBT (Tokyo, Japan)().
The mice were divided into six groups :(1)
Negative control group (control)fed a AIN-93G diet; (2)
Positive control group (HF/HS)fed a high-fat/high-sucrose diet; (3)a group (Yabukita 0. 2%)
Feeding HF/HS diet but containing 0. 2% Yabukita; (4)a group (Yabukita 1. 0%)
Feeding HF/HS diets containing 1. 0% Yabukita; (5)a group (Benifuuki 0. 2%)
Fed a HF/HS contains 0. 2% Benifuuki; and (6)a group (Benifuuki 1. 0%)
Feeding HF/HS diets containing 1. 0% Benifuuki.
The extract of Yabukita and benifuki is extracted from the leaves of the plant with boiling water and then frozendrying.
The composition of each extract powder is high-
High performance liquid chromatography (HPLC)().
The diet of green tea was added to 0. 2% and 1.
0% is equivalent to 1 human intake.
6 cups and 8 cups (2.
0g green tea/Cup)
According to the energy intake of 8,360 kilojoules per day, it is daily.
Feed mice three times a week and weigh them every week.
At the end of the 6-week feeding, the mice were given anesthesia under isocophonic vapor after food deprivation at night.
Blood samples were collected in test tubes. orbital sinus.
Collect plasma after Centrifuge (
2000 × 20 min at 4 °c)
Stored at-80 °c.
After blood collection, the mice were killed by an excessive amount of isocromine.
Visceral fat tissue (
Warehouse of abdominal cavity and ovary)
Harvest, rinse and weigh the liver.
From the central leaf of the liver ,~ 0.
Cut 5g of tissue from each mouse and place it in a plastic tube containing RNALater™Solution (
Ambion, Austin, Texas, United States of America).
Liver tissue in RNALater is maintained for 24 hours at 4 °c and then stored at-80 °c.
The experiment was conducted in accordance with the animal experiment guidelines of the Agricultural College of Kyushu University.
The study was approved by the Committee on Animal Care and Use at Kyushu University, Fukuoka, Japan.
Animal experiment approval number A22-146.
Preparation of tea samples for HPLC analysis.
Simply dip the sample into 50% ethanol containing 1% HPO, interrupt for 30 minutes using an ultrasonic device at 20 °c, and then filter through a membrane filter (pore size, 0. 45u2005μm).
HPLC using Shimadzu LC-
10A pump coupled to UV detector (SPD-M10Avp;
Shimizu, Kyoto, Japan)and a reverse-
Phase Wakopak navigation 18-5 column (4. 6u2005mm i. d. × 150u2005mm;
5 μm particle diameter;
Pure chemical industry, Wako, Kyoto, Japan).
Eluted by 2-45-
The minimum linear gradient of solvent B and solvent A is from 0 to 80%.
Solvent A is composed of distilled water/phosphate/acetone (400:10:1 )
Solvent B is composed of methanol/solvent (1:2 ).
Samples were eluted at 40 °c for 1 mL/min.
The detection wavelength is 272 nm.
By comparing the peak area of each catechin in tea extract and the peak area of standard preparations containing a fixed amount of caffeine, the amount of catechins and caffeine in tea extract was measured, EC, c. table TP, gallocatechin gallate (GCG)
, Map room, gallin gallate (CG)
EGCG3 "Me and gallocatechin-3--(3--methyl)gallate (GCG3″Me).
Plasma level of triacid gan oil (TG), non-
Fatty acids in ester (NEFAs)
, Amino acid (AST)
, Alt (ALT)
Total cholesterol (TC), high-
Cholesterol (HDL-C), low-
Density Cholesterolcholesterol (LDL-C)and very low-
Density Cholesterolcholesterol (VLDL-C)
Measurement using tg e-test, the non-
Fatty acids in ester (NEFA)C-
Transaminase-detection and Transaminasetest (each from Wako), HDL-C and LDL-C/VLDL-
C quantitative kit (
BioVision, Milpitas, California, USA), respectively.
Measurement of liver TG content using tg e-test (from Wako)
After extracting liver lipids with sirroform-respectivelymethanol (2:1).
All kits are used according to the manufacturer's instructions.
Extraction of total RNA from liver samples using RNeasy™Mini Kit (
Chigan, Valencia, California, United States of America).
Analysis of all total RNA samples using the Agilent 2100 biological analyzer (
Agilent Technology, Palo Alto, California, USA).
Nucleic acid-labeled anti-sense RNA synthesis and amplification in total RNA (1u2005μg)
Enhanced Amplification Kit using messagampiii Biotin (
Applied Biological Systems, Foster, California, USA)
According to the manufacturer agreement.
After the purification of ARNA, the bio-labeled aRNA (5u2005μg)
Crushing with 10 x crushing reagent (
Applied Biological system
Heat 7 at 70 °c. 5u2005min.
Hybridization was performed in 15 μ l hybrid buffer with a DNA chip (0. 12 µm Tris-HCl/month. 12u2005M NaCl/0. 05% Tween-
20 and 5 μg fragment biomarker aRNA)
Overnight at 65 °c.
After hybridization, the DNA chip was washed twice at 0. 12 µm Tris-HCl/month. 12u2005M NaCl/0. 05% Tween-
Wash for 20 minutes at 20 °c at 65 °c and then wash at 0 °c. 12 µm Tris-HCl/month.
12 m nac1 in 10 minutes.
Then a hybrid aRNA-was marked with 2 μg/mL of penicillin-Cy5 (
GE Healthcare (UK)in 0. 12 µm Tris-HCl/month.
12 m nac1 30 min at room temperature.
After fluorescent labeling, the DNA chip was washed 4 times in 0. 12 µm Tris-HCl/month. 12u2005M NaCl/0. 05% Tween-
20 at room temperature 5-min each.
Scan the DNA chip at multiple exposure times (0. 1–40u2005s)
Using a DNA chip reader (
Yokogawa motor, Tokyo, Japan)with multi-
Beam excitation technology.
Select the intensity value of each point with the best exposure condition.
Subtract the median value of the background spot from the strength value of each gene, and then standardize the value as an expression of an endogenous control (
β-acid nucleotide phosphate protein
Actin and glycerin-3-
Dehydrogen phosphate enzyme
A cDNA was synthesized from total RNA (1u2005μg)
Use the PrimeScript RT kit (
Takara Bio, Tokyo, Japan).
Analysis of gene expression by real-
Time Quantitative PCR using the Sybr green Program and the thermal cycling dice real-time system (Takara Bio).
The primer sequence is given in.
Expression of mRNA values was standardized relative to glycerin3-
Dehydrogen phosphate enzyme as internal control.
All statistical analyses are used-
The test of Tukey was analyzed for variance using Kyplot software.
Data refers to SEM. P < 0.
05 is considered to be meaningful.