CHEN HU (CN)
JIN BEI WEN (CN)
ZHOU RUI (CN)
WOUTERS BERT C (BE)
CHEN HU (CN)
JIN BEI WEN (CN)
ZHOU RUI (CN)
WOUTERS BERT C (BE)
| US4613672A | 1986-09-23 |
DATABASE WPI Section Ch Week 198711, Derwent World Patents Index; Class B04, AN 1987-076886, XP002164189
| 1. | Method of obtaining a tea pigment comprising theaflavin, thearubigin, theabrownin and catechins, characterised by (1) mixing tea leaves and ethanol, soaking and refluxing the obtained suspension; (2) centrifuging the suspension and discarding the pellet ; (3) adding the remaining samples to a gel filtration column, washing the column ; and (4) collecting the washing solution, extracting the tea pigment using a halogenated hydrocarbon having 13 carbon atoms, discarding the water phase, evaporating the halogenated hydrocarbon, and recovering a tea pigment powder containing less impurities and showing higher efficacy. |
| 2. | Method according to claim 1, characterised in that in step (1) the tea leaves and the ethanol are mixed in a w/wratio of 1: 120. |
| 3. | Method according to claim 2, characterised in that in step (1) the tea leaves and the ethanol are mixed in a w/wratio of 1: 10. |
| 4. | Method according to any of claims 13, characterised in that in step (3) the samples are added to a Sephadex column, preferably a Sephadox LH20 column. |
| 5. | Method according to any of the claims 14, characterised in that in step (4) the halogenated hydrocarbon is a chlorinated hydrocarbon having one carbon atom. |
| 6. | Method according to claim 5 characterised in that in step (4) the halogenated hydrocarbon is CH2C12. |
| 7. | The product obtained by carrying out the method according to any of the claims 1 6. |
| 8. | A pharmaceutical composition, comprising the product according to claim 7 as one of the active components. |
Detailed description of the invention The invention relates to a method comprising the following steps: (1) mixing tea leaves and thanol, soaking and refluxing the obtained suspension; (2) centrifuging the suspension and discarding the pellet; (3) adding the remaining samples to a gel filtration column, washing the column; and (4) collecting the washing solution, extracting the tea pigment using a halogenated hydrocarbon having 1-3 carbon atoms, discarding the water phase, evaporating the halogenated hydrocarbon and recovering a tea pigment powder containing less impurities and showing higher efficacy.
More in particular the method according to the invention comprises the following steps: (1) mixing tea leaves and ethanol in a w/w-ratio of 1: 1-20, preferably about 1: 10, soaking and refluxing the obtained suspension; (2) centrifuging the suspension and discarding the pellet; (3) adding the samples to a Sephadex column, washing the column ; and (4) collecting the washing solution, extracting tea pigment using a chlorinated hydrocarbon having one carbon atom, discarding the water phase, evaporating the chlorinated hydrocarbon and recovering a tea pigment powder containing less impurities and showing higher efficacy.
Most preferably the method according to the invention is carried out in the following way: 1. Mixing tea and 80% ethanol solution (w/w = 1 : 10), soaking the suspension at room temperature for 2 hours, heating and refluxing the suspension for 1 hour, filtrating the tea extraction solution, eliminating the tea residue; and
2. adjusting the suspension pH to 3.2, using lmol/L HC1,10,000 rpm centrifuge for 30 min, discarding the pellet; and 3. adjusting the supernatant solution pH to 7.0, using lmol/L NaOH, adding the sample to a Sephadex LH-20 column, washing the column with 40%-100% ethanol solution; and 4. collecting the washing solution, adjusting the washing solution pH to 8.0, using lmol/L NaOH, extracting tea pigment using CH2Cl2, discarding the water phase, evaporating the CH2C12 and obtaining a tea pigment powder having excellent pharmaceutical properties, in particular in applications against hyperlipidemia and related diseases.
Further the invention relates to the product obtained by means of the method defined above.
Another aspect of the convention relates to the pharmaceutical compositions comprising the product obtained by means of method defined above. The pharmaceutical composition is applicable in the field of applications against hyperlipidemia and related diseases.
Figure The production process according to the invention is illustrated by the following figure. Mixing, soaking and refluxing filter l. I Adjusting Eliminating solution pH tea residue centrifuge Discarding Adding supernatant pellet in column Washing column and extracting Evaporating Discarding organic phase water phase Obtaining tea pigment
Experimental Studies and Clinical Application 1. Animal experiments (1) Safety test Tea pigment obtained according to the method of the invention is poured directly into rats'stomachs. The dose for rats is 100 times higher than that for human. Observing these rats'behaviour for one week, we find that rats act normally. This test shows the tea pigment according to the invention is safe.
(2) Effect on serum lipids Tea pigment according to the invention can induce decrease of serum total cholesterol (TC), triglyceride (TG), low density lipoprotein (LDL), and raise high density lipoprotein cholesterol (HDL) in comparison with control groups.
Figure 2. effects of tea pigment on serum lipids 2. Population Studies (1) Hyperlipidemia study The following values of TC, TG, HDL, LDL are the standard for normal human: TC 3.1-5.7mol/L TG 0.56-1.7mol/L HDL 1.04-1.55mol/L LDL 1.80-3.36mol/L
A total of 1,696 patients participated in this trial, 920 males and 776 females, ages 35 to 81 with an average weight of 58.9 + 7.9. Patients took 125 mg tea pigment 3 times/day for 4 weeks.
Patients satisfied the following criteria, 1) did not suffer an acute heart attack, brain damage, no injury, no operations within the last 6 months; 2) no kidney diseases; 3) no diabetes mellitus; 4) no thyroid disease; 5) no phase III hypertension; 6) no drug induced hyperlipidemia; 7) no pregnant women.
Table 1 changes in blood TC, TG, HDL and LDL levels. Cases (n) Before treatment After treatment Rate of P value mol/L (mol/L) change (% TC 811 6. 71+0. 55 5. 65 + 0. 41-15.8 < 0, 01 TG 923 2.95 + 0. 59 2. 27 + 0. 31-23.1 < 0,01 LDL 154 4. 05 + 0. 37 3.35+0.34-17.3 < 0,01 HDL 276 1. 19 + 0. 28 1. 34+0. 19 +12. 6<0, 01 Fig. 3 changes in patients blood TC, TG, HDL and LDL levels Table 2 shows changes on patients with hyperlipidemia. Cases (n) Markedly Improved (%) Not improved (%) Total Improved (%) improvement TC 811 59. 8 (485) 12.5 (101) 27.7 (225) 72.3 TG 923 41. 7 (385) 24.1 (222) 34.2 (316) 65. 8 LDL 154 48. 7 (75) 31.2 (48) 20.1 (31) 79. 9 HDL 276 50. 7 (140) 23.9 (66) 25.4 (70) 74. 7
(2) Dose-Effect and Period of Treatment-Effect Relationships: 250mg versus 125mg Tea pigment 60 days versus 30 days of Tea pigment treatment.
A total of 521 patients participated in this trial, 310 males and 211 females, ages 28 to 79 with an average weight of 55.2 + 5.9.
Group A: 125 mg 3 times a day Group B: 250 mg 3 times a day Patients satisfied the following criteria, 1) did not suffer an acute heart attack, brain damage, no injury, no operations within last 6 months; 2) no kidney diseases; 3) no diabetes mellitus 4) no thyroid disease 5) no phase III hypertension 6) no drug induced hyperlipidemia 7) no pregnant women.
Table 3 shows changes in blood TC, TG, HDL and LDL levels. The period of treatment is 30 days. Group Cases (n) Before treatment After treatment Rate of P value mol/L) (mol/L) change (% TC A 210 6. 51 + 0. 97 5. 51 _0. 76-15.3 <0,01 TC B 156 6. 54 + 0. 88 5. 21 + 0. 61-20. 3 <0, 01 TG A 198 3.11 ~ 0.74 2.34 ~ 0. 46-24.8<0, 01 TG B 112 3.06 ~ 0.68 2.01 ~ 0. 71-34.3 <0,01 LDL A 165 4.32 ~ 0.76 3.41 ~ 0. 59-21.1 <0, 01 LDL B 120 4.26 ~ 0.81 3.03 ~ 0. 65-28.9 <0, 01 HDL A 171 1. 04 + 0. 35 1. 23 _ 0. 29 +18. 3 <0, 01 HDL B 104 0. 9 + 0. 26 1. 31 0. 22 +33. 7 <0,01 Fig. 4 changes in patient's blood TC, TG, HDL and LDL levels (30 days) Table 4 shows changes on patients with hyperlipidemia The period of treatment is 30 days.
Group Cases (n) Markedly Improved Not Improved Total Improved (%) (%) (%) improvement % TC A 210 55. 2 (116) 15.2 (32) 29.6 (62) 70.4 TC B 156 62. 8 (98) 14.1 (22) 23.1 (36) 76.9 TG A 198 39. 9 (79) 21.2 42) 38.9 (77 61.1 TG B 112 50. 9 (57) 19.6 (22) 29.5 (33) 70.5 LDL A 165 47. 3 (78) 28.5 (47) 24.2 (4) 75. 8 LDL B 120 50. 8 (61) 30.0 (36) 19.2 (23) 81. 8 HDL A 171 49. 7 (85) 21.6 (37) 28.7 (49) 71. 4 HDL B 104 55. 8 (58) 24.0 (25) 20.2 (21) 79. 8 Table 5 shows changes in blood TC, TG, HDL and LDL levels. The period of treatment is 60 days. Group Cases (n) Before treatment After treatment Rate of P value mol/L) (mol/L) change (% TC A 210 6.51 + 0.97 5.48 + 0. 73-15.8 <0,01 TC B 156 6. 54 + 0. 88 5. 14 + 0. 75-21.4 <0,01 TG A 198 3.11 ~ 0.74 2.30 ~ 0. 46-26.0 <0,01 TG B 112 3. 06 + 0. 78 1. 92 + 0. 53-37.2 <0,01 LDL A 165 4. 32 + 0. 76 3. 38 + 0. 46-21.8 <0,01 LDL B 120 4.26 ~ 0.81 2.89 ~ 0. 58-32.2 <0, 01 HDL A 171 1. 04 + 0. 35 1. 24 + 0. 24 +19. 2 <0, 01 HDL B 104 0. 98 + 0. 26 1. 34 + 0. 19 +36. 7 <0, 01
Fig. 5 changes in atients'blood TC, TG, HDL and LDL levels (60 days).
Table 6 shows changes on patients with hyperlipidemia.
The period of treatment is 60 days. Group Cases (n) Markedly Improved Not Improved Total Improved (%) (%) % improvement % TC A 210 59. 0 (124) 12.4 (26) 28.6 (60) 71. 4 TC B 156 67. 9 (106) 10.3 (16) 21.8 (34) 78.2 TG A 198 46. 5 92) 17.6 (35) 35.9 (71) 64.1 TG B 112 59. 8 (67) 13.4 (15) 26. 8 (30) 73.2 LDL A 165 57. 0 (94) 20.0 (33) 23.0 (38) 77.0 LDL B 120 65. 0 (78) 18.3 (22) 16.7 (20) 83.3 HDL A 171 54. 4 (93) 18. 1 (31) 27.5 (47) 72.5 HDL B 104 59. 6 (62) 21. 2 (22) 19.2 (20) 81. 8
Significant differences occurred in all lipid level measurements.
However, no significant differences (P>0.05) occurred in the same dose and 2 periods of treatment.