TWI536998B - A use of an extract of zingiber zerumbet thereof - Google Patents

Description

Use of red ball ginger extract

The invention relates to the use of a red ball ginger extract, in particular to a red ball ginger extract for preparing a medicament for improving diabetic retinopathy.

Diabetic retinopathy (diabetic retinopathy) is a complication of diabetes, which is caused by hyperglycemia-induced vascular wall insufficiency, thus affecting the function of the blood-retinal barrier and enhancing the blood vessels of the affected part. Symptoms caused by changes in retinal microcirculation, such as permeability, are the leading cause of blindness in diabetic patients.

Methods for the treatment of diabetic retinopathy include laser surgery, vitrectomy, and ocular injection of corticosteroids (cprtocpsterpod). All three are invasive medical behaviors, and still cannot achieve the effect of radical cure. You need to monitor your blood sugar at any time and keep your blood sugar in the normal range (euglycemia), and the administration of corticosteroids is more likely to cause side effects. Accordingly, there is still a need for improved methods for treating diabetic retinopathy.

The present invention provides a use of a red ball ginger extract as an active ingredient for improving diabetic retinopathy to improve the side effects caused by the conventional method for treating diabetic retinopathy.

The use of a red ball ginger extract for the preparation of a medicament for improving diabetic retinopathy, wherein the red ball ginger extract is administered to a desired organism to improve the diabetic retinopathy of the desired ascending object, wherein The red ball ginger extract is prepared by the following method for preparing red ball ginger extract: a red ball ginger sample is immersed in rice vinegar for 8 hours to obtain a sample of impregnated red ball ginger; the impregnated red ball ginger sample The vinegar red ball ginger sample is obtained by boiling at a temperature of 95-105 ° C and a pressure of 1.5-2.5 kg/m ² for 30 minutes; and extracting the vinegar red with an extraction solvent at a temperature of 70-90 ° C The ginger sample was incubated for 8 hours, and the extraction solvent was pure water or a 95% by volume ethanol solution to obtain the red ball ginger extract.

The use of the red ball ginger extract of the present invention, wherein the red ball ginger extract is administered to the desired individual at a dose of 600 mg per kg of the desired individual per day, and is administered continuously for 28 days.

The use of the red ball ginger extract of the present invention is a derivative of the red globin extract rich in flavonoid compound as an active ingredient for improving diabetic retinopathy, and preventing the side effects caused by conventional drugs, and is an effect of the present invention.

Figure 1a: Results of fundus photography of group C0 diabetic rats in this test.

Figure 1b: Results of PAS and hematoxylin staining in the retina of group C0 diabetic rats in this test.

Figure 2a: Results of fundus photography of Group C1 diabetic rats in this trial.

Figure 2b: Results of PAS and hematoxylin staining in the retina of the C1 group of diabetic rats.

Figure 3a: Results of fundus photography of Group C2 diabetic rats in this trial.

Figure 3b: Retinal PAS and hematoxylin staining in group C2 diabetic rats fruit.

Figure 4a: Results of fundus photography of Group C3 diabetic rats in this trial.

Figure 4b: Results of PAS and hematoxylin staining in the retina of group C3 diabetic rats.

Figure 5a: Results of fundus photography of Group C4 diabetic rats in this trial.

Figure 5b: Results of PAS and hematoxylin staining in the retina of group C4 diabetic rats in this test.

The above and other objects, features and advantages of the present invention will become more <RTIgt;"Gingersample" refers to the rhizome of Zingiber zerumbet , which is often used as a seasoning and flavoring ingredient. It can be used to treat symptoms such as toothache, indigestion, diarrhea, and also promote blood circulation.

The red ball ginger extract of the invention is extracted from a red ball ginger sample, and the flavonoid compound contained in the red ball ginger sample is easily released during the extraction process by impregnation with rice vinegar and high temperature cooking, so that the red Ginger extract is rich in flavonoids, which in turn can be used to repair damage to retinal cells caused by hyperglycemia.

In detail, the red ball ginger extract of one embodiment of the present invention can be obtained by a method comprising the steps of: providing a red ball ginger sample as described above, immersing the red ball ginger sample in rice vinegar, and continuing The red ball ginger extract can be obtained by cooking at a high temperature to obtain a sample of vinegar red ball ginger and extracting the vinegar red ball ginger sample.

The red ball ginger sample can be immersed in rice vinegar after removing impurities on the surface thereof to avoid impurities on the surface of the red ball ginger sample from affecting the subsequent steps. In this embodiment, The red ball ginger sample is placed in a closed container to prevent foreign matter from entering during the impregnation process, so that the rice vinegar can be smothered into the red ball ginger sample to form a sample of the impregnated red ball ginger. For example, in this embodiment, fresh roots of 50 kg of red ball ginger are obtained, washed and removed, and sliced and dried to obtain a 5.3 kg sample of red ginger (water content: 89.4%), and 5 kg of red ball ginger is taken. The sample was immersed in 5 L of rice vinegar for 8 hours to obtain the impregnated red ball ginger sample.

Continue to place the impregnated red ball ginger sample in a confined space (for example, a pressure cooker), and cook at a temperature of 95-105 ° C and a pressure of 1.5-2.5 kg/cm ² for 30 minutes until the impregnated red ball ginger The appearance of the sample is dark brown, and it can be judged that the sample of the impregnated red ball ginger has been vaporized, that is, a sample of vinegar red ball ginger is obtained. The vinegar red ball ginger sample can be further freeze-dried to reduce the water content of the vinegar red ball ginger sample to less than 10% to facilitate the preservation of the vinegar red ball ginger sample.

The vinegar red ball ginger sample can be further extracted by an extraction solvent to obtain the red ball ginger extract. The extraction solvent may be selected to be pure water or a 95% by volume ethanol solution, so that the flavonoids rich in the vinegar red ginger sample are released into the extraction solvent. The red ball ginger extract was obtained. In this embodiment, 500 grams of vinegar red ball ginger sample is taken, and 1 liter of pure water or the ethanol solution is refluxed at 70-90 ° C for 8 hours, and the extraction is repeated 3 times, and finally by vacuum filtration, The mixture was concentrated under reduced pressure and lyophilized to obtain the red ginger extract.

The red ball ginger extract of the invention can effectively repair the damage of retinal cells caused by hyperglycemia, and thus can be used as an active ingredient for improving diabetic retinopathy, for example, the red ball ginger extract can be applied to prepare and improve diabetes. A drug or health food for retinopathy, the red ball ginger extract is combined with a pharmaceutically acceptable carrier or excipient to form a pharmaceutical composition, wherein the red ball ginger extract can be prepared for any convenient consumption. The formula, such as a tablet, a capsule, a powder, a granule or a liquid, or the like, or the red ball ginger extract is combined with other foods or beverages, and is administered to the organism in an edible form for oral administration.

Further, the red ball ginger extract can be administered to a desired individual, for example, 600 mg of red ball ginger extract per kilogram of the desired individual body weight per day, and the desired individual is continuously administered for 28 days. The active ingredient such as a flavonoid can act in the desired individual to improve the diabetic retinopathy of the desired individual.

In order to confirm that the red ball ginger extract of the present invention does have an activity of improving the phenomenon of diabetic retinopathy, the following tests were carried out:

(A) Preparation of red ball ginger extract and detection of active ingredient content

Please refer to Table 1, the preparation method of each group of red ball ginger extract is as described above, each 500 grams of red ball ginger sample or vinegar red ball ginger sample, extracted with 1 liter of extraction solvent, after After filtration, concentration and lyophilization, each group of red ball ginger extract was obtained.

The content of flavonoids was determined by using quercetin as a standard, and its concentration and absorbance were used to prepare a standard curve. Aqueous aluminum chloride solution (2%, 1 mL) was added to the flavin standard (concentration range: 1 to 100 mg/mL, 1 mL) and the red ball ginger extract of each group (concentration: 1 to 100 mg/mL, 1 mL). In the case of standing at room temperature for 10 minutes, measuring the absorbance at a wavelength of 430 nm, and pushing back the content of the flavonoids contained in each group of red ginger extract according to the absorbance measurement result ( The content of quercetin contained in grams of red ball ginger extract is used as a benchmark. It is recorded in Table 2.

Table 2, the content of flavonoids such as red ball ginger extract in each group of the test.

Referring to the results of the above test, according to the above test results, the red ball ginger sample is immersed in rice vinegar, and then subjected to high temperature cooking, so that the flavonoid compound contained in the red ball ginger sample is easily released during the extraction process. It is dissolved in the extraction solvent (i.e., pure water or the alcohol solution) to obtain a red ball ginger extract rich in flavonoids.

(B) Survival effects on retinal cells caused by hyperglycemia

This experiment used retinal cells (ARPE-19cells) purchased from the Food Industry Development Institute. The retinal cell line was cultured with 1.2 g/L sodium bicarbonate and 2.5 mM _L -glutamate ( _L- Glutamine). ), 15 mM 4-hydroxyethylethanesulfonic acid (HEPES), 0.5 mM sodium pyruvate and 10% fetal bovine serum (FBS) in DME/F-12 (1:1) medium and placed in The fresh culture solution was changed every two days in an incubator at 37 ° C, a carbon dioxide concentration of 5%, and a humidity of 95%.

When the retinal cells are subcultured, the culture solution and the cells are centrifuged at 1000 rpm for 5 minutes, the supernatant is removed, a new culture solution is added, and the number of cells maintained in the 10 cm culture dish is about 1×10. ⁵ to 1 × 10 ⁶ cells/mL.

When the test is carried out, the 10 cm culture dish of the above retinal cells which has been 8 to 9 minutes old can be taken out, the discolored culture medium is removed, the cells are washed once with 8 mL of PBS buffer solution, and trypsin/ethylenediamine acetic acid is added. (Trypsin/EDTA) After 1~3 minutes of reaction, gently shake the culture dish to make the cells fall off the wall, add the warmed culture medium, and gently pour the cells. Disperse, then place the cell-containing culture solution into a centrifuge tube, centrifuge at 1500 rpm for 10 minutes, remove the supernatant, then add the serum-containing medium to disperse the cells evenly, and take 20 μL of the cells, add 20 μL 0.04 Trypan Blue was stained, placed in a cell counter, and the number of viable cells was counted under a microscope. The cell viability must be no less than 85% before subsequent experiments can be performed.

The concentration of the above retinal cells was adjusted to 1×10 ⁵ cells/mL with a serum-containing culture solution, and 100 μL of each retinal cell was seeded in a 96-well plate, so that each well contained 1×10 ⁴ cells at a temperature of 37° C. Incubate overnight in an incubator with a carbon dioxide concentration of 5%.

After 24 hours of incubation, as shown in Table 3, add glucose aqueous solution (to a final concentration of 6 mM) and 100 μL of red ball ginger extract (concentration of 1 mg / mL, dissolved in DMSO), and mix well, then Incubators with a temperature of 37 ° C and a carbon dioxide concentration of 5% were cultured overnight for 24 hours.

After 24 hours, the culture solution was removed, rinsed with phosphate buffer, and then 100 μL of fresh culture medium containing CCK-8 was added and placed in an incubator at a temperature of 37 ° C and a carbon dioxide concentration of 5% for 2 hours. After 5 minutes, the absorption of each slot at 450 nm is detected. The survival rate was calculated from the absorbance values of the control group without any drug treatment and is recorded in Table 4. The conversion formula for survival rate: "absorbance value of each group" / "absorbance value of control group without any drug treatment" × 100%.

Please refer to Table 4, according to the above test results, the red ball ginger extract extracted from the vinegar red ball ginger sample (ie, group B5, B6) does have the damage of retinal cells caused by repairing hyperglycemia. Ability.

(C) Effects on diabetic retinopathy in diabetic rats

In this experiment, Wistar male rats purchased from the National Experimental Animal Center of the National Experimental Research Institute of the Foundation were selected. The rats were all over 8 weeks old and weighed between 160 and 180 grams. The rats were housed in the Animal Center of Darien University of Science and Technology, and maintained in an animal room at room temperature of 25 ± 1 ° C. The light time and dark time were each 12 hours. The rats were free to eat and drink.

After 72 hours of fasting the rats, streptozotocin (STZ) was administered to the rats by intraperitoneal injection (administered at a dose of 60 mg/kg) to induce formation. Diabetic rats, after 72 hours, measured the blood sugar levels of the rats, blood sugar levels higher than 300mg / L, and there are more than three symptoms (eat more, drink more, more Rats with urine can be considered to be successfully induced into type 1 diabetic rats.

The diabetic rats were orally fed with the red ball ginger extract as shown in Table 5, fed twice a day for 28 days, before the experiment (day 0) and after the experiment (day 28). The assessment of intraocular pressure and retinal status of these diabetic rats was measured.

Please refer to the sixth table. After the anesthesia of each group of diabetic rats, the intraocular pressure of each group of diabetic rats on the 0th day and the 28th day was measured, and on the 28th day, the arsenic sulfate was added. After atropine (0.25%) eye drops, each group of diabetic rats was anesthetized with ether, and after 5 minutes, fundus photography of each group of diabetic rats was performed with an ophthalmoscope (MiiS Horus Scope DOC100), and sacrificed for each group of diabetes. In the rat, the retina was sectioned and stained with PAS and periodic acid-Schiff/hematoxylin.

Please refer to the sixth table, the intraocular pressure of the group C0 diabetic rats continued to rise, and the intraocular pressure of the red ball ginger extract (ie, group C3, C4) extracted from the vinegar red ball ginger sample was continuously fed. There was a significant decrease, and the trend of decreased intraocular pressure was positively correlated with the content of flavonoids shown in Table 2.

Continued, as shown in Figure 1a, the diabetic rat has retinal micro-vessels in the retina, and blood in the eye seeps out of the vitreous humour, and please refer to Figure 1b for the retina of the diabetic rat. There is also a diffuse fibrinogen-like protein accumulation and coverage, as well as cell edema, hemorrhage, vasodilation and angiogenesis; and continuous feeding from the vinegar red ball ginger sample red The retinopathy of diabetic rats in the ball ginger extract (ie, group C3, C4) has a mildening phenomenon, indicating that the red ball ginger extract of the present invention can indeed improve the condition of diabetic retinopathy, that is, by rice vinegar The flavonoids released by impregnation and high-temperature cooking can indeed repair the damage of retinal cells caused by hyperglycemia.

The use of the red ball ginger extract of the present invention is a derivative of the red globin extract rich in flavonoid compound as an active ingredient for improving diabetic retinopathy, and preventing the side effects caused by conventional drugs, and is an effect of the present invention.

While the invention has been described in connection with the preferred embodiments described above, it is not intended to limit the scope of the invention. The technical scope of the invention is protected, and therefore the scope of the invention is defined by the scope of the appended claims.

Claims (2)

The use of a red ball ginger extract for the preparation of a medicament for improving diabetic retinopathy, wherein the red ball ginger extract is administered to a desired organism to improve the diabetic retinopathy of the desired ascending object, wherein The red ball ginger extract is prepared by the following method for preparing red ball ginger extract: a red ball ginger sample is immersed in rice vinegar for 8 hours to obtain a sample of impregnated red ball ginger; the impregnated red ball ginger sample is Cooking at a temperature of 95-105 ° C and a pressure of 1.5-2.5 kg/m ² for 30 minutes to obtain a sample of vinegar red ginger; and extracting the vinegar red ball with an extraction solvent at a temperature of 70-90 ° C The ginger sample was 8 hours, and the extraction solvent was pure water or a 95% by volume ethanol solution to obtain the red ball ginger extract. The use of the red ball ginger extract according to claim 1, wherein the red ball ginger extract is administered to the individual in a dose of 600 mg per kg of the desired individual per day, and is continuously administered. 28 days.