JPH0565491B2 - - Google Patents

Description

[Detailed description of the invention]

[Industrial Application Field] The present invention relates to an anti-cancer active substance, and more particularly to an anti-cancer active substance whose active ingredient is a precipitate produced by adding an organic solvent to an aqueous extract of wakame sporophylls (bud stock). . [Prior Art and Problems to be Solved by the Invention] Many substances having anticancer activity have been known so far, including plant-derived polysaccharides. However, these have problems such as complicated manufacturing processes, low yields, low anticancer activity, and side effects, and so far no fully satisfactory products have been obtained. Previously, only the leaves of collected seaweed were used as food, and the sprouts were discarded. [Means for Solving the Problems] Therefore, the present inventors have made repeated studies to find anticancer active substances from plants, and found that the above-mentioned wakame bud stock contains a large amount of spores, and this part contains anticancer active substances. Focusing on the presence of active substances, as a result of research, it was discovered that polysaccharides obtained from wakame sporophylls (bud stocks) have anticancer activity, and based on this knowledge, the present invention was completed. . That is, the present invention provides an anticancer active substance whose active ingredient is a precipitate obtained by adding methanol or ethanol to an extract obtained by adding water to the sporophylls of wakame seaweed. The above precipitate, which is an anticancer active substance, can be obtained from sporophylls of wakame seaweed, for example, by the following method. Sporophylls of wakame are immersed in water for several to several tens of hours to obtain an extract, and if necessary, the extract is pulverized by a conventional method. When methanol or ethanol is added to the supernatant obtained by dissolving the extract or the extract powder in distilled water and performing solid-liquid separation such as centrifugation, a white thread-like precipitate is immediately formed. Next, this precipitate is subjected to solid-liquid separation such as centrifugation to separate it from the supernatant. Although it is possible to use the precipitate as it is as an anticancer active substance, it is preferable to dissolve it in distilled water, add methanol or ethanol again, leave it at a low temperature for a specified period of time, and then perform solid-liquid separation to separate the precipitate. Collect and dry in a heat insulator, etc. Water ranging from cold water to hot water can be used for water extraction of wakame sporophylls.The main components of the extract obtained by water extraction are polysaccharides (about 80%), as well as proteins (about 7%) and lipids (about 2%) and vitamins (A,
B ₁ , B ₂ , B ₆ ), contains minerals. There is no particular restriction on the amount of methanol or ethanol used for purifying the polysaccharide, but an amount equivalent to that of the extract is suitable. Polysaccharides partially purified with methanol or ethanol are water-soluble, and analytical tests show that
It contained 0.05% fucoidin and 0.13% uronic acid. Note that fucoidin was hydrolyzed with 1N sulfuric acid in a boiling bath for 2.5 hours, then fucose was measured by the Gibbons method, and the value was multiplied by 0.9, and uronic acid was calculated as glucuronic acid by the carbazole sulfuric acid method. In addition, this polysaccharide tested positive for carbohydrate stability using the Anthrone sulfuric acid method, and its infrared absorption spectrum is as shown in FIG. The polysaccharide according to the present invention has the effect of activating macrophages, causing Lewis lung cancer and
AKR It has anticancer activity against T-cell leukemia, etc. Additionally, this anticancer activity is thermostable;
No change in efficacy is observed even when the polysaccharide aqueous solution is heated to 100℃. Furthermore, no cytotoxicity of the polysaccharide was observed in vitro even at a high concentration of 260 μg/ml. [Example] Next, the present invention will be explained with reference to Examples, but the present invention is not limited thereto. Production example: After immersing wakame sporophylls in water at a temperature of about 2 to 10°C, 2 g of powder obtained by spray drying the extract was dissolved in 50 ml of distilled water, stirred at room temperature, and then centrifuged (10,000 rpm, 20 minutes). ) to obtain a supernatant. Add 50 ml of 95% ethanol to this supernatant, and centrifuge the white thread-like precipitate (10,000 rpm, 30
The supernatant liquid was separated from the supernatant liquid in 1 minute). After dissolving this precipitate (about 0.26 g as dry weight) in 25 ml of distilled water, 25 ml of 95% ethanol was added, and the mixture was left at 4° C. for 20 hours. Thereafter, centrifugation (10,000 rpm, 30 minutes) was performed to separate the supernatant and precipitate. The obtained precipitate was dried in a heat insulator at 37° C. for 2 days to obtain a precipitate (polysaccharide) with a dry weight of about 0.2 g. Example 1 Lewis lung cancer (solid tumor) obtained from the National Cancer Institute and maintained by the inventors in allogeneic mice (C57BL/6 strain) was injected into 10 ml of
Shredded in Hanks liquid, No. 80 screen and No. 21
A cell suspension was made through a syringe needle. 0.2 ml of the suspension (2 to 4 x ¹⁰⁵ cancer cells) was injected and transplanted intraperitoneally into allogeneic C57BL/6 or allogeneic DBA/2 mice (average weight 20 g). Starting from the day after the cancer cells were transplanted, a predetermined amount of the drug was intraperitoneally injected into the mice every day or every other day for a total of 10 or 6 times, and the average survival days, number of surviving mice, and survival rate were measured. The results are shown in Table 1 along with the results for standard chemical immunopotentiators.

【table】

[Table] Example 2 In Example 1, instead of injecting cancer cells into the peritoneal cavity of a mouse and administering a prescribed drug after transplantation, a prescribed drug was previously injected into the mouse peritoneal cavity prior to injecting and transplanting cancer cells. Except for this, the same procedure as in Example 1 was carried out. The results are shown in Table 2.

[Table] Example 3 Allogeneic and syngeneic mice were prepared in the same manner as in Example 1.
C57BL/6, allogeneic mouse DBA/2 and
After injecting and transplanting Lewis lung cancer cells into the peritoneal cavity of BALB/c, administering the polysaccharide of the present invention or a standard immunostimulant together with the anticancer drug cyclophosphamide,
We investigated the anticancer effects of polysaccharides on Lewis lung cancer in immunosuppressed mice. That is,
When cyclophosphamide is administered in a reduced dose, it has no anticancer effect and on the contrary promotes the progression of Lewis lung cancer. However, when the polysaccharide of the present invention was administered in such a state where the immune system was suppressed, cyclophosphamide It was confirmed that the cancer growth promoting effect of amide was suppressed and that it had anticancer effects. However, no anticancer effect was observed for isoprinosine. Example 4 In Example 3, 2-, which is selectively toxic to macrophages, was used instead of cyclophosphamide.
Chloradenosine was administered to test whether the anticancer effect of the polysaccharide of the present invention was affected. As a result, when the cancer cell killing ability of macrophages was eliminated by administration of 2-chloroadenosine, the polysaccharide of the present invention no longer exhibited anticancer effects. Similarly, isoprinosine also showed no anticancer effects. From the above results, it was revealed that the polysaccharides and isoprinosine of the present invention require macrophages having normal functions in the host in order to exert anticancer effects. Example 5 Cyclosporin, known as a powerful anti-rejection agent used after organ transplantation, has the effect of hastening the spontaneous onset of AKR T-cell leukemia. However, when the polysaccharide of the present invention was administered together with cyclosporin, it was found to have the effect of counteracting the leukemia-promoting effect of cyclosporin. On the other hand, immunostimulants such as isoprinosine, maleic anhydride divinyl ether copolymer, levamisole, and tyrolone were unable to counteract the effects of cyclosporin. From this, it has become clear that only the polysaccharide of the present invention specifically antagonizes cyclosporin and suppresses its carcinogenic effect. [Effects of the Invention] The anticancer active substance of the present invention activates the host and macrophages to enhance immunity, and is effective against Lewis lung cancer,
AKR Exhibits anticancer effects against T-cell leukemia, etc. Furthermore, there are no side effects such as destruction of normal tissue.

[Brief explanation of the drawing]

FIG. 1 is an infrared absorption spectrum (KBr tablet method) of the polysaccharide (precipitate-2) according to the present invention.

Claims (1)

[Scope of Claims] 1. An anticancer active substance whose active ingredient is a precipitate obtained by adding methanol or ethanol to an extract obtained by adding water to the sporophylls (bud stock) of wakame seaweed. 2. Anticancer active substances activate the host and macrophages to strengthen immunity, and are effective against Lewis lung cancer and AKR T.
- The anticancer active substance according to claim 1, which has anticancer activity against cellular leukemia.