JP4421684B2 - Anti-obesity agent and method for producing the same - Google Patents

Description

【００１７】
表１に示したように、ＳＲの根皮の水エキスに最も強い蔗糖の吸収阻害作用が認められた。一方、吸収阻害作用は蔗糖に対してのみ発揮され、ブドウ糖を負荷した場合には、対照薬として用いたアカルボース同様無効であった。
【００１８】
実施例２
ＳＲ抽出物とアカルボースとの作用強度を比較する実験は、次の方法により行った。２０〜２４時間絶食させたウィスター系雄性ラット(体重１４０〜１６０g)に被験物質を経口投与し、その３０分後に蔗糖(１g／kg)またはブドウ糖(０．５g／kg)を経口投与した。３０分後に無麻酔拘束下(採血時のみ)にて頚静脈より約０．４ml採血を行い、血糖値(mg／dl)を測定した。
【００１９】
【表２】

【００２０】
その結果を表２に示したように、ＳＲの根皮水エキスは、粗抽出物でありながら、単一成分の合成薬であるアカルボースに匹敵する強い作用を有することがわかった。
【００２１】
実施例３
ブドウ糖負荷の場合の過血糖は、ＳＲの根皮の水エキスを投与してもアカルボース同様、抑制効果が認められなかったことから、蔗糖分解酵素α−グルコシダーゼの阻害作用を特長とすることは推定されたが、この点を明らかとするべく、以下の検討を行った。
【００２２】
α−グルコシダーゼ阻害活性の検討は、被験薬物の表３に示す量を溶解した０．１Ｍリン酸緩衝液(pＨ７．０)をＡ液とし、Ａ液０．５mlに基質として０．０１Ｍ p−nitrophenyl−α−Ｄ−glucopyranoside(０．１Ｍリン酸緩衝液(ｐＨ７．０)に溶解)０．２５mlを加え、約５分間予備加温した。これにあらかじめラット空腸粘膜より調製した酵素液(０．０２unit, ０．２％牛血清アルブミンを含む０．０１Ｍリン酸緩衝液(pＨ７．０)に溶解したもの)を同様に予備加温し、その０．０５mlを加えて３７℃で約１５分間インキュベートした。０．２Ｍ炭酸ナトリウム液を０．２ml加えて反応を終了させ、生成したp−nitorophenolの量を４００nmの吸光度から算出した。ブランク液は、炭酸ナトリウム液を加えてから酵素液を加えたものを用いた。薬物濃度は、インキュベーション中の濃度で表した。この結果を表３に示す。５０％抑制濃度(ＩＣ₅₀値)の比較において、アカルボースと同程度、高濃度(５００μg／ml)における最大阻害率においてアカルボースを上回るα−グルコシダーゼ阻害作用を有することが明らかとなった。
【００２３】
【表３】

【００２４】
次に、α−グルコシダーゼ阻害活性とβ−グルコシダーゼ阻害活性の選択性の有無を検討した。β−グルコシダーゼ阻害は、α−グルコシダーゼ阻害試験において、α−グルコシダーゼをβ−グルコシダーゼに、p−nitrophenyl−α−Ｄ−glucopyranosideをp−nitorophenyl−β−Ｄ−glucopyranosideに変えて試験を行った。β−グルコシダーゼは、アーモンド由来のを用いた。その結果を表４に示す。すなわち、ＳＲ水エキスは、α−グルコシダーゼのみを非常に強く選択的に阻害することが明らかとなった。
【００２５】
【表４】

【００２６】
【発明の効果】
本発明によれば、実施例にも示した通り、蔗糖の吸収阻害に基づく優れた肥満防止作用を有する、天然由来の肥満防止剤およびその製造法を提供することができる。[0001]
BACKGROUND OF THE INVENTION
The present invention relates to an anti-obesity agent and a method for producing the same, and an object thereof is to provide a novel anti-obesity agent having an excellent sucrose degrading enzyme inhibitory effect.
[0002]
[Prior art]
Conventionally, naturally-derived antidiabetic drugs such as Gymnema leaves are known, but few have been evaluated for their effectiveness due to pharmacological support. In addition, there are many traditional drugs that are said to be effective for diabetes, but the current situation is that their features of action and effective doses are not fully elucidated.
[0003]
[Problems to be solved by the invention]
The object of the present invention is to provide a natural-derived anti-obesity agent that is effective and safe.
[0004]
[Means for Solving the Problems]
As a result of repeated systematic experimental research on natural resources for improving hyperglycemia symptoms, the present inventor has found that an extract from Salacia reticulata (hereinafter abbreviated as SR) has excellent sucrose-degrading enzymes (particularly, , Α-glucosidase) inhibitory effect was obtained, and the present invention was completed.
[0005]
It is well known that sucrose is broken down into glucose and fructose mainly by α-glucosidase present in the intestinal tract, and it moves into the blood and raises the blood sugar level. There is no such thing as hyperglycemia. Therefore, if the activity of α-glucosidase is inhibited, it is not decomposed into glucose and fructose, and an increase in blood glucose level is suppressed. Moreover, since sucrose is excreted without being decomposed, absorption of saccharides that cause obesity is inhibited, so that it also has an anti-obesity effect. Therefore, the anti-diabetic agent having α-glucosidase inhibition also has an anti-obesity action, and since the SR extract of the present invention is derived from natural products, it can also be incorporated into diet foods.
[0006]
The present invention is to establish an effective method for extracting SR, to clarify the drug efficacy and action characteristics using experimental animals, and to provide an effective and safe natural-derived anti-obesity agent.
[0007]
【Definition of terms】
The term “anti-obesity agent” of the present invention is derived from the effect of inhibiting glycolysis as described above.
[0008]
DETAILED DESCRIPTION OF THE INVENTION
SR is a plant belonging to the asterae family that grows mainly in Southeast Asia. SR can use leaves, bark, root bark, xylem, root xylem, etc., but root bark has an effect of preventing obesity.
[0009]
The solvent used for extraction of the active ingredient of SR is water or an organic solvent. Examples of organic solvents include alcohols such as methanol, isopropyl alcohol, butanol; esters such as ethyl acetate; ketones such as acetone; ethers such as methyl ethyl ether. These water or organic solvents may be used in combination of two or more, and a particularly preferred extraction solvent is water.
[0010]
An example of the method for producing the extract is as follows. After the collected SR site is dried, a solvent is added, and the mixture is heated and extracted for 2 to 4 hours. Then, the extract is filtered, and the filtrate is completely removed under reduced pressure of 50 ° C. or lower to obtain the desired extraction. You can get things. The amount of solvent added to each site varies depending on the site, but about 3 times the amount is appropriate for the site. In animal experiments, this extract is dissolved and used.
[0011]
The anti-obesity agent of the present invention comprises an extract of Salacia reticulata (SR) as an active ingredient. When used as a medicine, an SR extract may be blended together with an appropriate carrier or excipient. When blended in foods or the like, it can be added as it is or in the form of a concentrate or the like.
[0012]
The dosage is 5 to 200 mg / kg, preferably 50 to 200 mg / kg in the form of an extract powder. If it is less than 5 mg / kg, the medicinal effect is not remarkable, and it may be more than 200 mg / kg, but an increase in the effect cannot be expected.
[0013]
【Example】
The following examples illustrate the constitutional action of the anti-obesity agent of the present invention.
[0014]
Example 1
The effect on blood glucose was examined by the following method. That is, the test drug is orally administered to male Wistar rats (body weight 130 to 170 g) fasted for 20 to 24 hours, and 30 minutes later, sucrose and glucose are orally administered to different groups. Hyperglycemic symptoms were induced. The administered sucrose was 1 g / kg and glucose was 0.5 g / kg.
[0015]
After loading with sugar, about 0.4 ml of blood was collected from the jugular vein under anesthesia restraint (only during blood collection) at 30 minutes, and the blood glucose level (mg) was determined by enzymatic method (glucose CII-Test Wako, manufactured by Wako Pure Chemical Industries). / Dl) was measured. The results are shown in Table 1. In Table 1, the normal group represents those administered with water instead of administering the drug and sugar. In addition, those to which no drug was administered even when sugar was loaded were classified as a sugar load group. Further, for comparison, a similar experiment was also performed on a commercially available acarbose (commercially available from Bayer) that was orally administered at 10 mg / kg. In Table 1, N represents the number of individuals tested, and the blood glucose level was expressed as an average value and a standard error.
[0016]
[Table 1]

[0017]
As shown in Table 1, the strongest sucrose absorption inhibitory action was recognized in the water extract of SR root bark. On the other hand, the absorption inhibitory effect was exerted only on sucrose, and when glucose was loaded, it was not as effective as acarbose used as a control drug.
[0018]
Example 2
The experiment comparing the strength of action between the SR extract and acarbose was performed by the following method. The test substance was orally administered to male Wistar rats (body weight 140 to 160 g) fasted for 20 to 24 hours, and sucrose (1 g / kg) or glucose (0.5 g / kg) was orally administered 30 minutes later. After 30 minutes, about 0.4 ml of blood was collected from the jugular vein under anesthesia restraint (only during blood collection), and the blood glucose level (mg / dl) was measured.
[0019]
[Table 2]

[0020]
As shown in Table 2, the root skin water extract of SR was found to have a strong action comparable to that of acarbose, which is a single-component synthetic drug, while being a crude extract.
[0021]
Example 3
It was presumed that hyperglycemia in the case of glucose load is characterized by the inhibitory action of sucrose-degrading enzyme α-glucosidase, since the inhibitory effect was not observed as in the case of acarbose even after administration of the water extract of SR root bark. However, in order to clarify this point, the following examination was conducted.
[0022]
The α-glucosidase inhibitory activity was determined by using 0.1 M phosphate buffer solution (pH 7.0) in which the amount shown in Table 3 of the test drug was dissolved as A solution, and 0.5 mL of A solution as a substrate with 0.01 M p- 0.25 ml of nitrophenyl-α-D-glucopyranoside (dissolved in 0.1 M phosphate buffer (pH 7.0)) was added and pre-warmed for about 5 minutes. Similarly, an enzyme solution prepared in advance from rat jejunal mucosa (0.02 unit, dissolved in 0.01 M phosphate buffer (pH 7.0) containing 0.2% bovine serum albumin) was preliminarily heated in the same manner. The 0.05 ml was added and incubated at 37 ° C. for about 15 minutes. The reaction was terminated by adding 0.2 ml of 0.2 M sodium carbonate solution, and the amount of p-nitorophenol produced was calculated from the absorbance at 400 nm. As the blank solution, a solution obtained by adding a sodium carbonate solution and then an enzyme solution was used. The drug concentration was expressed as the concentration during incubation. The results are shown in Table 3. Comparison of 50% inhibitory concentration (IC ₅₀ value) revealed that α-glucosidase inhibitory activity was higher than that of acarbose at the maximum inhibition rate at a high concentration (500 μg / ml), similar to that of acarbose.
[0023]
[Table 3]

[0024]
Next, the presence or absence of selectivity between α-glucosidase inhibitory activity and β-glucosidase inhibitory activity was examined. In the α-glucosidase inhibition test, β-glucosidase inhibition was tested by changing α-glucosidase to β-glucosidase and p-nitrophenyl-α-D-glucopyranoside to p-nitorophenyl-β-D-glucopyranoside. β-glucosidase used was derived from almonds. The results are shown in Table 4. That is, it was revealed that the SR water extract inhibits only α-glucosidase very strongly and selectively.
[0025]
[Table 4]

[0026]
【The invention's effect】
According to the present invention, as shown in the Examples, a naturally-derived anti-obesity agent having an excellent anti-obesity action based on absorption inhibition of sucrose and a method for producing the same can be provided.

Claims (3)

An anti-obesity agent comprising an extract of Salacia reticulata as an active ingredient. A method for producing an anti-obesity agent, comprising extracting each part of dried Salacia reticulata using an organic solvent such as water or alcohol and then concentrating the extract. The method for producing an anti-obesity agent according to claim 2, wherein the site of Salacia reticulata is the root bark.