JP2002114703A - Antioxidant - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide an antioxidant safety, stable and easy to use. SOLUTION: This antioxidant comprises phycobiont or its extract of a unicellular Chlorophyceae such as Chlorella, Scenedesmus or Chlamydomonas or he like cultured in a dark and a heterotrophic condition. The antioxidant is cpable of suppressing production of lipoperoxide in low density lipoprotein(LDL) of serum, harmless and capable of carrying out an ingestion, usable for doods, medicines, cosmetics and their additives.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a material which has an antioxidant effect and can be widely used in foods and pharmaceuticals.

[0002]

2. Description of the Related Art In general, for example, tocopherols (vitamin E) are well known as antioxidants derived from natural products, and are currently widely used as pharmaceuticals, foods or additives thereof. However, since tocopherols are fat-soluble and the substance itself is oily, its use is limited to addition to oils and fats, gelatin capsules and the like. In addition, since a substance exhibiting such an antioxidant action itself is extremely easily oxidized by air, it is usually difficult to store it stably, and therefore, a product sealed in a gelatin capsule is widely used. .

[0003]

DISCLOSURE OF THE INVENTION The present invention is intended to solve the problems of antioxidants such as tocopherols and to make the dried powder usable as foods, pharmaceuticals and additives.

[0004]

Means for Solving the Problems As a result of research, the present inventor has
The inventors have found that algal cells cultured with Chlorella vulgaris E25 under heterotrophic conditions in the dark have strong antioxidant power and suppress the production of lipid peroxide, and based on this finding, complete the present invention. Reached.

[0005] The present invention relates to an antioxidant comprising a single-cell green alga alga body or an extract thereof cultured in a dark place under heterotrophic conditions.

[0006] Unicellular green algae that can be cultured under heterotrophic conditions in the dark include, for example, Chlorella
a), Chlamydomonas
s), and those belonging to Scenedesmus.

A preferred embodiment is Chlorella vulgaris E
25, which is currently industrially cultured and marketed under heterotrophic conditions in a completely dark place so that photosynthesis does not take place (JP-B-58-29047, JP-A-9-2386).
94).

The cultured and collected algal cells of the single-cell green algae of the present invention may be killed or not killed, or may be used as such or in the form of a dried product, or an extract with water or an aqueous solvent, or a mixture of methanol, ethanol, hexane, and the like. It can be used as an antioxidant in the form of an extract with an organic solvent.

The antioxidant contained in the single-cell green algae of the present invention is extremely stable even in dry powder.

Considering the reason why chlorophyll is stable, chlorophyll is stable in the dry powder of the single-cell green algae of the present invention, even though it is an extremely unstable natural organic pigment compound, and it is stable for several years in the dark. Even after the above, the content did not decrease, and almost no decomposition was performed. Chlorophyll is also susceptible to oxidative degradation, so oxidative degradation is hardly performed in the dried single-cell green algae of the present invention. May contain antioxidants in large amounts and in a stable form. This is supported by the extremely excellent antioxidant effect on human serum low-density lipoprotein (LDL) in the tests of the examples.

The present invention is extremely useful as an antioxidant which is safe, edible and stable even in the form of powder, and is useful as a food, drug, quasi-drug, cosmetic, and an additive thereof. It is intended to be widely used.

[0012]

The photosynthetic reaction does not take place in unicellular green algae cultured under heterotrophic conditions in the dark. Therefore, the antioxidant prepared in the alga body remains without being consumed in preparation for the oxidation reaction accompanying the reaction. The present invention utilizes the antioxidant.

[0013]

EXAMPLES The antioxidant effect of Chlorella vulgaris E-25 (E-25) on serum low density lipoprotein (LDL) was tested.

(1) Sample Chlorella vulgaris E-25 dried powder was added to distilled water at 10%.
(W / v) Suspended, heated on a boiling water bath for 20 minutes, extracted with hot water, centrifuged supernatant (E-25-W), chlorella.
Bulgaris E-25 dried powder in methanol at room temperature 10%
(W / v) Suspended and extracted by stirring for 10 minutes, and a centrifugal supernatant (E-25-M) was prepared as a sample.

In order to compare the antioxidant effect, a control group (Control) without addition of a sample and α-tocopherol (VE), whose antioxidant effect is known, are used.
A test plot containing 80 μmol was set.

(2) Method The concentration of serum low-density lipoprotein (LDL) fractionated and purified from healthy adult serum by specific gravity ultracentrifugation was desalted using a Sephadex G-25 column (Pharmacia PD-25). LD of each sample
The L concentration and the amount of protein contained in the micro BC4P
protein Assay Reagent Kit
(Manufactured by PIECE) and made uniform. In this example, the LDL concentration was adjusted to 200 μg
In 1 ml of the solution adjusted to / ml, the sample adjusted as described above was added to a final concentration of 50-fold dilution, and 5 μmol of copper sulfate was added as an oxidizing agent, followed by incubation at 37 ° C. for 2.5 hours. . To this was added 0.1 mmol of EDTA to stop the oxidation reaction, and the mixture was desalted with a Sephadex G-25 column (PD-25 manufactured by Pharmacia).

The degree of oxidation is determined by the amount of lipid peroxide (LPO),
The measurement was performed using a determiner-LPO kit (manufactured by Kyowa Medix). [This measurement technique utilizes the phenomenon in which a derivative of methylene blue reacts with hydroperoxide (lipid peroxide) using hemoglobin as a catalyst to form equimolar methylene blue. It quantifies oxidized lipids. ]

1 ml of LPO pretreatment solution (containing ascorbate oxidase, lipoprotein lipase, etc.) is added to 0.1 ml of the sample, mixed, and allowed to stand at 30 ° C. for 5 minutes. L
After adding and mixing 2 ml of a PO color developing solution (containing methylene blue derivative, MCDP, hemoglobin, etc.), the mixture is allowed to stand at 30 ° C. for 10 minutes. The amount of methylene blue generated in this way is 6
Quantification was performed using absorbance at 75 nm, and the amount of lipid peroxide was calculated from the value using the following formula. 0.1 ml of aqueous solution of cumene hydroperoxide (50.0 nmol / ml) was used as a standard solution. Lipid peroxide amount (nmol / ml) = Es / Estd x 5
0.0 Es: absorbance of sample Estd: absorbance of standard solution The results are shown in FIG.

(3) Results and Discussion As apparent from FIG. 1, the amount of lipid peroxide was extremely large in the control group where no sample was added, and the oxidation was suppressed in the test group where vitamin E was added. It was confirmed that it was. In the test group to which the above sample was added, remarkable oxidation suppression was observed as compared with the control group, and it was found that this oxidation suppression effect was particularly included in the hot water-extracted samples. Vitamin E used in this experiment was a high-purity (99% or more) reagent.
Chlorella vulgaris E-25 dry powder is extremely high because about 50% of the effect of μmol was observed in 10% (w / v) hot water extract of Chlorella vulgaris E-25 dry powder. It can be said that it has an antioxidant effect. The reason why the effect of the methanol extract is not as remarkable as that of the hot water extract may be due to the low antioxidant activity of pigments such as chlorophyll and carotenoids present as the main components of the methanol extract against LDL.

[Brief description of the drawings]

FIG. 1 In the tests in Examples,
It is a bar graph which measured and compared the amount of lipid peroxide in serum low density lipoprotein (LDL).

Claims (8)

[Claims] An antioxidant comprising a single-cell green alga alga body or an extract thereof cultured in a dark place under heterotrophic conditions. 2. The antioxidant according to claim 1, wherein the single-cell green algae is Chlorella, Scenedesmus or Chlamydomonas. 3. The antioxidant according to claim 1, wherein the single-cell green algae is Chlorella vulgaris. 4. The single-cell green algae is Chlorella vulgaris E.
The antioxidant according to claim 1, 2 or 3, which is 25. 5. A lipid peroxide production inhibitor comprising a single-cell green algae alga body or an extract thereof cultured in a dark place under heterotrophic conditions. 6. The lipid peroxide production inhibitor according to claim 5, wherein the single-cell green algae is Chlorella, Scenedesmus or Chlamydomonas. 7. The lipid peroxide production inhibitor according to claim 5, wherein the single-cell green algae is Chlorella vulgaris. 8. The single-cell green algae is Chlorella vulgaris E.
The lipid peroxide production inhibitor according to claim 5, 6 or 7, which is 25.