JP2754313B2 - Indene derivative and antibacterial agent containing the same as an active ingredient - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to an indene derivative and an antibacterial agent containing the same as an active ingredient.

[0002]

2. Description of the Related Art It is known that a dried product, a squeezed liquid or an extract of barley sprouts has an antibacterial activity, and monoglycerides of stearic acid or palmitic acid or derivatives thereof are suggested as compounds exhibiting such antibacterial activity. (JP-A-2-270825, JP-A-3-24047)
3).

[0003]

SUMMARY OF THE INVENTION The present invention provides a novel indene derivative which is isolated from pearl sprouts and has excellent antibacterial activity.

[0004]

Means for Solving the Problems Thus, the present inventors have
As a result of diligent research on the antibacterial activity of the dried barley sprouts, the squeezed liquid or the extract, when a predetermined treatment is applied to the barley sprouts, a new indene derivative is isolated, and the indene derivative shows excellent antibacterial activity I found that.

That is, the present invention relates to an indene derivative represented by the following formula 1 and an antibacterial agent containing the same as an active ingredient.

[0006]

(Equation 1)

The indene derivative of the formula 1 is isolated by treating barley sprouts as follows.
First, as described in detail later in the Examples, the seeds of the barley are dark-cultured to obtain barley sprouts. There are no particular restrictions on the varieties of the target barley seeds, such as Tokuda native, Nakazato native, Okayama native, and Kuroishi native. In addition, dark culture can be performed according to general sprouts dark culture conditions. For example, the pearl barley seeds may be dark-cultured at 25 ° C. for about 4 to 8 days. The thus obtained barley sprouts can use all the parts.

[0008] Next, as will be described later in detail in the examples, the above-mentioned barley sprouts, their crushed, freeze-dried, and further crushed products are extracted with a lower alcohol such as methyl alcohol or ethyl alcohol. The extract is subjected to liquid-liquid partition extraction with chloroform, acetone or the like to obtain acetone-soluble simple lipids.

Finally, as described later in detail in Examples, the above simple lipid is repeatedly subjected to chromatographic fractionation treatment to isolate a desired compound. The chromatographic fractionation treatment is preferably performed using a mobile phase having a different polarity, and a combination of gel permeation chromatography and high performance liquid chromatography.

The results of the structural analysis of the compound thus isolated are as follows. (1) molecular weight: 234 (C ₁₃ H ₁₄ O ₄ ) (2) infrared absorption spectrum: 1569, 1712, 3
401 cm ^-1 (3) Nuclear magnetic resonance spectrum ( ¹ H-NMR, δ):
2.01 (3H, s), 3.84 (3H, s), 4.0
0 (3H, s), 5.68 (1H, s), 7.08 (1
H, dd, J = 8.55, 2.67), 7.37 (1
H, d, J = 2.67), 7.57 (1H, d, J =
8.55) (4) Nuclear magnetic resonance spectrum (< ¹³ > C-NMR, [delta]): 3
1.6,55.9,57.1,76.0,99.1,1
09.5, 117.2, 128.7, 129.0, 13
7.0, 160.3, 168.3, 205.0

From the results of the above structural analysis, it was determined that the isolated compound was an indene derivative represented by the formula 1, and was 1-acetyl-1-hydroxy-3,5-dimethoxy-1H-indene.

As will be described later in detail in Examples, the indene derivative represented by the formula 1 exhibits excellent antibacterial activity against bacteria, and is attracting attention as a natural antibacterial agent for use in foods and cosmetics.

[0013]

【Example】

Test Category 1 (Isolation of Indene Derivatives) The harvested barley seeds of the native Tokuda are cultivated in the dark at 25 ° C for 4 days to obtain barley sprouts. Thus, a freeze-dried product was obtained.

6 liters of methyl alcohol was added to 150 g of the lyophilized product, homogenized, left at room temperature for 48 hours, and filtered to obtain an extract. 6 L of methyl alcohol was added to the residue, and an extraction treatment was performed in the same manner to obtain an extract, which was combined with the first extract. Then, the combined extract is heated at 40 to 45 ° C. under reduced pressure to evaporate methyl alcohol, thereby obtaining extract 3
0 g was obtained.

In 30 g of the above extract, methyl alcohol 1 was added.
00 ml and 200 ml of chloroform were added and mixed and stirred.
Further, 60 ml of a 0.8% aqueous potassium chloride solution was added, and the mixture was allowed to stand. The chloroform layer was separated, heated under reduced pressure at 40 to 45 ° C., concentrated to 1 ml, added with 10 ml of acetone and 0.2 ml of a 10% methyl alcoholic magnesium chloride solution, mixed and stirred, and cooled with ice for 1 hour. Thereafter, the mixture was centrifuged to separate an acetone solution of the supernatant. 10 ml of acetone and 10 ml
The mixture was treated in the same manner as above with 0.2 ml of a methyl alcoholic magnesium chloride solution to obtain an acetone solution, which was combined with the first acetone solution. 6. The combined acetone solution is heated at 40 to 45 ° C. under reduced pressure to evaporate the acetone, whereby the acetone is easily soluble simple lipid.
1 g was separated.

[0016] 7.1 g of the above simple lipid was dissolved in chloroform, and gel permeation chromatography was carried out using two pieces of JAIGEL 1H (trade name, manufactured by Nippon Kagaku Kogyo KK, 8.0 mmφ × 500 mm). At this time, chloroform was used as a mobile phase, and the chloroform was allowed to flow at a flow rate of 3.5 ml / min. Detected by RI (refractive index), four fractions showing major peaks were obtained. Using Bacillus subtilis, Saccharomyces cerevisiae and Aspergillus niger as test bacteria, an antibacterial test of each fraction was performed by a paper disc method, and R showing the strongest antibacterial activity was obtained.
t (retention time, the same applies hereinafter) = fraction of 55 to 64 minutes 1.8
g was obtained.

1.8 g of the above fraction was dissolved in methyl alcohol, and the mixture was dissolved in Capcell Pack C ₁₈ (CAPCELL).
PAK _C18 , trade name, manufactured by Shiseido, 20mmφ × 250
mm, the same applies hereinafter) using high performance liquid chromatography. At this time, the reaction was started with methyl alcohol / water = 20/80 (weight ratio) as a mobile phase, and 100/30 minutes later.
A linear gradient of 0 (weight ratio) was performed, and the mobile phase was allowed to flow at a flow rate of 3.0 ml / min. Detected by UV (ultraviolet absorbance, 254 nm, hereinafter the same), 6 fractions showing major peaks were obtained. According to the same antibacterial test as described above, Rt = 34-38 min fraction showing the strongest antibacterial activity 146.
4 mg were obtained.

The above fraction (146.4 mg) was dissolved in methyl alcohol, and subjected to high performance liquid chromatography using Capcell Pack _C18 . At this time, methyl alcohol / water = 50/50 (weight ratio) was used as the mobile phase, and the mobile phase was allowed to flow at a flow rate of 3.0 ml / min. Eight fractions detected by UV and showing major peaks were obtained. According to the same antibacterial test as described above, the high-performance liquid chromatography was again performed on the fraction having an Rt of 30 minutes showing antibacterial activity only against Bacillus subtilis among the test bacteria, using Capcell Pack _C18 . At this time, methyl alcohol / water = 25/75 (weight ratio) was used as the mobile phase, and the mobile phase was allowed to flow at a flow rate of 3.0 ml / min. 1.5 mg of compound with Rt = 23 min was isolated by UV detection.

The results of the structural analysis of the compound thus isolated are as described above, and the compound is 1-acetyl-1-hydroxy-3,5-dimethoxy-1H-
It was inden.

Test Category 2 (Antibacterial Activity of Indene Derivatives) The antibacterial activity was tested by the paper disk method using Bacillus subtilis as a test bacterium. At this time, the following tryptosoy agar medium (1.5% agar) was used.
5 ml was poured into a 90 mm diameter petri dish and solidified to prepare a plate medium. Tryptic soy agar medium (Nissui Pharmaceutical Co., Ltd.) 1.5% peptone, 0.5% soy peptone, NaCl
0.5% and agar 1.5%

The test bacteria were inoculated into 15 ml of the same agar medium as described above, and this was overlaid on the plate medium and solidified. Paper disk (Toyo filter paper, diameter 8m)
m, thin), 20 μl of the isolated compound (10 mg / ml) was dropped on the paper disc (200 μg / paper disc), and after culturing for 24 hours, the diameter of the growth inhibition circle was measured. The average value of five paper disks was taken, and the antibacterial activity was determined by the following equation 2. The antibacterial activity was 24.0 mm, and the isolated compound was tested to show excellent antibacterial activity against bacteria.

[Formula 2] Antibacterial activity (mm) = Diameter of blocking circle (mm)-Diameter of paper disc (8 mm)

[0022]

As is clear, the indene derivative of the present invention described above has an effect of being effective as an antibacterial agent.

──────────────────────────────────────────────────続 き Continuing on the front page (72) Inventor, Yoshimitsu Okamoto 47, Nishitoyama, Nishinasuno-cho, Nasu-gun, Tochigi Kagome-Nasu-Ryo 315 (72) Hideki Sakamoto 47, Kishimaki, Iguchi, Oguchi, Nisasu-cho, Nasu-gun, Tochigi 12 (58) Field surveyed (Int.Cl. ⁶ , DB name) C07C 49/527-49/583 A01N 35/04 A23L 3/3499 A61K 7/40 CA (STN) REGISTRY (STN)

Claims (2)

(57) [Claims] 1. An indene derivative represented by the following formula 1. (Equation 1) 2. An antibacterial agent comprising the indene derivative according to claim 1 as an active ingredient.