JPH07231792A - Lipid reduction active substance - Google Patents

Abstract

PURPOSE: To provide a new physiologically active substance produced by a specific microorganism, having LDL-incorporation promoting action on cultured HEPATOMA G2 cell and useful as a hypolipidemic agent.

CONSTITUTION: The objective substance is new hypolipidemic substances MC-151 (C₂₃H₃₆O₇), MC-152 (C₂₃H₃₆O₆) and MC-153 (C₂₅H₃₈O₇) produced by Sporotrichum pruinosum TF-0404 (FERM P-14056). The physical and chemical properties of MC-151 are as follows. (1) Molecular weight, 424; (2) color reactions, positive to iodine and sulfuric acid and negative to ninhydrin; (3) solubility, insoluble in water, scarcely soluble in chloroform and soluble in methanol, ethanol, ethyl- acetate and benzene; (4) nature, colorless oily substance; (5) specific rotation, a [α]D²³=+20.0 (C=0.02, methanol) ; (6) ultraviolet absorption spectrum, λmax 216 nm (ε=12,000), 258 nm (ε=5,000) and 292 nm (ε=3,000); (7) acidity, weakly acidic.

COPYRIGHT: (C)1995,JPO

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to a novel compound having a lipid lowering action.

[0002]

2. Description of the Related Art Hyperlipidemia is known as a factor associated with arteriosclerotic disease and has been proved to be a risk factor for ischemic disease. Conventionally, various drugs having an effective blood lipid-lowering effect on these pathological conditions have been discovered, but a drug that completely controls blood lipids in the clinic has not yet been found, and the development of a novel lipid-lowering agent Is desired.

[0003]

An object of the present invention is to find a novel substance having a lipid-lowering action from microorganisms isolated from the natural world.

[0004]

Means for Solving the Problems As a result of repeated exploratory research for achieving the above-mentioned object, the present inventors found that the specific microorganism found by the present inventors was HEPATOMA G2 cultured cells (human hepatoma cells; , HEP G2), and a novel physiologically active substance having an LDL uptake promoting action for HEP G2 was produced, and the present invention was completed.

That is, the present invention provides the lipid lowering agents MC-151 and MC-152 having the following physicochemical properties.
And MC-153. No lipid-lowering substance having the same physicochemical properties as this substance is known.

[1] Physicochemical properties of MC-151 (1) HRFAB mass spectrum: measured value; m / z 425.2509 (M + H) ⁺ theoretical value; m / z 425.2511 (C ₂₃ H ₃₇ O ₇ (2) FAB mass spectrum: FAB (+) m / z 425 (M + H) ⁺ FAB (−) m / z 423 (MH) ⁻ (3) molecular weight: 424 (4) molecular formula: C ₂₃ H ₃₆ O ₇ (5) Specific optical rotation: [α] _D ²³ = + 20.0 ° (c = 0.02, methanol) (6) Ultraviolet absorption spectrum: λmax 216 nm (ε = 12000) 258 nm (ε = 5000) 292 nm (Ε = 3000) (Measurement in methanol solution) (7) Infrared absorption spectrum: The result of measurement by Neat method is shown in FIG. 1 (8) ¹ H-NMR spectrum: 400 M in CD ₃ OD.
The result measured at Hz is shown in FIG. 2. (9) ¹³ C-NMR spectrum: 100 in CD ₃ OD
The results measured in MHz are shown in Fig. 3 (10) Solubility in solvent: insoluble in water, sparingly soluble in chloroform, soluble in methanol, ethanol, ethyl acetate and benzene (11) Color reaction: positive; iodine, sulfuric acid Negative; ninhydrin (12) Basic, acidic, neutral: weakly acidic (13) Appearance: colorless oil

[2] Physicochemical properties of MC-152 (1) HRFAB mass spectrum: measured value; m / z 447.2157 (M + K) ⁺ theoretical value; m / z 447.2160 (C ₂₃ H ₃₆ O ₆ (Calculated as K) (2) FAB mass spectrum: FAB (+) m / z 409 (M + H) ⁺ FAB (-) m / z 407 (MH) ^- (3) molecular weight: 408 (4) molecular formula: C ₂₃ H ₃₆ O ₆ (5) Specific optical rotation: [α] _D ²³ = + 10.0 ° (c = 0.02, methanol) (6) Ultraviolet absorption spectrum: λmax 216 nm (ε = 22000) 258 nm (ε = 10000) 291 nm (ε = 5000) (Measured in methanol solution) (7) Infrared absorption spectrum: The result measured by the Neat method is shown in FIG. 4 (8) ¹ H-NMR spectrum: CD ₃ OD + CDCl ₃
The result measured at 400 MHz in the medium is shown in FIG. 5. (9) ¹³ C-NMR spectrum: CD ₃ OD + CDCl ₃
6 shows the result of measurement at 100 MHz in (10) Solubility in solvent: insoluble in water, methanol,
Soluble in ethanol, ethyl acetate, chloroform and benzene (11) Color reaction: positive; iodine, sulfuric acid negative; ninhydrin (12) basic, acidic, neutral: weakly acidic (13) appearance: colorless oil

[3] Physicochemical properties of MC-153 (1) HRFAB mass spectrum: measured value; m / z 473.2505 (M + Na) ⁺ theoretical value; m / z 473.2506 (C ₂₅ H ₃₈ O ₇ (Calculated as Na) (2) FAB mass spectrum: FAB (+) m / z 451 (M + H) ⁺ FAB (-) m / z 449 (MH) ^- (3) Molecular weight: 450 (4) Molecular formula: C ₂₅ H ₃₈ O ₇ (5) Specific rotation: [α] _D ²³ = -130 ° (c = 0.02, methanol) (6) Ultraviolet absorption spectrum: λmax 215 nm (ε = 24000) 258 nm (ε = 9000) 291 nm (Ε = 5000) (Measurement in methanol solution) (7) Infrared absorption spectrum: The result measured by the Neat method is shown in FIG. 7 (8) ¹ H-NMR spectrum: CD ₃ OD + CDCl ₃
The result measured at 400 MHz in the middle is shown in FIG. 8. (9) ¹³ C-NMR spectrum: CD ₃ OD + CDCl ₃
9 shows the result measured at 100 MHz (10) Solubility in solvent: insoluble in water, methanol,
Soluble in ethanol, ethyl acetate, chloroform and benzene (11) Color reaction: positive; iodine, sulfuric acid negative; ninhydrin (12) distinction between basic, acidic and neutral: weakly acidic (13) Appearance: pale yellow oil

The novel lipid lowering substance MC-15 of the present invention
The strains producing 1, MC-152 and MC-153 are strains newly isolated by the present inventors from the natural world, and the name of the microorganism is " Sporotrichum pruino " .
sum TF-0404 ”and microorganism accession number“ FE ”
RM P-1405 6 "has been deposited at the Institute of Biotechnology, Institute of Industrial Science and Technology.

The mycological properties of this strain are shown below. 1) Morphology This strain grows extremely fast on all agar media tested, and spreads over the entire Petri dish at 26 ° C for about 4 days. Especially potato / glucose agar, malt extract agar, Yp
Both growth on Ss agar medium and sporulation are good, and medium growth and sporulation on Miura agar medium are observed.
Observation of a colony formed by this strain on Miura agar medium by culturing at 26 ° C. for 7 days under an optical microscope showed that the hypha had a septum (lacking a clamp), was highly branched, and had a diameter of 1.6. 〜7.0μm, wall thickness is usually from thin wall up to 2.0μm to slightly thick wall, but in some basal hypha, it becomes a thick wall of about 3.0μm, and the wall has large hyphae. Thick-walled hypha is also observed. The conidium formation pattern is that the hyphae are irregularly separated from the singularly-alero type at the tip of a short stalk (1.5 to 18.0 μm x 1.0 to 1.8 μm) branched from the side along the aerial hyphae. There are two types of segmental type that are formed. Aleuro-type conidia are elliptical to subspherical, with a width of 1.8 to 2.8 μm at the base and a slightly protruding cutout, 1 cell, colorless, smooth surface, length 4.0 to 11.0 μm, width Is 2.4 to 5.0 μm. Segmental conidia are mainly cylindrical, but the shape is extremely diverse, such as barrel, lemon, and L-shaped, with 1 cell, colorless, smooth surface, length 3.8-15.0 μm, width 1 .2 to 3.8 μm. In addition, a large number of chlamydospores are formed on the surface of the medium and in the medium, most of which are spherical to subspherical, rarely barrel-shaped, lemon-shaped, gourd-shaped, pear-shaped, atopic or mesenchymal, 1 cell, colorless , The surface is smooth, and the size is spherical, the diameter is 3.0 to 21.0 μm, and other shapes are 8.0 to 20.0 μm in length,
The width is 5.4 to 14.0 μm. Although the culture was extended to 3 weeks, formation of sexual reproductive organs was not observed.

2) Growth state on medium The following Table 1 shows the results of macroscopic observation when the cells were cultured on various mediums at 26 ° C. for 14 days. For the color display, the systematic color names of the JIS Standards Color Book (1985) of the Japanese Standards Association were cited.

[0012]

[Table 1]

3) Physiological Properties Growth temperature range and optimum temperature The strain of this strain is 12 in a Sabouraud liquid medium of pH 6.0.
It grows in the range of ~ 45 ° C, and the optimum temperature is 33 ~ 39 ° C. Growth pH range and optimum pH This strain has a pH of 2 to 7 at 26 ° C. in a YpSs liquid medium.
, And the optimum pH is 4-7. 4) Distinction between aerobic and anaerobic; aerobic

From the above morphological characteristics and culture properties, this strain was found to be Sporotrichu in the subphylum Incomplete.
It was clarified that it is a bacterium belonging to the genus m . A. Sta
lpers, “A revision of the
genus Sporotrichum 』Studie
s in Mycology, No. 24, 1st-10th
As a result of comparison and examination with many known bacterial species reported on page 1 (1984), this strain was found to be Sporotrichum.
pruinosum Gilman & Abbott
This strain is very similar to that of t.
richum pruinosum TF-0404 "
I named it.

MC-151, MC-152 and MC-
The production of 153 is similar to the case of producing a general fermented product, and Sporotri in a medium containing various nutrients.
chum prinosum TF-0404 strain is cultivated under aerobic conditions.

A liquid medium is mainly used as a medium, and a carbon source,
It consists of a nitrogen source and an inorganic salt, and vitamins, precursors and antifoaming agents can be added if necessary, and the pH is adjusted to around 6. As the carbon source, for example, glucose, maltose, dextrin, glycerin, starch and the like are used alone or in combination. Examples of nitrogen sources include yeast extract, peptone, meat extract, soybean powder, corn stee
Liquor, urea, ammonium salt, etc. may be used alone or in combination. Examples of the inorganic salt include monopotassium phosphate,
Magnesium sulfate, sodium chloride, calcium carbonate, etc. may be used alone or in combination. As the defoaming agent, adecanol, a silicon compound or the like can be used.

As the culturing method, aerobic culturing such as shaking culturing, aeration and stirring culturing is suitable, and the pH is 4 to 8, 25 to 3
2 to 4 days at 0 ° C, preferably pH 6 to 7, 26 to 28
Incubate at 4 ° C for 4 days. MC- produced by this culture
In order to isolate 151, MC-152 and MC-153, a general method for collecting a fermentation product may be used.

That is, MC-151 and MC-152 were isolated from the bacterial cells separated by centrifugation or filtration after the culture was completed.
And MC-153 were extracted with an organic solvent such as lower alcohol and acetone, and the extract was concentrated under reduced pressure to remove the organic solvent, and then transferred to a water-insoluble organic solvent such as ethyl acetate, benzene and chloroform. Is concentrated under reduced pressure to give a syrup. This syrup was again dissolved in an organic solvent such as ethyl acetate, benzene, chloroform, acetone, and methanol, and subjected to column chromatography using silica gel, high-performance liquid chromatography packed with silica gel ODS for reverse phase partition, and gel filtration column chromatography. By attaching it, MC-151, MC-152 and MC-153 can be purified and isolated.

[0019]

INDUSTRIAL APPLICABILITY The compound of the present invention has an LDL uptake promoting activity on HEP G2 cells (human hepatoma cells) and is therefore useful as a lipid lowering agent.

[0020]

EXAMPLES The present invention will be described in more detail with reference to Examples and Test Examples. Example (1) Glucose 2 g, polypeptone 0.5 g, yeast extract 0.3 g, and monopotassium phosphate 0.1 g per 100 ml.
PH 6 consisting of 1 g and 0.05 g of magnesium sulfate
In a 500 ml Erlenmeyer flask containing 100 ml of sterile liquid culture medium of Sporothrichum pruinosum
The TF-0404 strain was inoculated and cultured with shaking at 26 ° C. for 72 hours. Next, using a 50 L culture tank, 0.3 L of the above-mentioned seed culture solution was inoculated into 30 L of sterile medium having the same composition as that of the seed culture, and agitated and aerated culture was performed at 26 ° C. for 96 hours. Next 1000 L
Sterile medium with the same composition as the seed culture, using a volume culture tank
The above seed culture solution (6.0 L) was inoculated into 00 L and agitated and aerated for 96 hours at 26 ° C.

(2) After completion of the culture, the cells were separated into a supernatant and cells by a centrifuge. The obtained supernatant was adsorbed on 20 L of HP-20, washed with 40 L of purified water, and then eluted with 20 L of acetone. Next, the acetone eluate was concentrated under reduced pressure to remove acetone, and the residue was extracted 3 times with 10 L of ethyl acetate. The ethyl acetate extraction sections were combined, dehydrated with anhydrous sodium sulfate, and then concentrated under reduced pressure to dryness to obtain 57.8 g of a brown syrup.

(3) This brown syrup was mixed with chloroform 2
13 of silica gel [Kieselgel-60 (trade name, manufactured by Merck Ltd.)] dissolved in 00 ml and prepared with chloroform
It was adsorbed on a 00 ml column. Chloroform 2000m
After washing with l, chloroform-methanol (98: 2-8
5:15), eluting sequentially with a mixed solvent, 1 fraction 1
Fractionation of 000 ml was performed. These fractions were designated as Fr1 (mixed solvent ratio 92: 8), Fr2 and Fr3 (mixed solvent ratio 85:15) and concentrated under reduced pressure to dryness to give 3.32 g, 2.98 g and 2.51 g of brown substance. Got

(4) Fr3 obtained by silica gel column
Was dissolved in a small amount of methanol, and gel filtration was performed with methanol using a 310 ml column packed with Sephadex LH20 (trade name, manufactured by Pharmacia) prepared with methanol. The active fractions were collected to obtain 1.1 g of brown substance.

(5) The brown substance of the preceding paragraph was dissolved in methanol, and this solution was subjected to preparative medium pressure liquid chromatography using 55% methanol as a mobile phase [apparatus used; KP-6H manufactured by Kusano Kagaku Kikai Seisakusho; column: YMC -ODS-AQ12
Using 0A (50φ × 350mm), UV absorption 258nm
While monitoring at 1, flow rate 14.8 ml / min, 1
The peak eluting at 09 minutes was collected. After separating the obtained fractions and concentrating under reduced pressure to remove methanol,
It was extracted twice with half the volume of ethyl acetate. The ethyl acetate extraction sections were combined, dehydrated over anhydrous sodium sulfate, and concentrated under reduced pressure to dryness to obtain 61.3 mg of a brown substance.

(6) 5 ml of the brown substance of the preceding paragraph was added to methanol
And preparative high performance liquid chromatography using 60% methanol as a mobile phase [apparatus used: Waters M600; column: YMC-ODS-120T].
(10φ × 250 mm)], and UV absorption 258 nm
Monitored at 16.8 at a flow rate of 4.6 ml / min.
The peak eluting in minutes was collected. The separated fractions were combined, concentrated under reduced pressure to remove methanol, and then extracted twice with half the amount of ethyl acetate. The ethyl acetate extraction sections were combined, dehydrated with anhydrous sodium sulfate, and then concentrated under reduced pressure to dryness to obtain 20.0 mg of MC-151 as a colorless oily substance. ,

(7) Further, Fr1 obtained above was dissolved in a small amount of methanol, and packed with Sephadex LH20 (trade name, manufactured by Pharmacia) prepared with methanol 31.
Gel filtration was performed with methanol using a 0 ml column. The active fractions were collected to obtain 448 mg of brown substance.

(8) The brown substance of the preceding paragraph was converted to methanol 7.5.
Preparative high performance liquid chromatography using a solution of 65% methanol as the mobile phase [Dissolved device: Waters M600; Column: YMC-ODS-AQ]
(20φ × 250 mm)] and UV absorption 258 nm
The peak eluted at 17.8 minutes at a flow rate of 15 ml / min was collected while monitoring the above. The separated fractions were combined, concentrated under reduced pressure to remove methanol, and then extracted twice with half the amount of ethyl acetate. The ethyl acetate extraction sections were combined, dehydrated with anhydrous sodium sulfate, and concentrated under reduced pressure to dryness.
27.6 mg of a brown substance was obtained.

(9) 5 ml of the brown substance of the preceding paragraph was added to methanol
And preparative high performance liquid chromatography using 60% methanol as a mobile phase [apparatus used: Waters M600; column: YMC-ODS-AQ (1
0φ × 250 mm)] and the peak eluted at 26.1 minutes at a flow rate of 4.6 ml / min was collected while monitoring the UV absorption at 258 nm. The separated fractions were combined, concentrated under reduced pressure to remove methanol, and then extracted twice with half the amount of ethyl acetate. The ethyl acetate extraction sections were combined, dehydrated with anhydrous sodium sulfate, and concentrated under reduced pressure to dryness to obtain 11.5 mg of MC-152 as a colorless oily substance.

(10) Further, a small amount of Fr2 obtained above was dissolved in methanol, and the mixture was filled with Sephadex LH20 (trade name, manufactured by Pharmacia) prepared with methanol.
Gel filtration was performed with methanol using a 60 ml column. The active fractions were collected to obtain 677 mg of a brown substance.

(11) The brown substance of the preceding paragraph was added to methanol 10
Preparative high performance liquid chromatography using 60% methanol as the mobile phase [Dissolved device: Waters M600; column: YMC-ODS-AQ]
(20φ × 250 mm)] and UV absorption 258 nm
The peak eluted at 12.6 minutes at a flow rate of 15 ml / min was collected while monitoring the above. The separated fractions were combined, concentrated under reduced pressure to remove methanol, and then extracted twice with half the amount of ethyl acetate. The ethyl acetate extraction sections were combined, dehydrated with anhydrous sodium sulfate, and concentrated under reduced pressure to dryness.
20.0 mg of MC-153 was obtained as a pale yellow oily substance.

Test Example (LDL using HEP G2 cells
Uptake promoting action) (Sample) MC-151, MC-152 obtained in Examples
Each of 2.0 mg and MC-153 was dissolved in ethanol to prepare a target concentration. (Test cells) HEP G2 cells (human hepatoma cells) (Used culture medium) DMEM (including 10% FBS)

(Test method) 4 × 10 5 HEP G2 cells were used.
^The culture solution adjusted to a concentration of ⁵ / ml was used to
0.5 ml each was dispensed to a 4-well plate (manufactured by Corning) and cultured at 37 ° C. in a 5% carbon dioxide incubator for 48 hours. Then, 10μ of the sample diluted in advance to the target concentration
D containing 10% LPDS (International Bio Inc.)
The medium was replaced with 0.3 ml of MEM, and the cells were further cultured for 24 hours. After adding 1 μg of DiI-LDL (Funakoshi chemical) and culturing for 4 hours, the medium was removed and the cells were lysed with 0.4 ml of 2 mM SDS solution. 0.3 ml of the obtained cell lysate was diluted 2-fold with purified water and the fluorescence intensity was measured with a fluorometer (Shimadzu RF-5000).
The remaining 0.1 ml was subjected to protein quantification by the Lowry method.
The amount of DiI-LDL incorporated was calculated from the fluorescence intensity by a calibration curve, and the specific activity per unit protein amount relative to the control was expressed as the incorporation-promoting activity. (Results) The results are shown in Table 2.

[0033]

[Table 2]

[Brief description of drawings]

FIG. 1 shows an infrared absorption spectrum of MC-151 measured by a Neat method.

FIG. 2 MC-measured at 400 MHz in CD ₃ OD.
¹ shows the ¹ H-NMR spectrum of 151.

FIG. 3: MC-measured at 100 MHz in CD ₃ OD.
15 shows the ¹³ C-NMR spectrum of 151.

FIG. 4 shows an infrared absorption spectrum of MC-152 measured by a Neat method.

FIG. 5 shows the ¹ H-NMR spectrum of MC-152 measured at 400 MHz in CD ₃ OD + CDCl ₃ .

FIG. 6 shows the ¹³ C-NMR spectrum of MC-152 measured at 100 MHz in CD ₃ OD + CDCl ₃ .

FIG. 7 shows an infrared absorption spectrum of MC-153 measured by a Neat method.

FIG. 8 shows the ¹ H-NMR spectrum of MC-153 measured at 400 MHz in CD ₃ OD + CDCl ₃ .

FIG. 9 shows the ¹³ C-NMR spectrum of MC-153 measured at 100 MHz in CD ₃ OD + CDCl ₃ .

 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Inventor Tomoko Akama 3-24-1, Takada, Toshima-ku, Tokyo Taisho Pharmaceutical Co., Ltd. (72) Inventor Takuya Hamaguchi 3-24-1, Takada, Toshima-ku, Tokyo Taisho Pharmaceuticals Co., Ltd. (72) Inventor Ikuyo Taneoka 3-24-1 Takada, Toshima-ku, Tokyo Taisho Pharmaceutical Co., Ltd. (72) Inventor Akira Kawashima 3-24-1 Takada, Toshima-ku, Tokyo Taisho Pharmaceutical Incorporated (72) Inventor Shigeo Morimoto 3-24-1 Takada, Toshima-ku, Tokyo Taisho Pharmaceutical Co., Ltd.

Claims (3)

[Claims] 1. A hypolipidemic substance having the following physicochemical properties MC-151 (1) HRFAB mass spectrum: measured value; m / z 425.2509 (M + H) ⁺ theoretical value; m / z 425.2511 (C ₂₃ H ₃₇ O ₇ ) (2) FAB mass spectrum: FAB (+) m / z 425 (M + H) ⁺ FAB (−) m / z 423 (M−H) ⁻ (3) Molecular weight: 424 (4) Molecular formula: C ₂₃ H ₃₆ O ₇ (5) Specific rotation: [α] _D ²³ = + 20.0 ° (c = 0.02, methanol) (6) Ultraviolet absorption spectrum: λmax 216 nm (ε = 12000) 258 nm ( ε = 5000) 292nm (ε = 3000) ( measured in methanol solution) (7) infrared absorption spectrum: as shown in FIG. ¹ (8) 1 H-NMR spectrum: as shown in FIG. 2 (9) ¹³ C-NM Spectrum: As shown in Fig. 3 (10) Solubility in solvent: insoluble in water, poorly soluble in chloroform, soluble in methanol, ethanol, ethyl acetate and benzene (11) Color reaction: positive; iodine, negative sulfuric acid; ninhydrin (12) Basic, acidic, neutral: weakly acidic (13) Appearance: colorless oil 2. A hypolipidemic substance MC-152 (1) HRFAB mass spectrum having the following physicochemical properties: measured value; m / z 447.2157 (M + K) ⁺ theoretical value; m / z 447.2160 (C ₂₃ H ₃₆ O ₆ K) (2) FAB mass spectrum: FAB (+) m / z 409 (M + H) ⁺ FAB (−) m / z 407 (MH) ⁻ (3) Molecular weight: 408 (4 ) Molecular formula: C ₂₃ H ₃₆ O ₆ (5) Specific rotation: [α] _D ²³ = + 10.0 ° (c = 0.02, methanol) (6) Ultraviolet absorption spectrum: λmax 216 nm (ε = 22000) 258 nm (ε = 10000) 291nm (ε = 5000) ( measured in methanol solution) (7) infrared absorption spectrum: as shown in FIG. 4 (8) ¹ H-NMR spectrum: as shown in FIG. 5 (9) ¹³ C NMR spectrum: As shown in FIG. 6 (10) Solubility in solvents: Insoluble in water, methanol,
Soluble in ethanol, ethyl acetate, chloroform and benzene (11) Color reaction: positive; iodine, sulfuric acid negative; ninhydrin (12) basic, acidic, neutral: weakly acidic (13) appearance: colorless oil 3. A lipid-lowering substance having the following physicochemical properties MC-153 (1) HRFAB mass spectrum: measured value; m / z 473.2505 (M + Na) ⁺ theoretical value; m / z 473.2506 (C ₂₅ H ₃₈ O ₇ Na) (2) FAB mass spectrum: FAB (+) m / z 451 (M + H) ⁺ FAB (-) m / z 449 (MH) ^- (3) Molecular weight: 450 (4 ) Molecular formula: C ₂₅ H ₃₈ O ₇ (5) Specific rotation: [α] _D ²³ = -130 ° (c = 0.02, methanol) (6) Ultraviolet absorption spectrum: λmax 215 nm (ε = 24000) 258 nm ( ε = 9000) 291nm (ε = 5000) ( measured in methanol solution) (7) infrared absorption spectrum: as shown in FIG. 7 (8) ¹ H-NMR spectrum: as shown in FIG. 8 (9) ¹³ C NMR spectrum: As shown in FIG. 9 (10) Solubility in solvents: Insoluble in water, methanol,
Soluble in ethanol, ethyl acetate, chloroform and benzene (11) Color reaction: Positive; Iodine, Sulfuric acid negative; Ninhydrin (12) Distinction between basic, acidic and neutral: Weakly acidic (13) Appearance: pale yellow oil