JP3026864B2 - New sesquiterpene derivatives - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001] The present invention relates to a novel sesquiterpene derivative, and more specifically, to a general formula having the retroviral protease inhibitory activity:

[0002]

Embedded image Wherein R ¹ is a hydroxymethyl group and R ² is a hydroxyl group, or R ¹ is a hydroxymethyl group or a formyl group and R ² is a hydrogen atom, and a sesquiterpene derivative represented by the formula: Related to manufacturing method.

Wherein R ¹ is a hydroxymethyl group and R ² is a hydroxyl group, or R ¹ is a hydroxymethyl group or a formyl group and R ² is a hydrogen atom. , And a method for producing the same.

[0004] Acquired immunodeficiency syndrome (Aquired)
Immunodefinition Syndrom
e: AIDS) is a human immunodeficiency virus (Human Immunodeficici) which is a kind of retrovirus.
immunovirus (HIV).

[0005] With the progress of research on HIV, an enzyme specific to this virus has been clarified, and an inhibitor for an anti-HIV drug has been developed. In particular, the reverse transcriptase inhibitor azidothymidine (AZT) is one of the few anti-HIV drugs currently in clinical use. However, AZT has strong side effects, and the appearance of a resistance-acquiring virus has been a problem.

[0006] HIV protease is an enzyme essential for HIV maturation infection, and its structure has been almost elucidated, and it has been clarified that it belongs to acidic protease. Therefore, the development of new anti-HIV drugs has been promoted by finding their inhibitors [for example, Sham et al., Biochemical
and Biophysical Research Communications, vol. 17
5, 914-919 (1991), etc.].

Avian myeloblastosis virus (AMV), a kind of retrovirus
Proteases are similar to HIV proteases in terms of their enzymatic properties as well as their substrate recognition cleavage sites, and
It is applicable to screening of protease inhibitors [Skalka, Cell, Vol. 56, 911-913 (1989)]
Etc.].

The present inventors have sought to develop a new anti-HIV drug, seeking an AMV protease inhibitor for a secondary metabolite produced by a microorganism, and developing an AM produced by various microorganisms.
While continuing to search for a V protease inhibitor, it was found that a substance having AMV protease inhibitory activity was produced in a culture of a certain strain belonging to mold, and the active substance was isolated. The present invention has been completed by determining the properties and structure.

The general formula (I) provided by the present invention
Includes the following three compounds.

[0010]

Embedded image

Hereinafter, this compound is referred to as "NF5003-B substance".

[0012]

Embedded image

Hereinafter, this compound is referred to as "NF5003-E substance".

[0014]

Embedded image

Hereinafter, this compound is referred to as "NF5003-F substance".

The physicochemical properties of these compounds are as follows.

[A] A crystal containing 1/2 molecule of methylene chloride of NF5003-B substance has the following characteristics.

(1) Color and shape: white fine crystals (2) Melting point: Decomposes gradually from 215 ° C. and does not show a clear melting point.

[0019] (3) Molecular _{formula: C 23 H 32 O 6 ·} 1 / 2CH 2
Cl ₂ (4) mass spectrum (FAB-MS, matrix
3-nitrobenzyl alcohol): positive ion mode m / z 405 (M + 1) ⁺ negative ion mode m / z 403 (M-1) ^- (5) Specific rotation (24 ° C): [α] _D -71 ° ( c 0.
31, MeOH) (6) Ultraviolet absorption spectrum: shown in FIG.

Maximum absorption wavelength (nm) Molecular absorption coefficient 328 4,700 281 9,410 229 12,200 (7) Infrared external absorption spectrum: shown in FIG.

(8) Thin layer silica gel chromatography solvent Rf Ethyl acetate: hexane (2: 1) 0.03 Toluene: acetone (1: 1) 0.17 (9) Color reaction Negative: Gibbs reaction Positive: 2,4-dinitrophenylhydrazine reagent (10) ¹ H NMR spectrum: shown in FIG.

(11) ¹³ C NMR spectrum: shown in FIG.

(12) Solubility: methanol, acetone,
Soluble in chloroform, acetonitrile and dimethyl sulfoxide, insoluble in water and n-hexane.

(13) Distinguishing between basic, neutral and acidic: acidic substance [b] Crystals containing NF5003-E substance containing 1/2 molecule of methylene chloride have the following properties.

(1) Color and shape: colorless plate-like crystals (2) Melting point: Decomposes gradually from 173 ° C. and does not show a clear melting point.

(3) Molecular formula: C ₂₃ H ₃₂ O ₅ .1 / 2CH ₂
Cl ₂ (4) mass spectrum (FAB-MS, matrix
3-nitrobenzyl alcohol): positive ion mode m / z 389 (M + 1) ⁺ negative ion mode m / z 387 (M-1) ^- (5) Specific rotation (24 ° C.): [α] _D −68 ° ( c 0.
(27, MeOH) (6) Ultraviolet absorption spectrum: shown in FIG.

Maximum absorption wavelength (nm) Molecular absorption coefficient 328 5,430 286 10,800 225 15,100 (7) Infrared absorption spectrum: shown in FIG.

(8) Thin layer silica gel chromatography solvent Rf Ethyl acetate: hexane (2: 1) 0.16 Toluene: acetone (1: 1) 0.41 (9) Color reaction Negative: Gibbs reaction Positive: 2,4-dinitrophenylhydrazine reagent (10) ¹ H NMR spectrum: shown in FIG.

(11) ¹³ C NMR spectrum: shown in FIG.

(12) Solubility: methanol, acetone,
Soluble in chloroform, acetonitrile and dimethyl sulfoxide, insoluble in water and n-hexane.

(13) Distinguishing between basic, neutral and acidic: acidic substance [c] The NF5003-F substance has the following properties.

(1) Color and shape: pale yellow powder (2) Melting point: Decomposes gradually from 180 ° C. and does not show a clear melting point.

(3) Molecular formula: C ₂₃ H ₃₀ O ₅ (4) Mass spectrum (FAB-MS, matrix
3-nitrobenzyl alcohol): positive ion mode m / z 387 (M + 1) ⁺ negative ion mode m / z 385 (M-1) ^- (5) Specific rotation (24 ° C.): [α] _D −40 ° ( c 0.
(27, MeOH) (6) Ultraviolet absorption spectrum: shown in FIG.

Maximum absorption wavelength (nm) Molecular absorption coefficient 352 4,090 301 5,610 245 14,900 (7) Infrared absorption spectrum: shown in FIG.

(8) Thin layer silica gel chromatography solvent Rf Ethyl acetate: hexane (2: 1) 0.46 Toluene: acetone (1: 1) 0.63 (9) Color reaction Positive: Kibbs reaction 2, 4-Dinitrophenylhydrazine reagent (10) ¹ H NMR spectrum: shown in FIG.

(11) ¹³ C NMR spectrum: shown in FIG.

(12) Solubility: methanol, acetone,
Soluble in chloroform, acetonitrile and dimethyl sulfoxide, insoluble in water and n-hexane.

(13) Discrimination between basic, neutral and acidic: acidic substances NF5003-B, NF5003-E and N of the present invention
The F5003-F substance is a compound of the formula (Ia) or (Ib)
And (Ic), a group of compounds having a sesquiterpene skeleton. This group of compounds includes Stachybotrys Complementiy, nov. s
p. K-76 (Stachybotryscomple)
menti, nov. sp. K-76) has structural similarity to the known sesquiterbene compound K-76 which has been isolated from the culture of [Miyazaki et al., Mi.
crobiology and Immunolog
y, Vol. 24, 1091-1108 (1980) and Kaise et al., Journal of Chem.
ical Society Chemical Com
munication, Vol. 1979, 726-7
27 (1979)]. However, NF50O3
-B, NF5003-E and NF5003-F substances are new substances which are distinguished from the known compound K-76 by their molecular formula, physicochemical properties and structural characteristics.

The compound of the formula (I) of the present invention (NF
5003-B, NF5003-E and NF5003-
F) can be produced, for example, by culturing a microorganism having the ability to produce a sesquiterpene derivative represented by the general formula (I) belonging to the genus Stachybotrys in a medium, and collecting the sesquiterpene derivative from the culture.

The search for a microorganism capable of producing the sesquiterpene derivative of the general formula (I) can be carried out, for example, as follows. Synthetic peptide consisting of 10 amino acids (Thr-Phe-Gln-Ala-Tyr-Pr
o-Leu-Arg-Glu-Ala) is hydrolyzed by AMV protease, a culture solution of various microorganisms is added, and the progress of the hydrolysis reaction is detected by high performance liquid chromatography or the like. Compared to the hydrolysis reaction without adding a microbial culture, an active substance is isolated and confirmed from the microbial culture in which the reaction is inhibited, thereby obtaining a microorganism capable of producing the target enzyme inhibitor. be able to.

One of the microorganisms having the ability to produce the sesquiterpene derivative of the general formula (I) thus discovered is Stachybotrys sp. Mer-NF5003 isolated from the soil of Taketomi-cho, Yaeyama-gun, Okinawa Prefecture. (Hereinafter, abbreviated as Mer-NF5003).

The bacteriological properties of Mer-NF5003 are as follows.

1. Growth Forms in Various Media All cultures were performed at 26 ° C., and the growth forms on the agar plate medium were described.

(1) Tuapet agar medium The growth is very slow, and the colony has a diameter of 20 mm after culturing at 26 ° C. for 10 days. The growth of the colonies is thin, almost no aerial mycelium is observed, and the hypha grows only in the medium, but the amount of the hypha is small. The color of the colony is light yellow to light yellow brown, growth of only mycelia is observed, and almost no spores are formed. No sexual reproductive organs, chlamydospores are found.

(2) Malt extract agar medium The growth is very slow, and the colony has a diameter of 15 mm when cultured at 26 ° C. for 10 days. The growth of the colonies is thin, aerial hyphae is slightly observed, and the color of the colony surface is light yellow brown. Sporulation is observed, albeit a little, and a mucous, spherical conidial head develops on the tip of the conidiophore that has emerged from the medium surface. The conidial head is initially colorless, but turns gray-black when mature. No sexual reproductive organs, chlamydospores are formed.

(3) Oatmeal agar medium The growth is moderate, and the colony is 35 mm in diameter after culturing at 26 ° C. for 10 days. The growth of colonies is thin and the growth of aerial hyphae is small, but a large amount of sporulation is observed.More conidiophores do not branch out than the medium surface, they occur independently, and the tip is a mucous spherical conidium at the tip. To settle. The conidial head is initially colorless but turns dark gray when mature. The color of the colony surface is light yellow-brown, but when conidia mature, countless gray-black dots are observed. No sexual reproductive organs, chlamydospores are formed.

2. Morphological properties Many sporulations were observed on oatmeal agar medium,
The conidiophores stand upright from the mycelium on the medium surface.

The conidiophore occurs independently, has no branches, is colorless and has a septum. The size of the conidiophore is relatively small, 35-75 μ × 2.0-2.5 μ, and the tip is thin and the surface is smooth. The apical portion is bulging and exhibits a vesicle-like morphology.

Phyalides colonize 3 to 7 colonies on the apical part at the tip of the conidiophore, have a smooth and colorless club-shaped surface, and have a size of 2.5 to 3.0 μ × 6.0. 5 to 8.0 μ.

The conidium develops from the tip of the infarct, and a large number of oval cells form a spherical mucous conidial head having a diameter of 15 to 25 μm. Conidia are unicellular and colorless at the beginning of development, but become brownish gray when mature and have a slightly rough surface with a size of 3.0 to 3.0.
The size is 4.5 μ × 6.5 to 7.5 μ.

From the above mycological properties, Mer-NF500
The three fungi are fungi belonging to the incomplete fungus (Deuteromycotina) and are described in JAvon Arx “The Genera of Fungi Sporulating.
In Pure Culture, 1974, it was found to be a bacterium belonging to the genus Stachybotrys. GRBisby
“Stachybotrys” Trans. Brit. Mycol. Soc. 26, 133
143, MBEllis "Dematiaceous Hyphmycetes" 1
971, JCGilman “A Manual of Soil Fungi” 195
When compared to the description of known bacterial species with reference to 7, etc., it is similar to Stachybotrys atra, but the color of the conidiophore, showing a form with a swelling like a top capsule at the tip, the size of the needle There was a clear difference in pod shape, conidia size and morphology, etc., and none of the known bacterial species corresponded. Therefore, the bacterium Mer-NF5003 was transformed into Stachybotry sp.
ys sp. Mer-NF5003).

The present inventors have deposited this bacterium as Stachybotrys sp Mer-NF5003 with the Research Institute of Microbial Industry and Technology under the number FERM p-12344.

The culture of Mer-NF5003 can be carried out, for example, as follows. As a nutrient source of the medium, for example, as a carbon source, glucose, sucrose, starch syrup, molasses, dextrin, starch, glycerol, animal oil, vegetable oil, etc. can be used, and as a nitrogen source, soybean powder, wheat germ , Corn stay liquor, cottonseed meal, meat extract, peptone, yeast extract, ammonium sulfate, urea and the like can be used. Others
If necessary, it is effective to add sodium, potassium, calcium, magnesium, cobalt, chlorine, phosphoric acid, sulfuric acid, and other inorganic salts capable of generating ions.

As the culturing method, a culturing method under aerobic conditions is suitable, and the temperature suitable for the culturing is in the range of about 15 to 30 ° C, but in many cases, the culturing is performed at about 28 ° C. NF5
003-B, NF5003-E and NF5003-F
The production of the substance varies depending on the culture medium and culture conditions, but the maximum accumulation usually occurs in 2 to 10 days in both shaking culture and tank culture. NF5003- in culture
B, When the accumulation amount of the NF5003-E and NF5003-F substances reaches the maximum, the culture is stopped, and the target substance is isolated and purified from the culture.

For the collection of the NF5003-B, NF5003-E and NF5003-F substances from the culture, conventional separation means utilizing their properties, for example, solvent oil extraction, adsorption or distribution chromatography, Gel filtration,
A crystallization method or the like can be used alone or in an appropriate combination.

For example, NF5003 accumulated in a culture solution
-B, NF5003-E and NF5003-F substances are adsorbed on a synthetic adsorbent such as Diaion HP-20 (manufactured by Mitsubishi Kasei). Organic solvents that are immiscible with water,
For example, if extracted with ethyl acetate, butanol, etc., NF5
003-B, NF5003-E and NF5003-F
The substance is extracted into the organic solvent layer.

In order to further purify the substances NF5003-B, NF5003-E and NF5003-F, chromatography using an adsorbent such as silica gel or a gel filtration agent such as Sephadex LH-20 (manufactured by Pharmacia) is performed. Good. Also, reversed-phase high-performance liquid chromatography is an effective method. Further, NF5003-B, NF
The 5003-E and NF5003-F substances can also be purified by crystallization with a solvent system such as methylene chloride-hexane, acetone-hexane.

The sesquiterpene derivative of the general formula (I) of the present invention has excellent AMV protease inhibitory activity and is expected to be used as a retrovirus protease inhibitor.

The AMV protease inhibitory activity of the compound of the present invention can be measured as described below.

AMV protease (Molecula
r Genetic Resources) and A
Cleavage site in pol region of MV protease (RTα-
IN) and a synthetic peptide consisting of 10 amino acid residues (Thr- Phe- Gln-Ala-Tyr-Pro-
Using Leu-Arg-Glu-Ala) as a substrate, the inhibitory activity of the enzymatic reaction is measured by the following method.

30 μ of a 0.2 M sodium citrate buffer (pH 5.5) containing 2 M NaCl in 4 μl of the test sample
l and 6 μl of a 5 M aqueous solution of NaCl are added in order.
Add 20 μl of MV protease enzyme solution (50 μg / ml) and heat at 37 ° C. for 10 minutes.

10 μl of the above-mentioned synthetic substrate aqueous solution (1.2 mg / ml) containing 2 M NaCl is added to the mixture, and the mixture is further heated at 37 ° C. for 20 minutes.

The reaction is heated in boiling water for 3 minutes to inactivate the enzyme.

[0064] 140 µl of 0.1 M Tris-HCl buffer (pH 7.5) is added, and 40 µl is analyzed by high performance liquid chromatography.

As a high performance liquid chromatography analytical column, a reversed phase column (ODS-120T, manufactured by Tosoh Corporation) was used. At a flow rate of 1.0 ml / min, a 20% aqueous solution of acetonitrile (0.1% TFA) -40% aqueous solution of acetonitrile (0.1% TFA) was subjected to a linear gradient method for 9 minutes.
0% aqueous acetonitrile (0.1% TFA) for 1 minute, and further 1% with 20% aqueous acetonitrile (0.1% TFA)
Development was performed for 0 minutes, and the synthetic substrate was detected by absorption at 210 nm.

Under these conditions, the substrate eluted around 11 to 12 minutes and the degradation products of the enzyme were eluted earlier.
The peak area of the substrate on the chromatogram was measured, and the enzyme inhibition (%) of each sample was calculated according to the following equation.

[1- (reduced amount of substrate peak area by enzyme when sample is added / reduced amount of substrate peak area only by enzyme when sample is not added)] × 100 (%) NF5003-B, NF5003-E and NF500
The AMV protease inhibitory activity of the 3-F substance is shown in Table 1 below.

[0068]

[Table 1] The compounds of the present invention can be administered orally, topically or parenterally as retroviral protease inhibitors, with the compound alone or one or more pharmaceutically acceptable carriers or excipients. Can be administered in combination.

Depending on the intended dosage form, the compounds of the invention may be formulated in solid, semi-solid or liquid dosage forms, such as tablets, pills, capsules, powders, liquids, suspensions and the like. be able to.

Formulations containing a compound of the present invention include conventional pharmaceutical carriers or excipients and may further include other pharmaceutical compounds,
It may also contain a pharmaceutical agent or the like.

Next, the present invention will be described more specifically with reference to examples.

[0072]

EXAMPLES Example 1 As a seed culture medium, potato starch 2.0%, glucose 1.0.
%, Essan Meat 2.0%, Potassium Phosphate 0.1
%, Magnesium sulfate (heptahydrate) in a 500% Erlenmeyer flask using a medium having a composition of 0.05%.
Each was dispensed and sterilized. One platinum loop was inoculated from the slant medium (potato dextrose agar medium) of Mer-NF5003 bacteria, and cultured on a rotary shaker at 28 ° C. for 3 days to obtain a seed culture.

As a production medium, 2 kg of potato was used for 10
Using a medium in which 1.0% of glucose and 1.0% of potato starch were added to the supernatant extracted with 1 L of hot water (100 ° C.) for 1 hour,
50 ml was dispensed into a 500 ml Erlenmeyer flask and sterilized. The seed culture was inoculated in an amount of 1 ml into an Erlenmeyer flask containing a production medium, and cultured at 28 ° C. for 4 days on a rotary shaker to obtain a culture.

The culture solution (8 L) was separated into bacterial cells and supernatant by centrifugation, and 6.5 L of the culture supernatant was recovered. The culture supernatant was passed through a 500 ml column of Diaion HP-20 to adsorb the active ingredient to the resin. The column was washed with 1 L of water and 1 L of 30% methanol water, and then washed with 5 L of water.
The active ingredient was eluted with 0% -65% acetone water. After the active fraction was concentrated under reduced pressure and acetone was distilled off, the remaining aqueous layer was extracted twice with an equal volume of ethyl acetate, and the ethyl acetate layer was dried over anhydrous sodium sulfate. The ethyl acetate layer was filtered and concentrated to dryness to give 2.4 g of a yellow oil.

The yellow oil was applied to a 500 ml column of Sephadex LH-20 previously filled with methanol, developed with methanol to separate the active portion, and dried to give 1.4 g of a yellow solid. Obtained. Silica gel 60 (manufactured by Merck, A
rt7734) and developed with chloroform: methanol (50: 1-10: 1). The active ingredient was divided into two main parts.

The active portion previously eluted from the silica gel column was further subjected to preparative purification using reversed-phase high performance liquid chromatography. As a preparative column, YMC PACK S-343 I-15 ODS (manufactured by YMC Co., internal diameter 2 cm, length 25 cm) was used, and chromatography was performed at a flow rate of 7 ml / min using 55% acetonitrile water as a mobile phase. I went to Graphie. The active portion was concentrated to dryness, and the obtained pale yellow powder was crystallized from methylene chloride-hexane to obtain 18.2 mg of a colorless plate-like NF5003-E substance containing 1/2 molecule of methylene chloride.

The active portion eluted later from the silica gel column was further subjected to preparative purification using reversed-phase high-performance liquid chromatography. YMC Pack S-343 I-15 ODS was used as a preparative column, and chromatography was performed at a flow rate of 7 ml / min using 52% acetonitrile water as a mobile phase. The active portion was concentrated to dryness, and the obtained pale yellow powder was crystallized from methylene chloride-hexane to obtain 11.2 mg of a white NF5003-B crystal containing 1/2 molecule of methylene chloride.

Example 2 As a seed culture medium, potato starch 2.0%, glucose 1.0
%, Essan Meat 2.0%, Potassium Phosphate 0.1
%, Magnesium sulfate (heptahydrate) in a 500% Erlenmeyer flask using a medium having a composition of 0.05%.
Each was dispensed and sterilized. One platinum loop was inoculated from the slant medium (potato dextrose agar medium) of Mer-NF5003 bacteria, and cultured on a rotary shaker at 28 ° C. for 3 days to obtain a seed culture.

As a production medium, a medium obtained by adding 1.0% of glucose and 1.0% of potato starch to a supernatant obtained by extracting 2 kg of potato with 10 L of hot water (100 ° C.) for 1 hour is used. Five liters were dispensed into two jar armamenters and sterilized. 100 ml of the above seed culture was inoculated into a jar armmenter, and cultured with aeration and agitation at 28 ° C. for 65 hours (aeration rate 5 L / min , agitation 300 rpm).
To obtain a culture solution.

The culture solution (7 L) was separated into cells and supernatant by centrifugation, and 5.5 L of the culture supernatant was recovered. The culture supernatant was passed through a 500 ml column of Diaion HP-20 to adsorb the active ingredient to the resin. The column was washed with 1 L of water and 1 L of 30% methanol water, and then washed with 5 L of water.
The active ingredient was eluted with 0% -70% aqueous acetone. After the active fraction was concentrated under reduced pressure to remove acetone, the remaining aqueous solution was passed through a 150 ml column of silanized silica gel 60 (Art 7719, manufactured by Merck) to adsorb the active component on the carrier. This column is 50
After washing with 0 ml of water, the active ingredient was eluted using a linear gradient of 20% -65% acetonitrile in water. The active ingredient was divided into two main parts.

The active part previously eluted from the silanized silica gel column was further subjected to preparative purification using reversed-phase high-performance liquid chromatography. YMC Pack S-343 I-15 ODS was used as a preparative column, and 55% acetonitrile water was used as a mobile phase for 7 m.
Chromatography was performed at a flow rate of 1 / min. The active portion was concentrated to dryness, and NF5003-E in the form of a white powder was added to 2 portions.
2.5 mg were obtained.

The active portion eluted later from the silanized silica gel column was further subjected to preparative purification using reversed-phase high-performance liquid chromatography. YMC Pack S-343 I-15 ODS was used as a preparative column, and 7% using 60% acetonitrile water as a mobile phase.
Chromatography was performed at a flow rate of 1 / min. The active portion was concentrated to dryness and 1 pale yellow NF5003-F material was added.
0.2 mg was obtained.

[Brief description of the drawings]

FIG. 1: NF5003-B substance in acetonitrile (3
1 shows an ultraviolet absorption spectrum at 1 μg / ml).

FIG. 2 shows an infrared absorption spectrum of a potassium bromide tablet of NF5003-B substance.

FIG. 3. 4 of NF5003-B material in heavy methanol
¹ shows a 00 MHz ¹ H NMR spectrum.

FIG. 4. 1 of NF5003-B substance in heavy methanol
1 shows a 00 MHz ¹³ C NMR spectrum.

FIG. 5: NF5003-E substance in acetonitrile (3
3 shows an ultraviolet absorption spectrum at 3 μg / ml).

FIG. 6 shows an infrared absorption spectrum of a potassium bromide tablet of the NF5003-E substance.

FIG. 7. 40 of NF5003-E material in heavy acetone
¹ shows a 0 MHz ¹ H NMR spectrum.

FIG. 8. 10 of NF5003-E material in heavy acetone.
1 shows a 0 MHz ¹³ C NMR spectrum.

FIG. 9: NF5003-F substance in acetonitrile (3
3 shows an ultraviolet absorption spectrum at 3 μg / ml).

FIG. 10 shows an infrared absorption spectrum of a potassium bromide tablet of the NF5003-F substance.

FIG. 11: 4 of NF5003-F substance in heavy acetone
¹ shows a 00 MHz ¹ H NMR spectrum.

FIG. 12: NF5003-F substance 1 in heavy acetone
1 shows a 00 MHz ¹³ C NMR spectrum.

──────────────────────────────────────────────────の Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI // (C12P 17/04 C12R 1: 645) (72) Inventor Yoshio Watanabe 2-22-3 Fujigaoka, Fujisawa-shi, Kanagawa (72) Inventor Toshihiko Kumamoto 1-9-12 Kugenuma Sakuragaoka, Fujisawa City, Kanagawa Prefecture (72) Inventor Hiroshi Nishida 568, Tsukui, Yokosuka City, Kanagawa Prefecture Green Heights 11-3-503 (72) Inventor Rokuro Okamoto 2-18 Hanaki, Fujisawa City, Kanagawa Prefecture (72) 72) Inventor Hisayoshi Akakawa 2-10-35, Kami-Osaki, Shinagawa-ku, Tokyo, Japan (72) Inventor Saotoshi Mizuno 2-10-35, Kami-Osaki, Shinagawa-ku, Tokyo (56) References JP-A-54-106458 (JP, A) JP-A-57-83281 (JP, A) U.S. Pat. No. 4,981,980 (US, A) (58) Fields investigated (Int. Cl. ⁷ , DB name) C07D 307/94 A61K 31/343 CA ( TN) REGISTRY (STN)

Claims (6)

(57) [Claims] 1. A compound of the general formula Wherein R ¹ is a hydroxymethyl group and R ² is a hydroxyl group, or R ¹ is a hydroxymethyl group or a formyl group and R ² is a hydrogen atom. 2. The formula: The sesquiterpene derivative according to claim 1, which is an NF5003-B substance represented by the following formula: 3. The formula: The sesquiterpene derivative according to claim 1, which is an NF5003-E substance represented by the formula: 4. The formula: The sesquiterpene derivative according to claim 1, which is a NF5003-F substance represented by the following formula: 5. A microorganism having the ability to produce a sesquiterpene derivative represented by the general formula (I) according to claim 1, which belongs to the genus Stachybotrys, is cultured in a medium, and the sesquiterpene derivative is collected from the culture. A method for producing a sesquiterpene derivative represented by the general formula (I) according to claim 1. 6. A retroviral protease inhibitor comprising the sesquiterpene derivative represented by the general formula (I) according to claim 1 as an active ingredient.