JP2801491B2 - Epipolythiodioxopiperazine derivatives and their production - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

The present invention relates to an epipolythiodioxopiperazine derivative and a method for producing the same.

[0002]

BACKGROUND OF THE INVENTION An object of the present invention is to provide an epipolydioxopiperazine derivative having an excellent cell growth inhibitory action. Another object of the present invention is to provide a fermentative method for producing the epipolydioxopiperazine derivative. Still another object of the present invention is to provide a cell growth inhibitor containing the epipolythiodioxopiperazine derivative as an active ingredient.

[0003]

DISCLOSURE OF THE INVENTION The present inventors have collected various microorganisms living in the ocean and searched for metabolites thereof for the purpose of searching for a novel antitumor active substance, and found that Leptosphaeria Genus or sessilium (S
It has been found that a substance having a strong cell growth inhibitory activity is produced in a culture of a microorganism belonging to the genus Esquicillium. The present inventors have studied in detail the cytostatic substances and confirmed that they are a series of compounds having an epipolythiodioxopiperazine skeleton, and succeeded in isolating and purifying them.

[0004] These epipolythioxopiperazine derivatives (hereinafter collectively referred to as OUPS-80T) which are the object substances of the present invention are represented by the following general formula (I):

Embedded image Wherein R ₁ and R ₂ are the same or different and are lower alkyl or hydroxy-substituted lower alkyl, and R ₃ has the formula (a)

Embedded image Or equation (b)

Embedded image And R ₄ and R ₅ are the same or different and are lower alkyl or hydroxy-substituted lower alkyl, and m and n are the same or different and are an integer of 2 to 4). The lower alkyl part of the “lower alkyl” or “hydroxy-substituted lower alkyl” represented by the above symbol may be methyl, ethyl, propyl, butyl and the like. In the general formula (I),
Known substances are also included, and a compound of m = n = 2 when R ₃ is (a) and R ₁ ＲR ₂ ＲR ₄ ＲR ₅ ＣＨCH ₃ , wherein R ₃ is (a ) And R ₁ ＣＨCH ₂ OH;
And m = n = 2 when R ₂ ＲR ₄ ＲR _{5 、} CH ₃ , R ₃ is (a), R ₁ ＣＨCH ₂ OH, and R ₂ ＲR ₄ ＲR _When m = 2 and n = 3 when ₅ = CH ₃ and R ₃ is (a), R ₁ = R ₄ = CH ₂ OH
And m = n = 4 when R ₂ = R ₅ = CH ₃
The compounds of each are Verticillin
A, known as Verticillin B, Verticillin C and Chetracin A (Takao Saito et al., Chemical and Pharmaceutical Buretan (Chem. Pharm. Bull., 36 (6) 1942-1956 ( 198
8))). Substances other than these are considered to be new substances, and typical examples are shown in Table 1.

[Table 1]

[Table 2]

[Table 3]

[Table 4]

[Table 5]

[Table 6]

[Table 7]

[Table 8]

[0005] In order to produce the object substance OUPS-80T of the present invention, an OUPS-80T-producing bacterium belonging to the genus Leptosperia or Cesxylium, which is a marine microorganism, is cultured, and the substance is collected from the culture. I just need. Here, as an example of the microorganism of the genus Leptospheria used for the production of OUPS-80T, Leptosphaeria sp. OU, a fungus isolated from the seaweed Jolemoku collected on the coast of Tanabe Bay, Wakayama Prefecture, Japan
PS-80, which is provided to the Institute of Microbial Industry and Technology by the National Institute of Advanced Industrial Science and Technology
905). In addition, as an example of the microorganism of the genus Cesxylium, there is Sesquicillium canderabrum (Bono), a fungus isolated from the sea anemone collected from the coast of Osaka Bay.
rd.) W. Gams) OUPS-100, which is available from the Institute of Microbial Industry and Technology of the Institute of Industrial Science and Technology.
No. 1 (FERM P-13201).

The above-mentioned Leptosperia sp. OU
The bacteriological properties of PS-80 are as follows. (a) Culture (1) Growth state in each medium Tuapek agar medium Growth on a Tuapec agar medium does not occur at all at 5 ° C., and at 30 ° C., the diameter of a colony reaches about 2 cm in 7 days. The appearance is white floc. After 14 days of cultivation, the colonies reach a diameter of 3.5 to 4 cm and have a white flocculent surface but yellowish green inside. The back is black at the center,
The middle part is gray and the circumference is yellow. Potato-Sucrose Agar Medium Growth at 5 ° C does not occur on potato-sucrose agar. At 30 ° C., the diameter of the colony was 3.
Reaches 5-4 cm. The appearance is white cotton on the periphery and gray on the other parts. After 15 days of culturing, the colony reaches 6.5 cm in diameter, has a white flocculent surface, and turns black-green inside. The back of the colony is white at the periphery and black at the center. The formation of pseudomushrooms is observed in the black-green area. Malt Extract Agar Medium No growth occurs on Malt Extract Agar at 5 ° C. At 30 ° C., the colonies reach 2.4-2.6 cm in 7 days. The appearance is white floc. As in the case of the colonies on the potato-sucrose agar medium, the back surface of the colony is white at the periphery and black at the center. 1
After 5 days of cultivation, the diameter of the colony reaches 5 cm, whereby the whole becomes moist. Glucose / peptone / yeast extract / seawater agar medium Growth on glucose / peptone / yeast extract / seawater agar medium does not occur at 5 ° C at all. At 30 ° C., the colony reaches a diameter of 1 cm in 3 days and grows in a white flocculent state with the central part raised. The back of the colony is yellow-orange. Colony diameter 4.5c after 15 days of culture
m, and the entire back surface has an ocher color, and wrinkles in the form of black stripes are formed radially from the center. In addition, pseudoalcophorous shells are formed, and many ascos and ascospores are observed.

(2) Physiological and ecological properties Optimum growth conditions pH 6-7.5 Temperature 27-30 ° C. Growth range pH 4-8.5 Temperature 20-40 ° C. Other remarkable features glucose, peptone, yeast OUPS-80T is produced in the medium by culturing on an extract-seawater agar medium. (3) Microscopic findings Pseudo-assay shell: There is no marked hole. It is buried, oblate and has a size of 120 to 150 × 200 to 300 μm. Shell wall: Pseudo parenchymal thick film with brown color. Ascus: many, as club-cylindrical, sometimes bent. The false side thread is string-like and colorless. Ascospores: spindle-shaped, size 20-22 × 4 μm
With a yellow-brown to light brown color.

The above mycological properties were determined by A. Munk, Dansk Botan. Ark., (1957)
), Earl W. G. Dennis, British Ascomycetes (196)
8), and Keisuke Tsubaki, et al., Fungus illustration book (top) (1978)
And the strain was searched, and it was determined that this strain belongs to the genus Leptosperia. This strain resembles Leptosperia nodrum, except that the pseudoplastid shell is spherical in Leptosperia nodrum, whereas it is oblate in this strain. Furthermore, phyarotype conidia are formed in conidial shells in Leptosperia nodrum, but this is not observed in this strain. Based on the above results, this strain was identified as a new strain belonging to the genus Leptosperia and named Leptosperia sp. OUPS-80.

On the other hand, Cessilium candelabrum O
For UPS-100, potato sucrose agar,
The growth state, optimal growth temperature, optimal growth pH range, and observation results under a microscope on malt extract agar medium and Tzapek agar medium were examined, and the microbiological properties were examined by K.E.D.Domsch et al., Compendium.
Compendium of Soil Fungi
(1980), Toru Okuda et al., Trans. Mycol. Soc. Japan. (1977)
), Keisuke Tsubaki et al., "Fungi Encyclopedia (2)" (1978)
The strain was identified as a known cesxylium candelabrum.

[0010] The medium used for culturing the OUPS-80T-producing bacterium may be liquid or solid as long as it contains a nutrient that can be used by the bacterium.
In order to obtain 0T, it is preferable to use a liquid medium. The medium is appropriately mixed with a carbon source that can be assimilated by the bacterium, a nitrogen source that can be digested, and inorganic substances. As a carbon source,
Glucose, sucrose, maltose, lactose, starch and the like are used. As nitrogen sources, yeast extract, meat extract, peptone, polypeptone, malt extract, sodium nitrate and the like are used. Since the medium is usually prepared using artificial seawater, salts such as sodium, potassium, calcium, and magnesium are contained in the medium. The initial pH is suitably about 7.5, and the culture temperature is suitably in the range of 25 to 30 ° C.

[0011] OUPS-80T-producing bacteria have the property of proliferating by adhering to stationary objects.
In addition, if an object to which the bacterium adheres, for example, an aluminum foil is placed in a liquid medium, the proliferation is promoted. The cultivation period is not fixed, but it is desirable to culture until the concentration of OUPS-80T to be produced reaches the maximum. The number of days required for this is usually 3 to 4 in the case of static culture using a liquid medium.
Weeks are appropriate.

The obtained culture solution is separated into bacterial cells and culture solution by means such as centrifugation or filtration. Both of them show a cell growth inhibitory action, but OUPS-80T is more contained in the cells. OU from collected cells
To extract PS-80T, an organic solvent miscible with water,
For example, lower alcohols such as methanol and ethanol, acetone, and methyl ethyl ketone may be used. Further, an organic solvent immiscible with water, for example, chloroform, methylene chloride, ethyl acetate and the like may be used.
By evaporating the solvent from the thus obtained extract under reduced pressure, a crude extract containing OUPS-80T can be obtained.

In order to isolate and purify OUPS-80T from the crude extract, ordinary means for isolating and purifying a fat-soluble low-molecular substance can be applied. That is, gel filtration chromatography using Sephadex LH-20 (Pharmacia, trademark) or the like, adsorption chromatography using an adsorbent such as silica gel, high-performance liquid chromatography using a normal phase carrier such as silica gel alone or the like. What is necessary is just to implement in combination. When Sephadex LH-20 is used, it is generally eluted with a combination of a polar organic solvent and a non-polar organic solvent, for example, a mixed solvent of methanol and chloroform or methylene chloride. When using adsorption chromatography using silica gel, a mixed solvent such as hexane and chloroform or methylene chloride is preferably used as an elution solvent.
In the case of high performance liquid chromatography using silica gel as a carrier, a mixed solvent of methylene chloride and benzene or methanol or methylene chloride alone is used as an elution solvent. By applying such a purification means, one or more of OUPS-80T shown in Table 1 above is isolated. Compounds T-1 and T-2 are
In view of their physicochemical properties, they are considered to be stereoisomers.

The OUPS-80T obtained in the present invention generally exerts a cell growth inhibitory action. For example, as apparent from the test examples described later, the compound has a remarkable growth inhibitory effect on various malignant tumor cells of mice and humans. Therefore, OUPS-
80T is useful as a leukemia inhibitor or antitumor agent.

EXAMPLES Hereinafter, the present invention will be described with reference to Examples and Test Examples, which should not be understood as limiting the present invention. Note that “%” means “% by weight” unless otherwise specified. Example 1 Production of OUPS-80T (using Leptospheria sp.):-Glucose 2%, polypeptone 1%, yeast extract 0.5%
200 ml of an artificial seawater medium (pH 7.5) containing 200 ml of aluminum foil having a width of about 1 to 2 cm in advance in an S shape.
70 ml each was dispensed into a ml Erlenmeyer flask. After sterilization,
These are Leptospheria sp. OUPS-80
(Piercing No. 12905) inoculated in a loop amount of 1 platinum loop, 27 ° C
For 6 days. Next, 10 liters of the same medium
Similarly, 70 ml of the aluminum foil was dispensed into a 200 ml Erlenmeyer flask in which the aluminum foil was placed in an S shape, and after sterilization, the above-described inoculum was added.
Static culture was performed at 27 ° C. for about 4 weeks. After the culture, the cells were collected from the culture by filtration, and 5 liters of methanol was added thereto.
After stirring, the mixture was left overnight and filtered to obtain an extract. The methanol was distilled off under reduced pressure to obtain 25 g of a methanol extract. Hexane was added to remove the hexane-soluble part, and a mixed solvent of methanol / methylene chloride (1: 1) was added to the residue to obtain a mixed solvent. And the mixed solvent insoluble part. The solvent was distilled off from the mixed solvent-soluble portion under reduced pressure to obtain 10.2 g of an oily substance showing cytotoxicity. The obtained oily substance was subjected to gel filtration chromatography using Sephadex LH-20 and eluted with a mixed solvent of methanol / methylene chloride (1: 1) to give 4 g of a cytotoxic fraction. Obtained. This fraction was subjected to silica gel column chromatography, and hexane and then hexane / methylene chloride (8:
After elution was carried out with a mixed solvent of 2) to remove impurities, elution was carried out with a mixed solvent of hexane / methylene chloride (6: 4) to obtain fractions 1, T-2 and T-containing T-1. 1
1 containing fraction 2, containing T-3, T-4 and T-12 and fraction 4 containing T-5, T-6 and T-13, then hexane / methylene chloride ( The fraction 5 containing T-7 and T-8 was eluted with the mixed solvent of 4: 6), and the fraction 6 and T-10 containing T-9 were further eluted with methylene chloride as the elution solvent. Fraction 7 containing
I got Each fraction is silica gel (Nakarai Tesque, CO
(SMOSIL 10SL) using normal phase high performance liquid chromatography. Fractions 1 to 7 used methylene chloride as a developing solvent, and as a result, fractions 1 to T-1 (3
mg), T-2 (7 mg) and T-11 (5 mg
mg), fraction 3 to T-3 (4 mg), T-4 (5 mg)
And T-12 (3 mg) and fractions 4 to T-5
(24 mg), T-6 (4 mg) and T-13 (3 mg)
Was obtained as a single substance. In the case of fractions 5 to 7, methanol / methylene chloride (1: 9) was used as a developing solvent.
Using the mixed solvent of 9), fractions 5 to 7 (3 mg) and T-8 (40 mg), fractions 6 to T-9 (14 mg) and fractions 7 to T-10 (4 mg) Obtained as material.

Example 2 Production of OUPS-80T (using sessilium candelabrum): 2% glucose, 1% polypeptone, 0.5% yeast extract
200 ml of an artificial seawater medium (pH 7.5) containing 200 ml of aluminum foil having a width of about 1 to 2 cm in advance in an S shape.
70 ml each was dispensed into a ml Erlenmeyer flask. After sterilization,
These include Cessilium Candelabulum OUPS-1
00 (Piercing No. 13201) is inoculated with one platinum loop.
The cells were cultured at 27 ° C. for 6 days. Next, 20 liters of the same medium was dispensed in 70 ml portions into a 200 ml Erlenmeyer flask in which an aluminum foil was similarly placed in an S-shape, and after sterilization, the above-described inoculum was added, followed by stationary culture at 27 ° C. for about 4 weeks. . After the culture, the cells were collected by filtration from the culture, 10 liters of methanol was added thereto, and the mixture was stirred overnight, left to stand overnight, and filtered to obtain an extract. The methanol was distilled off under reduced pressure and methanol extract 128 g
Hexane was added to this to remove the hexane-soluble part,
A mixed solvent of methanol / methylene chloride (1: 1) was added to the residue to separate into a mixed solvent-soluble part and a mixed solvent-insoluble part.
The solvent was distilled off from the mixed solvent-soluble portion under reduced pressure to obtain 13.7 g of an oily substance showing cytotoxicity. The obtained oily substance was subjected to gel filtration chromatography using Sephadex LH-20 to give methanol / methylene chloride (1: 1).
The mixture was eluted with a mixed solvent of 0.8 g to obtain 0.8 g of a fraction showing cytotoxicity. This fraction was subjected to silica gel column chromatography, and elution was performed with hexane and then with a mixed solvent of hexane / methylene chloride (8: 2) to remove impurities, followed by a mixed solvent of hexane / methylene chloride (4: 6). To obtain a fraction 1 containing T-14 and T-15, followed by hexane / methylene chloride (2: 8).
By elution with a mixed solvent of No. 2, fraction 2 containing T-16 was obtained. Each fraction is silica gel (Nacalai Tesque,
(COSMOSIL 10SL) using normal phase high performance liquid chromatography. In the case of fraction 1, as a result of using benzene / methylene chloride (1: 9) as a developing solvent, T-14 (8.6 mg) and T-15 (11.9) were used.
mg) as a single substance. In the case of Fraction 2, as a developing solvent, acetone / methylene chloride (3:97) was used, and as a result, T-16 (5.3 mg) was obtained as a single substance.

Test Example 1 Inhibition of cell growth of OUPS-80T (P in vitro
388 mouse leukemia cell proliferation inhibitory action): 10% FBS to which compounds T-1 to 14 were added at final concentrations of 0.001, 0.01 and 0.1 μg / ml
P388 mouse leukemia cells are seeded at 10 × 10 ⁴ cells / ml on a plastic plate containing the containing Eagle MEM medium, and cultured at 37 ° C. for 72 hours in a 5% carbon dioxide incubator. After completion of the culture, the cells in the culture solution were calculated using a hemocytometer, and a 50% effective dose (ED ₅₀ ) was calculated. The results are shown in Table 2.

[Table 9]

Test Example 2 Inhibition of Cell Growth of OUPS-80T (Inhibition of Mouse and Human Tumor Cell Growth in Test Tube) Various tumor cells were suitable for each cell at 2 to 10 × 10 ⁴ cells / ml. The cells are seeded on a plastic plate containing a medium, and cultured at 37 ° C. for 24 hours in a 5% carbon dioxide incubator. Then, the compound T-5 diluted with the culture solution was used.
Or T-9 at a final concentration of 0.01, 0.03, 0.1,
Add to 0.3 and 1.0 μg / ml, add 3
Incubate in a 5% CO 2 incubator at 7 ° C. for 48 to 72 hours. After the culture, the cells were detached from the plate by trypsin digestion, and the number of cells was counted using a hemocytometer.
The% growth inhibitory concentration (IC ₅₀ ) was calculated. The results are shown in Table 3.

[Table 10]

The survival in Tangan mice Test Example 3 Oups-80T effects: over mouse sarcoma sarcoma (Sarcoma) 180 10 ⁶ cells / mouse were transplanted into the abdominal cavity of ICR mice, the 24 hours after compound T-5 or T-9 at 0.25, 0.5 and 1.0 mg / kg, 5-FU (positive control) 10 mg / kg
Alternatively, a physiological saline solution containing 10% DMSO (negative control) was administered once intraperitoneally, and the average survival days were calculated from the number of days until death of the bile cancer mouse, and compared with the negative control. Table 4 shows the results.

[Table 11]

Furthermore, mouse sarcoma sarcoma
From the day after transplantation of compound 180, 0.01, 0.
Table 5 shows the results obtained by intraperitoneal administration of 03 and 1.0 mg / kg / day for 9 consecutive days.

[Table 12]

Test Example 4 Inhibitory effect of OUPS-80T on tumor cell growth in bile cancer mice: mouse sarcoma (Sarcoma) 180 5 × 10 ⁴ cells /
The mice were implanted intradermally into the back skin of ICR mice, and once every four days from the next day, 0.25 of compound T-5,
0.5 and 1.0 mg / kg were administered subcutaneously or intravenously in the vicinity of the tumor cell transplant, and the estimated tumor weight was calculated from the measured values of the major axis and minor axis of the tumor. Further, on the last day of the experiment (18 or 19 days after transplantation), the tumor was excised and its wet weight was weighed. In addition, 10-FU of 5-FU was used as a positive control.
mg / kg, physiological saline containing 10% DMSO was used as a negative control. The results are shown in Tables 6 and 7.

[Table 13]

[Table 14]

Effect of the Invention As is clear from the test examples, the target substance OU of the present invention
PS-80T generally has a remarkable cytostatic effect. Therefore, it is useful as a growth inhibitor for leukemia cells and other various malignant tumor cells. For example, OUP
S-80T can be topically applied to skin cancer as an antitumor agent and intravenously applied to other cancers.

──────────────────────────────────────────────────続 き Continued on the front page (72) Inventor Yutaka Murahashi 282-4 Horikamicho, Sakai City, Osaka Prefecture (72) Inventor Chituko Kitagawa 2-529-3 Gakuen Asahimotomachi, Nara City, Nara Prefecture Excel Heights B104 (72) Inventor Hideo Iwaki 3-6-19 Minamitsukaguchi-cho, Amagasaki-shi, Hyogo (72) Inventor Hiromasa Araki 1-12, Shinohara-cho, Yamatokoriyama-shi, Nara (56) References Chem. Pharm. Bull. , 36 (6), 1942-1956 (1988) (58) Fields investigated (Int. Cl. ⁶ , DB name) C07D 513/22 A61K 31/495 C12P 17/18 CA (STN) REGISTRY (STN)

Claims (12)

(57) [Claims] 1. A compound of the general formula (I): Wherein R ₁ and R ₂ are the same or different and are lower alkyl or hydroxy-substituted lower alkyl, and R ₃ is a group represented by the formula (b): Wherein n is an integer of 2 to 4. In producing the epipolythiodioxopiperazine derivative represented by the formula (1), a microorganism having an ability to produce an epipolythiodioxopiperazine derivative belonging to the genus Leptosperia or the genus Cesxylium is cultured, and from the culture, the derivative is obtained. A method of collecting a sample. 2. The method according to claim 1, wherein the microorganism capable of producing an epipolythiodioxopiperazine derivative belonging to the genus Leptosperia is Leptosperia sp. 3. The microorganism having an ability to produce an epipolythiodioxopiperazine derivative belonging to the genus Cesxylium is Cesxylium candelabulum.
The manufacturing method as described. 4. A compound of the general formula (I): Wherein R ₁ and R ₂ are the same or different and are lower alkyl or hydroxy-substituted lower alkyl, and R ₃ is a group represented by the formula (b): Wherein n is an integer of 2 to 4. An epipolythiodioxopiperazine derivative represented by the formula: 5. The compound according to claim 4, wherein R ₁ is isopropyl, R ₂ is methyl, and n is 2. 6. The compound according to claim 4, wherein R ₁ is isopropyl, R ₂ is methyl, and n is 3. 7. The compound according to claim 4, wherein R ₁ is isopropyl, R ₂ is methyl and n is 4. 8. The compound according to claim 4, wherein R ₁ is ethyl, R ₂ is methyl and n is 3. 9. The compound according to claim 4, wherein R ₁ is ethyl, R ₂ is methyl, and n is 4. 10. The compound according to claim 4, wherein R ₁ and R ₂ are methyl and n is 4. 11. A compound of the general formula (I): (In the above formula, R ₃ is the formula (a) Is a group represented by An epipolythiodioxopiperazine derivative represented, selected from the following compounds: (i) R ₁ and R ₄ are isopropyl, R ₂ and R ₅ are methyl,
a compound wherein m is 2 and n is 2; (ii) R ₁ and R ₄ are isopropyl, R ₂ and R ₅ are methyl, m
Is a compound wherein n is 3; and (iii) R ₁ and R ₄ are isopropyl, R ₂ and R ₅ are methyl,
m is 2, n is 4 compound; (iv) R ₁ is isopropyl, methyl R ₂ and R _5, R ₄ is hydroxymethyl, compounds m is 2, n is 2; (v) R ₁ is Isopropyl, a compound in which R ₂ and R ₅ are methyl, R ₄ is hydroxymethyl, m is 2, and n is 3; (vi) R ₁ is isopropyl, R ₂ and R ₅ are methyl, R ₄ is hydroxymethyl, m (Vii) a compound wherein R ₁ is isopropyl, R ₂ and R ₅ are methyl, R ₄ is hydroxymethyl, m is 3, and n is 2; (viii) R ₁ is isopropyl A compound wherein R ₂ and R ₅ are methyl, R ₄ is hydroxymethyl, m is 4, and n is 2; (ix) R ₁ is isopropyl, R ₂ and R ₅ are methyl, R ₄ is hydroxymethyl, and m is 4. A compound wherein n is 3. 12. A cell growth inhibitor comprising an epipolythiodioxopiperazine derivative selected from the following compounds as an active ingredient: General formula (I): (Wherein, R ₃ is a group represented by the formula (b): Is a group represented by (1) a compound wherein R ₁ is isopropyl, R ₂ is methyl, and n is 2; (2) R ₁ is isopropyl, R ₂ is methyl, and n is an epipolythiodioxopiperazine derivative represented by the following formula: (3) a compound wherein R ₁ is isopropyl, R ₂ is methyl and n is 4; (4) a compound wherein R ₁ is ethyl, R ₂ is methyl and n is 3; (5) R _A compound wherein ₁ is ethyl, R ₂ is methyl and n is 4; (6) a compound wherein R ₁ and R ₂ are methyl and n is 4 or a compound of the general formula (I): (In the above formula, R ₃ is the formula (a) Is a group represented by An epipolythiodioxopiperazine derivative represented by: (i) R ₁ and R ₄ are isopropyl, R ₂ and R ₅ are methyl,
a compound wherein m is 2 and n is 2; (ii) R ₁ and R ₄ are isopropyl, R ₂ and R ₅ are methyl, m
Is a compound wherein n is 3; and (iii) R ₁ and R ₄ are isopropyl, R ₂ and R ₅ are methyl,
m is 2, n is 4 compound; (iv) R ₁ is isopropyl, methyl R ₂ and R _5, R ₄ is hydroxymethyl, compounds m is 2, n is 2; (v) R ₁ is Isopropyl, a compound in which R ₂ and R ₅ are methyl, R ₄ is hydroxymethyl, m is 2, and n is 3; (vi) R ₁ is isopropyl, R ₂ and R ₅ are methyl, R ₄ is hydroxymethyl, m (Vii) a compound wherein R ₁ is isopropyl, R ₂ and R ₅ are methyl, R ₄ is hydroxymethyl, m is 3, and n is 2; (viii) R ₁ is isopropyl A compound wherein R ₂ and R ₅ are methyl, R ₄ is hydroxymethyl, m is 4, and n is 2; (ix) R ₁ is isopropyl, R ₂ and R ₅ are methyl, R ₄ is hydroxymethyl, and m is 4. A compound wherein n is 3.