Methods of treatment using extracts of Anisomeles heyneana

Abstract

The present invention describes the use of pharmaceutical compositions including compounds of formula 2 (below) for treatment of infections related to M. tuberculosis and for anti-proliferative activity. Also, the present invention discloses a process of extraction of compounds of formula 2 from the extract of aerial parts of Anisomeles heyneana: ##STR00001##

Claims

1. A method of inhibiting cell proliferation in human Thp-1 cell line by administering to the cell a composition of a compound extracted from Anisomeles heyneana of formula: ##STR00007##

2. The method of claim 1, wherein the compound of the formula is present in the range of 0.1 to 99% w/w of the pharmaceutical composition.

3. The method of claim 1, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable excipient is selected from the group comprising diluents, binders, lubricants, wetting agents, disintegrants and combinations thereof.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) FIG. (1a) depicts .sup.1HNMR spectra.

(2) FIG. (1b) depicts .sup.13CNMR spectra.

(3) FIG. (1c) depicts DEPT spectra of compound of formula 1.

(4) FIG. (2a) depicts .sup.1HNMR spectra.

(5) FIG. (2b) depicts .sup.13CNMR spectra.

(6) FIG. (2c) depicts DEPT spectra of compound of formula 2.

(7) FIG. (3a) depicts .sup.1HNMR spectra.

(8) FIG. (3b) depicts .sup.13CNMR spectra.

(9) FIG. (3c) depicts DEPT spectra of compound of formula 3.

(10) FIG. 4 depicts I-V (current vs voltage) plot for compound of formula 3.

(11) FIG. 5 is Scheme 1 representing a chromatographic separation of A. heyneana.

SUMMARY OF THE INVENTION

(12) Accordingly, present invention provides pharmaceutical composition comprising at least one compound selected from formula 1 and formula 2 along with pharmaceutically acceptable excipients for anti proliferative activity.

(13) ##STR00002##

(14) Additionally, the present invention provides a pharmaceutical composition comprising compound of formula 1 along with pharmaceutically acceptable excipients for the treatment of infections caused due to M. tuberculosis.

(15) Further, the present invention provides a compound of formula 3 for the conversion of solar energy into electric current in dye sensitized solar cells.

(16) ##STR00003##

(17) In yet another embodiment; present invention provides a process for isolation of compound of formula 1, formula 2 and formula 3 from the extract of aerial parts of Anisomeles heyneana

DETAILED DESCRIPTION OF THE INVENTION

(18) Accordingly, the present invention relates to pharmaceutical composition comprising at least one compound selected from formula 1 and formula 2 along with pharmaceutically acceptable excipients for the treatment of leukemia.

(19) The present invention further relates to pharmaceutical composition comprising compound of formula 1 along with pharmaceutically acceptable excipients for the treatment of infections caused due to M. tuberculosis.

(20) The present invention further describes the use of compound of formula 3 for the conversion of solar energy into electric current in dye sensitized solar cells.

(21) According to one aspect of the invention, the compound of formula 1 or formula 2 are present in the range of 0.1 to 99% w/w in the pharmaceutical composition.

(22) According to yet another aspect, the pharmaceutically acceptable excipients are selected from the group comprising diluents, binders, lubricants, wetting agents, and disintegrants and their combinations thereof.

(23) The pharmaceutically acceptable excipients are the excipients well known in the art and those which have been conventionally used in the formulation of various compositions.

(24) Present invention also relates to process for the isolation of non-toxic, economically viable and stable diterpenoids of formula 1 and 2 and phenethyl cinnamoyl glycoside of formula 3 with solar cell sensitizing activity isolated from extract of Anisomeles heyneana.

(25) In an aspect, the present invention provide process for isolation of non-toxic, cheap/cost-effective, stable, eco-friendly and biocompatible diterpenes from extract of aerial parts of Anisomeles heyneana plant, wherein the diterpene compounds are selected from the group consisting of cembrane(formula 1), phyllocladane(formula 2) exhibit substantial anti-mycobacteria and anti-proliferation activities respectively and phenethylglycoside (formula 3) which is effective in conversion of solar energy into electric current in dye-sensitized solar cells.

(26) Anisomeles heyneana, the entire mature plants in flowering, are collected from Purandar fort area, District, Pune. A herbarium is deposited in Botanical Survey of India, Western Circle, Pune (Voucher No. SPJ5). Plant material is cleaned off adhering dust and unwanted plant material. Roots are separated and aerial parts are dried in shade and pulverized.

(27) The present invention provides process for isolation of compounds of formula 1, 2 and 3 from the extract of aerial parts of Anisomeles heyneana plant comprising the steps of: i. powdering aerial parts of A. heyneana followed by extraction with acetone at temperature in the range of 25 to 30° C., filtering acetone solubles and concentrating under reduced pressure at 50 to 100 mm of Hg to obtain a greenish acetone extract; ii. separating the extract of step a) by using column chromatography (CC) with increasing polarity of the polar solvent to afford 18 fractions (AH-1 to AH-18); iii. Subjecting Fraction AH7 to CC in acetonitrile: chloroform gradient from 2 to 10% to collect six fractions (AH7a-AH7f). iv. Subjecting Fraction AH8 to CC in acetonitrile: chloroform gradient from 2 to 10% to collect eight fractions (AH8a-AH8h); v. Combining Fractions AH8a, AH8b and AH7b based on similarity of their TLC profile and to obtain compound 1 (80 mg) after CC in 1 to 5% methanol in chloroform; vi. subjecting fraction AH-11 of step ii) to column chromatography in 1 to 5% of methanol in chloroform to collect eleven fraction. Sub fraction 7, which is further subjected to column chromatography with 6% methanol in chloroform, to collect 8 fractions. Sub-fraction 2 subsequently purified by washing with chloroform to obtain compound 2 (14 mg) i.e. (phyllocladan-16α,17-dihydroxy-19-oic acid); vii. subjecting fraction AH-14 of step b) to column chromatography in chloroform with acetonitrile from 5 to 50% gives fifteen fractions. Sub-fraction 11 is further subjected to preparative thin layer chromatography in 20-30% methanol in chloroform to afford brown amorphous powder of compound 3 (35 mg) i.e. verbascoside.

(28) The chromatographic separation of diterpene compounds 1 and 2 and phenethylglycoside 3 from aerial parts of A. heyneana is disclosed in Scheme 1.

(29) The diterpenoid compound 1 (ovatodiolide) isolated from extract of A. heyneana is belonging to class cembrane, having chemical name (1R,3E,5S,9R,12E)-3,12-Dimethyl-8-methylene-6,18-dioxatricyclo[14.2.1.05,9]nonadeca-3,12,16(19)-triene-7,17-dione and molecular formula C.sub.20H.sub.24O.sub.4.

(30) ##STR00004##

(31) According to the process of the present invention, the aerial parts of A. heyneana is powdered and extracted with acetone (3×3 L) for 14 hrs at room temperature 25 to 30° C., the mixtures is further filtered and concentrated under reduced pressure to obtain greenish acetone extract, which is then separated by using column chromatography (CC) with increasing polarity of acetone in petroleum ether to afford 18 fractions of A. heyneana. The fraction 7 is subjected to column chromatography in chloroform with acetonitrile from 2 to 10%. Also fraction 8 is subjected to column chromatography in chloroform with acetonitrile from 2 to 10%. From fraction 8, sub-fractions 1 and 2 are pooled with sub-fraction 2 of fraction 7 from above column chromatography and compound 1 i.e. (ovatodiolide) is obtained from this pool after rechromatography in 2% methanol in chloroform.

(32) The diterpenoid compound 2 (phyllocladan-16α,17-dihydroxy-19-oic acid) isolated from A. heyneana is belonging to phyllocladane class.

(33) ##STR00005##

(34) 18 fractions of A. heyneana are separated by column chromatography as described hereinabove, wherein fraction 11 is subjected to column chromatography in 3% methanol in chloroform to obtain sub-fraction 7 which is again subjected to column chromatography in 6% methanol in chloroform. From fraction 2 of this column, compound 2 i.e. (phyllocladan-16α,17-dihydroxy-19-oic acid) is precipitated as small crystals which are purified by washing with chloroform.

(35) In another embodiment, the compound 3 (verbascoside) isolated from extract of A. heyneana is belonging to class phenethylglycoside, which is also known as TJC 160; Acetoside; Kusaginin; 2-(3,4-dihydroxyphenyl)ethyl 3-O-(6-deoxy-alpha-L-mannopyranosyl)-4-O-[(2E)-3-(3,4-dihydroxyphenyl)prop-2-enoyl]-beta-D-gluco pyrano side; beta-D-glucopyranoside, 2-(3,4-dihydroxyphenyl)ethyl 3-O-(6-deoxy-alpha-L-mannopyranosyl)-4-O-[(2E)-3-(3,4-dihydroxyphenyl)-1-oxo-2-propenyl] having molecular formula C.sub.29H.sub.36O.sub.15.

(36) ##STR00006##

(37) 18 fractions of A. heyneana are separated by column chromatography as described hereinabove, wherein fraction 14 is subjected to column chromatography in chloroform with acetonitrile from 5 to 50% to obtain sub-fraction 11, from which compound 3 i.e. (verbascoside) is isolated as brown amorphous powder after preparative thin layer chromatography in 25% methanol in chloroform.

(38) The diterpenoid compound of formula 1 and compound of formula 2 are used for the preparation of pharmaceutical composition for anti-proliferative activity-MTT cell proliferation assay on human monocytic leukaemia Thp-1 cell lines. The diterpenoid compound 1 is used for the preparation of pharmaceutical composition having anti-tubercular activity. The effect of compound 1 and compound 2 on the viability of Thp-1 cell line at 100 μg/ml and IC.sub.90.sup.b value against M. tuberculosis respectively is described herein below in Table 1.

(39) TABLE-US-00001 TABLE 1 Effect of compounds 1 and 2 on the viability of Thp-1 cell line and M. tuberculosis bacilli. IC.sub.90.sup.b value determined against % Inhibition on Thp-1 M. tuberculosis Compound cell line at 100 μg/ml (μg/ml) 1  96.4 ± 0.4592 6.53 ± 0.893 2 59.02 ± 0.8725 Not active.sup.c Isoniazid — 0.05 ± 0.003 Paclitaxel 61% ± 0.501 — Vehicle Control.sup.a No inhibition No inhibition .sup.a1% Dimethyl sulfoxide (DMSO), .sup.bConcentration of compounds exhibiting 90% inhibition against growth of M. tuberculosis. .sup.cat 100 μg/ml

(40) The present invention provides a pharmaceutical composition comprising diterpenoid compound isolated from of A. heyneana consisting of at least one compound selected from the group ovatodiolide of formula 1 and phyllocladan-16α,17-dihydroxy-19-oic acid of formula 1 together with at least one pharmaceutical acceptable excipient, useful in the treatment of mycobacterial tubercular infections and for leukemia. The pharmaceutical acceptable excipients are selected from diluents, binders, lubricants, wetting agents and disintegrant etc. Further the active ingredients and excipients can be formulated into compositions and dosage forms according to methods known in the art.

(41) The pharmaceutical composition of the present invention may be formulated in form of tablets, capsules, pellets, granules, powder, suspension, syrup, liquid, intravenous, intramuscular injections, topical creams, ointments, gels, etc.

(42) The compound 3 (verbascoside) is obtained from the fraction 14 of the A. heyneana extract of the present invention is found to be effective as dye-sensitizer in dye-sensitized solar cells (DSSCs), wherein solar energy is converted in electric current by using the doctor blading method. The conversion is carried out in the presence of electrolyte such as 0.5M to 0.8M 1 propyl-2,3-dimethyl-imidazolium iodide, 0.05 to 0.2 M LiI, 0.03 to 0.07 M 1.sub.2, and 0.3 to 0.6 M 4-tertbutylpyridine in acetonitrile. The I-V (current vs voltage) characteristics is measured which is depicted in FIG. 4 herein below. Further the measurements of the incident-photon-to-current conversion efficiency (IPCE) are performed by changing the excitation wavelength, and it is observed that compound 3 have ability to convert solar energy into electric energy, which subsequently gives 0.28% conversion efficiency which is described in Table 2 herein below.

(43) TABLE-US-00002 TABLE 2 Solar cell properties of compound 3 Sample Voc (V) Isc (mA) FF (%) Eff. (%) 3 0.49 0.15 59.1 0.28

(44) The compounds of the current invention isolated from A. Heyneana find use in various areas including, but not limited to therapeutics, herbicide, converting solar energy to electricity in dye sensitized solar cells.

(45) In one embodiment, present invention provides a method of treating a subject in need of anti proliferative agent comprising administrating a pharmaceutical composition comprising the compound of formula 1 or compound of formula 2 along with pharmaceutically acceptable excipients.

(46) In yet another embodiment, present invention provides a method of treating a subject suffering from mycobacterial tuberculosis infections comprising administrating a pharmaceutical composition comprising the compound of formula 1 along with pharmaceutically acceptable excipients.

(47) In another embodiment of the present invention, compounds of formula 1 and 2 are useful for anti proliferative activity.

(48) In yet another embodiment, compounds of formula 1 and 2 exhibit 59 to 97% inhibition on Thp-1 cell line at 100 μg/ml.

(49) In yet another embodiment, compounds of formula 1 is useful for the treatment of infections caused due to M. tuberculosis.

(50) In yet another embodiment, compound of formula 1 exhibit an IC.sub.90 of 6.53±0.893 μg/ml in treatment of infections caused due to M. tuberculosis.

(51) In yet another embodiment, compound of formula 3 is useful for the conversion of solar energy into electric current in dye sensitized solar cells.

(52) In another embodiment of the present invention, compound of formula 3 exhibits 0.28% efficiency in conversion of solar energy into electric current in dye sensitized solar cells.

EXAMPLES

(53) The following examples are given by the way of illustration and therefore should not be construed to limit the scope of the invention.

Example 1

(54) Powdered aerial parts of A. heyneana (1.8 kg) were extracted with acetone (3×3 L, 14 h) at 27° C. temperature. The mixture was filtered and concentrated under reduced pressure to provide a greenish acetone extract (45.0 g). Acetone extract, (42.0 g), was separated by column chromatography (CC) with increasing polarity of acetone in petroleum ether to gave 18 fractions (AH1-AH18).

Example 2

(55) Fraction AH7 (1.8 g) was subjected to column chromatography in chloroform with acetonitrile from 2 to 10%. Fraction AH8 (5.2 g) was subjected to column chromatography in chloroform with acetonitrile from 2 to 10%. From fraction 8, sub-fractions 1 and 2 were pooled with sub-fraction 2 of fraction 7 from above column chromatography and compound 1 was obtained from this pool after rechromatography in 2% methanol in chloroform (80 mg).

Example 3

(56) Fraction AH11 (3.4 g) was subjected to column chromatography in 3% methanol in chloroform. Sub-fraction 7 was again subjected to column chromatography in 6% methanol in chloroform. From fraction 2 of this column, compound 2 was precipitated as small crystals (14 mg) which were purified by washing with chloroform.

Example 4

(57) Fraction AH14 (15.6 g) was subjected to column chromatography in chloroform with acetonitrile from 5 to 50%. From sub-fraction 11, compound 3 was isolated as brown amorphous powder (25 mg) after preparative thin layer chromatography in 25% methanol in chloroform.

Example 5

(58) Anti-Mycobacterial Activity

(59) Compounds 1 and 2 were evaluated for their inhibition on M. tuberculosis by following an established protocol published earlier using isoniazid as positive control. The compound 1 exhibited anti-mycobacterial activity with IC.sub.90 of 6.53 μg/ml.

Example 6

(60) Anti Proliferative Activity-MTT Cell Proliferation Assay on Human Thp-1 Cell Line

(61) Compounds 1 and 2 were tested for their inhibitory effect on Thp-1 cells. About 10,000 cells were taken per well in 96-well tissue culture plates and treated with test samples at 100 μg/ml for 72 h. Vehicle control (DMSO, 1%) and positive control (Paclitaxel, 100 μg/ml) was run simultaneously. Cell proliferation was assessed with 10 μl samples from 5 mg/ml stock solution of tetrazolium salt (MTT) was dissolved in cell culture medium and subsequently incubated for additional 1 h at 37° C., 5% of CO.sub.2 and 95% humidity in incubator. The violet coloured formazan crystals formed were solubilized in 200 μl of isopropanol and incubated for another 4 h. The optical density was read on a micro plate reader (Spectramax plus384 plate reader, Molecular Devices Inc) at 490 nm filter against a blank prepared from cell-free wells. Absorbance given by cells treated with the carrier DMSO alone was taken as 100% cell growth. All experiments were performed in triplicate, and the quantitative value was expressed as the average±standard deviation (as shown in Table 1).

Example 7

(62) Solar Cell Sensitizing Activity

(63) Fabrication and Testing of DSSCs. To make and study the DSSC solar cells, present inventors were employed a doctor blading method. After making the films they were annealed at 450° C. for 30 min. For sensitization, the films were dipped in solution of verbascoside for 24 h at room temperature (in separate experiments) and also with 0.5 mM N719 dye in ethanol for comparison. The samples were then rinsed with methanol to remove excess dye on the surface and air-dried at room temperature. This was followed by redox electrolyte addition and top contact of Pt coated FTO. The electrolyte used was 0.6M 1-propyl-2,3-dimethyl-imidazolium iodide, 0.1 M LiI, 0.05 M 12, and 0.5 M 4-tertbutylpyridine in acetonitrile.27 The I-V characteristics were measured under exposure with 100 mW/cm.sup.2 (450W xenon lamp, Newport Instruments), 1 sun AM 1.5, simulated sunlight as a solar simulator. The current was measured using a Kiethley 2400 source. Measurements of the incident-photon-to-current conversion efficiency (IPCE) were performed by changing the excitation wavelength (Newport Instruments).

(64) The compound 3 was evaluated for its ability to convert solar energy into electric current in dye sensitized solar cells. It gave 0.28% conversion efficiency (FIG. 4: voltage vs current plot for compound 3).

Examples 8-21 Examples of Pharmaceutical Composition

Example 8

(65) Composition

(66) TABLE-US-00003 Phyllocladan-16α,17-dihydroxy-19-oic acid 10.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(67) Procedure: Dissolve mannitol in water, add color and flavor to it.

(68) Evaporate water to adsorb color and flavor on mannitol.

(69) Mix active ingredient and Magnesium stearate and fill in pouch or bottle.

(70) Mode of administration:

(71) Disperse the powder in water/juice.

Example 9

(72) Composition

(73) TABLE-US-00004 Phyllocladan-16α,17-dihydroxy-19-oic acid 20.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(74) Procedure: Dissolve mannitol in water, add color and flavor to it.

(75) Evaporate water to adsorb color and flavor on mannitol.

(76) Mix active ingredient and Magnesium stearate and fill in pouch or bottle.

(77) Mode of administration:

(78) Disperse the powder in water/juice.

Example 10

(79) Composition

(80) TABLE-US-00005 Phyllocladan-16α,17-dihydroxy-19-oic acid 10.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(81) Procedure: Dissolve mannitol in water, add color and flavor to it.

(82) Evaporate water to adsorb color and flavor on mannitol.

(83) Mix active ingredient and Magnesium stearate and may be filled in a capsule of suitable size.

(84) Mode of administration: The capsule may be had with water of juice

Example 11

(85) Composition

(86) TABLE-US-00006 Phyllocladan-16α,17-dihydroxy-19-oic acid 20.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(87) Procedure: Dissolve mannitol in water, add color and flavor to it.

(88) Evaporate water to adsorb color and flavor on mannitol.

(89) Mix active ingredient and Magnesium stearate and may be filled in a capsule of suitable size.

(90) Mode of administration;

(91) Mode of administration: The capsule may be had with water of juice

Example 12

(92) Composition

(93) TABLE-US-00007 Phyllocladan-16α,17-dihydroxy-19-oic acid 10.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(94) Procedure: Dissolve mannitol in water, add color and flavor to it.

(95) Evaporate water to adsorb color and flavor on mannitol.

(96) Mix active ingredient and Magnesium stearate and may be compressed as tablet.

(97) Mode of administration;

(98) Mode of administration: The tablet may be taken with water or juice

Example 13

(99) Composition

(100) TABLE-US-00008 Phyllocladan-16α,17-dihydroxy-19-oic acid 5.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(101) Procedure: Dissolve mannitol in water, add color and flavor to it.

(102) Evaporate water to adsorb color and flavor on mannitol.

(103) Mix active ingredient and Magnesium stearate and may be compressed as tablet.

(104) Mode of administration: The tablet may be taken with water or juice

Example 14

(105) Composition

(106) TABLE-US-00009 Phyllocladan-16α,17-dihydroxy-19-oic acid 5.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(107) Procedure: Dissolve mannitol in water, add color and flavor to it.

(108) Evaporate water to adsorb color and flavor on mannitol.

(109) Mix active ingredient and Magnesium stearate and fill in pouch or bottle.

(110) Mode of administration:

(111) Disperse the powder in water/juice.

Example 15

(112) Composition

(113) TABLE-US-00010 Ovatodiolide 10.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(114) Procedure: Dissolve mannitol in water, add color and flavor to it.

(115) Evaporate water to adsorb color and flavor on mannitol.

(116) Mix active ingredient and Magnesium stearate and fill in pouch or bottle.

(117) Mode of administration:

(118) Disperse the powder in water/juice.

Example 16

(119) Composition

(120) TABLE-US-00011 Ovatodiolide 20.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(121) Procedure: Dissolve mannitol in water, add color and flavor to it.

(122) Evaporate water to adsorb color and flavor on mannitol.

(123) Mix active ingredient and Magnesium stearate and fill in pouch or bottle.

(124) Mode of administration:

(125) Disperse the powder in water/juice.

Example 17

(126) Composition

(127) TABLE-US-00012 Ovatodiolide 10.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(128) Procedure: Dissolve mannitol in water, add color and flavor to it.

(129) Evaporate water to adsorb color and flavor on mannitol.

(130) Mix active ingredient and Magnesium stearate and may be filled in a capsule of suitable size.

(131) Mode of administration: The capsule may be had with water of juice

Example 18

(132) Composition

(133) TABLE-US-00013 Ovatodiolide 20.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(134) Procedure: Dissolve mannitol in water, add color and flavor to it.

(135) Evaporate water to adsorb color and flavor on mannitol.

(136) Mix active ingredient and Magnesium stearate and may be filled in a capsule of suitable size.

(137) Mode of administration;

(138) Mode of administration: The capsule may be had with water of juice

Example 19

(139) Composition

(140) TABLE-US-00014 Ovatodiolide 10.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(141) Procedure: Dissolve mannitol in water, add color and flavor to it.

(142) Evaporate water to adsorb color and flavor on mannitol.

(143) Mix active ingredient and Magnesium stearate and may be compressed as tablet.

(144) Mode of administration;

(145) Mode of administration: The tablet may be taken with water or juice

Example 20

(146) Composition

(147) TABLE-US-00015 Ovatodiolide 5.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(148) Procedure: Dissolve mannitol in water, add color and flavor to it.

(149) Evaporate water to adsorb color and flavor on mannitol.

(150) Mix active ingredient and Magnesium stearate and may be compressed as tablet.

(151) Mode of administration: The tablet may be had with water of juice

Example 21

(152) Composition

(153) TABLE-US-00016 Ovatodiolide 5.0% w/w Color Amaranth 0.3% w/w Raspberry Flavor 0.7% w/w Magnesium stearate 2.0% w/w Mannitol q.s. to 100.0% w/w

(154) Procedure: Dissolve mannitol in water, add color and flavor to it.

(155) Evaporate water to adsorb color and flavor on mannitol.

(156) Mix active ingredient and Magnesium stearate and fill in pouch or bottle.

(157) Mode of administration:

(158) Disperse the powder in water/juice.

(159) The structures of compounds 1, 2 and 3 were elucidated using .sup.1H NMR and .sup.13C NMR.

(160) TABLE-US-00017 TABLE 3 .sup.1H NMR and .sup.13C NMR data for compound 1 (in CDCl.sub.3) No. .sup.13C (δ) .sup.1H (δ) 1 42.58 2.42(m)** 2 23.60 2.83(m), 2.12(m) 3 24.76 2.5(m), 2.42(m) 4 131.25 — 5 124.87 4.85 (bt, 5.69) 6 33.14 1.64(m) 7 36.13 2.19(d, 14.61), 2.04(m) 8 139.63 — 9 147.39 7.02(bs) 10 77.85 4.81 (dd, 10.17, 1.42) 11 40.11 2.84(dd, 10.34, 3.61), 2.29(dd, 10.41, 3.65) 12 134.12 — 13 128.88 5.10(d, 10.43) 14 78.72 5.08 (bs) 15 134.30 — 16 172.96 — 17 122.59 5.57(d, 1.5), 6.18(d, 1.5) 18 14.96 1.59(bs) 19 170.20 — 20 19.16 1.72 (d, 0.88)

(161) TABLE-US-00018 TABLE 4 .sup.13C and .sup.1H NMR data of compound 2: (in CD.sub.3OD) 2 (in CD.sub.3OD) No δ.sub.H (J in Hz) δ.sub.C 1 1.70 (eq)*, 41.0 0.89 dt (4.1, 13.3) (ax)* 2 1.37 m (eq), 1.86 m (ax) 20.1 3 2.11m (eq), 1.00m (ax) 39.1 4 — 44.6 5 1.08m 58.3 6 1.83m (eq), 1.70m (ax) 23.1 7 1.66 m, 1.46 dt 42.8 (3.78, 13.45) 8 — 44.9 9 1.1m 57.7 10 — 39.4 11 1.59m(eq), 1.31m(ax) 20.7 12 1.71m (eq), 1.43m (ax) 27.9 13 1.88m 44.8 14 2.10m, 1.04m 49.4 15 2.04 d (11) (eq), 45.2 1.19 bd (11) (ax), 16 — 85.6 17 3.67 d (11), 66.1 3.56 d (11), 18 1.18 s 29.6 19 — 181.7 20 0.84 s 13.9

(162) TABLE-US-00019 TABLE 5 .sup.1H NMR and .sup.13C NMR data for compound 3 in (CD.sub.3OD) .sup.13C .sup.1H Caffeoyl 1  131.42 — 2  117.10 6.68 m 3  146.06 — 4  144.61 — 5  116.29 6.68 m 6  121.25 6.56 bd (7.4) 7  36.50 2.79 bt (9.5) 8  72.24 4.03 q (8.56), 3.72 q (7.8) Gucose 104.12 4.38 d (7.58) 76.14 3.4 bt (8.31) 81.67 3.82 t (9.0) 70.39 4.94 o 75.92 3.6-3.5 m 62.27 3.6-3.5 m Rhamnose 103.00 5.19 bs 72.29 3.93 bs 71.98 3.6-3.5 m 73.73 3.30 o 70.48 3.6-3.5 m 18.44 1.09 d(6.1) Cinnamoyl 1′ 127.38 2′ 115.08 7.06 bs 3′ 150.18 — 4′ 146.94 — 5′ 116.53 6.79 d(7.09) 6′ 123.30 6.95 bd (7.58) 7′ 148.12 7.6 d(15.9) 8′ 114.41 6.275 d(15.9) 9′ 168.36

ADVANTAGE OF THE INVENTION

(163) The compounds isolated are active against proliferative cells

(164) The compounds isolated are active against mycobacterium infection

(165) The invention has applications in solar cell.

(166) Compound 2 has anti proliferative activity comparable to well known drug, paclitaxel.