FIELD OF THE INVENTION:

The present invention relates to a novel bio-active compound. More particularly, the present invention relates to an anti-oxidative, anti-angiogenic, anti-cancer and anti-diabetic properties exhibiting compound for treating angiogenic as well as cancer and diabetic diseases.

BACKGROUND OF THE INVENTION AND PRIOR ART:

Over the last few years, there has been emerging interest in the exploration of natural organisms (terrestrial and marine) for the identification of novel bio-active compounds for drug development. Extensive survey reveals that more than 60% of available approved anti-cancer drugs were either of natural origin or were modeled on natural product lead compounds (Cragg et al Nat. Prod. 1997, 60, 52-60). Therefore, there exists an urgent need to intensify natural product drug discovery research in the forthcoming years.

Bioactive compounds in plants are compounds produced by plants having pharmacological effects in man and animals. The typical bioactive compounds in plants are produced as secondary metabolites. The main group of bioactive compounds present in plants are glycosides, flavonoids, tannins, terpenoids, resins, lignans, alkaloids, proteins, peptides etc. These compounds may elicit a long range of different effects in man and animals eating the plants dependent on plant species and amount eaten. Plants with potent bioactive compounds are often characterized as medicinal, and beneficial.

US 7709031 discloses presence of anti-angiogenic agents from plant extracts of Chinese blackberry (Rubus suavissimus).

US 7416744 discloses a pharmaceutical compositions containing an extract obtainable from a plant of the genus Trichocaulon or Hoodia having anti-diabetic activity; and to the use of such extracts and its analogues for the manufacture of medicaments having anti¬diabetic activity.

US 7410656 discloses a compound or group of compounds present in an active principle derived from plants of species Euphorbia peplus, Euphorbia hirta and Euphorbia drummondii, and to pharmaceutical compositions comprising these compounds. Extracts from these plants have been found to show selective cytotoxicity against several different cancer cell lines. The compounds are useful in effective treatment of cancers, particularly malignant melanomas and squamous cell carcinomas (SCCs).

US 7763287 relates to extract of Cercis chinensis having anti-oxidant activity and anti-aging activity containing compound and cosmetical composition for anti-oxidation, skin-aging protection and wrinkle improvement containing the extract as effective ingredient.

The prior arts discussed above disclose various plant extracts, exhibiting various medical values. From this, it is evident that plants are excellent resources for extracting various bio-active compounds for drug development. So, there exists an immediate need to exploit various plants for the extraction of novel bioactive compounds exhibiting various medicinal properties.

OBJECT OF THE INVENTION:

The main object of the present invention is to provide a novel bioactive compound for treating angiogenic as well as cancer and diabetic diseases.

Another object of the present invention is to provide an anti-oxidative, anti-angiogenic anti-cancer and anti-diabetic properties exhibiting compound for treating angiogenic as well as cancer and diabetic diseases.

Yet another object of the present invention is to provide Mazapotin [ 6-(tetrahydro-3, 4, 5-trihydroxy-6-methyl-2H-pyran-2-yl)-3,5,7-trihydroxy-2-(4-hydroxy-3-(3-methylbut-2-enyl)phenyl)-4H-chromen-4-one ; Quercetin-2-enyl-6-rhamnose ] compound for treating angiogenic as well as cancer and diabetic diseases.

Yet another object of the present invention is to formulate a pharmaceutical composition comprising of Mazapotin.

Further object of the present invention is to utilize the formulated pharmaceutical composition for treating angiogenic as well as cancer and diabetic diseases.

BRIEF DESCRIPTION OF THE DRAWINGS:

FIG.1 depicts the UV spectrum of the novel flavonoid Mazapotin (Formula 1) in various shift reagents. FIG.2 depicts the FT-IR spectrum of the novel flavonoid Mazapotin (Formula 1) FIG.3 depicts the 1H-NMR spectrum of the novel flavonoid Mazapotin (Formula 1) FIG.4 depicts the 1H-NMR spectrum in D20 of the novel flavonoid Mazapotin (Formula-!) FIG.5 depicts the 13C-NMR spectrum of the novel flavonoid Mazapotin (Formula 1) FIG.6 depicts the Mass spectrum of the novel flavonoid Mazapotin (Formula 1)

FIG.7 depicts the DPPH radical scavenging assay of novel flavonoid Mazapotin (Formula 1) FIG.8 depicts the antiangiogenic activity of flavonoid Mazapotin(Formula 1) FIG.9 depicts the anti-cancer activity of novel flavonoid Mazapotin(Formula 1) FIG.10 depicts the a-amylase enzyme inhibition activity of novel flavonoid Mazapotin (Formula 1)

SUMMERY OF THE INVENTION:
The present invention discloses a novel anti-oxidative, anti-angiogenic, anti-cancer and anti-diabetic properties exhibiting compound for treating angiogenic as well as cancer and diabetic diseases. The compound is Mazapotin [ 6-(tetrahydro-3, 4, 5-trihydroxy-6-methyl-2H-pyran-2-yl)-3,5,7-trihydroxy-2-(4-hydroxy-3-(3-methylbut-2-enyl)phenyl)-4H-chromen-4-one ; Quercetin-2-enyl-6-rhamnose ] with the structure of formula 1. The invention further discloses a pharmaceutical composition comprises of a therapeutically effective amount of compound of formula 1 and a pharmaceuticaly acceptable carrier, diluent, or excipient.

DETAILED DESCRIPTION OF THE INVENTION:

The present invention discloses a novel anti-oxidative, anti-angiogenic, anti-cancer and anti-diabetic properties exhibiting compound Mazapotin [ 6-(tetrahydro-3, 4, 5-trihydroxy-6-methyl-2H-pyran-2-yl)-3,5,7-trihydroxy-2-(4-hydroxy-3-(3-methylbut-2-enyl)phenyl)-4H-chromen-4-one ; Quercetin-2-enyl-6-rhamnoside ] with the structure of formula 1 for treating angiogenic as well as cancer and diabetic diseases.

The compound Mazapotin is extracted from Manilkara zapota.

The physical characteristics of the compound are determined and found to be yellow powder in nature with melting point of 285±2°C . The compound is soluble in methanol and water and exhibits UV MeoH Amax at 226 nm, 263 nm and 349 nm.

The UV spectrum of the novel flavonoid Mazapotin (Formula 1) was studied in various shift reagents (FIG. 1). The UV spectrum in methanol was observed at 226, 263, 349 nm. The UV absorption showed the unsaturated aromatic nature and presence of chromophore groups of the novel flavonoid Mazapotin

The IR absorption regions of the novel flavonoid Mazapotin (Formula 1) are given in TABLE 1. Absorption region at 3259.85 cm"1 suggested the presence of OH group (FIG. 2).

TABLE 1. Absorption regions in FT-IR spectrum.

FIG.3 and 4 are the 1H-NMR spectrum of the novel flavonoid Mazapotin (Formula 1). A signal ranging from 6 at 0.82-12.68 indicated that the novel compound Mazapotin (Formula 1) was aromatic. Signals were observed in the aromatic region and therefore the compound was confirmed to be aromatic. The D2O exchange spectrum also supports the presence of aromatic OH groups. A signal 5 at 12.96 indicated the presence of 5th OH group in the aromatic ring. A signal 5 at 10.96 indicated the presence of 7th OH group in the aromatic ring. A signal 5 at 9.32 indicated the presence of 4' OH group in the aromatic ring. The resonance of 11 sugar protons as multiplet, 5 at 0.82 - 3.97 was observed and the presence of equatorial anomeric proton 5 at 4.61 showed the presence of the sugar moiety.

FIG.5 is the 13C-NMR spectrum of the novel flavonoid Mazapotin (Formula 1). From the 13C-NMR spectrum, it was observed that the compound Mazapotin (Formula 1) contain 24 signals. The values 5 at 17.42, 69.93, 70.29, 70.51 and 71.17 showed the presence of rhamnose sugar moiety. The values 6 at 177.7 showed the presence of carbonyl group.

FIG.6 is the Mass spectrum of the novel flavonoid Mazapotin (Formula 1). The molecular weight of the novel flavonoid Mazapotin (Formula 1) was found to be 499, which was supported by the mass spectrum.
Based on the comparison of the physical characteristics and spectral data, the isolated compound Mazapotin (Formula 1) was conformed to be a flavonoid. It was identified as 6-(tetrahydro-3, 4, 5-trihydroxy-6-methyl-2H-pyran-2-yl)-3,5,7-trihydroxy-2-(4-hydroxy-3-(3-methylbut-2-enyl)phenyl)-4H-chromen-4-one ; Quercetin-2-enyl-6-rhamnoside and it is a novel compound and it is named as Mazapotin (Formula 1) . This is the first report from the leaves of Manilkara zapota. The structure of Mazapotin (Formula 1) is shown below:


6-(tetrahydro-3, 4, 5-trihydroxy-6-methyl-2H-pyran-2-yl)-3,5,7-trihydroxy-2-(4-hydroxy-3-(3-methylbut-2-enyl)phenyl)-4H-chromen-4-one ;

Quercetin-2-enyl-6-rhamnoside
Biological activities of the compound were studied to ascertain its anti-oxidative, anti-angiogenic, anti-cancer and anti-diabetic properties.

Antioxidant activity of novel flavonoid Mazapotin was determined by DPPH free radical scavenging activity. The activity was carried out at various concentrations of Mazapotin.

DPPH' is a purple colored chromophore which is reduced to DPPH on a concentration dependant manner upon addition of the novel flavonoid Mazapotin. This is visualized as a discoloration from purple to yellow.
TABLE 2 and FIG.7 are the results of DPPH free radical scavenging activity of the Mazapotin and the standard. For Mazapotin at 7 ug/mL concentration, the percentage of DPPH free radical scavenging activity was 98.34 and for the standard Quercetin, it was 67.17. The IC50 value of Mazapotin was 1.93 (ug/mL), whereas it was 4.06 (ug/mL) for the standard Quercetin.

TABLE 2. DPPH free radical scavenging activity of novel flavonoid Mazapotin.

The results showed that the novel flavonoid Mazapotin was capable of donating hydrogen as well as electron to free radicals in order to remove odd electrons which are responsible for radical's reactivity. This invention provides a definitive report about the free radical scavenging capacity of novel flavonoid Mazapotin, since the antioxidant activity of a drug may depend on the free radical scavenging activity. Therefore, such phenomena suggest that the novel flavonoid Mazapotin act as free radicals scavenger and react with radicals to convert them to more stable products and terminate radical chain reaction.
CAM assay (chorioallantoic membrane) was used to evaluate anti-angiogenic activity. In the present invention on reduction of vascularization, the novel flavonoid Mazapotin was examined for vascularization inhibitory activity on CAM model. TABLE 3 and FIG.8 are the results of vascular inhibitory activity of the novel flavonoid Mazapotin. At 0.5 mg/pellet concentration, the percentage of angiogenesis inhibition of the flavonoid Mazapotin was 57.25.

TABLE 3. Antiangiogenic activity of flavonoid Mazapotin.

The results indicated that the novel flavonoid Mazapotin inhibit angiogenesis in vivo in a dose dependent manner. The results showed that novel flavonoid Mazapotin may also inhibit the tumor spread by the action of blocking angiogenesis.

The effect of the novel flavonoid Mazapotin on breast cancer cell line MCF 7 was carried out. TABLE 4 and FIG.9 are the results of anticancer activity of the novel flavonoid Mazapotin. At 25 ug/ mL concentration of Mazapotin, the MCF7 cell death was 85.42%. The IC5o value of cell death was at 7.61 ug/mL of Mazapotin. At 25 ug/ mL concentration of standard (Doxorubicin), the MCF 7 cell death was 96.29%. The IC50 value of cell death was 4.97 ug/mL of Doxorubicin. At the same concentration of standard (Quercetin), the MCF 7 cell death was 68.75%. The IC5o value of cell death was 17.49 µg/mL of Quercetin.

TABLE 4. Anti-cancer activity of the novel flavonoid Mazapotin.

From the results, it is inferred that the novel flavonoid Mazapotin prevents the binding of carcinogens to DNA and strengthens the connective tissue, which may keep cancer cells from spreading. The novel flavonoid Mazapotin has the ability to inhibit mutations within the cell's DNA. Furthermore, the novel flavonoid Mazapotin is considered to be a cancer inhibitor which has the ability to cause apoptosis in cancer cells.

In this invention on reduction of diabetes risk, Mazapotin was examined for a-amylase inhibitory activity. Porcine pancreas a-amylase was used to evaluate anti-diabetic activity of Mazapotin. TABLE 5 and FIG.10 are the results of a-amylase inhibitory activity of the novel flavonoid Mazapotin and the standard Acarbose. For the novel flavonoid Mazapotin at 35 µg/mL concentration, the inhibition of a-amylase activity was 66.66% and for the standard, it was 98.16%. The IC5o value of Mazapotin was 23.57 µg/mL, whereas it was 5.23 µg/mL for the standard Acarbose.

TABLE 5. a-amylase enzyme inhibition activity of novel flavonoid Mazapotin.

The results showed that novel flavonoid Mazapotin was inhibiting the action of a-amylase enzyme activity on starch in a concentration dependant manner.

In one of the preferred embodiment, the present invention shall disclose an anti-oxidative, anti-angiogenic, anti-cancer and anti-diabetic properties exhibiting compound Mazapotin [ 6-(tetrahydro-3, 4, 5-trihydroxy-6-methyl-2H-pyran-2-yl)-3,5,7- trihydroxy-2-(4-hydroxy-3-(3-methylbut-2-enyl)phenyl)-4H-chromen-4-one ; Quercetin-2-enyl-6-rhamnoside ] of formula 1 for treating angiogenic as well as cancer and diabetic diseases.


As per the invention.the compound Mazapotin has an anti-oxidant activity in the range of 35.29% to 98.34% for 1-7 µg per mL concentration.

According to the invention, the compound Mazapotin has an anti-angiogenic activity of 82 branch points of blood vessels for 0.25 mg of the compound per pellet and 53 branch points of blood vessels for 0.50 mg of the compound per pellet.

As per the invention the compound Mazapotin has 85.42% of MCF7 cell death at 25 ug/ mL concentration.

In accordance with the invention, the compound Mazapotin has an anti-diabetic activity in the range of 18.18% to 66.66% for 5-35 ug per mL concentration of the compound Mazapotin.

In another preferred embodiment, the present invention shall disclose a pharmaceutical composition exhibiting anti-oxidative, anti-angiogenic, anti-cancer and anti-diabetic properties for treating angiogenic as well as cancer and diabetic diseases. The composition comprises of a therapeutically effective amount of compound Mazapotin [ 6-(tetrahydro-3, 4, 5-trihydroxy-6-methyl-2H-pyran-2-yl)-3,5,7- trihydroxy-2-(4-hydroxy-3-(3-methylbut-2-enyl)phenyl)-4H-chromen-4-one ; Quercetin-2-enyl-6-rhamnoside ] of formula 1
and a pharmaceuticaly acceptable carrier, diluent, or excipient.

In accordance with the invention, the composition is formulated as at least one member of the group comprising of tablet, a pill, a capsule, oral syrup, a caplet, a troche, a pouch, or sprinkles.

Although preferred embodiments have been depicted and described in detail herein, it will be apparent to those skilled in the relevant art that various modifications, additions, substitutions, and the like can be made without departing from the spirit of the invention and these are therefore considered to be within the scope of the invention as defined in the claims which follow.

We Claim,

1. An anti-oxidative, anti-angiogenic, anti-cancer and anti-diabetic properties exhibiting compound Mazapotin [ 6-(tetrahydro-3, 4, 5-trihydroxy-6-methyl-2H- pyran-2-yl) 3,5,7- trihydroxy-2-(4-hydroxy-3-(3-methylbut-2-enyl)phenyl)-4H-chromen-4-one ; Quercetin-2-enyl-6-rhamnoside ] for treating angiogenic as
well as cancer and diabetic diseases, the said compound comprises of

2. The compound as claimed in claim 1 has an anti-oxidant activity in the range of 35.29 to 98.34% for 1-7 ug per mL concentration of the compound Mazapotin.

3. The compound as claimed in claim 1 has an anti-angiogenic activity of 82 branch points of blood vessels for 0.25 mg of the compound Mazapotin per pellet and of 53 branch points of blood vessels for 0.50 mg of the compound Mazapotin per pellet,

4. The compound as claimed in claim 1 has 85.42% of MCF7 cell death at 25 ug/ mL concentration of Mazapotin,

5. The compound as claimed in claim 1 has an anti-diabetic activity of in the range of 18.18 to 66.66% for 5-35 ug per mL concentration of the compound Mazapotin.

6. A pharmaceutical composition exhibiting anti-oxidative, anti-angiogenic, anti-cancer and anti-diabetic properties for treating angiogenic as well as cancer and diabetic diseases, the said composition comprises of a therapeutically effective amount of compound Mazapotin [ 6-(tetrahydro-3, 4, 5-trihydroxy-6-methyl-2H-pyran-2-yl)-3,5,7- trihydroxy-2-(4-hydroxy-3-(3-methylbut-2-enyl)phenyl)-4H-chromen-4-one ; Quercetin-2-enyl-6-rhamnoside ] of formula 1

and a pharmaceutical acceptable carrier, diluent, or excipient.

7. The composition as claimed in claim 1 is formulated as at least one member of the group comprising of tablet, a pill, a capsule, oral syrup, a caplet, a troche, a pouch, or sprinkles.