Title of Invention

A PROCESS FOR THE ISOLATION OF CAMBOGIN FROM GARCINIA COWA

Abstract A process for the isolation of cambogin from Garcinia The present invention relates to a process for the isolation of cambogin from Garcinia cowa. Among the species Garcinia cowa is found in Northeastern part of India and Andaman Islands. In upper Assam, it is often cultivated in homesteads for its acid fruit. The fruits are edible, dried slices of the fruit are used in curry in place of tamarind/lemon. The process relates to the isolation, purification and identification of cambogin from the fruit rinds of Garcinia cowa. Any of the extracts obtained with the organic solvents can be used as the healthy foods or antioxidants. For the first time, in this invention cambogin from G. cowa. Has been isolated, purified and identified. The process is simple and the solvents used in this process can be regenerated for further use. The left over spent can be further used for the extraction of (-)-Hydroxycitric acid.
Full Text The present invention relates to a process for the isolation of cambogin from Garcinia cowa.
Numerous xanthones and their derivatives have been reported from the various parts of the plant of different Garcinia species (Bennet G.J. and H.H. Lee. 1989. Xanthones from Guttiferae. Phytochemistry 28: 967-998) with wide range of biological and pharmacological activities (antioxidant activity: Minami, H., M. Kinoshita, Y. Fukuyama, M. Kodama, T. Yoshizawa, M. Sugiura, K. Nakagawa and H. Tago. 1994. Antioxidant xanthones from Garcinia subelliptica. Phytochemistry 36: 501-506; cytotoxicity and inhibitory activity of microtubule disassembly: Roux, D., H. A. Hadi, S. Thoret, D. Guenard, O.Thoison, M. Pais, and T. Sevenet. 2000. Structure - activity relationship of poly isoprenyl benzophenones from Garcinia pyrifera on the tubulin/microtubule system. Journal of Natural Product 63: 1070-1076; Thoison, O., J. Fahy, V. Dumontet, A. Chiaroni, C. Riche, M.V. Tri, and T. Sevenet. Cytotoxic prenylxanthones from Garcinia bracteata. Journal of Natural Product 63: 441-446; antimalarial activity: Likhitwidtayawuid, K., W. Chanmahasathien, N. Ruangrungsi, and J. Krungkrai. Xanthones with antimalarial activity from Garcinia dulci. Planta Medica 64: 281-282). A polyisoprenylated benzophenone known as garcinol, isolated from stem bark of G. huillensis has been shown to have chemotherapeutical activity against Gram positive and Gram negative cocci, mycobacteria but inactive against Gram negative enteric bacilli (Bakana, P., M. Claeys, J. Totte, L.A.C. Pieters, L. Van Hoof, Tamba-Vemba van Den, D.A. Berghe and A.J. Vlie-Tink. 1987. Structure and chemotherapeutical activity of a polyisoprenylated benzophenone from the stem bark of Garcinia huillensis. Journal of Ethnopharmacology 21: 75-84) . Alpha-mangostin, rubraxanthone, and xanthochymol isolated from G. mangostana, G. diocia and G. subelliptica respectively showed strong antibacterial activity against both methicillin-resistant and methicillin-sensitive Staphylococcus aureus (linuma, M., H. Tosa, T. Tanaka, F. Asai, Y. Kobayashi, R. Shimano and K. MIyauchl. 1996. Antibacterial activity of xanthones from guttiferaeous plants against methicillin-resistant Staphylococcus aureus. Journal of Pharmacy Pharmacology 48: 861-863; linuma, M., H. Tosa, T. Tanaka, S. Kanamaru, F. Asai, Y. Kobayashi, K. Miyauchi and R. Shimano. 1996. Antibacterial activity of some Garcinia benzophenone derivatives against
methicillin-resistant Staphylococcus aureus. Biological & Pharmaceutical Bulletin 19:311-314).
Garcinia (Family: Guttiferae) is a large genus of polygamous trees or shrubs, distributed in the tropical Asia, Africa and Polynesia. It consists of 180 - 200 species, out of which about 30 species are found in India {The Wealth of India -Raw Materials, Vol. IV, CSIR, New Delhi, India, 1956, pp.99-108). Among the species Garcinia cowa is found in Northeastern part of India and Andaman Islands. In upper Assam, it is often cultivated in homesteads for its acid fruit. The fruits are edible, dried slices of the fruit are used in curry in place of tamarind/lemon.
In this context, the present invention provides a process for the isolation of cambogin from Garcinia cowa, wherein the process relates to the isolation, purification and identification of cambogin from the fruit rinds of Garcinia cowa.
The main object of the present invention is to provide a process for the isolation of cambogin from Garcinia cowa.
In an another object of the present invention, the isolated compound is to be structurally identified.
The inventors have carried out the study to solve the aforementioned problems.
The plants containing the polyisoprenylated benzophenone derivatives described above are some kind of tropical plants belonging to Guttiferae family, for example Garcinia cowa. The dry fruits of Garcinia cowa are currently produced in large amounts as acidulants for cooking in various parts of India, and thus these may be available. In recent years, hydroxycitric acid (an antiobesity agent) is extracted from the dry fruits of G. cowa on large industrial scale, and thus the extraction residue occurring, as the industrial waste can also be available for the present purpose.
The pure compound can be obtained by the synthetic method, which requires a complicated procedure and thus it is desirable to extract the derivatives from G. cowa with a conventional technique for example with organic solvent. The organic solvents include for example (hydrous) methanol, ethanol, acetone and ethyl acetate as well as chloroform, dichloromethane, pentane, hexane, and heptane. Among these solvents, methanol, ethanol, acetone, cyclohexane, benzene and hexane are preferred.
Accordingly, the present invention relates to a process a process for the isolation of cambogin from Garcinia cowa. Comprising:
a) selection of rinds from the species of Garcinia cowa,
b) cutting the rinds of Garcinia cowa manually to a size ranging from 1.5 x 9 mm to 5 x 15 mm,
c) extracting of the above said material as obtained in (i) and (ii) in a Soxhlet extractor at a temperature ranging from 50-66 °C for a period of 0.5-10 h,
d) the solvents selected is from group of cyclohexane, pentane, benzene, acetone, and ethanol,
e) filtering the above solvent mixture extract as obtained in step iii) obtain the particle free extract,
f) concentrating the particle free extract at a temperature in the range of 30 - 40 °C under vacuum in the range of 10-25 mm of mercury to recover the solvent to an extent of 80-95%,
g) drying the above concentrated extract under vacuum oven at the temperature in the range of 30 - 40 °C and vacuum at 05-25 mm of mercury to get dried product,
h) elution of different fractions of dried extract as obtained in step g) using an adsorbant non-polar and medium polar solvents selected from group of hexane, cyclohexane, benzene and acetone to get 500 ml of ten fractions,
i) concentrating different elutes under vacuum and crystallized conditions to obtain cambogin.
In an embodiment of the process the isolated compound is identified as campogin having the structural formula/ structural formule.
The novelty of the process is
1. For the first time, isolation, purification and identification of cambogin from G. cowa.
The advantages of the process is
1. The process is simple and the solvents used in this process can be regenerated for further use.
2. The left over spent can be further used for the extraction of (-)-Hydroxycitric acid.t
(Table Removed)

As described above, any one of the extracts obtained with the organic solvents can be used as the healthy foods or antioxidants according to the present invention.
While the extraction product thus obtained may be used for food and pharmaceutical application the organic solvent is more preferably be removed by the conventional method, for example by a rotary evaporator. Furthermore, the product having the organic solvent removed there from may be subjected to the conventional treatments such as lyophilization and drying with heating.
The polyisoprenylated benzophenone derivative as the active component of the present invention has a vanety of activities useful for maintaining health.
The following examples are given by way of illustration of the present invention and therefore should not be constructed to limit the scope of the present invention.
Example -1
Rinds (100 g) were cut into 1.5 x 5 mm size manually, and it was extracted with 600 ml of hexane in a Soxhiet extractor for 30 min. The extract was filtered by suction through a Buchner filter and washed with hexane until the total amount of the filtrate reached 600 ml. The extract was concentrated under vacuum at 30 °C under 25 mm vacuum. The dried extract was loaded to silica gel column chromatography and it was eluted with hexane, hexane: ethyl acetate, ethyl acetate with increasing polarity. A total ten fractions were collected. Compound (1) was obtained by fraction 7. It was crystallized from chloroform and subjected to spectral analysis. The crystalline product has a melting point of 242-243 °C and a specific rotation of -209.9 degree. UV absorptions at 250 nm and 365 nm, and a molecular weight measured by mass spectrometry of 602. 1H and 13C NMR data of compound (1) have been presented in Table 1. On the basis of 1H, 13C NMR and mass spectral data, the compound (1) has been identified as cambogin.

(Formula Removed)
Table 1. 1H and 13C NMR data of cambogin
(Table Removed)








We Claim:
1) A process for the isolation of cambogin from Garcinia cowa. Comprising:
a. selection of rinds from the species of Garcinia cowa,
b. cutting the rinds of Garcinia cowa manually to a size ranging from
1.5 x 9 mm to 5 x 15 mm,
c. extracting of the above said material as obtained in (a) and (b) in a
Soxhlet extractor at a temperature ranging from 50-66 °C for a
period of 0.5 -10 h, the solvents selected from group of cyclohexane,
pentane, benzene, acetone, and ethanol,
d. filtering the above solvent mixture extract as obtained in step c) to
obtain the particle free extract,
e. concentrating the particle free extract at a temperature in the range
of 30 - 40 °C under vacuum in the range of 10-25 mm of mercury
to recover the solvent to an extent of 80-95%,
f. drying the above concentrated extract under vacuum oven at the
temperature in the range of 30 - 40 °C and vacuum at 05-25 mm of
mercury to get dried product,
g. eluting the different fractions of dried extract as obtained in step f)
using an adsorbant non-polar and medium polar solvents selected
from group of hexane, cyclohexane, benzene and acetone to get
500 ml of ten fractions,
h. concentrating different elutes under vacuum and crystallized conditions to obtain cambogin.
2. A process as claimed in claim 1, the isolated compound is identified as campogin having the structural formula/ structural formule as given below,
(Formula Removed)
3. A process for the isolation of cambogin from Garcinia substantially as herein described with reference to the examples.

Documents:

519-DEL-2004-Abstract-(19-02-2010).pdf

519-del-2004-abstract.pdf

519-DEL-2004-Claims-(19-02-2010).pdf

519-del-2004-claims.pdf

519-DEL-2004-Correspondence-Others-(19-02-2010).pdf

519-del-2004-correspondence-others.pdf

519-del-2004-correspondence-po.pdf

519-del-2004-correspondence.pdf

519-DEL-2004-Description (Complete)-(19-02-2010).pdf

519-del-2004-description (complete).pdf

519-del-2004-description.pdf

519-del-2004-form-1.pdf

519-del-2004-form-18.pdf

519-del-2004-form-2.pdf

519-del-2004-form-3.pdf

519-del-2004-form-5.pdf

519-del-2004-form1.pdf

519-del-2004-form2.pdf

519-del-2004-form3.pdf

519-del-2004-form5.pdf


Patent Number 242799
Indian Patent Application Number 519/DEL/2004
PG Journal Number 38/2010
Publication Date 17-Sep-2010
Grant Date 13-Sep-2010
Date of Filing 19-Mar-2004
Name of Patentee COUNCIL OF SCIENTIFIC AND INDUSTRIAL RESEARCH
Applicant Address RAFI MARG, NEW DELHI-110001, INDIA.
Inventors:
# Inventor's Name Inventor's Address
1 GUDDADARANGAVVANAHALLY KRISHNAREDDY JAYAPRAKASHA CENTRAL FOOD TECHNOLOGICAL RESEARCH INSTITUTE, MYSORE.
2 LINGAMALLU JAGAN MOHAN RAO CENTRAL FOOD TECHNOLOGICAL RESEARCH INSTITUTE, MYSORE.
3 MANDYAM CHAKRAVARATHY VARADARAJ CENTRAL FOOD TECHNOLOGICAL RESEARCH INSTITUTE, MYSORE.
4 BHABANI SANKAR JENA CENTRAL FOOD TECHNOLOGICAL RESEARCH INSTITUTE, MYSORE.
PCT International Classification Number A 61 K 31 /35
PCT International Application Number N/A
PCT International Filing date
PCT Conventions:
# PCT Application Number Date of Convention Priority Country
1 NA