Drug – bio-affecting and body treating compositions – Plant material or plant extract of undetermined constitution... – Containing or obtained from a tree having matured height of...
Reexamination Certificate
2002-05-13
2003-07-01
Tate, Christopher R. (Department: 1651)
Drug, bio-affecting and body treating compositions
Plant material or plant extract of undetermined constitution...
Containing or obtained from a tree having matured height of...
C424S725000, C514S935000
Reexamination Certificate
active
06586021
ABSTRACT:
FIELD OF THE INVENTION
The invention relates to biologically active extracts obtained from the plant
Salvadora persica
Linneaus 1753. The invention also provides a process for obtaining the crude extract. Further, the invention provides pharmaceutical composition exhibiting biological activity, especially tocolytic activity.
PRIOR ART REFERENCES
The associated mangrove plants,
Salvadora persica
Linneaus 1753, belong to the order Salvadoracea and are shrubs or small trees with white flowers frequent in degraded mangrove swamps and saline banks all over the west coast of India. Large numbers of marine plants have been examined for bioactive substances. Nazarine et al., Indian Journal of Marine Sciences 27:499-501 (1998) have reported promising pharmacological activities in marine organisms from Indian waters.
There are several patents available from all over the world related to processes and compounds from nature for various purposes. Kwak et al. patented a process of extracting and purifying biologically effective ingredients from combined medicinal plants, along with the extract compositions (U.S. Pat. No.5,910,307 issued on Jun. 8, 1999). D'Amelio et al. disclosed therapeutic compositions and methods for treating skin using an extract from the
Centipeda cunninghami
plant (U.S. Pat. No. 5,804,206 issued on Sep. 8, 1998). Zimmerman et al. isolated compounds from the methanolic extract of the eelgrass
Zostrea marina
having significant antifouling aquatic properties (U.S. Pat. No. 5,607,741 issued on Mar. 4, 1997).
Betulinic acid, which is prepared from the compound betulin, has many pharmaceutical potentials. Pezzuto et al. disclosed methods of manufacturing betulinic acid from betulin (U.S. Pat. No. 5,804,575 issued on Sep. 8, 1998). The betulinic acid has been intensively investigated as a potential therapeutic agent for a variety of diseases (U.S. Pat. No. 5,962,527 issued Oct. 5, 1999). Pisha et al., in Nature Medicine 1:1046-1051 (1995), disclosed that betulinic acid has an unexpected selective anti-tumour activity against human melanoma, e.g., MEL-1, MEL-2 and MEL-4. In addition, Fujioka et al.,
J Nat. Prod.
57(2):243-247 (1994), disclosed that betulinic acid has anti-HIV activity in H9 lymphocytic cells. These inventors mentioned that the research directed to betulinic acid as a therapeutic agent is hindered because the betulinic acid is available in very limited quantities and at a very high cost. Ramadoss et al., in U.S. Pat. No. 6,048,847, which issued on Apr. 11, 2000, describes uses of betulinic acid and its derivatives for inhibiting cancer growth and a method of monitoring this. Kang et al. disclosed flavoured product additives in U.S. Pat. No. 5,948,460 issued on Sep. 7, 1999 in which ursolic acid was one of the compounds amongst a group of three compounds that was added to a flavoured product to reduce aftertaste in the product and enhance its sweetness like in a diet drink. It was also used as a constituent in a preparation for inhibition of skin Tumorigenesis.
Herman (U.S. Pat. No. 5,190,979 issued on Mar. 2, 1993) disclosed lupeol also as a compound that can make pharmacologically active terpene ozonides which have medicinal value.
OBJECTS OF THE INVENTION
The main object of the present invention is to provide a process for obtaining a crude extract from stem, leaves, and flowers of
Salvadora persica
, a commonly available shrub in mangrove swamps, and screening to study its bioactivity.
Another object of the invention is to isolate naturally occurring compounds from the plant
Salvadora persica
and identify their molecular weights, molecular formulae, melting points and their structural formulae; which will be helpful in chemical synthesis of these compounds.
Yet another object of the invention is pharmacological screening of crude extract, its fractions and purified compounds to check that the activities shown by the crude extract and fractions are maintained throughout.
Another object of the invention is to provide pharmaceutical compositions containing extracts from the plant
Salvadora persica
and exhibiting biological activity, especially, tocolytic activity.
SUMMARY OF THE INVENTION
The present invention seeks to overcome the drawbacks inherent in the prior art by providing the highly efficient and selective means for processing of active crude extract including fractionation, isolation and purification of the active compounds.
Furthermore, the invention provides pharmaceutical compositions containing the extract obtained from the plant
Salvadora persica
and useful in relieving pains in the uterine muscles.
DETAILED DESCRIPTION OF THE INVENTION
The present invention discloses for the first time the methods of isolation, purification and pharmacological screening of all these above said commercially important compounds from a commonly available plant from mangrove swamps of the west coast of India. The said plant, identified as
Salvadora persica
, is an associated mangrove plant. It is a shrub, and the twigs with leaves of flowers can be hand picked. Even the crude extract and fractions exhibit therapeutic value. The important point is that the biological activity shown by the extract is maintained in the purified compounds such as methyl palmitate and betulin.
This disclosure points towards future potential clinical uses of the extract and fractions for treatment of diseases such as smooth muscle relaxant, bronchial asthma, renal colics and prevention of premature delivery. It further relates to the use of methyl palmitate and betulin in motion sickness and abdominal cramps.
The invention provides methods whereby a biologically active crude extract of an associated mangrove plant identified as
Salvadora persica
Linneaus 1753 is prepared.
Salvadora persica
Linneaus (Salvadoraceae) are shrubs or small trees with white flowers frequent in degraded mangrove swamps and saline banks all over the west coast of India and west Asia. The process disclosed in the invention further relates to the extraction, fractionation and purification of active constituent metabolites of the said plant. The invention is also concerned with the spectral identification of the compounds such as &bgr;-amyrin (non-steroidalpolycyclic triterpene), betulin, ursolic acid (triterpenic acid), methyl palmitate (aliphatic ester) and lupeol (non-steroidalpolycylic triterpene). The invention also deals with molecular formulae, molecular weights, melting points and structural formulae of the said compounds. The invention provides a highly efficient and selective means for processing of active crude extract obtained from
Salvadora persica
, including its fractionation, isolation and purification. As used herein, the term “fractionation” means separating the crude extract. The terms “isolation” and “purification” mean separating the fractions into pure compounds.
The invention further relates to methods of screening pharmacological activities of the said compounds in mammalian tissues. The applicant has found that the crude extract obtained from the plant
Salvadora persica
can be separated into two fractions, i.e., chloroform and aqueous fractions. The aqueous fraction was found to exhibit tocolytic activity, which is described in detail in the present invention.
Accordingly, the invention provides a process of extracting and purifying biologically useful molecules from an associated mangrove plant which comprises the steps of:
i) collecting and processing the plant parts of
Salvadora persica,
ii) preparing a crude extract from the plant parts of
Salvadora persica,
iii) testing the crude extract using methods of pharmacology,
iv) fractionating the crude extract,
v) testing the fractions using methods of pharmacology,
vi) isolating the pure compounds by column chromatography,
vii) testing the pure compounds by using methods of pharmacology, and
viii) identifying the compounds by spectroscopy.
As said earlier, the aqueous fraction of the crude extract obtained from the mangrove plant
Salvadora persica
showed tocolytic activity, while the remaining 2 fractions (
Fernandes Nazarine
Goswami Usha
Council of Scientific & Industrial Research
Piper Rudnick LLP
Tate Christopher R.
Winston Randall
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