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
2002-06-26
2003-10-28
Lankford, Jr., Leon B. (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
Reexamination Certificate
active
06638546
ABSTRACT:
FIELD OF INVENTION
The present invention relates to a novel muscurine antagonist obtained from an associated mangrove plant called
Salvadora persica
Linneaus 1753 collected from estuarine mangrove swamps of Goa, India. Particularly, the present invention relates to the use of methyl palmitate isolated from the plant
Salvadora persica
as a muscurine antagonist.
BACKGROUND OF THE INVENTION
Methyl palmitate is a fatty acid methyl ester. It is prepared commercially from naturally occurring edible fats and oils. These compounds are widely used indirectly in a wide range of food, pharmaceutical, cosmetic and industrial applications (Pearson, R., Report of the FAME (Fatty Acid Methyl Esters) Task Force, Jan. 9, 1997). In the same report, data on methyl esters including methyl palmitate, which were tested for oral toxicity in rats, is given. Methyl palmitate was proved to be nontoxic. The report also described that administration of an emulsion of methyl palmitate to mice by oral intubation or intraperitoneal injection produced no alteration in organ weight or phagocytic function of the reticuoendothelial system. In Environment Mutagen 8(Suppl 7):1-119 (1986) (quoted from Pearson, R., supra), it was reported that methyl palmitate was devoid of any mutagenic effect based on the tests on Salmonella and mammalian microsome assays. The methyl palmitate though showed sensitivity of rabbit's skin to the compound, the results with human skin proved to have a very mild effect. Methyl esters of fatty acids can be used as supplementary source of fat for animal feeds.
“Muscarinic actions” are the actions produced as a result of acetylcholine released from the post-ganglionic parasympathetic nerve endings or the actions resulting from exogenously administered acetylcholine on the receptors of organs with post-ganglionic parasympathetic nerve supply. The muscarinic actions of both endogenously released, as well as exogenously administered, acetylcholine are blocked by atropine. The designation “muscarinic actions” comes from the fact that these actions are similar to those produced by the poisonous mushroom alkaloid muscarine (R. S. Satoskar and S. D. Bhandarkar, Pharmacology and pharmacotherapeutics, Vol.1., Popular Prakashan, Bombay (1990).
Acetylcholine acts on two types of receptors, namely muscarinic and nicotinic (Goodman & Gilman's: The Pharmacological Basis of Therapeutics, 9
th
edition, McGraw-Hill Health Professions Division, N.Y.; pp. 278-279 (1997)). Since the activity of the crude extract of the
Salvadora persica
plant was seen on smooth muscle, only muscarinic receptors have been studied (Nazarine, F., Ph.D. Thesis, Goa University, 1998).
A large number of binding and functional studies have indicated the existence of 5 subtypes of muscarinic receptors namely, M
1
, M
2
, M
3
, M4 and M
5
(N. J. M. Birdsall and F. C. Hulme,
Trends In Pharmacological Sciences
, 4:459-461 (1983)). The most important are the M
1
, M
2
and M
3
muscarinic receptor subtypes. M
1
receptors are found in the stomach and mediate gastric secretions, whereas M
2
receptors are found in the myocardium of the heart and M
3
are found in the ileum and bladder detrusor muscles and cause smooth muscle contractions.
Thompson et al., in PCT Patent Application No. WO 97/16192, published on May 9, 1997, disclose the antimuscarinic activity of derivatives of piperdin and describe their use in the treatment and/or prevention of myopia, commonly known as near-sightedness.
Haertel et al., in DE Patent No. 4136811 published on May 13, 1993, disclose a skin cleanser especially for removing paint containing methyl, ethyl and/or isopropyl palmitate with some solvents and emulsifiers. Jeanne et al., in U.S. Pat. No. 5,109,022, issued on Apr. 28, 1992, describe a composition which contains methyl palmitate and some other compounds and is useful for repelling flying, biting and stinging insects. U.S. Pat. No. 6,039,950, issued on Mar. 21, 2000, describes formulations of several pharmaceutical grade compounds including fatty esters from the Saw Palmetto, which is a small palm found in the southern United States. U.S. Pat. No. 6,049,013, issued on Apr., 11, 2000, discloses that in methods of producing alcohols, the fatty acids (one example is of methyl palmitate) could be extracted from coconut, palm and palm kernel for this use.
U.S. Pat. No. 5,821,249, which issued on Oct. 13, 1998, describes production of anhydroecgonine methylester (MEG) by heating cocaine base (“crack”) and their use as anticholinergic agents. MEG alone and in combination with cocaine was tested for action on isolated tracheal rings stimulated to contact with acetylcholine.
There is no patent available on the antimuscarinic activity of methyl palmitate. The present invention for the first time discloses the effect of methyl palmitate on M
1
receptors by observing gastric acidity by pyloric ligation under basal conditions. Like atropine, the compound methyl palmitate decreased free acidity. However, in the studies conducted, it was 50 times less potent than the standard antagonist atropine. M
2
muscarinic receptors found in the conducting tissue and the myocardium of the heart mediate both the bradycardia and the negative inotropic effect of cholinergic stimulation. The present invention relates to the comparative effect of methyl palmitate and atropine on the rate and force of contraction of the guinea pig atria. It describes the inhibitory effect of atropine. The invention describes that the compound methyl palmitate had no such effect in the dosage range used showing that it probably does not block M
2
receptors. Though the reason for this effect is not known yet, the present invention for the first time describes the ability of the compound methyl palmitate to distinguish between the muscarinic receptors in the atrium and elsewhere in the body. The very reason that methyl palmitate does not affect the M
2
receptors of the atrium confirms the heterogeneity of muscarinic receptors.
As used herein the term “antimuscarinic activity” means the antagonistic activity on muscarinic receptors, where “antagonistic activity” refers to the inhibitory effect of atropine and the said compound. The “heterogeneity of muscarinic receptors” means the ability of the said compound to distinguish between the muscarinic receptors in the atrium and elsewhere in the body. The “selectivity of the compound” means its inhibitory action only on M
1
and M
3
muscarinic receptors. The “competitive antagonism” means the compound produced a shift to the right in the dose response curves of the acetylcholine with the maximum response remaining the same. In gastric acidity experiments, the compound was injected subcutaneously as per the body weight of the rat measured in kilograms and written as Kg. s.c. where “s.c.” means subcutaneous.
The approach adopted for antagonistic activity screening by pharmacological methods is described for the first time for this compound. The invention describes that methyl palmitate produces a shift to the right in the dose response curves of acetylcholine with the maximal response remaining the same. It further discloses that the competitive antagonism shown by the said compound is reversible in nature. Methyl palmitate blocks M
3
receptors in the same way as atropine, though it is 2.2 times less potent.
OBJECTS OF THE INVENTION
The main object of the present invention is to study the compound methyl palmitate purified from the extract of the plant
Salvadora persica
for its antimuscarinic activity.
Another object of the present invention is to study the heterogeneity of methyl palmitate as a muscarinic receptor.
Still another object of the present invention is to study the comparative effect of methyl palmitate and atropine during in vivo and in vitro experiments.
One more object of the present invention is to detect the effect of methyl palmitate on the dose response curve of acetylcholine.
One other object of the present invention is to detect the effect of methyl palmitate in rats.
Another object of the present invention is to
Fernandes Nazarine
Goswami Usha
Council of Scientific & Industrial Research
Davis Ruth A.
Kelber Steven B.
Lankford , Jr. Leon B.
Piper Rudnick LLP
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