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Description  |
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The present invention relates to a new pharmaceutical composition
containing a combination of products used in tissue-metabolic and
energetic biochemical process, such as Coenzyme Q.sub.10 and carnitine or
acetylcarnitine. Preferably, the composition according to the invention
contains carnitine (or acetylcarnitine) in L form.
As it is well-known, carnitine is .beta.-hydroxy-.gamma.-trimethylamino
butirric acid, whereas Coenzyme Q.sub.10 stands for
2,3-dimethoxy-5-methyl-6-decaprenyl-benzoquinone.
The composition according to the invention is able to develop an effective
therapeutic action in the prevention and treatment of atherosclerotic
phenomena, in myocardial and coronary insufficiency, as well as in
hypertension and in general in pathological conditions due to tissue
anoxia.
L-carnitine is derived from lysine through steps including the
transformation of .epsilon.-N-trimethyl-lysine into .gamma.-butyrrobetaine
with the intervention of hydroxylase and proteino-methyl-transferase
systems.
Already in 1952 it was observed (Carter et al.-Arch. Biochem. Biophys.
38-405-1952) that in the absence of carnitine Tenebrio moliter larvae died
because of fat accumulation, and the explanation of this phenomenon can
now be found through the biochemical observations concerning the carnitine
function. Carnitine can be in fact acetylated at the expense of acetyl-CoA
and it can stimulate the oxidation of fatty acids (Friedman S., Fraenkel
G.-Arch. Biochem. Biophys., 59-491-1955).
Since carnitine is able to receive the acyl group of acyl-COA, is also able
to provide this coenzyme for its catalytic functions and particularly for
the Krebs cycle and thereby catalytically exerting an energetic function
(Siliprandi N.-Biochem. J., 95-777-1965).
Coenzyme Q.sub.10 also plays an important role in the cell energetic
metabolism.
Coenzyme Q.sub.10, in fact, serves as high mobility electrons carrier
between the flavoproteins and the cytochromes of the respiratory system
(Morton R. A., Nature 182-1764-1958; Gale P. et al., Arch. Biochem.
Biophys., 93-211-1961).
The beta-oxidation processes of fatty acids take place in the mitochondrion
and they need as enzymes FAD and NAD that are oxidized again by means of
the respiratory system, where Coenzyme Q.sub.10 acts as an essential
element.
In turn fatty acids, in order to be able to cross the mitochondrial
membrane, must be transformed into acyl-carnitine.
In short, therefore, carnitine lets the acyle groups cross the miochondrial
membrane.
The carnitine system is therefore a shuttle system through which acyl-CoAs
can go into and out of the mitochondrion without having to use energy.
Once they are inside the mitochondrion, acyl-CoA are oxidized by means of
.beta.-oxidation process.
The biochemical processes binding at the different metabolic steps the
action both of carnitine and of Coenzyme Q and that show the close
interdependence of these two different systems for the cellular and
tissutal and metabolic energetic function, also correlate with the
biologic observation among the different activities that both carnitine
and Coenzyme Q are able to carry out.
By means of the composition of the present invention it has been possible
to notice the synergism that these two substances exert at the level of
the myocardial functional capacity and coronaric circulation, and at the
level of fat metabolism alterations and at vascular level.
The following pharmacological tests show the activity of the new
pharmaceutical composition, and they point out the synergic effect
obtained by the use of this composition.
Another aspect of the synergism resulting from the carnitine-Coenzyme Q
association, is the one relating to the different activity of the two
compounds in the prevention of many pathological conditions, such as, for
example, tissue anoxia. On the one hand, in fact, carnitine, through the
beta-oxidation of fatty acids, is able to restore the energy supplies
necessary to cell-life, whereas Coenzyme Q is able to restore the ATP
supplies necessary for the energetic metabolic processes of the cell.
The latter mechanism connected with Coenzyme Q action can be possibly
attributed to its intervention at calcium ions level and to a block of
their entry at mitochondrial level, which would imply a reduction of the
ATP levels and also the activation of the proteinases subsequently leading
to the destruction of cell-structures.
The effectiveness of the present invention is not, however, bound to the
control of the biologic mechanisms so far exposed.
TOXICOLOGICAL TESTS
Both carnitine and Coenzyme Q do not possess a toxicity of their own if
administered in association per os in the animal, as it had been possible
to ascertain administering very high doses of Coenzyme and L-carnitine to
Swiss mice and Wistar rats of both sexes. In fact, the administration per
os of 1 g of Coenzyme Q associated to 1 g, 2 g, 5 g of carnitine did not
cause any death in the animals thus treated.
Even the parenteral administration of high doses of associated carnitine
and Coenzyme Q did not show any toxic effect or any kind of intolerance,
and it was not possible in the same way, to ascertain a significant
mortality in the animals with the injection of a composition containing mg
400 of Coenzyme Q and g 2 of carnitine.
Also the chronic toxicological tests (3 months) carried out both on
Sprague-Dawley rats and in Beagle dogs (100 mg of Coenzyme Q and 500 mg of
carnitine/day per os) confirmed the good tolerance and the lack of
toxicity of the association Coenzyme Q-carnitine.
PHARMACOLOGICAL TEST
Tests on the Pitrexine-Induced Myocardial Anoxia
These tests showed how myocardial anoxia induced by the coronaric spasm
caused by the venous administration of pitrexine in the rat could be
inhibited by the administration of the Coenzyme Q-carnitine combination in
comparison with the two individual components of the combination itself.
The venous injection of 1 unit/kg of pitrexine in the rat causes a
coronaric spasm with subsequent decreased coronaric vascularization and
myocardial oxygenization that can be electrocardiographically recorded
through the characteristic T wave (Lindler's T wave).
From the tests that were carried out it was possible to ascertain that the
administration of Coenzyme Q, contrary to the administration of carnitine
only, decreases the amplitude of the T wave induced by the pitrexine
injection, but that with the association of the two drugs (Coenzyme
Q.sub.10 mg 50+carnitine mg 250 i.p.) it is possible to obtain the virtual
disappearance of this wave, that is the inhibition of the myocardial
anoxic tissue processes.
It is therefore evident the synergism existing between the activity of
carnitine and the activity of Coenzyme Q.
As a matter of fact, no rat treated with the Coenzyme Q-carnitine shows
alterations against the T wave after administration of pitrexine, whereas
more than 90% of the rats treated with carnitine only and more than 60% of
the rats treated with Coenzyme Q only show this electrocardiographic sign
due to the myocardial anoxia.
EVALUATION OF THE ANTIANOXIC CEREBRAL ACTIVITY
In order to evaluate the antianoxic activity also at a cerebral level,
rabbits were used that were put into a close, airtight cage in which the
air was subsequently substituted through nitrogen admission.
The re-admission of air in the cage can let the animal recover and keep it
back to normality.
It is possible to evaluate the antianoxic activity of a substance by the
comparison of the times necessary between control animals and treated
animals for the anoxic electro-encepalographic signs to appear, as well as
by the recovery-time necessary to achieve a normal electro-encephalogram
after the air has been again re-admitted in the cage. The anoxia caused by
the progressive substitution of the air with nitrogen produces
electro-encephalographic alterations until electric silence is reached;
these alterations are easily detectable by the electro-encephalogram.
In these tests we could ascertain that Coenzyme Q only (mg 50/kg i.p. or mg
200 per os) is able to induce a significant difference, between control
groups and treated groups, of the time necessary for cerebral anoxia
phenomena to appear, as well as for the recovery time necessary to go back
to a normal electro-encephalogram: these times become thus much more
significant, in case that carnitine is associated to Coenzyme Q (mg 250/kg
i.p. or mg 500 per os), that by itself, in these tests, does not result to
cause any significant shortening of the anoxia times.
From these tests the synergism between Coenzyme Q.sub.10 and carnitine is
therefore evident. Also the state of tissue anoxia caused by the
serotonine injected at the root of the rat's tail, causing gangrene and
necrosis of the tail itself, can be an effective test to measure the
antianoxic capacity of different substances.
During these experiences, a group of Wistar rats were injected daily with
10 mg/kg of serotonine sulphate at the root of the tail. Whereas a group
of animals was used as control-group, other groups of animals received
simultaneously either Coenzyme Q (mg 100/kg) only per os by the parenteral
route (mg 25/kg), or carnitine per os only (mg 100/kg) or by the
parenteral route (mg 250/kg). Another group of animals received the two
products associated both per os and by the parenteral route.
14 Days after the beginning of the treatment, both the time of appearance
of cutaneous dyschromasia in the tail, and the appearance of gangrene and
necrosis were calculated.
In none of the Coenzyme Q.sub.10 -carnitine combination treated animals,
signs of gangrene or necrosis in the tail appeared, after fourteen days,
whereas both in the Coenzyme Q.sub.10 treated rats and in the rats treated
with carnitine these signs were evident already seven days after the
beginning of serotonine sulphate administration.
Also in this case, it is evident the synergism exerted by the two
associated substances, Coenzyme Q and carnitine.
ANTIHYPERTENSIVE ACTIVITY
Rats which had been made experimentally hypertensive with the daily
administration of mg 10 of DOCA (deoxycorticosterone acetate) five days a
week for three consecutive weeks were used.
At the same time a 1% saline solution was administered to the animals.
A group of these animals, simultaneously to the DOCA solution, was treated
with Coenzyme Q only, with carnitine only, and with the two products
associated at the same doses as indicated in the previous tests.
After three weeks of treatment, the group of animals treated with DOCA
showed arterial pressure values higher than 120 and the groups of animals
treated with the two drugs, Coenzyme Q and carnitine associated did not
show significant changes of the arterial pressure as compared with normal
rats, whereas changes were evident both in the group of animals treated
with Coenzyme Q and in the group of animals treated with carnitine; above
all, no difference was detectable between the group treated with carnitine
and the hypertensive rats control group.
TESTS ON THE INHIBITION OF THE EXPERIMENTAL ATHEROSCLEROTIC LESION
In these tests the capacity of the carnitine-Coenzyme Q combination was
pointed out, in comparison with the two products administered
individually, to prevent the incidence of tissue vasal lesions
experimentally induced in the rat administering an atherogenic
hypercholesteraemic diet having the following composition: casein 24%,
cotton oil 10%, salt 5%, sugar 61%, cholesterol 0,5%, vitamin D.sub.2
200mUST/g diet).
After six weeks of this diet, all the animals were sacrificed, after dosing
their plasma cholesterol, whereas aorta and heart were fixed and submitted
to hystologic test.
Whereas the rats treated with the hypercholesteraemic and vitamina D.sub.2
diet only, as well as the rats treated with this same diet and carnitine
showed clear increases of hypercholesteraemia and serious lesions both at
aortic and at myocardial level, the group of animals treated with the
carnitine-Coenzyme Q combination (Coenzyme Q mg 50/kg per os, carnitine mg
500/kg per os) showed a definitely lower extent of such diet-induced
atherosclerotic lesions.
The present invention also relates to all the aspects applicable in the
industry connected with the use of the ubiquinon-carnitine combination in
cardiovascular therapy.
A basic aspect of the present invention is therefore represented by
pharmaceutical compositions containing an ubiquinoninc coenzyme,
preferably Coenzyme Q.sub.10, and carnitine or acetylcarnitine preferably
in L forms, in ratios ranging from 1:100 to 1:2, in addition to other
suitable eccipients in the form of, for example, capsules, tablets, pills,
granulates, syrups, injectable solutions.
Some non-limitative examples of pharmaceutical compositions, that can be
obtained with conventional methods of pharmaceutical procedure, are
hereinafter reported.
Tablets containing 500 mg of acetyl L-carnitine and 10 mg of coenzyme
Q.sub.10.
Operculate capsules containing 500 mg of L-carnitine and 5 mg of Coenzyme
Q.sub.10.
Operculate capsules containing 250 mg of L-carnitine and 50 mg of Coenzyme
Q.sub.10.
Pills containing 50 mg of L-carnitine and 25 mg of Coenzyme Q.sub.10.
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