U.S. patent application number 13/576780 was filed with the patent office on 2013-01-03 for combination composition, comprising as active ingredients l-carnitlne or propionyl l-carnitlne, for the prevention or treatment of chronic venous insufficiency.
This patent application is currently assigned to SIGMA-TAU INDUSTRIE FARMACEUTICHE RIUNITE S.P.A.. Invention is credited to Aleardo Koverech, Mohamed Ashraf Virmani.
Application Number | 20130005670 13/576780 |
Document ID | / |
Family ID | 41820403 |
Filed Date | 2013-01-03 |
United States Patent
Application |
20130005670 |
Kind Code |
A1 |
Virmani; Mohamed Ashraf ; et
al. |
January 3, 2013 |
COMBINATION COMPOSITION, COMPRISING AS ACTIVE INGREDIENTS
L-CARNITlNE OR PROPIONYL L-CARNITlNE, FOR THE PREVENTION OR
TREATMENT OF CHRONIC VENOUS INSUFFICIENCY
Abstract
It is described a combination composition comprising as active
ingredients L-carnitine or propionyl L-carnitine, troxerutine,
diosmine and hesperidine, useful for the prevention and/or
treatment of chronic venous diseases.
Inventors: |
Virmani; Mohamed Ashraf;
(Rome, IT) ; Koverech; Aleardo; (Rome,
IT) |
Assignee: |
SIGMA-TAU INDUSTRIE FARMACEUTICHE
RIUNITE S.P.A.
Rome
IT
|
Family ID: |
41820403 |
Appl. No.: |
13/576780 |
Filed: |
February 7, 2011 |
PCT Filed: |
February 7, 2011 |
PCT NO: |
PCT/IB2011/000200 |
371 Date: |
August 28, 2012 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
61322532 |
Apr 9, 2010 |
|
|
|
Current U.S.
Class: |
514/27 |
Current CPC
Class: |
A61P 7/02 20180101; A61P
7/10 20180101; A61K 31/205 20130101; A61P 9/14 20180101; A61P 39/06
20180101; A61P 9/12 20180101; A61P 3/02 20180101; A61K 31/221
20130101; A61K 31/221 20130101; A61K 45/06 20130101; A61K 31/7048
20130101; A61P 29/00 20180101; A61P 17/00 20180101; A61K 31/7048
20130101; A61K 31/205 20130101; A61P 9/00 20180101; A61K 2300/00
20130101; A61K 2300/00 20130101; A61K 2300/00 20130101 |
Class at
Publication: |
514/27 |
International
Class: |
A61K 31/7048 20060101
A61K031/7048; A61P 9/12 20060101 A61P009/12; A61P 9/00 20060101
A61P009/00 |
Foreign Application Data
Date |
Code |
Application Number |
Feb 2, 2010 |
EP |
10152363.7 |
Claims
1. Composition comprising as active ingredients L-carnitine or a
derivative or a salt thereof, troxerutine, diosmine and
hesperidine, and optionally one or more pharmaceutically acceptable
excipients.
2. Composition of claim 1, wherein the derivative of L-carnitine is
propionyl L-carnitine.
3. Composition of claim 1 comprising: L-carnitine or propionyl
L-carnitine, in a dose of from 10 to 3000 mg; troxerutine, in a
dose of from 900 mg to 50 mg; diosmine, in a dose of from 900 mg to
50 mg; and hesperidine in a dose of from 10 mg to 500 mg.
4-5. (canceled)
6. Composition of claim 1, further comprising co-enzymes, mineral
substances, antioxidants, vitamins and anticlotting agents.
7. (canceled)
8. Method of preventing or treating diseases of the vein comprising
administering to a human subject in need thereof a composition
comprising as active ingredients L-carnitine or a derivative or a
salt thereof, troxerutine, diosmine and hesperidine.
9. Method according to claim 8 in which the derivative of
L-carnitine is propionyl L-carnitine.
10. Method according to claim 8, wherein the disease of the veins
is selected from the group consisting of: chronic venous
insufficiency, chronic venous disease and complication thereof.
11. Method according to claim 10, wherein said complication is
selected from the group consisting of: swelling and inflammation of
veins in the rectum, anus and vulva; venous hypertension; increased
permeability; oedema; capillary damage; skin changes; venous leg
ulcers; swelling ankles; heavy legs; varicose veins; swelling leg;
ulcers; vein thrombosis; phlebitis; thrombo-phlebitis; pulmonary
embolus and hemorrhoids.
12. Method according to claim 8, wherein the salt of L-camitine or
propionyl L-carnitine is selected from the group consisting of:
chloride, bromide, orotate, aspartate, acid aspartate, acid
citrate, magnesium citrate, phosphate, acid phosphate, fumarate and
acid fumarate, magnesium fumarate, lactate, maleate and acid
maleate, oxalate, acid oxalate, pamoate, acid pamoate, sulphate,
acid sulphate, glucose phosphate, tartrate and acid tartrate,
glycerophosphate, ethanesulphonate, mucate, magnesium tartrate,
2-amino-ethanesulphonate, magnesium 2-amino-ethanesulphonate,
methanesulphonate, choline tartrate, trichloroacetate, and
trifluoroacetate.
13. Method according to claim 8, for oral, parenteral, intravenous,
topical and/or transdermal administration.
14. Method according to claim 8, for oral administration.
15. Composition of claim 1 comprising L-carnitine or propionyl
L-carnitine in a dose of from 50 mg to 400 mg.
16. Composition of claim 1 comprising L-carnitine or propionyl
L-carnitine in a dose of 136 mg.
17. Composition of claim 1 comprising troxerutine in a dose from
200 mg to 400 mg.
18. Composition of claim 1 comprising troxerutine in a dose of 300
mg.
19. Composition of claim 1 comprising diosmine in a dose from 200
mg to 400 mg.
20. Composition of claim 1 comprising diosmine in a dose of 300
mg.
21. Composition of claim 1 comprising hesperidine in a dose of from
50 mg to 200 mg.
22. Composition of claim 1 comprising hesperidine in a dose of 100
mg.
Description
FIELD OF THE INVENTION
[0001] The present invention relates to a combination composition
comprising as active ingredients L-carnitine or a derivative
thereof, or a salt thereof, troxerutine, diosmine and hesperidine,
useful for the prevention and/or treatment of chronic venous
insufficiency (CVI).
BACKGROUND OF THE INVENTION
[0002] In humans, arteries bring oxygen-rich blood from the heart
to the rest of the body and veins return oxygen-poor blood back to
the heart. When human leg veins cannot pump enough blood back to
heart, there is the onset of chronic venous insufficiency (CVI).
CVI is also sometimes called chronic venous disease, or CVD.
[0003] Humans have three kinds of veins: superficial veins, which
lie close to the skin, deep veins, which lie in groups of muscles,
and perforating veins, which connect the superficial to the deep
veins. Deep veins lead to the vena cava, human body's largest vein,
which runs directly to the heart.
[0004] When humans are in the upright position, the blood in their
leg veins must go against gravity to return to our heart. To
accomplish this, leg muscles squeeze the deep veins of their legs
and feet to help move blood back to our heart. One-way flaps,
called valves, in the veins keep blood flowing in the right
direction. When the leg muscles relax, the valves inside the veins
close. This prevents blood from flowing in reverse, back down the
legs. The entire process of sending blood back to the heart is
called the venous pump.
[0005] When humans walk and the leg muscles squeeze, the venous
pump works well. But when patient sit or stand, especially for a
long time, the blood in the leg veins can pool and increase the
venous blood pressure. Deep veins and perforating veins are usually
able to withstand short periods of increased pressures. However,
sitting or standing for a long time can stretch vein walls because
they are flexible. Over time, in susceptible individuals, this can
weaken the walls of the veins and damage the vein valves, causing
CVI.
[0006] During CVI ankles may swell and calves may feel tight. The
legs may also feel heavy, tired, restless, or achy. Patient may
feel pain while walking or shortly after stopping.
[0007] CVI may be associated with varicose veins. Varicose veins
are swollen veins that we can see through the skin. They often look
blue, bulging, and twisted. Large varicose veins can lead to skin
changes like rashes, redness, and sores.
[0008] CVI can also cause problems with leg swelling because of the
pressure of the blood pooling in the veins. The lymphatic system
may also produce fluid, called lymph, to compensate for CVI. Human
leg tissues may then absorb some of this fluid, which can increase
the tendency for the legs to swell. In severe cases, CVI and the
leg swelling can cause ulcers to form on the lower parts of the
leg.
[0009] Over the long-term, blood pressure that is higher than
normal inside the leg veins causes CVI. Other causes of CVI include
deep vein thrombosis (DVT) and phlebitis, both of which cause
elevated pressure in our veins by obstructing the free flow of
blood through the veins.
[0010] DVT occurs when a blood clot (properly called a thrombus)
blocks blood from flowing toward the heart, out of a deep or
perforating vein. The blood trying to pass through the blocked
veins can increase the blood pressure in the vein, which, in turn,
overloads our valves. Vein valves that do not work properly are
called incompetent because they stretch and no longer work
efficiently, and incompetent valves contribute to CVI. DVT is a
potentially serious condition that causes leg swelling and requires
immediate medical attention because sometimes the blood clots in
the veins can break off and travel to the lungs. This condition is
called a pulmonary embolus.
[0011] Phlebitis occurs when a superficial or deep vein becomes
swollen and inflamed. This inflammation causes a blood clot to
form, which can also lead to DVT.
[0012] Factors that can increase the risk for CVI include a family
history of varicose veins, being overweight, being pregnant, not
exercising enough, smoking, and standing or sitting for long
periods of time. Although CVI can affect anyone, age and sex can
also be factors that may increase the tendency to develop CVI;
women older than 50 most often get CVI.
[0013] For mild cases of CVI, physician may recommend compression
stockings. Compression stockings are elastic stockings that squeeze
the veins and stop excess blood from flowing backward. In this way,
compression stockings can often also help heal skin sores and
prevent them from returning. Patient may need to wear compression
stockings daily for the rest of the life.
[0014] More serious cases of CVI may be treated with injections,
called sclerotherapy, or with surgical procedures. Fewer than 10
percent of patient with CVI require surgery to correct the problem.
Surgical treatments include ablation, vein stripping, bypass
surgery, valve repair, and angioplasty or stenting of a vein.
[0015] The flavonoids, troxerutine, diosmine and hesperidine, also
referred to as rutins oxerutins or rutosides, are well known for
their antioxidant properties, in addition they also display other
properties that impact upon the circulatory system i.e. blood and
microvascular endothelial cells (G.I.O.T. 2009; 35:23-33).
[0016] Many epidemiological studies suggest that these flavonoids
are associated with improvement of circulation and a general
reduced risk of cardiovascular diseases. The mechanisms underlying
these protective actions include antithrombotic, anti-ischemic,
anti-oxidant, and vasorelaxant properties. In particular these
flavonoids have actions that improve circulation by vasodilatation,
decreasing platelets clotting and preventing low-density
lipoproteins (LDLs) from oxidizing (Journal of Agricultural and
Food Chemistry 2008 56 (15), pp 6185-6205).
[0017] These flavonoids also demonstrate strong anti-inflammatory
properties that are thought to be the result of the inhibition of
the synthesis and biological activities of different
pro-inflammatory mediators, mainly the arachidonic acid
derivatives, prostaglandins E2, F2, and thromboxane A2. These
processes improve venous tone and lymphatic drainage, and reduce
capillary hyperpermeability by protecting the microcirculation from
inflammatory processes.
[0018] This effect has been shown also in clinical studies. The
study of Belcaro et al (Angiology, 59; 5S) showed that these
flavonoids were effective in the control and treatment of oedema
and increased capillary filtration in venous hypertension and
diabetic microangiopathy.
[0019] Another study by Belcaro et al. (Angiology, Mar. 1, 2008; 59
(1) suppl. 7S-13S) also looked at the use of these flavonoids in
the treatment of chronic venous disease and signs and symptoms of
chronic venous insufficiency (CVI), varicose veins, and deep venous
disease.
[0020] In Rev Fr Gynecol Obstet. 1991 Feb. 25; 86 (2 Pt 2): 209-12,
is reported that troxerutine in women (half in the context of
premenstrual syndrome and half in pregnant) was useful for the
treatment of vulval varicosities and venous insufficiency of the
lower limbs.
[0021] In Br. J. Surg. 2000, 87; 868-872, it is reported that
diosmine has anti-inflammatory actions that play a role in
protecting the blood vessels and contributes to the maintenance of
good circulation of the blood and maintains the venous tone.
[0022] In Farmaco, 40 (11); 709-712, it is reported that
hesperidine has an antioxidant action and together with troxerutine
and diosmine promotes healthy vessel functions.
[0023] In U.S. Pat. No. 4,255,449 it is reported that L-carnitine
is useful for increasing the HDL cholesterol and for treating
diseases liked to high cholesterol level.
[0024] In WO04091602 9 it is reported that L-carnitine is useful
the treatment of cardiovascular diseases.
[0025] In U.S. Pat. No. 5,811,457 it is reported that propionyl
L-carnitine is useful the treatment of chronic arteriosclerosis
obliterans
[0026] In WO 2007045639 it is reported that propionyl L-carnitine
is useful the treatment of left ventricular hypertrophy in dialysed
patients.
[0027] In U.S. Pat. No. 4,343,816 it is reported that propionyl
L-carnitine is useful the treatment of peripheral vascular
diseases.
[0028] Furthermore, while there are other publications available in
which is shown that the compounds of the invention are useful for
the treatment of diseases of the veins, none of them mention nor
suggest about the unexpected synergistic effect shown by the
composition of the invention.
DESCRIPTION OF THE INVENTION
[0029] It has now been found that a combination composition
comprising as active ingredients L-carnitine or a derivative
thereof (e.g. propionyl L-carnitine) or a salt thereof,
troxerutine, diosmine and hesperidine, is endowed with a
surprisingly synergistic effect for the prevention and/or treatment
of diseases of the veins selected from Chronic venous insufficiency
(CVI) and chronic venous disease (CVD) or their complications.
[0030] An example of complications or diseases of the veins (due to
CVI or CVD) are reported in the following: swelling and
inflammation of veins in the rectum, anus and vulva; venous
hypertension; increased permeability; oedema; capillary damage;
skin changes; venous leg ulcers; swelling ankles; heavy legs;
varicose veins; swelling leg; ulcers; vein thrombosis; phlebitis;
thrombo-phlebitis; pulmonary embolus; hemorrhoids.
[0031] It is therefore one object of the present invention a
combination composition comprising as active ingredients
L-carnitine or a salt thereof, troxerutine, diosmine and
hesperidine.
[0032] It is a further object of the present invention a
combination composition comprising as active ingredients propionyl
L-carnitine or a salt thereof, troxerutine, diosmine and
hesperidine.
[0033] The compositions mentioned above may further comprise other
active ingredients useful for treating diseases of the vein.
[0034] It is a further object of the present invention a
composition comprising:
[0035] (a) L-carnitine or propionyl L-carnitine, in a dose of from
10 to 3000 mg, preferred doses is of from 50 mg to 400 mg, the most
preferred dose is 136 mg.
[0036] (b) troxerutine, in a dose of from 900 mg to 50 mg,
preferred doses is of from 400 mg to 200 mg, the most preferred
dose is 300 mg;
[0037] (c) diosmine, in a dose of from 900 mg to 50 mg, preferred
doses is of from 400 mg to 200 mg, the most preferred dose is 300
mg; and
[0038] (d) hesperidine in a dose of from 10 mg to 500 mg, preferred
doses is of from 50 mg to 200 mg, the most preferred dose is 100
mg.
[0039] It is a further object of the present invention the
composition mentioned above, for use as anti-chronic venous
insufficiency and complications thereof.
[0040] It is a further object of the present invention the
composition mentioned above, for use as anti-chronic venous disease
and complications thereof.
[0041] It is a further object of the present invention the use of
the composition mentioned above, for preparing a medicament for the
prevention or treatment of chronic venous insufficiency, chronic
venous disease and complications thereof, in which said
complications are selected from the group comprising: swelling and
inflammation of veins in the rectum, anus and vulva; venous
hypertension; increased permeability; oedema; capillary damage;
skin changes; venous leg ulcers; swelling ankles; heavy legs;
varicose veins; swelling leg; ulcers; vein thrombosis; phlebitis;
thrombo-phlebitis; pulmonary embolus or hemorrhoids.
[0042] It is a further object of the present invention the use of
the composition mentioned above, for preparing a dietary supplement
for the prevention or treatment of chronic venous insufficiency,
chronic venous disease and complications thereof.
[0043] The composition of the invention may further comprise
co-enzymes, mineral substances, antioxidants, vitamins, anticloting
agents and agents useful for treating diseases of the veins.
[0044] What is meant by salt of L-carnitine is any salt of the
latter with an acid that does not give rise to toxic or side
effects.
[0045] Non-limiting examples of such salts are: chloride, bromide,
orotate, aspartate, acid aspartate, acid citrate, magnesium
citrate, phosphate, acid phosphate, fumarate and acid fumarate,
magnesium fumarate, lactate, maleate and acid maleate, oxalate,
acid oxalate, pamoate, acid pamoate, sulphate, acid sulphate,
glucose phosphate, tartrate and acid tartrate, glycerophosphate,
mucate, magnesium tartrate, 2-amino-ethanesulphonate, magnesium
2-amino-ethanesulphonate, methanesulphonate, choline tartrate,
trichloroacetate, and trifluoroacetate.
[0046] A list of FDA-approved pharmaceutically acceptable salts is
given in the publication Int. J. of Pharm. 33 (1986), 201-217.
[0047] L-carnitine, propionyl L-carnitine, troxerutine, diosmine
and hesperidine according to the present invention can be
administrated in a "co-ordinated manner". What is meant by
"co-ordinated manner" of the aforesaid compounds is, indifferently,
either the co-administration, i.e. the substantially concomitant or
sequential supplementation of L-carnitine or propionyl L-carnitine
and at least one troxerutine, diosmine, hesperidine, or the
administration of a composition comprising the aforesaid active
ingredients in combination and in a mixture optionally further
comprising one or more excipients or diluents pharmaceutically
acceptable.
[0048] The composition of the present invention can be administered
orally, parenterally, intravenously, topically and/or
transdermally, in any suitable form. The oral administration is
preferred.
[0049] An example of form of administration is in a liquid,
semi-liquid or solid form in sachets, pills, vials, ointment, gel
or liposome.
[0050] L-carnitine and propionyl L-carnitine are known compounds
and their preparation process is described in U.S. Pat. No.
4,254,053.
[0051] Troxerutine, diosmine and hesperidine are widely sold on the
market and their CAS-No. are: 7085-55-4; 520-27-4 and 520-26-3
respectively.
[0052] The pharmaceutical composition according to the present
invention is composed of active ingredients which are familiar to
operators in the medical field and already in use.
[0053] Their procurement therefore is very easy, inasmuch as these
are products which have been on the market now for a long time and
are of a grade suitable for human administration.
[0054] For any compound, the therapeutically effective dose can be
estimated initially either in cell culture assays or in animal
models, usually mice or rats.
[0055] The animal model may also be used to determine the
appropriate concentration range and route of administration. Such
information can then be used to determine useful doses and routes
for administration in humans.
[0056] The precise effective dose for a human subject will depend
upon the severity of the disease state, general health of the
subject, age, weight, and gender of the subject, diet, time and
frequency of administration, drug combination(s), reaction
sensitivities, and tolerance/response to therapy. This amount can
be determined by routine experimentation and is within the
judgement of the clinician.
[0057] The following non limiting examples further illustrate the
invention.
Example 1
[0058] Reduction of the Swelling of the Recto-Anus, in Croton
Oil-Induced Hemorrhoid Model in Rats.
[0059] Male SD rats (6 weeks old, approximately 140 g) were
purchased from Harlan Sprague Dawley and allowed to acclimate for 1
week. The rats were maintained in a pathogen-free facility in
accordance with the National Research Council of Laboratory Animal
care and use guidelines. Each experiment was preformed with
age-matched rats 7-8 weeks old. The croton oil-induced hemorrhoid
model in rats was performed according to the method published by
Nishiki (Nishiki et al. (1988) Folia Pharmacology Japan 92:215-225;
Nishiki et al. (1988) Folia Pharmacol. Japan 92:227-240). Briefly,
a cotton swab with a diameter of 4 mm soaked with 0.16 mL of
inducer (deionized water: pyridine: ethyl ether: 6% croton
oil/ethyl ether (1:4:5:10) was applied to the rat's anus for 12
seconds. The final concentration of croton oil was 3%. The edema
developed linearly until 7-8 hours after application and the
severity of the edema was sustained for more than 24 hours.
Twenty-four hours later, recto-anus tissue (approx. 10 mm long) was
isolated after the rats were euthanized. The weights of rat body
and recto-anus were measured. The recto-anus coefficient (RAC) was
calculated using the formula: weight of recto-anus (mg)/body weight
(g).
[0060] The compounds of the invention were administered orally
suspended in saline (1 mL) once per day for five days. Last
treatment with the compounds of the invention was made the day of
induction of oedema with croton oil.
[0061] The compounds of the invention were administered alone or in
combination at the following doses: 20 mg/kg propionyl L-carnitine
or L-carnitine; 30 mg/kg troxerutine, diosmine or hesperidine.
[0062] Twenty-four hours after hemorrhoid induction with croton oil
the RAC of rats was determined.
[0063] The results obtained are reported in the following Table
1.
TABLE-US-00001 TABLE 1 n = 5 rats per group; degrees of freedom =
8. RAC SE P< 1 Vehicle alone 1.28 0.14 -- 2 No treatment 0.97
0.11 -NS vs 1 3 L-carnitine + Troxerutine + 1.33 0.12 -NS vs 1
diosmine + hesperidine. 20 mg/kg + 30 mg/kg + 30 mg/kg + 30 mg/kg.
4 Propionyl L-carnitine + Troxerutine + 1.31 0.11 -NS vs 1 diosmine
+ hesperidine. 20 mg/kg + 30 mg/kg + 30 mg/kg + 30 mg/kg. 5
crotonoil (3%) 253 0.18 0.001 vs 1 6 L-carnitine + Troxerutine +
1.34 0.09 -NS vs 1 diosmine + hesperidine + croton oil. -NS vs 3
-NS vs 4 -0.001 vs 5 -0.001 vs 8 -0.001 vs 9 -0.001 vs 10 -0.001 vs
11 -0.001 vs 12 -0.05 vs 21 7 Propionyl L-carnitine + Troxerutine +
1.32 0.11 -NS vs 1 diosmine + hesperidine + -NS vs 3 croton oil.
-NS vs 4 -0.001 vs 5 -0.001 vs 8 -0.001 vs 9 -0.001 vs 10 -0.001 vs
11 -0.001 vs 12 -0.05 vs 21 8 Troxerutine + croton oil 2.11 0.19
0.05 vs 1 9 Diosmine + croton oil 2.02 0.18 0.05 vs 1 10
Hesperidine + croton oil 2.07 0.20 0.05 vs 1 11 L-carnitine +
croton oil 1.99 0.18 0.05 vs 1 12 Propionyl L-carnitine + croton
oil 1.96 0.16 0.05 vs 1 13 Troxerutine + diosmine + croton oil 2.00
0.20 0.05 vs 1 14 Troxerutine + hesperidine + croton 1.99 0.18 0.05
vs 1 oil 15 troxerutine + L-carnitine + croton 1.91 0.17 0.05 vs 1
oil 16 Troxerutine + propionyl l-carnitine + 1.89 0.17 0.05 vs 1
croton oil 17 Diosmine + hesperidine + croton oil 2.01 0.21 0.05 vs
1 18 Diosmine + L-carnitine + croton oil 1.87 0.15 0.05 vs 1 19
Diosmine + propionyl L-carnitine 1.83 0.13 0.05 vs 1 20 Hesperidine
+ L-carnitine + croton 1.84 0.14 0.05 vs 1 oil 20 Hesperidine +
propionyl L-carnitine + 1.84 0.13 0.05 vs 1 croton oil 21
Troxerutine + hesperidine + 1.65 0.14 NS vs 1 diosmine + croton oil
-0.05 vs 5
[0064] The results reported in Table 1 show that using the
compositions of the invention results statistically significant
more active (respect to the use of single component) were
obtained.
[0065] In fact, the results reported in Table 1 show that the
compositions of the invention completely inhibited the swelling of
the recto-anus induced by croton oil.
Example 2
[0066] Relaxant Effect on Aortic Rings of the Compounds Tested.
[0067] The objective of this study was to investigate the
vasoarelaxation effects of the compositions of the invention using
isolated aorta from spontaneously hypertensive rats.
[0068] Materials and Methods
[0069] The compounds tested were used at a concentration expressed
as final concentration in the organ chamber.
[0070] Animals
[0071] Male spontaneously hypertensive rats (SHR) 10-12 weeks old,
weighing 250-300 g, were housed at 24.+-.2.degree. C. with
60.+-.20% relative humidity, on a 12-h light-dark cycle. Rats were
given free access to a diet of standard food and water. All
experiments were performed according to the guidelines for the
ethical treatment of animals of the European Union. The rats were
killed by cervical dislocation and the aortae were rapidly
dissected.
[0072] Aortic Ring Preparation
[0073] The descending thoracic aorta was placed in a modified
Krebs-Henseleit solution (PSS) containing (mM): NaCl 118, KCl 4.75,
NaHCO.sub.3 25, MgSO.sub.4 1.2, CaCl.sub.2 1.8, KH.sub.2PO.sub.4
1.2 and glucose 11.
[0074] After excess fat and connective tissue were removed, the
aortae were cut into 2-3-mm rings. Aortic rings were mounted under
a basal tension of 2 g in 20 mL organ baths containing PSS and
attached to an isometric transducer (Harvard UF-1); the signal was
recorded by a Powerlab data acquisition system (AD-Instruments).
The tissue bath was maintained at 37.degree. C. and bubbled with a
95% O.sub.2 and 5% CO.sub.2 gas mixture.
[0075] Relaxant Effect of the Compounds Tested.
[0076] The relaxant effect of compounds tested was assessed by
adding the compound to aortic rings precontracted by phenylephrine
(1 micromolar).
[0077] After the aortae were contracted with phenylephrine and when
the contractile response was reached, the test compound was added
to the bath. All the results were expressed as a percentage of the
maximal contraction of phenylephrine-induced responses.
[0078] Statistical Analysis
[0079] Results are expressed as percentages from the initial
precontraction level.
[0080] The significance was calculated on the actual contraction
values means.+-.SE (n=8 aorta preparations per group). Eight
preparations were studied from a single aorta. The Student t-test
was used as the comparison test for statistical analysis.
[0081] p<0.05 Values were considered to represent a significant
difference.
[0082] The results obtained are reported in the following Table
2.
TABLE-US-00002 TABLE 2 % TREATMENT RELAXATION P< 1 Saline 0 -- 2
L-carnitine + troxerutine + 52 -0.001 vs 1 hesperidine + diosmine
-0.01 vs 4 (100 micro-molar + 100 micro- -0.01 vs 5 molar + 100
micro-molar + 100 -0.01 vs 6 micro-molar). -0.05 vs 7 -0.05 vs 8
-0.05 vs 18 3 Propionyl L-carnitine + 53 -0.001 vs 1 troxerutine +
hesperidine + -0.01 vs 4 diosmine. -0.01 vs 5 (100 micro-molar +
100 micro- -0.01 vs 6 molar + 100 micro-molar + 100 -0.05 vs 7
micro-molar). -0.05 vs 8 -0.05 vs 18 4 Troxerutine 10 0.05 vs 1
(100 micro-molar). 5 Diosmine (100 micro-molar). 10 0.05 vs 1 6
Hesperidine (100 micro-molar). 9 0.05 vs 1 7 L-carnitine (100
micro-molar). 25 0.01 vs 1 8 Propionyl L-carnitine 28 0.01 vs 1
(100 micro-molar). 9 Troxerutine + diosmine 12 0.05 vs 1 10
Troxerutine + hesperidine 13 0.05 vs 1 11 Troxerutine + L-carnitine
27 0.01 vs 1 12 Troxerutine + propionyl L- 29 0.01 vs 1 carnitine
13 Diosmine + hesperidine 14 0.05 vs 1 14 Diosmine + L-carnitine 26
0.01 vs 1 15 Diosmine + propionyl L- 28 0.01 vs 1 carnitine 16
Hesperidine + L-carnitine 26 0.01 vs 1 17 Hesperidine + propionyl
L- 29 0.01 vs 1 carnitine 18 Troxerutine + hesperidine + 35 0.001
vs 1 diosmine
[0083] The results reported in Table 2 show that the addition of
compositions of the invention to aortic rings precontracted with
phenylephrine elicited endothelium-dependent relaxation which was
statistically significant more active respect to the single
components.
[0084] In the following are reported some non limiting examples of
the compositions of the invention.
[0085] Composition 1
TABLE-US-00003 L-Carnitine tartrate 200 mg Equivalent to
L-carnitine 136 mg Troxerutine NEC .RTM. 700 mg Comprising
Troxerutina 300 mg Diosmine 300 mg Hesperidine 98% 100 mg
Equivalent to Hesperidine 98 mg.
[0086] Composition 2
TABLE-US-00004 Propionyl L-Carnitine tartrate 250 mg Equivalent to
propionyl L-carnitine 136 mg Troxerutine NEC .RTM. 700 mg
Comprising Troxerutina 300 mg Diosmine 300 mg Hesperidine 98% 100
mg Equivalent to Hesperidine 98 mg.
[0087] Composition 3
TABLE-US-00005 L-Carnitine fumarate 200 mg Equivalent to
L-carnitine 136 mg Troxerutine NEC .RTM. 700 mg Comprising
Troxerutina 300 mg Diosmine 300 mg Hesperidine 98% 100 mg
Equivalent to Hesperidine 98 mg.
[0088] Composition 4
TABLE-US-00006 Propionyl L-Carnitine fumarate 250 mg Equivalent to
propionyl L-carnitine 136 mg Troxerutine NEC .RTM. 700 mg
Comprising Troxerutina 300 mg Diosmine 300 mg Hesperidine 98% 100
mg Equivalent to Hesperidine 98 mg.
* * * * *