Composition useful as a dietary supplement

Abstract

Compositions comprising L-carnitine or a salt thereof, acetyl L-carnitine or a salt thereof, vitamin B9 (folic Acid), vitamin A, vitamin B12, vitamin B5 (pantothenic acid), vitamin B6 (pyridoxine), arginine, vitamin C, vitamin E (-Tocopherol), selenium, zinc, copper, iron, vitamin D and N-acetyl cysteine, and optionally one or more pharmaceutically acceptable excipients are provided. Dietary supplements comprising these compositions are also provided.

Claims

1. A combination composition comprising as active ingredients: L-carnitine or a salt thereof in a dose of from 2589 to 287.6 mg (corresponding to 1500-166.6 mg L-carnitine inner salt), acetyl L-carnitine or a salt thereof in a dose of from 750 to 25.0 mg, vitamin B9 (folic Acid) in a dose of from 600 to 66.6 g, vitamin A in a dose of from 2400 to 266.6 g, vitamin B12 in a dose of from 7.5 to 0.833 g, vitamin B5 (pantothenic acid) in a dose of from 18.0 to 2.0 mg, vitamin B6 (pyridoxine) in a dose of from 6.0 to 0.666 mg, arginine in a dose of from 1500 to 166.6 mg, vitamin C in a dose of from 270.0 to 30.0 mg, vitamin E (-Tocopherol) in a dose of from 90.0 to 10.0 mg, selenium in a dose of from 150 to 16.6 g, zinc in a dose of from 30.0 to 3.33 mg, copper in a dose of from 4.95 to 0.55 mg, iron in a dose of from 42.0 to 4.66 mg, vitamin D in a dose of from 15.0 to 1.66 g and N-acetyl cysteine in a dose of from 150 to 16.6 mg, and optionally one or more pharmaceutically acceptable excipients.

2. The combination composition of claim 1, further comprising co-enzymes, mineral substances, antioxidants, and vitamins.

3. A dietary supplement comprising the combination composition of claim 1.

4. The combination composition of claim 1, in which the salt of L-carnitine or acetyl L-carnitine is selected from the group consisting of chloride, bromide, orotate, aspartate, citrate, phosphate, fumarate, lactate, maleate, oxalate, pamoate, sulphate, tartrate and mucate.

5. The combination composition of claim 1, for oral administration.

6. The combination composition of claim 1, which comprises: L-carnitine fumarate in a dose of from 1726 to 431.5 mg (corresponding to 1000-250 mg of L-carnitine inner salt); Acetyl L-carnitine in a dose of from 500 to 125 mg; folic acid (vitamin B9) in a dose of from 400 to 100 g; vitamin A from beta carotene in a dose of from 1600 to 400 g; vitamin B12 in a dose of from 5.0 to 1.250 g; vitamin B5 (pantothenic) in a dose of from 12.0 to 3.0 mg; vitamin B6 (pyridoxine) in a dose of from 4.0 to 1.0 mg; arginine in a dose of from 1000.0 to 250.0 mg; vitamin C in a dose of from 180.0 to 45.0 mg; vitamin E (-Tocopherol) in a dose of from 60.0 to 15.0 mg; selenium in a dose of from 100.0 to 25.0 g; zinc in a dose of from 20.0 to 5.0 mg; copper in a dose of from 3.3 to 0.825 mg; iron in a dose of from 28.0 to 7.0 mg; vitamin D in a dose of from 10.00 to 2.50 g; and N-acetyl cysteine in a dose of from 100.0 to 25.0 mg.

7. The combination composition of claim 6, which comprises: L-carnitine fumarate in a dose of 863 mg (corresponding to 500 mg of L-carnitine inner salt); Acetyl L-carnitine in a dose of 250 mg; folic acid (vitamin B9) in a dose of 200 g; vitamin A from beta carotene in a dose of 800 g; vitamin B12 in a dose of 2.5 g; vitamin B5 (pantothenic) in a dose of 6 mg; vitamin B6 (pyridoxine) in a dose of 2 mg; arginine in a dose of 500 mg; vitamin C in a dose of 90 mg; vitamin E (-Tocopherol) in a dose of 30 mg; selenium in a dose of 50 g; zinc in a dose of 10 mg; copper in a dose of 1.65 mg; iron in a dose of 14 mg; vitamin D in a dose of 5 g; and N-acetyl cysteine in a dose of 50 mg.

8. A method of promoting female fertility, the method comprising administering the combination composition of claim 1 to a women in need thereof.

9. A method of promoting female fertility during drug stimulation, hormonal stimulation, or drug and hormonal stimulation, the method comprising administering the combination composition of claim 1 to a women in need thereof.

10. A method of promoting female fertility during drug stimulation, hormonal stimulation, or drug and hormonal stimulation for supporting ovulation, the method comprising administering the combination composition of claim 1 to a women in need thereof.

11. The method of claim 10, in which the administration of the combination composition is started at least a month before the beginning of the drug stimulation, hormonal stimulation, or drug and hormonal stimulation.

12. The method of claim 10 in which the administration of the combination composition is started at least a week before the beginning of the drug stimulation, hormonal stimulation, or drug and hormonal stimulation.

13. A method of promoting female fertility during during drug stimulation, hormonal stimulation, or drug and hormonal stimulation for promoting oocyte fertilization, the method comprising administering the combination composition of claim 1 to a women in need thereof.

14. A method of promoting female fertility during drug stimulation, hormonal stimulation, or drug and hormonal stimulation for promoting oocyte fertilization in vivo, the method comprising administering the combination composition of claim 1 to a women in need thereof.

15. A method of promoting female fertility during drug stimulation, hormonal stimulation, or drug and hormonal stimulation for promoting oocyte fertilization in vitro, the method comprising administering the combination composition of claim 1 to a women in need thereof.

16. A method of reducing side effects due to the use of hormones, drugs, or hormone and drugs useful for promoting female fertility, the method comprising administering the combination composition of claim 1 to a women in need thereof.

Description

EXAMPLE 1

(1) Materials and Methods

(2) Animals.

(3) CD1 mice, 60 female and 60 male; 6 weeks age; weight 16.2 g; 5 mice per group, were used for the study. Mice were allowed to acclimate for a week prior to weighing. Female mice began superovulation 2 to 3 day after achieving the correct weight. All mice were maintained under SPF conditions under a 12:12-h light:dark cycle (lights on, 0700 to 1900) at temperatures of 21 to 24 C. Mice were housed in static microisolation caging with ad libitum access to food and water.

(4) Treatment.

(5) All the female mice, except the Control group, were treated orally (0.5 ml, gastric gavage, twice a day) for 7 days (included the days in which the mice were treated with the hormones for the superovulation) with the composition of the invention (DNCO64) having the following composition: L-carnitine fumarate 0.4 mg+ Acetyl-L-carnitine HCl. 0.12 mg+ Vitamin B12 0.0012 mcg+ Vitamin B9 0.09 mcg+ Vitamin B6 0.0009 mg+ Vitamin B5 0.003 mg+ Vitamin A 0.4 mcg+ Vitamin D 0.0022 mcg+ Selenium 0.022 mcg+ Vitamin C 0.04 mg+ Vitamin E 0.014 mg+ Zinc 0.004 mg+ Copper 0.0008 mg+ Iron 0.007 mg+ L-Arginine 0.24 mg+ N-acetyl cysteine 0.24 mg, combination thereof, as reported in Table 1.

(6) Superovulation and Mating.

(7) The superovulation technique used is described in J. Am. Assoc. Lab. Anim. Sci. 2011, July 50(4); 471-478. In short, female mice were treated with 5 IU (0.1 mL intra peritoneum) of Pregnant Mares Serum Gonadotropin (PMSG). These mice received 5 IU (0.1 mL IP) of Human Chorionic Gonadotropin (HCG) 47 to 49 h after their last PMSG injection. Immediately after HCG injection, female mice were mated 1:1 to male mice. All female mice underwent this superovulation treatment.

(8) Oocyte Collection and Analysis.

(9) The day after mating, all female donors were euthanized by cervical dislocation and oviducts were collected from all female mice and placed into 2 mL M2 media (Sigma-Aldrich) in a 35-mm culture dish (Fisher Scientific). Each oviduct then was moved to a dish containing 2 mL M2 media (Sigma-Aldrich) and 75 L hyaluronidase (10 mg/mL; Sigma-Aldrich), where the ampulla was torn open to release the oocytes.

(10) After all the oviducts for that group had been processed, all of the oocytes were collected and placed into a 100-L drop of KSOM (Millipore, Billerica Mass.) under embryo-tested mineral oil (Sigma-Aldrich) that had been equilibrated to 37 C. at 5% CO.sub.2. Oocytes were allowed to incubate for 24 h, after which the drop was scored for the number of 2-cell of fertilized oocytes.

(11) The results obtained are reported in the following Table 1.

(12) TABLE-US-00001 TABLE 1 Superovulation treatment (Groups 1-12; 5 + 5 IU PMSG + HCG) Mean no. of oocytes/ Mean no. of fertilized female oocytes/female Groups mouse mouse (5 mice per group) (SE) (SE) N.sup.o Treatment P < 1 Control (no treatment 29.1 1.7 10.7 1.3 with the composition of the invention) 2 Complete composition 30.3 1.3 18.5 0.9 0.001 vs 1 of the invention (DNC064) 3 (Carnitines) L-carnitine fumarate 30.6 1.7 13.1 1.6 0.05 vs 2 0.4 mg + Acetyl- NS vs 1 L-carnitine HCl. 0.12 mg 4 (Vitamins) 29.6 1.3 12.7 2.1 0.05 vs 2 Vitamin B12 0.0012 mcg + NS vs 1 Vitamin B9 0.09 mcg + Vitamin B6 0.0009 mg + Vitamin B5 0.003 mg + Vitamin A 0.4 mcg + Vitamin D 0.0022 mcg 5 (Antioxidants) Selenium 0.022 mcg + 30.0 1.5 13.6 1.8 0.05 vs 2 Vitamin C 0.04 mg + NS vs 1 Vitamin E 0.014 mg 6 (Micro-elements) Zinc 0.004 mg + 29.6 1.7 14.0 1.7 0.05 vs 2 Copper 0.0008 mg + NS vs 1 Iron 0.007 mg 7 (Amino acids) L-Arginine 0.24 mg + 28.6 2.0 13.9 1.6 0.05 vs 2 N-acetyl cysteine 0.24 mg NS vs 1 8 (Group 3 + Group 4) 29.6 1.8 13.8 1.5 0.05 vs 2 NS vs 1 9 (3 + 5) 29.6 2.1 14.1 1.3 0.05 vs 2 NS vs 1 10 (3 + 6) 28.1 1.7 14.0 1.3 0.05 vs 2 NS vs 1 11 (3 + 7) 30.3 2.2 13.8 1.5 0.05 vs 2 NS vs 1 12 (4 + 5 + 6 + 7) 28.6 1.7 13.4 2.0 0.05 vs 2 NS vs 1

(13) Statistical Analysis

(14) The significance was calculated using the Student t-test, p<0.05 Values were considered to represent a significant difference.

(15) The results reported in Table 1 show that the treatment of the invention statistically increased the number of fertilized oocytes with respect to the single components or different combination thereof.

(16) The composition of the present invention can be administered in any suitable form for oral administration.

(17) An example of form of administration is in a liquid, semi-liquid or solid form in sachets, pills, vials, gel or liposome.

(18) L-carnitine and acetyl L-carnitine are known compounds and their preparation process is described in U.S. Pat. No. 4,254,053.

(19) Vitamin A, vitamin B9, vitamin B12, vitamin B5 (pantothenic acid), vitamin B6 (pyridoxine), arginine, vitamin C, vitamin E (-Tocopherol), selenium, zinc, copper, iron, vitamin D and N-acetyl cysteine are very long time widely sold on the market for human use.

(20) The composition according to the present invention is composed of active ingredients which are familiar to operators in the medical field and already in use.

(21) 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.

(22) For any compound, the therapeutically effective dose can be estimated initially either in cell culture assays or in animal models, usually mice or rats.

(23) 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.

(24) 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.

(25) In the following a non-limiting example of the compositions of the invention are reported.

(26) TABLE-US-00002 Composition 1 L-carnitine fumarate 863 mg (500 mg L-Carnitine base) acetyl-L-carnitine 250 mg folic Acid (vitamin B9) 200 mcg vitamin A from Beta Carotene 800 mcg vitamin B12 2.5 mcg vitamin B5 (pantothenic) 6 mg vitamin B6 (pyridoxine) 2 mg arginine 500 mg vitamin C 90 mg vitamin E (-Tocopherolo) 30 mg selenium 50 mcg zinc 10 mg copper 1.65 mg iron 14 mg vitamin D 5 mcg N-acetyl cysteine 50 mg.

(27) TABLE-US-00003 Composition 2 L-carnitine fumarate 430 mg acetyl-L-carnitine 125 mg folic Acid (vitamin B9) 200 mcg vitamin A from Beta Carotene 800 mcg vitamin B12 2.5 mcg vitamin B5 (pantothenic) 6 mg vitamin B6 (pyridoxine) 2 mg arginine 500 mg vitamin C 90 mg vitamin E (-Tocopherolo) 30 mg selenium 50 mcg zinc 10 mg copper 1.65 mg iron 14 mg vitamin D 5 mcg N-acetyl cysteine 50 mg.

(28) TABLE-US-00004 Composition 3 L-carnitine fumarate 863 mg acetyl-L-carnitine 125 mg folic Acid (vitamin B9) 200 mcg vitamin A from Beta Carotene 800 mcg vitamin B12 2.5 mcg vitamin B5 (pantothenic) 6 mg vitamin B6 (pyridoxine) 2 mg arginine 500 mg vitamin C 90 mg vitamin E (-Tocopherolo) 30 mg selenium 50 mcg zinc 10 mg copper 1.65 mg iron 14 mg vitamin D 5 mcg N-acetyl cysteine 50 mg.

(29) TABLE-US-00005 Composition 4 L-carnitine fumarate 430 mg acetyl-L-carnitine 250 mg folic Acid (vitamin B9) 200 mcg vitamin A from Beta Carotene 800 mcg vitamin B12 2.5 mcg vitamin B5 (pantothenic) 6 mg vitamin B6 (pyridoxine) 2 mg arginine 500 mg vitamin C 90 mg vitamin E (-Tocopherolo) 30 mg selenium 50 mcg zinc 10 mg copper 1.65 mg iron 14 mg vitamin D 5 mcg N-acetyl cysteine 50 mg.