METAL SALTS OF CARNITINES USEFUL AS DIETARY/NUTRITIONAL SUPPLEMENT OR DRUGS AND COMPOSITIONS CONTAINING SUCH SALTS The present invention relates to novel metal salts of carnitines with an organic polybasic acid useful as dietary/nutritional supplements or drugs, the processes for preparing same and the compositions containing such salts.

More particularly, the present invention relates to stable, non- hygroscopic zinc salts of carnitines with galactaric acid. Therefore, these salts especially lend themselves to the production of solid compositions suitable for oral administration.

According to a more specific aspect thereof, the present invention relates to improvements to the compositions suitable to enhance sperm motility and/or concentration in the seminal fluid of mammals and treat individuals suffering from idiopathic asthenospermia, disclosed in US 6,090,848 (Sigma-Tau) whose disclosures are incorporated herein by reference.

According to the present invention, the term"carnitines"means, collectively, both L-carnitine and alkanoyl L-carnitines wherein the alkanoyl group, straight or branched-chain, contains 2-5 carbon atoms.

Such alkanoyl shall be briefly referred to hereinbelow as" (C2-C5) alkanoyl".

With the term"non-hygroscopic"reference is herein made to the ability endowed by certain carnitine salts, when they occur as powders or granules, to withstand a relative humidity of at least 60%, at 25°C, for 24 hours, without giving rise to adverse phenomena of clotting, agglomeration or even deliquescence which result in loss of their flowability.

With the term"hygroscopic"reference is herein made to the property shown by most of carnitine salts (particularly by their"inner salts") to undergo, when they occur as powders or granules, significant alteration of their flowability due to their clotting, agglomeration or even deliquescence, following exposure to an environment of relative humidity lower than 50-60%, at 25°C, for 24 hours.

The problems of storage and processing brought about by the high hygroscopicity of L-carnitine and alkanoyl L-carnitine inner salts have long since been known. This high hygroscopicity renders the manufacture and storage of orally administrable solid presentation forms particularly troublesome.

However, administration forms such as tablets and capsules represent the preferred presentation forms inasmuch as they make it particularly easy for users to take the active ingredient and comply with optimal dosage regimens.

The problem of L-carnitine and alkanoyl L-carnitine inner salts hygroscopicity has been solved by converting these inner salts into salts of pharmacologically acceptable acids, based on the assumption that such salts maintain the same therapeutical/nutritional activities of the inner salts and do not exhibit unwanted toxic or side effects.

Although there is now an extensive body of literature, particularly patents, disclosing the production of allegedly stable, non-hygroscopic carnitine salts, actually only L-carnitine acid fumarate (US 4,602,039, Sigma-Tau) L-carnitine L- (+)-tartrate (US 5,703,376, Lonza) and, more recently, acetyl L-carnitine galactarate (US 5,952,379, Sigma-Tau) have been developed on an industrial scale and marketed to date.

For a detailed review of the various pathological consequences induced by carnitine and zinc deficiencies (which may be brought about by an insufficiently balanced diet: for instance, high milk consumption, poor in zinc, may be responsible for such deficit) reference is made to"The Merck Manual of Diagnosis and Therapy", 17th Edition (Centennial Edition), 1999, pages 32 and 53, whose contents is incorporated herein by reference.

With specific reference to the treatment of male infertility provoked by disturbances in sperm motility, inadequate sperm concentration in the seminal fluid and altered sperm morphology, the beneficial therapeutical effects achieved with the administration of L-carnitine (see, e. g., Vitali G. et al., Drugs Exptl. Clin. Res. XXI (4), 157-159, 1995; Costa M. et al., Andrologia, 26: 155-159,1994) and acetyl L- carnitine (Moncada M. L. et al., Acta. Eur. Fertil. 23 (5) 221-224,1992), have long since been known.

The aforesaid US patent 6,090,848 discloses the unexpected synergistic effect on sperm motility and concentration in the seminal fluid of sub-fertile males, and the therapeutical effect in the treatment of idiopathic asthenospermia, achieved by administering a combination composition of L-carnitine/acetyl L-carnitine wherein their molar ratio ranges from 2.8: 1 to 4.0: 1 and is preferably about 3: 1, in contrast with the monopharmacological treatment.

Recently, a number of nutritional therapies entailing the supplementation of the diet with mineral elements such as zinc and selenium have been shown to improve sperm motility and sperm counts (see, e. g., S. Sinclair,"Male infertility: Nutritional and environmental considerations", Altern. Med. Rev. 2000; 5 (1), 28-38).

The human body contains 2 to 3 grams of zinc and in males a relevant part hereof is found in testes. The sign and symptoms of zinc deficiency include, among others,"delayed sexual maturation, hypogonadism and hypospermia" (The Merck Manual, loc. cit. page 53).

Zinc is an essential mineral for proper prostate gland function whose secretions comprise approximately 40% of seminal fluid wherein zinc levels are directly related to sperm motility and concentration.

Deficiency of dietary zinc reduces both sperm count and seminal plasma volume and results in delayed spermatozoal maturation and impaired sperm motility. Diet supplementation with zinc has been shown to reverse these phenomena.

Zinc contained in prostatic secretions of healthy males exerts a potent antibacterial action and thus prevents infections ascending from the urethra. Lower than normal zinc levels have been shown in prostatic secretions of males with bacterial prostatitis (see Current Medical Diagnosis & Treatment, 38th Edition, 1991, page 900).

It is, therefore, felt the need to have available stable and non- hygroscopic zinc salts of carnitines with pharmacologically acceptable acids in order to allow the preparation of orally administrable solid compositions to be used as dietary/nutritional supplements in all those situations wherein zinc supplementation reverses or prevents the onset of the prejudicial effects brought about by zinc deficiency.

Moreover, in the light of the firmly established therapeutical properties of L-carnitine, acetyl L-carnitine and propionyl L-carnitine (such as, e. g., L-carnitine protective action on the cardiovascular tract), zinc and carnitine-containing compositions can also be used as drugs.

It is advantageous, from an industrial viewpoint, that the aforesaid solid compositions lend themselves to be prepared with conventional apparatuses, and avoiding to resort to dehumidified facilities. This object can only be achieved if the aforesaid salts are stable and non- hygroscopic.

It has now been found that carnitine and zinc galactarates having formula (I): wherein: R = H or a straight or branched-chain alkanoyl group having 2-5 carbon atoms; n is 1 or 1/2; mis lor2 ; and (a) if m = 1 COOY is COO-if n = 1 ; or COOY is COOH if n = 1/2; or (b) if m=2 the COOYs are both COO-if n = 1 ; or one COOY = COOH, the other is COO-if n = 1/2 are stable and non hygroscopic, thus fully complying with the aforesaid prerequisites.

It is surprising that the salts of formula (I) are non-hygroscopic insofar as it could have been foreseen that, although L-carnitine and (C2-C5) alkanoyl galactares are non-hygroscopic, the corresponding zinc salts were hygroscopic. Indeed, although L-carnitine acid fumarate and L- (+)-tartrate are non-hygroscopic, the corresponding zinc salts, i. e. L- carnitine zinc fumarate and L-carnitine zinc tartrate are hygroscopic.

It is therefore, surprising that in the formation of zinc and carnitines salts with polybasic organic salts, galactaric acid's behaviour is different from that of fumaric and L- (+)-tartaric acid.

Preferably, the alkanoyl group of the galactarates of the invention is selected from the group consisting of acetyl, propionyl, butyryl, valeryl and isovaleryl.

With one of the codes"1: 1: 1","1 : 1/2 : 1,"2: 1: 1" and"2 : 1/2: 1" placed after the name of a galactarate, it is concisely represented the carnitine: zinc: galactaric acid molar ratio characterizing that specific galactarate.

Particularly preferred galactarates of formula (I) are the following: L-carnitine and zinc galactarate 1: 1: 1; L-carnitine and zinc galactarate 1: 1/2: 1; acetyl L-carnitine and zinc galactarate 1: 1: 1; acetyl L-carnitine and zinc galactarate 1: 1/2: 1; -propionyl L-carnitine and zinc galactarate 1: 1: 1; propionyl L-carnitine and zinc galactarate 1: 1/2: 1; isovaleryl L-carnitine and zinc galactarate 1: 1: 1; isovaleryl L-carnitine and zinc galactarate 1: 1/2: 1; L-carnitine and zinc galactarate 2: 1: 1; L-carnitine and zinc galactarate 2: 1/2: 1; acetyl L-carnitine and zinc galactarate 2: 1: 1; propionyl L-carnitine and zinc galactarate 2: 1: 1; and isovaleryl L-carnitine and zinc galactarate 2: 1: 1.

The galactarates of formula (I) can be prepared via one of the alternative procedures which are illustrated hereinbelow. The average- skilled expert shall be able to easily select the most suitable procedure in the light of the available apparatuses.

According to a first process, suitable molar amounts of galactaric acid, carnitine and zinc oxide, finely powdered, are thoroughly mixed and 15 to 35% of water is added to the mixture. The resulting mixture is rapidly heated to about 70-100°C and upon reaching the consistency of a pasty semi-liquid slurry, it is vigorously stirred for a few minutes and then discharged into collection tanks. On cooling the mass takes on the consistency of a dough which, mechanically stirred, under heating and by blowing thereon a current of dry, warm air, rapidly solidifies. The solid product thus obtained is then brought to the wanted water content by means of conventional drying apparatuses and then ground to the wanted particle size.

Alternatively, the aforesaid semi-liquid slurry is poured into a heated ball-mill, wherein a current of dry, warm air is circulated; the solid product which thus forms is brought to the wanted water content and then ground to the selected particle size.

According to a further process, suitable molar amounts of galactaric acid, carnitine and zinc oxide finely powdered are thoroughly mixed and 20 to 40% of water is added to the mixture. The resulting mixture is then rapidly heated to about 70-100°C and vigorously stirred for a few minutes. A ten-fold amount (in mL) of acetone with respect to the weight of the salt to be prepared is then added. Two phases form, the more viscous and heavier of which comprises the salt which is being prepared. This phase, due to the extracting action of acetone, becomes progressively lower in water, increasing in viscosity and hardening, until, after 5 to 15 minutes, it is completely transformed into a crystalline solid mass.

In order to avoid a hard crust formation on the reactor bottom, the reactor should be equipped with a potent stirrer. If such a stirrer is not available, the previously described procedure should be preferably adopted.

The following non-limiting examples show the preparation and the physico-chemical characteristics of some compounds of the present invention. The elementary analyses relate to the anhydrous compounds. Depending on the relative humidity the compounds are exposed to, they absorb small amounts of water resulting in the formation of stable hydrates.

Example 1 L-carnitine and zinc galactarate 1: 1: 1 To 21 g (0.1 moles) of galactaric acid, 16.1 g (0.1 moles) of L-carnitine inner salt and 8.14 g (0.1 moles) of zinc oxide, finely powdered and thoroughly mixed, 17 mL of water were added and the resulting mixture was rapidly brought under stirring to a temperature from 70°C to 80°C with an outer heating bath at 100-110°C. As soon as the mixture was sufficiently fluid, it was vigorously stirred for about two minutes and then discharged into collection tanks. The solidification process was aided by moving the reaction mixture, blowing a current of warm, dry air onto the mixture and also by heating the collection tanks.

The yield was quantitative.

IR (KBr), cm-1 3500-2900 (OH); 1598 (COO-) ; 1395 (COO-) ; 1120,1060 (C-O) Analysis for Cl3H23NOllZn Calc. C 35.92; H 5.33; N 3.22; Zn 15.04 Found C 35.45; H 5.40; N 3.20; Zn 14.98 Example 2 L-carnitine and zinc galactarate 1: 1: 1: To 21 g (0.1 moles) of galactaric acid, 16.1 g (0.1 moles) of L-carnitine inner salt and 8.14 g (0.1 moles) of zinc oxide, finely powdered and thoroughly mixed, 17 mL of water were added and the resulting mixture was rapidly brought under stirring to a temperature from 70°C to 80°C with an outer heating bath at 100-110°C. As soon as the mixture was sufficiently fluid, it was vigorously stirred for about two minutes and then 500 ml of acetone were added thereto. Two phases thus formed: the heavier and more viscous one was kept dispersed into the other by means of a vigorous stirring.

In about ten minutes the whole mass turned, via a progressive increase in viscosity and consistency, into a crystalline solid which was filtered and dried under vacuum.

The characteristics of the resulting product were the same as those of the compound of Example 1.

Example 3 L carnitme and zinc galactarate 1.1/2.1 The procedure of Example 1 was repeated, substituting 4.07g (0.05 moles) of zinc oxide for the double amount.

The yield was quantitative.

IR (KBr), cm-1 3500-2900 (OH); 1730 (COOH); 1598 (COO-) ; 1395 (COO-) ; 1100,1065 (C-O) Analysis for C13H23NOIIZnO5 Calc. C 38.74; H 6.00; N 3.48; Zn 8.11 Found C 38.70; H 6.06; N 3.41; Zn 8.01 Examples 4 and 5 Acetyl L-carnitine and zinc galactarate 1: 1: 1 Starting from 21 g (0.1 moles) of galactaric acid, 20.3 g (0.1 moles) of acetyl L-carnitine inner salt and 8.14 g (0.1 moles) of zinc oxide, finely powdered and thoroughly mixed, and 16 ml of water, the same procedures as those described in Example 1 (Example 4) and, respectively in Example 2 (Example 5) were repeated.

The yields were quantitative.

IR (KBr), cm-1 3500-3000 (OH); 1740 (COCHs) ; 1598 (COO-); 1385 (COO-); 1240,1100,1065 (C-O) Analysis for C15H25NO12Zn Calc. C 37.79; H 5.29; N 2.94; Zn 13.71 Found C 37.55; H 5.34; N 2.91; Zn 13.70 Examples 6 and 7 Propionyl L-carnitine and zinc galactarate 1.1 1 Starting from 21 g (0.1 moles) of galactaric acid, 21.7 g (0.1 moles) of propionyl L-carnitine inner salt and 8.14 g (0.1 moles) of zinc oxide, finely powdered and thoroughly mixed, and 17 mL of water, the same procedures as those described in Example 1 (Example 6) and, respectively in Example 2 (Example 7) were repeated.

The yields were quantitative.

IR (KBr), cm-1 3500-3000 (OH); 1738 (COCH2CH3) ; 1598 (COO-) ; 1385 (COO-) ; 1240,1100,1065 (C-O) Analysis for CisH27NOizZn Calc. C 39.16; H 5.55; N 2.85; Zn 13.32 Found C 39.00; H 5.58; N 2.80; Zn 13.30 Example 8 L-carnitine and zinc galactarate 2: 1: 1 The procedure of Example 1 was repeated, substituting 32.2 g (0.2 moles) of L-carnitine inner salt for 0.1 moles thereof.

The yield was quantitative.

IR (KBr), cm-1 3500-2900 (OH); 1598 (COO-) ; 1395 (COO-) ; 1120,1060 (C-O) Analysis for C20H38N204ZZn Calc. C 40.31; H 6.43; N 4.70; Zn 10.97 Found C 40.02; H 6.45; N 4.66; Zn 10.85 Example 9 L-carnitine and zinc galactarate 2: 1: 1 The procedure of Example 2 was repeated, substituting 32.2 g (0.2 moles) of L-carnitine inner salt for 0.1 moles thereof and adding 650 mL of acetone instead of 500 mL.

The characteristics of the product were the same as those of the compound of Example 8.

Example 10 L-carmtme and zinc galactarate 2: 1/2: 1 The procedure of Example 8 was repeated, substituting 4.07 g (0.05 moles) of zinc oxide for 0.1 moles.

The yield was quantitative.

IR (KBr), cm-1 3500-2900 (OH); 1730 (COOH); 1598 (COO-); 1395 (COO-) ; 1100,1065 (C-O) Analysis for C20H3sN2Ol4Znos Calc. C 42.58; H 6.97; N 4.96; Zn 5.79 Found C 42.39; H 7.03; N 5.00; Zn 5.70 Examples 11 and 12 Acetyl L-carnitine and zinc galactarate 2: 1: 1 Starting from 21 g (0.1 moles) of galactaric acid, 40.6 g (0.2 moles) of acetyl L-carnitine and 8.14 g (0.1 moles) of zinc oxide, finely powdered and thoroughly mixed, and 19 mL of water, the same procedures as those of Example 1 (Example 11) and, respectively Example 2 (Example 12) were repeated.

The yields were quantitative.

IR (KBr), cm-1 3500-3000 (OH); 1740 (COCHs) ; 1598 (COO-); 1385 (COO-) ; 1240, 1100,1065 (C-O) Analysis for C24H42N2OisZn Calc. C 42.39; H 6.23; N 4.12; Zn 9.61 Found C 42.29; H 6.30; N 4.06; Zn 9.55 Following the same procedures, propionyl L-carnitine and zinc galactarate 2: 1: 1: and isovaleryl L-carnitine and zinc galactarate 2: 1: 1: were also prepared.

The present invention also relates to compositions which comprise as active ingredient at least one of the aforesaid carnitine and zinc galactarates, and a pharmacologically acceptable excipient.

The compositions can present themselves as pharmaceuticals, OTC compositions, nutritional supplements, dietary supplements, veterinary products or fodders.

The compositions according to the present invention can also comprise further nutritional of pharmacological active ingredients. In particular, the compositions can comprise other pharmacologically acceptable salts of L-carnitine and/or (C2-C5) alkanoyl L-carnitines.

The compositions can also comprise fillers, binders, lubricants, mold- release agents, flow-regulating agents, dispersing agents, colorants, flavoring agents and the like as it will be apparent to any expert in pharmaceutical technology or pharmacy.

The orally administrable, solid forms comprise tablets, chewable tablets, pills, troches, lozenges, capsules, powders or granulates. In case of powders or granulates the presentation form can occur as sachets.

Since zinc's Recommended Dietary Allowance (RDA) is 12-15 mg/day for adults (Goodman and Gilman's,"The Pharmacological Basis of Therapeutics", Eight Edition, 1990, page 1525) the compositions of the present invention, in unit dosage form, suitably contain the whole zinc's RDA, i. e. 12-15 mg, or a lower amount such as e. g. about 4-8 mg of zinc as galactarate of formula (I), for those individuals who stay on a multiple dose administration regimen.

Compositions which are particularly suitable to contain at least one of the galactarates of formula (I) are compositions like those disclosed in the aforesaid US patent 6,090,848. The improved composition of the present invention may also comprise a selenium compound.

At least half the selenium in the male body is found in the semen.

Recent studies indicate that reduced daily selenium intake due to an unbalanced diet may be linked to an increased risk of male infertility.

A connection between selenium and sperm production has been reported. Selenium potent anti-oxidant activity in combination with vitamin E has long since been known.

Since selenium RDA is 55-57 ig/day (for adults) (see Goodman and Gilman's, loc. cit.), it is advantageous that the compositions of the invention in unit dosage form also comprise about 20-35 llg/day of selenium (preferably as selenomethionine).

Some non-limiting examples of improved compositions are given hereinbelow, wherein all the carnitine salts are non-hygroscopic and, therefore, lend themselves to the production not only of sachets, but also tablets, chewable tablets, pills, troches, lozenges and capsules. L- carnitine/acetyl L-carnitine molar ratio is about 3 in each composition.

The following abbreviations shall be used: Ga = galactarate; LC = L- carnitine; ALC = acetyl L-carnitine.

Composition 1 ALC Ga 0.600 g ALC: Zn: Ga 1: 1: 1 (Zn = 5.5 mg) 0.040 g LC fumarate 1.725 g Composition 2 ALC Ga 0.600 g ALC: Zn: Ga 1: 1/2: 1 (Zn = 5. 5 mg) 0.075 g LC fumarate 1.725 g Composition 3 ALC Ga 0.640 g LC: Zn: Ga 2: 1: 1 (Zn = 7 mg) 0.064 g LC fumarate 1.700 g Composition 4 ALC Ga 0.640 g LC: Zn 2: 1/2: 1 (Zn = 6 mg) 0.100 g LC fumarate 1.700 g In the light of zinc's RDA, in each of the aforesaid compositions 1-4, the amount of the carnitine: zinc: galactarate salt can be twice as much that indicated.

Advantageously, any composition may contain an effective amount of a selenium compound, e. g. 70 ig of selenomethionine corresponding to 28 g of selenium.

Optionally, further active ingredients, antioxidants and nutrients may supplement the compositions of the invention such as Vitamin C, Vitamin E, B Vitamins (B6, Bl2 and folic acid), Coenzime Qlo and ferulic acid.

As it will be apparent to any expert in pharmaceutical technology or pharmacy, the compositions for sachets may comprise suitable excipients such as fructose, citric acid, saccharin sodium, tonic water flavour, D-mannitol and colloidal silicon dioxide.

The compositions for tablets and chewable tablets may comprise excipients such as mint essence, saccharin sodium, sorbitol solution, sorbitol, magnesium stearate, talc, pregelatinized corn starch, mannitol and saccharose.

Thanks to the stability and non-hygroscopicity of the aforesaid salts of L-carnitine and alkanoyl L-carnitines, the compositions for capsules can be entirely free of excipients, in view of the chemical inertness of the ingredients towards the gelatinous material comprising the capsules.