ORAL COMPOSITION COMPRISING S-ADENOSYL METHIONINE AND A PROBIOTIC AND ITS USE FOR TREATING AND/OR PREVENTING MOOD DISORDERS

Abstract

The present invention refers to oral compositions containing S-adenosyl methionine (or S-adenosyl-1-methionine or SAMe) in combination with at least one probiotic, at least one physiologically acceptable excipient and optionally at least one additional ingredient and use thereof in the treatment and/or prevention of mood disorders preferably in case of anxiety or depression.

Claims

1. A composition comprising S-adenosyl methionine and at least one probiotic, in which said at least one probiotic is present in a microencapsulated form.

2. A solid oral composition comprising of S-adenosyl methionine and at least one probiotic according to claim 1, and at least one physiologically acceptable excipient.

3. The solid oral composition according to claim 2, wherein said solid oral composition is orosoluble.

4. The solid oral composition according to claim 2, wherein said at least one probiotic is selected from Lactobacillus helveticus, Bifodobacterium longum or a mixture thereof.

5. The solid oral composition according to claim 3, wherein said probiotic is a mixture of Lactobacillus helveticus and Bifidobacterium longum.

6. The composition according to claim 1, wherein said S-adenosyl methionine is S-adenosyl methionine disulfate p-toluensulfonate.

7. The composition according to claim 1, wherein said S-adenosyl methionine is contained in an amount from 50 to 500 mg, expressed in mg of SAMe ion.

8. The composition according to claim 1, further comprising at least one additional ingredient selected from minerals, vitamin group B or a mixture thereof.

9. A composition according to claim 1, wherein said composition is in the form of a tablet, capsule, powder, granule.

10. A composition according to claim 1, for use in the treatment of anxiety, depression, psychological stress, mild postpartum syndromes and/or stress related intestinal disorders.

11. The composition according to claim 10, wherein said depression is minor, mild or subthreshold depression.

12. The solid oral composition according to claim 3, wherein said probiotic is a mixture of Lactobacillus helveticus Rosell®-52 and Bifidobacterium longum Rosell®-175.

13. The composition according to claim 1, wherein said S-adenosyl methionine is contained in an amount from 100 to 400 mg, expressed in mg of SAMe ion.

14. The composition according to claim 1, wherein said S-adenosyl methionine is contained in an amount of about 200 mg, expressed in mg of SAMe ion.

15. The composition according to claim 1, further comprising a magnesium oxide and vitamin B6.

16. A composition according to claim 1, wherein said composition is in the form of a tablet or granulate.

17. A composition according to claim 1, wherein said composition is in the form of an orosoluble granulate.

18. A composition according to claim 1, for use in the treatment and/or prevention of mood disorders.

Description

DESCRIPTION

[0042] Surprisingly, it has now been found that a new composition comprising S-adenosyl-1-methionine (SAMe and/or salts thereof) and at least one probiotic, specifically formulated in solid oral form, preferably orosoluble, provides a simultaneous supplementation of these components without nullifying the stability of the two components. Thanks to this new formulation it is now possible to obtain one or more of the following advantages at the level of processing/production, of finished product and/or at the level of treatment, and that is i) overcoming the technical difficulties linked to the formulation of these substances singularly and even more in combination with each other; ii) the obtainment of a product containing stable, palatable, highly bioavailable SAMe; iii) the obtainment of a product containing alive probiotics, capable of arriving unaltered to the intestine; iv) the maintenance of probiotics in active form, without degradation; it is in fact surprising how, despite the acidity of SAMe, there is no degradation of probiotics thanks to the new formulation of the invention; v) an integrated effect of substances formulated together in a single composition; vi) an improved therapeutical adherence (compliance) by the patient and vii) a synergistic effect between SAMe and said at least one probiotic. Such advantages eventually lead to an improved efficacy on mood disorders, with no side effects.

[0043] An object of the present invention is therefore a combination of S-adenosyl methionine (or S-adenosyl-1-methionine or SAMe) and at least one probiotic. An object of the present invention is further a solid oral composition, preferably orosoluble, comprising S-adenosyl methionine (SAMe) in combination with at least one probiotic and at least one physiologically acceptable excipient.

[0044] Further object of the present invention is a combination of S-adenosyl methionine (or S-adenosyl-1-methionine or SAMe) and at least one probiotic, for use in the treatment and/or prevention of mood disorders, preferably for use in case of anxiety and/or depression (primary or secondary).

[0045] Further object of the present invention is a solid oral composition, preferably orosoluble, comprising S-adenosyl methionine (SAMe) in combination with at least one probiotic and at least one physiologically acceptable excipient for use in the treatment and/or prevention of mood disorders, preferably for use in case of anxiety and/or depression (primary or secondary).

[0046] Further object of the present invention is a solid oral composition, preferably orosoluble, comprising S-adenosyl methionine (SAMe) in combination with at least one probiotic and at least one physiologically acceptable excipient where said SAMe is in stable and/or palatable form.

[0047] Further object of the present invention is a solid oral composition, preferably orosoluble, comprising S-adenosyl methionine (SAMe) in combination with at least one probiotic and at least one physiologically acceptable excipient, where said at least one probiotic is alive and active, and reaches the intestine in an alive and active form.

[0048] Further object of the present invention is a solid oral composition, preferably orosoluble, comprising S-adenosyl methionine (SAMe) in combination with at least one probiotic and at least one physiologically acceptable excipient where said composition improves patient therapeutical adherence to the treatment (compliance).

[0049] Further object of the present invention is a solid oral composition, preferably orosoluble, comprising S-adenosyl methionine (SAMe) in combination with at least one probiotic and at least one physiologically acceptable excipient where said SAMe and at least one probiotic remain stable in the composition.

[0050] Unless otherwise specified, within the scope of the present invention a range of values indicated for a quantity, for example the weight or percentage content of a component, includes the lower limit and the upper limit of the range. For example, if the weight or volume content of a component A is indicated as “from X to Y”, where X and Y are numerical values, A may be X or Y or anyone of the intermediate values.

[0051] Unless otherwise specified, within the scope of the present invention the indication that a composition “comprises” one or more components or substances means that other components or substances may be present beside that, or those, specifically indicated. The terms “comprising”, “having”, “including” and “containing” are to be intended as open terms (i.e., meaning “comprising, but not limited to”) and are to be considered as a support also for terms such as “to consist essentially of”, “consisting essentially of”, “to consist of” or “consisting of”.

[0052] Unless otherwise indicated, within the scope of the present invention the term “physiologically acceptable excipient” refers to a substance lacking in any pharmacological effect of its own and which does not produce adverse reactions when administered to a mammal, preferably a human being. Physiologically accepted excipients are well known in the art and are described, for example, in the Handbook of Pharmaceutical Excipients.

[0053] Unless otherwise indicated, within the scope of the present invention the term “simultaneous” refers to the contemporaneous administration of active substances, in the same composition, such that the compounds act together in the body, giving origin to a contemporaneous integration thereof.

[0054] Unless otherwise indicated, within the scope of the present invention the term S-adenosyl methionine refers to the S-adenosyl methionine (or S-adenosyl-l-methionine or SAMe) molecule and/or to anyone of the pharmaceutical acceptable salts thereof. In particular, S-adenosyl-methionine salts are selected from S-adenosyl methionine disulfate p-toluensulfate, S-adenosyl methionine butanediene sulfate, S-adenosyl methionine phytate. Preferably, S-adenosyl methionine disulfate p-toluensulfate, and more preferably S-adenosyl methionine disulfate p-toluensulfate titrated at 50%.

[0055] Said at least one probiotic according to the present invention preferably belongs to the Lactobacillus and/or Bifidobacterium class and is more preferably selected from Lactobacillus helveticus, Bifidobacterium longum or a mixture thereof. Even more preferably, said at least one probiotic according to the present invention is a Lactobacillus helveticus and Bifidobacterium longum mixture.

[0056] According to a preferred embodiment, the composition of the present invention contains a mixture of Lactobacillus helveticus Rosell®-52 and Bifidobacterium longum Rosell®-175. The Lactobacillus helveticus Rosell®-52 strain belongs to the genre of Lactobacilli, it was isolated in 1990 and its genomic sequence was entirely mapped and it was deposited in France with the identification code I-1722-CNCM. In accordance with the present invention, it is important to note that said strain is capable of surviving well along the gastrointestinal tract, adhering to the intestinal epithelium cells, displacing pathogens and increasing the barrier function.

[0057] The Lactobacillus helveticus Rosell®-175 strain belongs to the genre of Bifidobacteria, it was isolated in 1988 and its genomic sequence was entirely mapped and it was deposited in France with the code I-3470-CNCM. In accordance with the present invention, it is also important to note here that such strain survives well in the gastrointestinal tract and carries out an antimicrobial activity. Such probiotic strain proves to be active against a great number of intestinal pathogens (such as, for example, E. Coli, C. difficile, S. aureus) and is capable of reducing the production of inflammatory cytokines, therefore positively modulating the immune response and decreasing intestinal permeability.

[0058] Unless otherwise specified the term “orosoluble” according to the present invention refers to compositions capable of dissolving and releasing immediately the active substance contained therein, when in contact with the oral mucosa. In this way the active substance can be directly absorbed in the oral mucosa, thus avoiding the hepatic circle.

[0059] The term orosoluble according the invention is therefore aimed at compositions to insert in the oral cavity, where the active substance S-adenosyl methionine becomes immediately orodispersible and available.

[0060] Thanks to this specific formulation the S-adenosyl methionine is absorbed right away, ensuring an improved bioavailability thereof. According to a preferred embodiment, in fact, the absorption is ensured by the orosoluble form, which allows to solve the problems of scarce bioavailability which generally characterize the oral forms of this substance.

[0061] The term “oral” according to the present invention refers to a composition aimed at administration by mouth. The oral composition according to the invention is in solid form, preferably orosoluble. Preferably, said solid oral form of the invention is in form of a tablet, capsule, powder, granulate or microgranulate, more preferably granulate or tablet.

[0062] According to a preferred embodiment, the solid oral form of the invention is orosoluble granulate.

[0063] According to another preferred embodiment, the solid oral form of the invention is orosoluble tablet or modified-release multilayer swallowable tablet.

[0064] According to a further preferred embodiment the solid oral form of the invention is effervescent orosoluble and/or soluble granulate.

[0065] According to a further preferred embodiment, the composition according to the present invention is packaged in sachet, preferably single-dose sachet or bipartite sachet.

[0066] According to a more preferred embodiment the composition of the present invention is in form of an orosoluble granulate, packaged in single-dose sachet (stick pack).

[0067] The probiotic strains are present in the composition according to the present invention in a preferably gastro-resistant form, such to go beyond the gastric district and to reach the intestine where they must colonize and perform their action. More preferably, probiotic strains are present in the composition in microencapsulated form. The microencapsulated form according to the invention ensures the probiotic strains a greater stability, such to allow going beyond the stomach and reaching the intestine in alive and active form. Thanks to the microencapsulation the probiotics of the invention are therefore gastro-resistant, stable from the moment of intake up to the site of action.

[0068] As said above, the probiotics are very delicate substances or living organisms and going beyond the stomach, with the reaching of the intestine in alive and active form represents one of the formulation hurdles to overcome. Gastro-resistance becomes a crucial aspect in the processing of the probiotic, as preferably comprised in the composition of the invention. Thanks to their new microencapsulated form it is possible to obtain such result. Thanks to this specific composition, probiotic strains arrive to the intestine alive and active, ensuring the best efficacy.

[0069] Said at least one physiologically acceptable excipient according to the present invention can be selected from diluents, aggregants or binders, lubricants, glidants, disaggregants, solubilizers, sweeteners, flavoring agents, pH adjusters, or a mixture thereof.

[0070] The composition according to the present invention may further comprise one or more additional ingredients, preferably selected from minerals and/or vitamins. More preferably, said minerals are chosen from magnesium, potassium, calcium, phosphorus, iron, zinc, copper, manganese, fluorine, selenium, chromium, molybdenum, iodine, boron, chlorine, sodium and silicon or other mineral comprised in the corresponding Table of the Italian Ministry of Health or a mixture thereof, whereas said vitamins are vitamins group B, preferably selected from Vitamin B1, Vitamin B6, Vitamin B12, or Vitamin B9 or other vitamin such as Vitamin A, Vitamin D, Vitamin E, Vitamin K, Vitamin C, riboflavin (Vitamin B2), Niacin, Folic Acid, Biotin, Pantothenic Acid, or other Vitamin comprised in the corresponding Table of the Italian Ministry of Health or a mixture thereof.

[0071] According to a preferred embodiment of the present invention, the composition comprises magnesium and vitamin B6 as additional ingredients. Preferably, the magnesium of the present invention is magnesium oxide. Magnesium oxide and vitamin B6 contribute synergistically to rebalance the nervous system and normalize the psychological function.

[0072] The synergistic effect given by the composition of the invention is further amplified by the combined effect of magnesium oxide and vitamin B6, which flanks the integrated effect of the simultaneous administration of SAMe and probiotics.

[0073] The S-adenosyl methionine (SAMe) is contained in the composition of the present invention in an amount that varies from 50 mg to 500 mg, preferably from 100 mg to 400 mg. More preferably, the S-adenosyl methionine is contained in the composition of the present invention in an amount of about 200 mg. The amounts of SAMe in the composition are here expressed as mg of SAMe ion; the amounts here expressed are intended for single composition, preferably expressed in mg for sachet (mg/stick pack), always expressed as mg of SAMe ion.

[0074] Said at least one probiotic is contained in the composition of the present invention in an amount that varies from 0.5×10.sup.9 CFU and 30×10.sup.9 CFU (Colony-Forming Units), preferably from 1×10.sup.9 CFU to 5×10.sup.9 CFU. More preferably said at least one probiotic is contained in the composition of the present invention in an amount of about 3×10.sup.9 CFU. According to a preferred embodiment of the invention, the probiotic is represented by the one mixture of probiotics which preferably corresponds to a Lactobacillus and Bifidobacterium mixture, more preferably as discussed above a Lactobacillus helveticus and Bifidobacterium longum mixture. Therefore, the probiotic mixture is contained in the composition of the present invention in an amount that varies from 0.5×10.sup.9 CFU and 30×10.sup.9 CFU (Colony-Forming Units), preferably from 1×10.sup.9 CFU to 5×10.sup.9 CFU. More preferably said at least one probiotic is contained in the composition of the present invention in an amount of about 3×10.sup.9 CFU. The amounts here expressed are intended for single composition, preferably expressed in sachet (stick pack), or for daily dose.

[0075] The vitamin group B is present in the composition of the present invention in an amount that varies from 5% VNR to 1250% VNR (Value Nutritional Rate), preferably from 15% to 300% VNR. More preferably said vitamin is contained in an amount of about 100% VNR. According to a preferred embodiment of the present invention said vitamin is vitamin B6 (Pyridoxine Hydrochloride powder). The mineral is contained in the composition of the present invention in an amount that varies from 5% VNR to 300% VNR (Value Nutritional Rate), preferably from 10% VNR to 100% VNR. More preferably, said mineral is contained in an amount of about 15% VNR. According to a preferred embodiment of the present invention the mineral is magnesium oxide, the dosage of which corresponds to a daily dose of magnesium of about 15% VNR. The amounts here expressed are intended for single composition, preferably expressed in sachet (stick pack), or for daily dose. The composition according to the present invention is indicated for use in the treatment and/or prevention of mood disorders, preferably in case of anxiety and depression (both generalized Anxiety Disorder, Minor Depression, reactive Depression and Unipolar Major Depression). More preferably, Minor or Subthreshold Depression (mild-depression). The depression of the invention can be primary and/or secondary.

[0076] With the term “mood disorders” according to the present invention, a wide series of disorders and/or pathologies which involve the central nervous system, with respect to the emotional and psychological balance, are meant to be taken into account. In particular, the composition of the present invention is useful in case of anxiety, depression, psychological stress (of various origin such as, for example, work, family, study or other), mild post-partum syndromes (such as post-partum Blues Syndrome) and/or intestinal disorders correlated to stress (such as for example irritable bowel syndrome, leaky gut syndrome) and burn-out symptoms. The depression can be primary depression or secondary depression, where for secondary a widespread condition among many patients already affected by chronic diseases (for example diabetes or hypertension) or acute diseases (for example cancer, TBC, HIV) is intended. Preferably, the composition of the present invention is useful in case of anxiety or depression, where the depression can be minor depression (or subthreshold—mild depression) or major depression (in combination with AD); more preferably, the composition of the invention is indicated in case of minor depression (or subthreshold—mild depression).

[0077] The depression, whether minor or subthreshold, can also be correlated to changes of the hormonal component, in particular for women during the transition phase toward menopause (perimenopause) or during menopause itself. Menopause and depression can also be an association altering the optimal functioning of many women. It is an evidence base medicine datum that the risk of depression increases during perimenopause, in case of early menopause and in the period after menopause. The correlation between menopause and depression appears to be closely linked to hormonal changes which characterize this phase of a woman's life. At the same time, however, the stress and culture with which this physiological change is faced and experienced can also affect our mood, even going so far as to affect the subtle biochemical balances that regulate it.

[0078] “Minor” depression in menopause is therefore an eventuality to evaluate with care and awareness to prevent and treat a physical and mental problem which can limit a woman's life and which represents an important risk factor for her health. The composition of the invention can be particularly advantageous in these cases of hormonal imbalance, with also an improvement of the dysbiosis with consequent attenuation of inflammatory and degenerative phenomena of the mucous membranes.

[0079] According to a preferred embodiment, the composition of the present invention is administered with a posology of one single-dose sachet (stick-pack) per day, preferably one sachet in the morning after the first main meal. More preferably for at least two weeks.

[0080] The single daily administration of the composition of the invention is correlated to an improved acceptance of the treatment by the patient who in this way overcomes the possible distress of multiple intakes (compliance). Having to take different compositions or products, possibly several times a day and/or at different times of the day, is one of the problems often encountered in the patient affected by emotional imbalance and mood disorder. It is important that the patient adheres in a precise and continuous way to the treatment, and the administration of a single composition, preferably once a day, solves the problem. This datum is based on EBM: in chronic therapies, the patient who already has to take multiple drugs a day prefers the single administration. The patient's subjective perception of taking numerous (“too many” or as such perceived) drugs often leads the patient to a reduced compliance. The fixed combination of active substances and/or ingredients according to the present invention, with respect to the current individual administrations of multiple products, even several times a day, leads to the advantage of an improved compliance.

[0081] Such type of composition, administration and/or packaging of the product therefore improves compliance of the patient, to whom, thanks to the composition of the invention, the necessary intake of both SAMe and probiotics is provided, optionally with the addition of one or more further ingredients. As already said above, patient adherence to the pharmacological treatment (compliance) is of fundamental importance for the success of therapeutic intervention in chronic diseases, including anxiety and depression (both primary and secondary).

[0082] The present composition in form of orosoluble granulate is packaged in single-dose sachet (stick-pack). According to a preferred embodiment, the composition of the present invention is a food supplement.

Experimental Part

1. Compatibility Study

[0083] A compatibility study intended to demonstrate the possibility to mix SAMe (or its salt) and probiotics was executed. The probiotics to be tested have been selected in accordance with the composition of the invention such as Lactobacillus helveticus Rosell®-52, Bifidobacterium longum Rosell®-175 and a mixture of the two. The study was further conducted using such probiotics in both not microencapsulated and microencapsulated forms.

[0084] The study was carried out in a first phase with SAMe+Lactobacillus helveticus R0052, not microencapsulated, through a liquid compatibility method. The initial population tested was 15.10.sup.E9 CFU/g units at 500 mg of SAMe. For this evaluation a pHmeter with 8 probes was used, and values were verified every 15 minutes.

[0085] The strains are inoculated into a liquid medium suitable for their growth, only under aerobic conditions (reason why in this way the Bifidobacterium is not tested), and the active substance is inoculated into the sample and not into the control. The compatibility evaluation was executed monitoring the pH, as following: [0086] if an acidification is found: a multiplication of the strain (natural fermentation) occurs and therefore no negative interaction with the active substance is produced; [0087] if a light acidification of the medium is found: a multiplication of the strain occurs, but slower than it should be; the method of analysis must be adequate to the time of the microorganisms count on the finished product; [0088] if no acidification is found: the active substance shows bacteriostatic or lethal effect on the strain.

[0089] From an operational point of view: the vial is filled with the liquid medium (50 ml RCM); the active, which should have been tested directly in its granulated form in the vial, is added, and it is mixed. Then the pH of the medium in the presence of the active is verified, if necessary, the pH is adjusted so that it is comprised between 6.5 and 7. R0052 strain is therefore inoculated (batch U12171600, 500.10.sup.E9 CFU/g) into the vial. The sample is diluted 1/100 with Dilumat to obtain a dilution to 5.10.sup.E9 CFU/ml and finally it is inoculated with 3 ml into the 50 ml RCM vial, obtaining 15.10.sup.E9 CFU/ml in vial. The vial is placed in warm bath, at 37°, and the pH started to be recorded for 18 hours.

[0090] Conclusion: it was seen that pH of the R0052 strain decreases from 6.1 to 4.2: it is the natural multiplication of the bacteria that acidifies the medium. The pH of the active (SAMe) remains essentially unchanged, stable and it is neutral (7-7.5). When R0052 is put in contact with the SAMe sample, the pH decreases from 7.2 to 4.2. Consequently, it is demonstrated that the L. helveticus R0052 strain appears compatible with the SAMe.

[0091] 1.2 MICROBIOLOGICAL TEST: within the bio-compatibility study a microbiological evaluation was carried out. R0052 and R0175 probiotic strains were cultured in a Petri dish, in the presence of the active (SAMe, marketed formulation) at a 500 mg dose (as above). A sample was also analyzed as a reference of probiotic strain only, so that it could be compared with the addition of the active on the result in Colony-Forming Units (CFU). The obtained results showed that the insertion of the SAMe in the presence of each probiotic strain (R0052 o R0175) modifies the CFU count only up to 5%. This is easily justified by the simple analytical error, known in the microbiological field. These results therefore confirm what already seen with the compatibility test, demonstrating again that the probiotic strains are compatible with SAMe.

[0092] 2.SAMe—MIVO® STUDY: in vitro study of absorption of S- Adenosil-L-Metionina (SAMe), through the buccal epithelial tissue and MIVO® device.

[0093] The in vitro study of absorption was carried on with the utilization of reconstructed oral epithelial tissue Reconstructed Human Oral Epithelium (HOES5 kit of 12 tissues) of EpiSkin Company mounted on the MIVO® Bulk2 Box (12 MIVO®) device of React4life Company.

[0094] Given the orosoluble characteristic of the composition as described below in Example 1, in a first experimental phase the oral digestion of the active is mimicked through incubation of the samples with solutions of artificial saliva.

[0095] Subsequently to the first digestive phase, the assay of the absorption of SAMe through HOES5 follows, a 3D model of epithelial tissue free of the stratum corneum, which histologically resembles the mucosa of the oral cavity, usable for tests of toxicity, metabolic, absorption and efficacy of the compounds.

[0096] The absorption assay is executed in triplicate mounting the transwell of the HOES5 tissue on the MIVO® device, which combines 3D tissues and fluidic circulation, reproducing physiological conditions for the tissues at the following experimental timepoints: T10 (10 minutes after the insertion of the digested sample in contact with HOES5 tissue), T20 (20 minutes after the insertion) and T30 (30 minutes after the insertion).

[0097] Subsequently, the quantitative analysis of the analyte SAMe ion, present in the formulation of the Example 1 pre-digested, digested and after each experimental timepoint, is carried out through RP-HPLC chromatographic analytical technique, including fluorometric and UV detector.

[0098] Each sample is analyzed at a non-toxic concentration for the cells of the buccal tissue, the same is derived starting from the present SAMe quantity and from the posology of the finished product.

[0099] The permeability of the tissue to each active is expressed in terms of percentage of permeation with respect to the applied quantity.

[0100] All the experiments are carried out in triplicate (biological replicates) and the results expressed as mean±SEM (standard error of the mean). The statistical significance between the groups is evaluated through two-tailed Student t test (p<0.05), after evaluation of the normal distribution of the data.

[0101] The expected results for such in vitro study demonstrate the absorption of the SAMe according to the invention through the buccal epithelial tissue.

3.Clinical Efficacy Study

[0102] A study on the efficacy of the composition according to the present invention is under evaluation, as reported in example 1, in patients with minor or subthreshold depression (mild-depression): the clinical study is single-center, controlled, randomized, parallel-group, cross-over, with run-in and follow-up periods, double-blind vs placebo.

[0103] Primary objective of the clinical study is to evaluate the efficacy of the composition of the present invention as food supplement based on S-adenosyl methionine (SAMe), Lactobacillus helveticus and Bifidobacterium longum in improving and/or preventing the mood tone disorders.

[0104] Such evaluation will be measured through the administration to the enrolled subjects of validated questionnaires: [0105] Hamilton Depression Rating Scale—HAM-D (administered by the doctor), [0106] General Health Questionnaire—GHQ-1 (self-administered in the doctor's office).

[0107] Perceived Stress Scale—PSS Test (self-administered in the doctor's office). Secondary objective of the study is to evaluate the efficacy of the composition of the present invention as food supplement based on S-adenosyl methionine (SAMe), Lactobacillus helveticus and Bifidobacterium longum in cases of anxiety disorders. Such evaluation will be measured through the administration to the enrolled subjects of the validate questionnaire: [0108] Hamilton Anxiety Rating Scale (HAM-A) (administered by the doctor); and [0109] Self-administered tests to carefully evaluate the psychic condition of the investigator.

[0110] The experimental groups will be the following: [0111] Group 1: subjects who will take the treatment daily; and [0112] Group 2: subjects who will take the placebo daily. [0113] The total duration of the study will be 12 months: 1 month for patients enrollment, 3 months of treatment, 1 month of wash-out, a cross-over design and finally 6 weeks of follow-up.

4. Stability Study

[0114] The stability of the composition as described in example 1 has been evaluated for a period of 5 days in the oven at 50°. Control parameters were: [0115] Appearance [0116] Mean weight [0117] % H.sub.2O determined through KF (Karl Fischer) method [0118] Assay of SAMe active [0119] Degradation products

[0120] The product showed to be stable for 5 days, at 50° temperature, as it can be inferred from the table below. Such accelerated stability test is an industry standard with a well-established use for products based on SAMe.

TABLE-US-00001 SAMe + PROBIOTICS BATCH: 1518-7 ANALYSIS TIME ZERO 5 DAYS APPEARANCE White powder Unchanged MEAN WEIGHT 1.889 g/bst 1.886 g/bst K.F. 0.732% 0.744% ASSAY 187.96 mg/bst 169.74 mg/bst DEGRAD. PROD. 1.46% A 3.88% A (0.04 W/W) (0.09 W/W)

[0121] The following examples are included herein to better describe the present invention, without limiting it in any way.

EXAMPLES

[0122]

TABLE-US-00002 EXAMPLE N1 PHARMACEUTICAL FORM OROSOLUBLE GRANULATE IN STICK PACK INGREDIENTS mg/stick pack S-adenosyl-L-methionine disulfate p- 400 toluensulfonate (SAMe) of which SAMe ion 200 Mixture of lactic ferments (Lactobacillus 3 × 10.sup.9 CFU helveticus Rosell-52, Bifidobacterium longum Rosell -175) Magnesium oxide 93.3 of which magnesium 56.25 Vitamin B6 (Pyridoxine Hydrochloride) powder 1.7 of which Vitamin B6 1.4 Xylitol 660.68 L-Arginine 150 Mannitol 100 Fatty acids 80 Flavour 18.79 Silicon dioxide 9.15 Sucralose 1.37 Acesulfame k 0.85 Neohesperidine DC 0.16 Total per stick pack 1900

TABLE-US-00003 EXAMPLE N2 PHARMACEUTICAL FORM OROSOLUBLE GRANULATE IN SACHET INGREDIENTS mg/sachet S-adenosyl-L-methionine disulfate p- 500 toluensulfonate (SAMe) of which SAMe ion 250 Mixture of lactic ferments (Lactobacillus 6 × 10.sup.9 CFU helveticus Rosell-52, Bifidobacterium longum Rosell -175) Magnesium oxide 186.6 of which magnesium 112.5 Vitamin B6 (Pyridoxine Hydrochloride) powder 11.53 of which Vitamin B6 9.5 Xylitol 1237.37 L-Arginine 200 Mannitol 1000 Fatty acids 50 Flavour 25 Silicon dioxide 15 Sucralose 5 Acesulfame k 1 Neohesperidine DC 0.5 Total per sachet 4000

TABLE-US-00004 EXAMPLE N3 PHARMACEUTICAL FORM MODIFIED- RELEASE MULTILAYER SWALLOWABLE TABLET INGREDIENTS mg/tbl S-adenosyl-L-methionine disulfate p- 400 toluensulfonate (SAMe) of which SAMe ion 200 Mixture of lactic ferments (Lactobacillus 3 × 10.sup.9 CFU helveticus Rosell-52, Bifidobacterium longum Rosell -175) Magnesium oxide 93.3 of which magnesium 56.25 Vitamin B6 (Pyridoxine Hydrochloride) powder 1.7 of which Vitamin B6 1.4 Maltodextrin 116 Microcrystalline cellulose 100 Fatty acids 80 red iron oxide 5 Silicon dioxide 10 Magnesium stearate 5 Ethylcellulose 5 Total per tablet 1200

TABLE-US-00005 EXAMPLE N4 PHARMACEUTICAL FORM OROSOLUBLE TABLET INGREDIENTS mg/tbls S-adenosyl-L-methionine disulfate p- 250 toluensulfonate (SAMe) of which SAMe ion 125 Mixture of lactic ferments (Lactobacillus 1.5 × 10.sup.9 CFU helveticus Rosell-52, Bifidobacterium longum Rosell -175) Magnesium oxide 93.3 of which magnesium 56.25 Vitamin B1 2.5 Vitamin B2 2.5 Vitamin C 80 Xylitol 717.19 L-Arginine 100 Mannitol 80 Fatty acids 60 Flavour 15 Silicon dioxide 5 Sucralose 1.5 Acesulfame k 0.85 Neohesperidine DC 0.16 Total per tablet 1600

TABLE-US-00006 EXAMPLE N5 PHARMACEUTICAL FORM EFFERVESCENT OROSOLUBLE GRANULATE IN BIPARTITE SACHET INGREDIENTS mg/sachet CHAMBER A S-adenosyl-L-methionine disulfate p- 500 toluensulfonate (SAMe) of which SAMe ion 250 L-Arginine 200 Fatty acids 100 Silicon dioxide 10 CHAMBER B Mixture of lactic ferments (Lactobacillus 6 × 10.sup.9 CFU helveticus Rosell-52, Bifidobacterium longum Rosell -175) Magnesium oxide 186.6 of which magnesium 112.5 Vitamin B6 (Pyridoxine Hydrochloride) powder 11.53 of which Vitamin B6 9.5 Xylitol 177.37 Sodium Bicarbonate 500 Flavour 25 Silicon dioxide 15 Sucralose 5 Acesulfame k 1 Neohesperidine DC 0.5 Total per bipartite sachet 2500

5.Pilot Clinical Study

[0123] A pilot clinical study entitled: “Efficacy study of the food supplement based on S-adenosyl methionine (SAMe), and probiotics for the maintenance of the normal mood tone through the reduction of subthreshold depression, as risk factor of major depression” was conducted. With this pilot study the efficacy of the composition of the invention based on S-adenosyl methionine (SAMe), Lactobacillus helveticus e Bifidobacterium longum as reported in Example 1 was evaluated in the improvement of the mood tone in healthy subjects, with a mild alteration of the mood tone, potential risk factor for the development of major depression. The duration of the treatment was two months.

[0124] The participants underwent two office visits. During the first visit, carried out at the enrollment (t0), sociodemographic, clinical parameters and scores obtained after the completion of the following two questionnaires, chosen as primary outcomes of the study, were recorded on the patient card (Case report form—CRF): 1) SCORE PATIENT HEALTH QUESTIONNAIRE-9 (PHQ-9), which investigates the presence “in the last two weeks” of the 9 symptoms of depression and the functional impairment caused by depression on the normal development of patient's life, and 2) HAMILTON RATING SCALE FOR DEPRESSION (HAM-D), which evaluates the severity of depressive symptoms and it is useful to document its modifications, e.g. following treatment. In the second visit, carried out after 10 weeks (t1), the above-mentioned questionnaires were again administered to the subjects and the obtained values were reported on the CRF.

[0125] In total 10 subjects were enrolled in the pilot clinical study, with the following inclusion and exclusion criteria: [0126] INCLUSION CRITERIA: male and female subjects aged between 18 and 65 years with scores less than or equal to 19 in the HAM-D and to 17 in the PHQ-9 (corresponding to subjects with a slightly altered mood tone, unsuitable for antidepressant drugs because with mild or moderate depression), capable of understanding and satisfying the protocol requirements. [0127] EXCLUSION CRITERIA: subjects severely depressed or who suffered from severe depression over the last years; individuals who took drugs or food supplements with activity on the psychological function over the 12 weeks before intake; pregnant women and those who took monoamine oxidase inhibitors (anti-MAO) over the 14 days before intake.

Results

[0128] In Table 1 the demographic characteristics and scores at baseline of HAM-D and PHQ-9 questionnaires have been reported.

TABLE-US-00007 TABLE 1 Characteristics Study population (n = 10) Age  39 ± 19 Ethnic origin: European HAM-D* 14 ± 5 PHQ-9** 10 ± 4 *The total score of the HAM-D to 21 questions was given by the sum of the first 17 items, considered those “cardinal” of depression: 0-7 = subjects with normal mood; 8-16 = mild depression; 17-23 = moderate depression; >24 = severe depression **The score of the PHQ-9 was considered a range between 0 and 27. The scores between 5 and 9 indicates the presence of a subthreshold depression. The score of 10 is the optimal cut-off to highlight depressions of clinical relevance with three different levels of severity depending on the score. 0-4 = Absent; 5-9 = Subthreshold depression; 10-14 = Mild major depression; 15-19 = Moderate major depression; >20 = Severe major depression.

[0129] In Table 2 the scores of HAM-D and PHQ-9 questionnaires recorded for each patient have been reported.

TABLE-US-00008 TABLE 2 Subject's code Time HAM-D PHQ-9 MRF t0 18 11 t1 11 4 MR t0 16 12 t1 10 5 AS t0 7 9 t1 5 8 AR t0 19 17 t1 12 6 A.S. t0 13 6 t1 9 3 ML t0 9 6 t1 5 4 MD t0 14 6 t1 8 4 TLAV t0 19 7 t1 11 4 ORRS t0 12 10 t1 8 8 GGED t0 8 12 t1 2 8

[0130] In Table 3 the descriptive statistics (mean±SD, range) of the score of the two questionnaires measured at t0 and t1 is reported.

TABLE-US-00009 TABLE 3 Questionnaires t0 t1 HAM-D 13 ± 4 (7-19)  8 ± 3 (2-12) PHQ-9 10 ± 4 (6-17) 5 ± 2 (3-8)

Discussion

[0131] The results demonstrate that in all the enrolled subjects a reduction of the symptomatology of depressive state occurred.

[0132] As far as the HAM-D questionnaire is concerned, for the 60% of the subjects (6 out of 10) a decrease of the depressive state level was recorded. Specifically: [0133] 3 subjects out of 10 moved from a moderate depression state to a mild depression state; [0134] 3 subjects out of 10 moved from a mild depression state to the absence of depression; [0135] 4 subjects out of 10 remained in the same range of mild depression, even with a reduction of 6/4 points within the range;

[0136] As far as the PHQ-9 questionnaire is concerned, for the 90% of the subjects (9 out of 10) there was a decrease of the depressive state level. Specifically: [0137] 4 subjects out of 10 moved from a subthreshold depression state to the absence of depression; [0138] 4 subjects out of 10 moved from a mild depression state to a subthreshold depression state; [0139] 1 subject out of 10 moved from having a mild depression to the absence of depression; [0140] 1 subject out of 10 remained in the same range of “subthreshold depression”, even with a reduction of 1 point withing the range.

[0141] With regard to tolerability and compliance, limited to the subjects enrolled in the pilot study, there were no side effects, the food supplement was well tolerated and no subject introduced changes of dosage.

Conclusions

[0142] The results of the pilot study demonstrate how the composition of the invention based on S-adenosyl methionine (SAMe), Lactobacillus helveticus and Bifidobacterium longum is effective in reducing the depressive state or in restoring the normal mood tone, with optimal tolerability by the subjects who took the product.