DESC:Technical field:
The invention relates to stable pharmaceutical compositions of Probiotics and Melatonin for oral administration. More particularly, the invention relates to a pharmaceutical composition of Probiotic and Melatonin for the treatment of Irritable bowel syndrome (IBS).

Background and prior art of the invention:
Irritable bowel syndrome (IBS) is a functional gastrointestinal disorder (FGID). According to Rome III definition, it is characterized by chronic and recurrent abdominal pain/discomfort associated with disturbed defecation and also Gut-Brain interaction disorder associated with altered microbiota.

Functional GI disorders are disorders of gut–brain interaction and classified by GI symptoms related to any combination of motility disturbance, visceral hypersensitivity, altered mucosal and immune function, altered gut microbiota, and altered central nervous system processing.

IBS is conventionally treated with antispasmodics, psychopharmacological treatments, psychotherapy, and newer pro-kinetic drugs with inconsistent results.

The three main subtypes of IBS are IBS-C (predominant constipation), IBS-D (predominant diarrhea), and IBS-M (IBS with mixed bowel habits). People whose symptoms do not fit into any of the above categories are considered to have IBS unclassified.

Over all, IBS is common in the society with worldwide prevalence ranging from 5% to 15%. Clinically, IBS is not only confined to the colon but may also extend to other organs and systems since IBS individuals usually have multiple comorbidities such as dyspepsia, gastro-esophageal reflux disease, interstitial cystitis, fibromyalgia, chronic fatigue, insomnia, headache/migraine and psychiatric disturbances.
IBS is believed to be a biopsychosocial dysfunction since it manifested through biological factors like genetics, immune, gut mirobiota, motility, brain gut interaction, psychological factor like mood, anxiety, depression, somatisation and social factor like family background, illness behavours. This updated pathogeneses and managements of IBS need multi-dimensional looking and multidisciplinary approaches.

Complementary and alternative medicine (CAM) is the term for medical products and practice that are not belonging to current conventional medicine with therapeutic effect determined. The gut microbiota and microbial metabolites are important for maintaining healthy bowels. The gut microbiota can influence the scope and quality of the immune system response; in turn, the immune system participates in regulating the localization and composition of the gut microbiota.

A series of immune diseases, such as paediatric spondyloarthritis (SpA), IBS, IBD, and colon and GI cancer are associated with the gut microbiota. Contribution to intestinal immune function by Tryptan (Trp) metabolites from resident microbiota should not be ignored.

Recent studies have emphasized the profound effects of diet and nutrients on the localization and composition of the gut microbiota as well as on the connection between the gut microbiota and immunological pathways.

Specific gut bacteria determine the availability of Trptophan to the host and then regulate serotonin and subsequent melatonin synthesis. Moreover, melatonin can alleviate the increase in gut permeability and immune activation induced by Escherichia coli.

Melatonin acts as a powerful anti-inflammatory molecule, and the role of melatonin in the gut, especially its role in gut permeability, has recently been established. Melatonin release in the gut is 400-fold higher than that in the pineal gland, with a peak after food intake. Melatonin has positive impact on gut disorders, including inflammatory bowel disease (IBD) and GI cancer.

Melatonin is a significant positive regulator. Melatonin can reduce the incidence of sepsis by decreasing the activation of NLRP3 which is inflammasomes, important effectors of gut permeability and interactions with gut bacteria.
Several potential mediators that connect melatonin, gut bacteria, and gut immunity have been discovered.
Ingested Trptophane is converted to serotonin (5-HT) and melatonin via the serotonin pathway. Approximately 90% of total serotonin in humans is located in enterochromaffin cells in the GI tract, where it can promote intestinal peristalsis. Melatonin (N- acetyl-5-methoxytryptamine) is derived from 5-HT via two-step enzymatic conversion reactions (acetylation and methylation) mainly in the pineal gland but also in other tissues such as the retina, GI tract, skin, and leukocytes. Melatonin play important roles in regulating gut immune homeostasis in mammals.

The results of this meta-analysis of studies showed that sleep disorders/disturbance were common in IBS and the prevalence rate was 37.6%.

The gut–brain axis plays an important role in the pathogenesis of IBS. Central nervous system (CNS), autonomic nervous system (ANS), enteric nervous system (ENS), and hypothalamic pituitary adrenal (HPA) axis are thought to be involved. The symptoms of IBS, such as abdominal pain, may activate sympathetic nervous system and reduce sleep efficiency. The impact of altered microbiota on the development of IBS has been considered more seriously. Similar mechanism may exist between sleep and gut microbiota. Gut microbes may affect sleep status via degradation products such as muramyl peptides (MPs), lipopolysaccharide (LPS), and melatonin. These degradation products could activate immune cells leading to the release of cytokines, which could affect sleep.
Overall, although the reason for sleep disorders seen commonly among IBS patients is obscure, gut–brain–microbiota axis disorder may underlie this association.

The prevalence of sleep disorders among IBS varied slightly in different geographical locations. We also found that the prevalence of sleep disorders among female IBS patients was higher than male IBS patients.

Research studies reveal the association between melatonin metabolite (6-HMS) urinary excretion and the symptoms intensity in IBS. It indicates the significant role of Melatonin deficiency in the complex pathogenesis of IBS and justifies usefulness of its therapeutic administration in this disease.

Probiotics are immune boosting and health enhancing live microbial supplements and hence the administration of the same improves gut microbial balance. Probiotic cocktail had potent anti-inflammatory properties of suppressing mucosal inflammation and restoring cytokine balance. Multi species preparation are probable the best to treat IBS.

There is ample prior art available on individual use of melatonin and probiotics for the treatment of variety of diseases and also prior arts regarding use of the combination or probiotics optionally with melatonin in addition with other active agents as a topical gel and oral dosage forms for variety of indications as discussed below.

WO2017208172 discloses a composition in the form of a gel for topical use comprising an effective amount of an active substance selected from the group comprising or, alternatively, consisting of:
(i) at least one microorganism selected from the group comprising or, alternatively, consisting of: live lactic bacteria; live bifidobacteria, biologically active bacteria or active bacterial components, extracts or cell components, bacterial enzymes, tyndallized bacteria, lysed bacteria, sonicated bacteria and peptidoglycans; and
(ii) melatonin and/or the natural and/or synthetic derivatives thereof and topical carrier.

The micro-organism species disclosed are Lactobacillus acidophilus, Lactobacillus rhamnosus, Bifidobacterium breve, and Streptococcus thermophbus. The composition is for topical transdermal use in the preventive and/or curative treatment of pathologies, disorders or diseases associated with/deriving from alterations of the immune system selected from the group comprising allergies, atopy, allergic rhinitis, food hypersensitivity, dermatitis, atopic dermatitis, eczema, psoriasis, asthma and immunodeficiencies.

WO2018217588 discloses a composition having an application in food and beverage industries comprising a carrier and one or more bioactive agents, wherein said one or more bioactive agents are incorporated into said carrier, wherein said one or more bioactive agents exists in nature in their unincorporated form as a mixture with sugars, and wherein sugars from said mixture are preferentially excluded out of said carrier in said composition.

WO2018169297 discloses compositions comprising tryptophanase-negative microorganisms selected from lactic bacteria, bifidobacteria, streptococci and mixtures thereof, in combination with microorganisms inducing IFN gamma selected from lactic bacteria, bifidobacteria, streptococci and mixtures thereof.

WO2011161501 discloses solid compositions in form of powders or granulates wherein an active ingredient insoluble or poorly soluble in water and/or thermolabile and/or having unpleasant organoleptic properties is dispersed in a lipid matrix containing a triglyceride, a polyoxyethylene sorbitan ester and ascorbyl palmitate. The active ingredient is selected from among polyphenols with molecular weight below 1,000 daltons and insoluble or poorly soluble in water, omega-3 fatty acids, fat-soluble vitamins, amino acids poorly soluble in water, flavonoids, xanthines, boswellic acids, essential oils, capsaicin, melatonin, probiotic yeasts and mixtures thereof. The probiotics present in the composition are Lactobacillus rhamnosus (LGG), Lactobacillus casei (LC), Lactobacillus acidophilus (LA) and mixtures thereof (claim 9).

WO2000078322 discloses combination of lactic acid bacteria which comprises (a) a first component consisting of at least one strain of H202-producing lactic acid bacteria, and (b) a second component consisting of at least one strain of arginine-utilizing lactic acid bacteria. The combination of lactic acid bacteria specifically encompasses Lactobacillus acidophilus, Lactobacillus casei, Lactobacillus plantarum, Bifidobacterium infantis, Bifidobacterium longum, Bifidobacterium breve, Streptococcus thermophilus.

It is evident from the cursory review of the prior arts, as above, the combinations of Prebiotic & Probiotic compositions and optionally with melatonin are known in the literature.

Further, Melatonin with probiotics is also currently available in the market under various brands. Although, these products contain sufficient amounts of melatonin however content of probiotics is very low thereby limiting the efficiency of these marked products is reduced to mere sleeping pills with no other therapeutic efficacy.

In the light of the above, the present invention aims to provide pharmaceutical composition which comprises a combination of melatonin and specific probiotics in effective amounts, for efficient treatment of irritable bowel syndrome.

Summary of the invention:
In line with the above objective, the present invention provides synergistic pharmaceutical composition which comprises a combination of melatonin and specific probiotics in effective amounts, for efficient treatment of irritable bowel syndrome.

Detailed description of the invention:
The invention will now be described in detail in connection with certain preferred and optional embodiments, so that various aspects thereof may be more fully understood and appreciated.

Accordingly, the present invention provides synergistic pharmaceutical composition which comprises a combination of melatonin and specific probiotics in effective amounts, for efficient treatment of irritable bowel syndrome.

In an embodiment, Melatonin is contained in said composition in a concentration range of from about 0.1mg to 10mg.
In a preferred embodiment, the composition contains melatonin in amount of 0.5mg to 5mg.
In a most preferred embodiment, the composition contains melatonin in amount of 3mg.

In an embodiment, the probiotics according to the invention are selected from but not limited to Lactobacillus acidophilus, Lactobacillus rhamnosus, Lactobacillus casei, Lactobacillus bulgaricus, Lactobacillus plantarum, Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, Streptococcus thermophiles.

According to the invention the composition contains Lactobacillus acidophilus in a concentration range of from about 6 million CFU to 6000 million CFU, preferably 100 million CFU to 1000 million CFU, more preferably 700 million CFU.

According to the invention the composition contains Lactobacillus rhamnosus in a concentration range of from about 6 million CFU to 6000 million CFU, preferably 100 million CFU to 1000 million CFU, more preferably 600 million CFU.

According to the invention the composition contains Lactobacillus casei in a concentration range of from about 6 million CFU to 6000 million CFU, preferably 100 million CFU to 1000 million CFU, more preferably 600 million CFU.

According to the invention the composition contains Lactobacillus bulgaricus in a concentration range of from about 6 million CFU to 6000 million CFU, preferably 100 million CFU to 1000 million CFU, more preferably 400 million CFU.

According to the invention the composition contains Lactobacillus plantarum in a concentration range of from about 6 million CFU to 6000 million CFU, preferably 100 million CFU to 1000 million CFU, more preferably 600 million CFU.

According to the invention the composition contains Bifidobacterium longum in a concentration range of from about 6 million CFU to 6000 million CFU, preferably 100 million CFU to 1000 million CFU, more preferably 500 million CFU.

According to the invention the composition contains Bifidobacterium breve in a concentration range of from about 6 million CFU to 6000 million CFU, preferably 100 million CFU to 1000 million CFU, more preferably 600 million CFU.

According to the invention the composition contains Bifidobacterium infantis concentration range of from about 6 million CFU to 6000 million CFU, preferably 100 million CFU to 1000 million CFU, more preferably 500 million CFU.
According to the invention the composition contains Streptococcus thermophilus concentration range of from about 6 million CFU to 6000 million CFU, preferably 100 million CFU to 1000 million CFU, more preferably 600 million CFU.

Accordingly, in an embodiment, the pharmaceutical composition comprises;
a) Melatonin in a concentration range of from about 0.1mg to 10mg,
b) contains Lactobacillus acidophilus in a concentration range of from about 6 million CFU to 6000 million CFU;
c) Lactobacillus rhamnosus in a concentration range of from about 6 million CFU to 6000 million CFU;
d) Lactobacillus casei in a concentration range of from about 6 million CFU to 6000 million CFU;
e) Lactobacillus bulgaricus in a concentration range of from about 6 million CFU to 6000 million CFU;
f) Lactobacillus plantarum in a concentration range of from about 6 million CFU to 6000 million CFU;
g) Bifidobacterium longum in a concentration range of from about 6 million CFU to 6000 million CFU;
h) Bifidobacterium breve in a concentration range of from about 6 million CFU to 6000 million CFU;
i) Bifidobacterium infantis in a concentration range of from about 6 million CFU to 6000 million CFU; and
j) Streptococcus thermophilus in a concentration range of from about 6 million CFU to 6000 million CFU.

Accordingly, in another embodiment, the pharmaceutical composition comprises;
a) Melatonin in a concentration range of from about 0.5mg to 5mg,
b) contains Lactobacillus acidophilus in a concentration range of from about 100 million CFU to 1000 million CFU;
c) Lactobacillus rhamnosus in a concentration range of from about 100 million CFU to 1000 million CFU;
d) Lactobacillus casei in a concentration range of from about 100 million CFU to 1000 million CFU;
e) Lactobacillus bulgaricus in a concentration range of from about 100 million CFU to 1000 million CFU;
f) Lactobacillus plantarum in a concentration range of from about 100 million CFU to 1000 million CFU;
g) Bifidobacterium longum in a concentration range of from about 100 million CFU to 1000 million CFU;
h) Bifidobacterium breve in a concentration range of from about 100 million CFU to 1000 million CFU;
i) Bifidobacterium infantis in a concentration range of from about 100 million CFU to 1000 million CFU; and
j) Streptococcus thermophilus in a concentration range of from about 100 million CFU to 1000 million CFU.

In a preferred embodiment, the pharmaceutical composition comprises;
a) Melatonine in a concentration range of from about 3mg;
b) Lactobacillus acidophilus in a concentration of 700 million CFU;
c) Lactobacillus rhamnosus in a concentration of 600 million CFU;
d) Lactobacillus casei in concentration of 600 million CFU;
e) Lactobacillus bulgaricus in a concentration of 400 million CFU;
f) Lactobacillus plantarum in a concentration of 600 million CFU;
g) Bifidobacterium longum in a concentration of 500 million CFU;
h) Bifidobacterium breve in a concentration of 600 million CFU;
i) Bifidobacterium infantis in a concentration of 500 million CFU; and
j) Streptococcus thermophilus in a concentration of 600 million CFU.

In an additional embodiment, the present invention provides pharmaceutical compositions in the form of powders, pellets, granules or tablets or capsules etc.
In a further embodiment, the invention provides a process for preparation of the pharmaceutical composition of melatonin and probiotics, which process comprises;
a) preparing melatonin tablets by direct compression or wet granulation method through addition of pharmaceutical diluents, binder, lubricant etc. or preparing melatonin granules by wet granulation method;
b) preparing a blend of probiotics consisting of Lactobacillus acidophilus, Lactobacillus rhamnosus, Lactobacillus casei, Lactobacillus bulgaricus, Lactobacillus plantarum, Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, Streptococcus thermophilus, by admixing with microcrystalline cellulose, colloidal silicon dioxide; and
c) filling the contents of step a) and step b) into a capsule or a sachet.

The suitable pharmaceutical diluents(fillers) can be selected from the group consisting of natural diluents such as starches, hydrolyzed starches, partially pregelatinized starches, anhydrous lactose, lactose monohydrate, and sugar alcohols such as sorbitol, xylitol or mannitol.

The suitable pharmaceutical binders include sucrose, gelatin and starch; polymers such as cellulose derivatives and polyvinylpyrrolidone, which have improved adhesive properties; and dry binders such as cross-linked polyvinylpyrrolidone and microcrystalline cellulose.

The suitable pharmaceutical lubricants include talc or silica, magnesium stearate or stearic acid.
The combination of Melatonin along with specific Probiotics as disclosed in the present invention has been proven to relieve abdominal pain, regulate GI motility and correct bowel habits. The positive interaction between Melatonin and probiotics corrects gut microbiota, improves melatonin metabolism.

The pharmaceutical composition of the present invention corrects the Gut brain interaction disorder as it contains Melatonin & Pre-Probiotics and also corrects altered microbiota as it offers Gut specific probiotics.

The synergistic interaction between Melatonin and probiotics in the composition of the present invention as correcting gut microbiota results in improved melatonin metabolism and improved melatonin levels which further leads to increased beneficial gut microbiota.
Melatonin also promotes the sleep and relieves stress associated with abdominal pain.
The present invention is exemplified by the following examples which are provided for illustration only and, should not be construed to limit the scope of the invention.

EXAMPLES:
Example 1
Each Veg. capsule contains:
Melatonin 3mg (as tablet) + Probiotics 5 billion CFU (Lactobacillus acidophilus 700 Million CFU CFU, Lactobacillus rhamnosus 600 Million CFU CFU, Lactobacillus casei 600 Million CFU CFU, Lactobacillus bulgaricus 400 Million CFU CFU, Lactobacillus plantarum 600 Million CFU CFU, Bifidobacterium longum 500 Million CFU CFU, Bifidobacterium breve 500 Million CFU CFU, Bifidobacterium infantis 500 Million CFU CFU, Streptococcus thermophilus 600 Million CFU CFU) along with excipients like diluents, anticaking agents, Prebiotic (FOS-Fructooligosaccharide)

Example:2
Each capsule contains:
Melatonin 3mg (as granules) + Probiotics 5 billion CFU (Lactobacillus acidophilus 700 Million CFU CFU, Lactobacillus rhamnosus 600 Million CFU CFU, Lactobacillus casei 600 Million CFU CFU, Lactobacillus bulgaricus 400 Million CFU CFU, Lactobacillus plantarum 600 Million CFU CFU, Bifidobacterium longum 500 Million CFU CFU, Bifidobacterium breve 500 Million CFU CFU, Bifidobacterium infantis 500 Million CFU CFU, Streptococcus thermophilus 600 Million CFU CFU) along with excipients like diluents, anticaking agents, Prebiotic (FOS-Fructooligosaccharide)

Example:3
Each sachet contains:
Melatonin 3mg (as granules) + Probiotics 5 billion CFU (Lactobacillus acidophilus 700 Million CFU CFU, Lactobacillus rhamnosus 600 Million CFU CFU, Lactobacillus casei 600 Million CFU CFU, Lactobacillus bulgaricus 400 Million CFU CFU, Lactobacillus plantarum 600 Million CFU CFU, Bifidobacterium longum 500 Million CFU CFU, Bifidobacterium breve 500 Million CFU CFU, Bifidobacterium infantis 500 Million CFU CFU, Streptococcus thermophilus 600 Million CFU CFU) along with excipients like diluents, anticaking agents, Prebiotic (FOS-Fructooligosaccharide).

Manufacturing process for examples 1 to 3:
The manufacturing process for preparation of pharmaceutical composition comprising the following steps.
Step 1:
Melatonin tablets were prepared by direct compression or wet granulation method by the addition of suitable pharmaceutical diluents, binders, lubricants etc. As an alternate method, the melatonin granules were prepared by wet granulation.
Step 2:
A blend of probiotics consisting of Lactobacillus acidophilus, Lactobacillus rhamnosus, Lactobacillus casei, Lactobacillus bulgaricus, Lactobacillus plantarum, Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, Streptococcus thermophiles was prepared by admixing the same with microcrystalline cellulose, colloidal silicon dioxide.

Step 3:
The contents of contents of step 1 and step 2 were filled into a capsule or a sachet.

,CLAIMS:1. A pharmaceutical composition comprising combination of melatonin in a concentration range of from about 0.1mg to 10mg and probiotics selected from the group consisting of Lactobacillus acidophilus, Lactobacillus rhamnosus, Lactobacillus casei, Lactobacillus bulgaricus, Lactobacillus plantarum, Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, Streptococcus thermophiles, in effective amounts, for treatment of irritable bowel syndrome.
2. The pharmaceutical composition as claimed in claim 1, wherein, the composition comprises;
a) Melatonin in a concentration range of from about 0.1mg to 10mg,
b) Lactobacillus acidophilus in a concentration range of from about 6 million CFU to 6000 million CFU;
c) Lactobacillus rhamnosus in a concentration range of from about 6 million CFU to 6000 million CFU;
d) Lactobacillus casei in a concentration range of from about 6 million CFU to 6000 million CFU;
e) Lactobacillus bulgaricus in a concentration range of from about 6 million CFU to 6000 million CFU;
f) Lactobacillus plantarum in a concentration range of from about 6 million CFU to 6000 million CFU;
g) Bifidobacterium longum in a concentration range of from about 6 million CFU to 6000 million CFU;
h) Bifidobacterium breve in a concentration range of from about 6 million CFU to 6000 million CFU;
i) Bifidobacterium infantis in a concentration range of from about 6 million CFU to 6000 million CFU; and
j) Streptococcus thermophilus in a concentration range of from about 6 million CFU to 6000 million CFU.
3. The pharmaceutical composition as claimed in claim 1, wherein, the composition comprises;
a) Melatonin in a concentration range of from about 0.5mg to 5mg,
b) Lactobacillus acidophilus in a concentration range of from about 100 million CFU to 1000 million CFU;
c) Lactobacillus rhamnosus in a concentration range of from about 100 million CFU to 1000 million CFU;
d) Lactobacillus casei in a concentration range of from about 100 million CFU to 1000 million CFU;
e) Lactobacillus bulgaricus in a concentration range of from about 100 million CFU to 1000 million CFU;
f) Lactobacillus plantarum in a concentration range of from about 100 million CFU to 1000 million CFU;
g) Bifidobacterium longum in a concentration range of from about 100 million CFU to 1000 million CFU;
h) Bifidobacterium breve in a concentration range of from about 100 million CFU to 1000 million CFU;
i) Bifidobacterium infantis in a concentration range of from about 100 million CFU to 1000 million CFU; and
j) Streptococcus thermophilus in a concentration range of from about 100 million CFU to 1000 million CFU.
4. The pharmaceutical composition as claimed in claim 1, wherein, the composition comprises;
a) Melatonine in a concentration range of from about 3mg;
b) Lactobacillus acidophilus in a concentration of 700 million CFU;
c) Lactobacillus rhamnosus in a concentration of 600 million CFU;
d) Lactobacillus casei in concentration of 600 million CFU;
e) Lactobacillus bulgaricus in a concentration of 400 million CFU;
f) Lactobacillus plantarum in a concentration of 600 million CFU;
g) Bifidobacterium longum in a concentration of 500 million CFU;
h) Bifidobacterium breve in a concentration of 600 million CFU;
i) Bifidobacterium infantis in a concentration of 500 million CFU; and
j) Streptococcus thermophilus in a concentration of 600 million CFU.
5. The pharmaceutical composition as claimed in claim 1, wherein, the composition is provided in the form of powders, pellets, granules or tablets or capsules.
6. The pharmaceutical composition as claimed in claim 1, wherein, the composition corrects the Gut brain interaction disorder and also corrects altered microbiota as it offers Gut specific probiotics.
7. A process for preparation of pharmaceutical composition comprising combination of melatonin and probiotics as claimed in claim 1 comprising the steps of;
a) preparing melatonin tablets by direct compression or wet granulation method by employing pharmaceutical diluents, binder, lubricant etc. or preparing melatonin granules by wet granulation method;
b) preparing a blend of probiotics consisting of Lactobacillus acidophilus, Lactobacillus rhamnosus, Lactobacillus casei, Lactobacillus bulgaricus, Lactobacillus plantarum, Bifidobacterium longum, Bifidobacterium breve, Bifidobacterium infantis, Streptococcus thermophilus, by admixing with microcrystalline cellulose, colloidal silicon dioxide; and
c) filling the contents of step a) and step b) into a capsule or a sachet.

8. The process as claimed in claim 1, wherein, the diluents are selected from the group consisting of natural diluents such as starches, hydrolyzed starches, partially pregelatinized starches, anhydrous lactose, lactose monohydrate, and sugar alcohols such as sorbitol, xylitol or mannitol.
9. The process as claimed in claim 1, wherein, the binders are selected from the group consisting of sucrose, gelatin and starch; polymers such as cellulose derivatives and polyvinylpyrrolidone, which have improved adhesive properties; and dry binders such as cross-linked polyvinylpyrrolidone and microcrystalline cellulose.
10. The process as claimed in claim 1, wherein, the lubricants are selected from the group consisting of talc or silica, magnesium stearate or stearic acid.