Indian Patents. 250137:DIETARY ANTI-BACTERIAL COMPOSITION

Title of Invention	DIETARY ANTI-BACTERIAL COMPOSITION
Abstract	A synergistic dietary antibacterial composition comprising the combination of proteolytic enzymes and one or more antibiotics effective against Helicobacter pylori, useful for the prevention or treatment of gastrointestinal disorders like peptic ulcer, gastric cancers and gastritis in the intestine caused by the Helicobacter pylori is disclosed in the present invention.

Full Text	FORM 2 THE PATENTS ACT 1970 (39 of 1970) & The Patents Rules, 2003 PROVISIONAL SPECIFICATION (See section 10 and rule 13) 1. TITLE OF THE INVENTION: "DIETARY ANTI-BACTERIAL COMPOSITIONS" 2. APPLICANT: (a) NAME: ADVANCED ENZYME TECHNOLOGIES LIMITED (b) NATIONALITY: Indian Company incorporated under the Indian Companies ACT, 1956 (c) ADDRESS: Above Navneet Motors, Gokul Nagar, P.O. Box 182, Thane (W) 400 601, Maharashtra, India 3. PREAMBLE TO THE DESCRIPTION The following specification particularly describes the invention. Technical field of the invention: The present invention relates to dietary anti-bacterial compositions comprising of plant extracts and proteolytic enzymes, for the treatment and prevention of Helicobacter pylori (H. pylori) infections and associated disorders such as peptic ulcers, gastric cancer and chronic active gastritis in intestine. The present invention specifically relates to dietary compositions having synergistic effect when administered along with antibiotic used against H. pylori infections. Background and prior art: Helicobacter pylori is a gram negative spiral rod shaped bacterium having flagella at one end and colonizing in the human gastric mucosa. B.J. Marshall and J. R. Warren in Australia reported in 1983 that this bacterium was frequently detected in stomach biopsy specimen from patients with gastritis or gastric ulcers. At that time this bacterium was named Campylobacter pylori since it resembles Campylobacter in morphology and growth characteristics. Later it was found that the bacterium is different from Campylobacter in the fatty acid composition of its outer membrane and sequence of ribosomes 16S-RNA. Therefore, the bacterium is now referred to as H. pylori and belongs to the newly established genus of Helicobacter. Strains of H. pylori are main cause of gastrointestinal diseases for instance peptic ulcers, gastric cancer and chronic active gastritis in intestine has been a major breakthrough in the gastroenterology. H. pylori strains are sensitive to various antibacterial agents intrinsically sulphonamide trimethoprim, polymixin, nalidixic acid and vancomycin, but in current treatment system two antibiotics amoxicillin and clarithromycin along with the proton inhibitors are given for a week. In other treatment bismuth salt, meteronidazole, amoxicillin, rifamycin and tetracycline are also used. However this treatment may fail for several reasons 2 as such as poor patient compliance, low gastric pH, and high bacterial load in humans and multi drug resistance (MDR) in the H. pylori strains. To overcome these side effect several compositions based on plant extracts having bactericidal actions on H. pylori have also been developed. JP2001122793 discloses a natural product having bactericidal actions on Helicobacter pylori regarded as a causative bacterium of gastric cancer and gastric ulcer without adverse effects, i.e., a substance derived from a stevia and having anti-Helicobacter pylori actions. US 5618564 discloses a method for the treatment of Helicobacter pylori infection wherein a composition containing protease and an antibacterial agent as active ingredients, which can remove Helicobacter pylori from a stomach at high probability without causing side effects or the occurrence of resistant bacteria, and which can treat and prevent peptic ulcer caused by Helicobacter pylori infection and can prevent the recurrence of peptic ulcer is employed. Currently there are no standard sensitivity testing for these fastidious organisms; moreover interpretative criteria for the susceptibility or resistance have yet to be standardized. It is very important however, to assess antimicrobial resistance when therapy failure has occurred. Susceptibility testing is required before administering a second course of treatment. Modern concepts in biotechnology have extended the role of enzyme to new application including bio-therapeutics. Today, enzymes are increasingly being used in cosmetics for the removal of dead cells from skin and in tooth paste owing to their antibacterial and antifungal properties. The use of enzymes such as streptokinase and urokinase for dissolving clots in patients with cardiovascular diseases is well documented and is in the common practice. There is a remote probability in stimulating the evolution of drug resistance development in pathogenic microorganisms when using enzyme based treatment that operate in a manner different from antibiotics. 3 With this background the aim of the present invention is to provide dietary compositions exhibiting inhibitory activity against H. pylori with reduced multiple drug resistance (MDR) as well as drug associated side effects. The present invention also evaluates dietary compositions exhibiting inhibitory antibacterial activity against H. pylori. Object of the invention: The main object of the invention is to provide synergistic dietary anti-bacterial compositions containing plant extracts, proteolytic enzymes and vitamins having synergistic effect. It is yet another object of the present invention to provide dietary anti-bacterial compositions for the treatment and prevention of H. pylori infections and associated disorders such as peptic ulcers, gastric cancer and chronic active gastritis in intestine. It is yet another object of the invention to provide molecular identification of clinical isolates of H. pylori by polymerase chain reaction (PCR). Yet another object of the present invention is to provide compositions of plant extracts and enzymes having synergistic effect when administered along with antibiotic used against H. pylori infections. Further the object of the present invention is to provide lower dosages of antibiotic during the treatment of H. pylori infection for effective synergistic action. Further the object of the present invention is to reduce the risk and rate of drug resistance as well as drug associated side effects in infections caused by the bacterium H. pylori and thus providing improvement in the health of the patient during the chemotherapy of infectious diseases. 4 Summary of the Invention: The present invention discloses dietary anti-bacterial compositions exhibiting a high degree of inhibitory activity against the bacterium H. pylori. The present invention provides synergistic compositions comprising plant extracts and proteolytic enzymes. The anti bacterial compositions of the present invention provide a synergistic action when administered with antibiotics against H. pylori. The present invention provides compositions for the treatment and prevention of pathological conditions associated with H. pylori. The compositions, typically provides treatment against infections and H. pylori related gastrointestinal disorders such as peptic ulcers, gastric cancer and chronic active gastritis in intestine. The compositions of the present invention comprise of one or more enzymes selected from serratiopeptidase, papain, protease, lipase, amylase and one or more plant extracts selected from amla (Emblica officinalis), rutin which is a natural bioflavonoid and pineapple extracts (Ananas comosus). The dietary composition of the present invention further relates to the synergistic effect with other agents having inhibitory activity against H. pylori. Furthermore the composition reduces multiple drug resistance (MDR) as well as drug associated side effects and toxicity in humans and animal thereby providing natural dietary supplements for health benefits. Detailed description of the invention: The present invention describes compositions for the treatment and prevention of pathological conditions associated with H. pylori. The present invention describes synergistic dietary compositions active against bacteria H. pylori and their use in the treatment of gastrointestinal disorders such as peptic ulcers, gastric cancer and chronic active gastritis in intestine. 5 The dietary composition comprising of proteolytic enzymes along with plant extracts act as antibacterial for the treatment of H. pylori related infections having synergistic action with antibiotic exhibiting anti H. pylori activity. The compositions of the present invention comprises of one or more enzymes selected from serratiopeptidase, papain, protease, lipase, amylase and one or more plant extracts selected from amla {Emblica officinalis), rutin which is a natural bioflavonoid and pineapple extracts {Ananas comosus). The dietary compositions exhibit synergistic effect when administered along with antibiotics effective against the bacterium H. pylori. For effective synergistic action lower dosages of antibiotic shall be administered during the treatment of H. pylori infection. The compositions reduce multiple drug resistance (MDR) and drug associated side effects and thereby providing health benefits. Sensitivity profiles of dietary composition against H. pylori along with the clinical isolates are performed by the disc diffusion assays to study antibacterial effect against H. pylori strains. Also studies for synergistic action with antibiotics effective against H. pylori are been conducted. The present invention further deals with molecular identification of clinical isolates of H. pylori by polymerase chain reaction (PCR) and antibacterial activity of individual component comprising the dietary composition. The present invention further deals with sensitivity of standard antibiotics for the treatment of H. pylori related disorders against H. pylori strains viz. wild type and clinical isolates. The present investigation is more specifically explained by following examples. However, it should be understood that the scope of the present invention is not limited by the examples in any manner. It will be appreciated by any person skilled in this art that the present investigation includes the following examples and further can be modified and altered within the technical concept of the present investigation. 6 Example 1: Dietary antibacterial compositions contains proteolytic enzymes selected from a group consisting of serratiopeptidase, papain, protease, lipase, amylase and plant extracts selected from a group consisting of amla, rutin and pineapple extracts at various concentrations. The dietary composition comprises at least one or more enzymes and plant extracts separately and entire composition acts antibacterial against H. pylori and exhibits synergistic effect with antibiotic agents against H. pylori. Example 2: Dietary composition for the prevention of peptic ulcer and gastrointestinal disorder induced by the H. pylori is enclosed: Acid stable protease (Aspergillus niger) usually in the range of about 1-3 mg, Serratiopeptidase in the range of about 0.5-2 mg, Papain in the range of about 0.5-2 mg, Bromelain in the range of about 1-4 mg and Amla in the range of about 3-7 mg. Example 3: Dietary composition showed the antibacterial activity against H. pylori (Wild Type - ATCC-26695) and two clinical isolates (HP-1734 and HP-1735) by disc diffusion assays (in-vitro). The effective concentration of antibacterial dietary composition used is 1-10 mg/ml and it showed the activity of in term zone of growth inhibition in Table 1. 7 Tablel: Antibacterial sensitivity profile of dietary composition against H. pylori along with the clinical isolates by the disc diffusion assays. S.No. Bacteria strains (Helicobacter pylori) Growth of condition at various concentration of Dietary composition 1 mg/ml 5 mg/ml 10 mg/ml 1. Wild Type + ± - 2. Clinical isolates -1734 + ± - 3. Clinical isolates -1735 ± - - + = Resistance, - = Sensitive, ± = Inhibitory Dr. Gopakumar G. Nair Agent for the Applicant Dated this 11th day of July 2006

Full Text

FORM 2
THE PATENTS ACT 1970
(39 of 1970)
&
The Patents Rules, 2003
PROVISIONAL SPECIFICATION
(See section 10 and rule 13)
1. TITLE OF THE INVENTION:
"DIETARY ANTI-BACTERIAL COMPOSITIONS"
2. APPLICANT:
(a) NAME: ADVANCED ENZYME TECHNOLOGIES LIMITED
(b) NATIONALITY: Indian Company incorporated under the Indian
Companies ACT, 1956
(c) ADDRESS: Above Navneet Motors, Gokul Nagar, P.O. Box 182,
Thane (W) 400 601, Maharashtra, India
3. PREAMBLE TO THE DESCRIPTION
The following specification particularly describes the invention.

Technical field of the invention:
The present invention relates to dietary anti-bacterial compositions comprising of plant extracts and proteolytic enzymes, for the treatment and prevention of Helicobacter pylori (H. pylori) infections and associated disorders such as peptic ulcers, gastric cancer and chronic active gastritis in intestine. The present invention specifically relates to dietary compositions having synergistic effect when administered along with antibiotic used against H. pylori infections.
Background and prior art:
Helicobacter pylori is a gram negative spiral rod shaped bacterium having flagella at one end and colonizing in the human gastric mucosa. B.J. Marshall and J. R. Warren in Australia reported in 1983 that this bacterium was frequently detected in stomach biopsy specimen from patients with gastritis or gastric ulcers. At that time this bacterium was named Campylobacter pylori since it resembles Campylobacter in morphology and growth characteristics. Later it was found that the bacterium is different from Campylobacter in the fatty acid composition of its outer membrane and sequence of ribosomes 16S-RNA. Therefore, the bacterium is now referred to as H. pylori and belongs to the newly established genus of Helicobacter.
Strains of H. pylori are main cause of gastrointestinal diseases for instance peptic ulcers, gastric cancer and chronic active gastritis in intestine has been a major breakthrough in the gastroenterology. H. pylori strains are sensitive to various antibacterial agents intrinsically sulphonamide trimethoprim, polymixin, nalidixic acid and vancomycin, but in current treatment system two antibiotics amoxicillin and clarithromycin along with the proton inhibitors are given for a week. In other treatment bismuth salt, meteronidazole, amoxicillin, rifamycin and tetracycline are also used. However this treatment may fail for several reasons
2

as such as poor patient compliance, low gastric pH, and high bacterial load in humans and multi drug resistance (MDR) in the H. pylori strains.
To overcome these side effect several compositions based on plant extracts having bactericidal actions on H. pylori have also been developed.
JP2001122793 discloses a natural product having bactericidal actions on Helicobacter pylori regarded as a causative bacterium of gastric cancer and gastric ulcer without adverse effects, i.e., a substance derived from a stevia and having anti-Helicobacter pylori actions.
US 5618564 discloses a method for the treatment of Helicobacter pylori infection wherein a composition containing protease and an antibacterial agent as active ingredients, which can remove Helicobacter pylori from a stomach at high probability without causing side effects or the occurrence of resistant bacteria, and which can treat and prevent peptic ulcer caused by Helicobacter pylori infection and can prevent the recurrence of peptic ulcer is employed.
Currently there are no standard sensitivity testing for these fastidious organisms; moreover interpretative criteria for the susceptibility or resistance have yet to be standardized. It is very important however, to assess antimicrobial resistance when therapy failure has occurred. Susceptibility testing is required before administering a second course of treatment.
Modern concepts in biotechnology have extended the role of enzyme to new application including bio-therapeutics. Today, enzymes are increasingly being used in cosmetics for the removal of dead cells from skin and in tooth paste owing to their antibacterial and antifungal properties. The use of enzymes such as streptokinase and urokinase for dissolving clots in patients with cardiovascular diseases is well documented and is in the common practice. There is a remote probability in stimulating the evolution of drug resistance development in pathogenic microorganisms when using enzyme based treatment that operate in a manner different from antibiotics.
3

With this background the aim of the present invention is to provide dietary compositions exhibiting inhibitory activity against H. pylori with reduced multiple drug resistance (MDR) as well as drug associated side effects. The present invention also evaluates dietary compositions exhibiting inhibitory antibacterial activity against H. pylori.
Object of the invention:
The main object of the invention is to provide synergistic dietary anti-bacterial compositions containing plant extracts, proteolytic enzymes and vitamins having synergistic effect.
It is yet another object of the present invention to provide dietary anti-bacterial compositions for the treatment and prevention of H. pylori infections and associated disorders such as peptic ulcers, gastric cancer and chronic active gastritis in intestine.
It is yet another object of the invention to provide molecular identification of clinical isolates of H. pylori by polymerase chain reaction (PCR).
Yet another object of the present invention is to provide compositions of plant extracts and enzymes having synergistic effect when administered along with antibiotic used against H. pylori infections.
Further the object of the present invention is to provide lower dosages of antibiotic during the treatment of H. pylori infection for effective synergistic action.
Further the object of the present invention is to reduce the risk and rate of drug resistance as well as drug associated side effects in infections caused by the bacterium H. pylori and thus providing improvement in the health of the patient during the chemotherapy of infectious diseases.
4

Summary of the Invention:
The present invention discloses dietary anti-bacterial compositions exhibiting a high degree of inhibitory activity against the bacterium H. pylori. The present invention provides synergistic compositions comprising plant extracts and proteolytic enzymes. The anti bacterial compositions of the present invention provide a synergistic action when administered with antibiotics against H. pylori. The present invention provides compositions for the treatment and prevention of pathological conditions associated with H. pylori. The compositions, typically provides treatment against infections and H. pylori related gastrointestinal disorders such as peptic ulcers, gastric cancer and chronic active gastritis in intestine.
The compositions of the present invention comprise of one or more enzymes selected from serratiopeptidase, papain, protease, lipase, amylase and one or more plant extracts selected from amla (Emblica officinalis), rutin which is a natural bioflavonoid and pineapple extracts (Ananas comosus). The dietary composition of the present invention further relates to the synergistic effect with other agents having inhibitory activity against H. pylori. Furthermore the composition reduces multiple drug resistance (MDR) as well as drug associated side effects and toxicity in humans and animal thereby providing natural dietary supplements for health benefits.
Detailed description of the invention:
The present invention describes compositions for the treatment and prevention of pathological conditions associated with H. pylori. The present invention describes synergistic dietary compositions active against bacteria H. pylori and their use in the treatment of gastrointestinal disorders such as peptic ulcers, gastric cancer and chronic active gastritis in intestine.
5

The dietary composition comprising of proteolytic enzymes along with plant extracts act as antibacterial for the treatment of H. pylori related infections having synergistic action with antibiotic exhibiting anti H. pylori activity.
The compositions of the present invention comprises of one or more enzymes selected from serratiopeptidase, papain, protease, lipase, amylase and one or more plant extracts selected from amla {Emblica officinalis), rutin which is a natural bioflavonoid and pineapple extracts {Ananas comosus). The dietary compositions exhibit synergistic effect when administered along with antibiotics effective against the bacterium H. pylori. For effective synergistic action lower dosages of antibiotic shall be administered during the treatment of H. pylori infection. The compositions reduce multiple drug resistance (MDR) and drug associated side effects and thereby providing health benefits.
Sensitivity profiles of dietary composition against H. pylori along with the clinical isolates are performed by the disc diffusion assays to study antibacterial effect against H. pylori strains. Also studies for synergistic action with antibiotics effective against H. pylori are been conducted.
The present invention further deals with molecular identification of clinical isolates of H. pylori by polymerase chain reaction (PCR) and antibacterial activity of individual component comprising the dietary composition. The present invention further deals with sensitivity of standard antibiotics for the treatment of H. pylori related disorders against H. pylori strains viz. wild type and clinical isolates.
The present investigation is more specifically explained by following examples. However, it should be understood that the scope of the present invention is not limited by the examples in any manner. It will be appreciated by any person skilled in this art that the present investigation includes the following examples and further can be modified and altered within the technical concept of the present investigation.
6

Example 1:
Dietary antibacterial compositions contains proteolytic enzymes selected from a group consisting of serratiopeptidase, papain, protease, lipase, amylase and plant extracts selected from a group consisting of amla, rutin and pineapple extracts at various concentrations.
The dietary composition comprises at least one or more enzymes and plant extracts separately and entire composition acts antibacterial against H. pylori and exhibits synergistic effect with antibiotic agents against H. pylori.
Example 2:
Dietary composition for the prevention of peptic ulcer and gastrointestinal disorder induced
by the H. pylori is enclosed:
Acid stable protease (Aspergillus niger) usually in the range of about 1-3 mg,
Serratiopeptidase in the range of about 0.5-2 mg,
Papain in the range of about 0.5-2 mg,
Bromelain in the range of about 1-4 mg and
Amla in the range of about 3-7 mg.
Example 3:
Dietary composition showed the antibacterial activity against H. pylori (Wild Type - ATCC-26695) and two clinical isolates (HP-1734 and HP-1735) by disc diffusion assays (in-vitro). The effective concentration of antibacterial dietary composition used is 1-10 mg/ml and it showed the activity of in term zone of growth inhibition in Table 1.
7

Tablel:
Antibacterial sensitivity profile of dietary composition against H. pylori along with the
clinical isolates by the disc diffusion assays.

S.No. Bacteria strains (Helicobacter pylori) Growth of condition at various concentration of Dietary composition
1 mg/ml 5 mg/ml 10 mg/ml
1. Wild Type + ± -
2. Clinical isolates -1734 + ± -
3. Clinical isolates -1735 ± - -
+ = Resistance, - = Sensitive, ± = Inhibitory

Dr. Gopakumar G. Nair Agent for the Applicant
Dated this 11th day of July 2006

DIETARY ANTI-BACTERIAL COMPOSITION

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