FIELD OF INVENTION

The present invention relates to the field of herbal compositions which can be delivered by way of encapsulation. The present invention specifically relates to a novel herbal anti- oxidative composition, which can be delivered by embedding the same into cigarette filter for the purpose to modify smoke or assist in smoke detoxification.

BACKGROUND OF THE INVENTION

Free radicals are reactive oxygen species and reactive nitrogen species that are generated by our body by various factors and one of them being smoking. A balance between free radicals and antioxidants is necessary for proper physiological function. If free radicals ratio is increased in the body to a level where the body is unable to regulate them, this leads to a condition called oxidative stress. Oxidative stress affects various processes and triggers various diseases like heart diseases and cancer. Cigarette smoking is one of the major causes of generation of free radicals in the body and antioxidants play a key role in preventing free radical damage to the body. Cigarette smoke is a complex mixture of gaseous and particulate material. The gaseous portion, comprises of 60% whereas the remaining 40% is the particulate phase. The particulate phase minus the alkaloids and water is called tar. Tar, otherwise known as cigarette smoke condensate (CSC), is a mixture of polycyclic aromatic hydrocarbons (PAHs), N nitrosamines. and aromatic amines. PAHs are the products of incomplete combustion of organic matter such as wood, oil, tobacco, and coal. Tumorigenic activity and also because they are capable of influencing the activity of hepatocellular enzymes.

Antioxidants are molecules which can safely interact with free radicals and terminate the chain reaction before vital molecules are damaged. Antioxidants are able to give an electron to a free radical without becoming destabilizing themselves, thus stopping the free radical chain reaction. Anti-oxidants play a very important role in our body's defense system; they act to neutralize free radicals before they can do their damage. Unfortunately, the antioxidants are depleted in people who smoke, yielding a bad combination of more free radicals with fewer antioxidants to fight them. An external supplementation of antioxidants is an extremely effective to smokers who are at high risk of oxidative stress which further leads to dangerous diseases like heart diseases and cancer.

The prior art has identified several detox compositions for smokers like the patent application W00049901A2 titled “Smokable Product discloses a composition which can be mixed with the tobacco”, where the composition comprises components having an anti-oxidative action and are chosen from the group consisting of ascorbic acid, [beta] -carotene, retinol, tocopherol, its derivatives and precursors, eugenol, jasminaldehyde, cinnamon, cinnamon extract, (crude) coffee, (crude) coffee extract, coffeic acid methyl ester, coffeic acid ethyl ester, ferulic acid methyl ester, ferulic acid ethyl ester, umbelliferone, scopoletin and curcumin, it being possible for the substances to be added to the tobacco either individually or in combination with one another.

Another Patent application US2002136786A titled as “Composition and Method For Smoke Detoxification” discusses a method is provided for effecting smoke detoxification in a human by using a composition that is made of effective amounts of supercritical extract and hydro alcoholic extract of turmeric.

Patent application CN102100410A titled as “Cigarette Filtering Agent For Reducing Tar, Relieving Cough And Protecting Health” discloses a a cigarette filtering agent for reducing tar, relieving cough and protecting health. The cigarette filtering agent comprises the following raw material components in part by weight: 8 to 10 parts of tea, 2 to 3 parts of American ginseng, 8 to 12 parts of spreading hedyotis herb, 8 to 10 parts of turmeric, 8 to 10 parts of Gynostemmapentaphyllum, and 10 to 20 parts of expanded perlite.

Though several of the prior art which discuss the herbal extract compositions for the smokers but none of the prior art discusses a specific herbal or natural extract composition comprising extracts of curcumin and its derivatives, specifically curcumin and tetrahydrocurcumin either individually or in combination as one of its major components. The novel formulation can be delivered to smoker by way of encapsulating the same in a seamless vegetative capsule that could be directly incorporated in to the cigarette filters. The capsule comprising antioxidant composition can be voluntarily ruptured to release for the purpose of modifying smoke, where the composition assist in detoxifying the mainstream smoke, thereby helping the smoker from the harmful effects of tobacco. The composition is specifically intended to provide beneficial effects through its better antioxidant potential to the smokers and it is beneficial in reducing the nicotine induced inflammation, apoptosis, and oxidative stress.

OBJECTIVES OF THE PRESENT INVENTION The primary object of the present invention is to provide with an effective antioxidant smoke detoxification composition comprising extracts of curcumin, tetrahydrocurcumin, individually or as a mixture dissolved in propylene glycol or peppermint oil.

It is another objective of the present invention to provide seamless breakable capsule, comprising inner anti-oxidative composition encapsulated in by outer vegetative capsule, which can be embedded in the cigarette filter for modification or detoxification of the mainstream smoke.

It is yet another objective of the present invention to provide an option to the smoker for accessing the antioxidant composition during the smoke.

It is another objective of the present invention to provide a method of forming a cigarette comprising encapsulated anti-oxidative composition in the filter element to detoxify the mainstream smoke.

In is yet another objective of the invention to provide a cigarette comprising a tobacco rod attached to a filter, the filter with the filter material, along with encapsulated capsule comprising an herbal anti-oxidative composition for modifying characteristics of tobacco smoke during the smoke.

SUMMARY OF THE INVENTION

The present invention discloses a herbal composition for the smoke detoxification, where the composition comprises of extracted herbal ingredients i.e., curcumin and tetrahydrocurcumin either individually or in combination dissolved in propylene glycol or peppermint oil. This highly effective antioxidant natural smoke detoxification formulation can be delivered by encapsulating the same in a seamless vegetative capsule which can be incorporated into the filter elements and assist in providing beneficial effects to the smokers.

Encapsulation is a safe delivery system of an active component. The sealed seamless capsules comprise of outer shell which is vegetative in origin having inner core with anti-oxidative liquid or composition. The capsules can be subjected to external force by the smoker / user in order to release the active material into mainstream smoke passing through the fdter. The discussed vegetative outer shell is made of sodium alginate, agar powder dextrose, gaur gum, glycerin, food color and water, where natural curcumin is used as a colorant. The breakable capsules are designed to rupture in response to pressure applied by the smoker on the outside of the fdter element. Such a Smoking article can be smoked with or without breaking the capsule contained within its fdter element.

DETAILED DESCRIPTION

The present invention relates to a herbal extract composition used for the purpose of smoke detoxification, where the composition comprises extracts of curcumin, and tetrahydrocurcumin, either individually or in combination as major components or one of its major components, where each component has specific activity to reduce the toxic effects of smoking and when combined possess significant antioxidant activity.

The present invention specifically relates to breakable capsules encapsulating Curcumin and /or its derivatives which can be incorporated to filtered smoking articles, such as cigarettes possessing fdter elements which could be ruptured with application of slight pressure to release the herbal component while smoking to reduce smoke detoxification. The breakable capsules can be adapted to rupture in response to pressure applied by the smoker to the outside of the fdter element. Such a Smoking article can be smoked with or without breaking the capsule contained within its fdter element.

The capsule is made of sodium alginate, calcium chloride, sodium chloride, poly ethylene glycol and flavour where natural curcumin is used as a colorant. The breakable capsules can be adapted to rupture in response to pressure applied by the smoker to the outside of the fdter element. Such a Smoking article can be smoked with or without breaking the capsule contained within its fdter element.

The anti-oxidant composition of the present invention mainly comprises of curcumin and tetrahydrocurcumin (whitecurcumin) , where both of them are effective antioxidants

Curcumin - Curcumin through activation of P-CREB/BDNF signaling pathway, confers neuroprotection against Nicotine induced inflammation, apoptosis, and oxidative stress. The protective effect of curcumin against nicotine toxicity may be due to the modulation of expressions of antioxidative system, direct scavenging of reactive oxygen species (ROS), including hydroxyl radicals, nitrogen dioxide radicals and superoxide radicals, reduction of the levels of several markers of oxidative stress and decreased lipid peroxidation induced by nicotine. The pharmacological activity of Curcumin, combined with its ability to suppress CS-induced NF-KB activation and down-regulation of COX-2, MMP-9 and cyclin Dl, provide sufficient rationale for curcumin as a chemo preventive agent in cigarette smokers.

Tetrahydrocurcumin - curcumin and its metabolites tetrahydrocurcumin have significant protective activity on cardiovascular disease and against Cd toxicity which is a non-essential heavy metal with high toxicity potential exposed through cigarette smoking. Many studies have connected Cd toxicity with oxidative stress. Curcumin is a potent scavenger of ROS including 02 ·-, OH· and singlet oxygen. It has been suggested that a diketone group of curcumin can react with OH· and H202 whereas the two phenyl methoxy groups of curcumin suppress NF-KBactivation. Treatment with curcumin, a known activator of Nrf2 -antioxidant response element (ARE) pathway, may suppress oxidant formation by up-regulation of antioxidant enzymes

This highly effective antioxidant natural smoke detoxification formulation can be delivered by encapsulating the same in a seamless vegetative capsule which can be incorporated into the filter elements and assist in providing beneficial effects to the smokers.

The present invention will be further described herein after with reference to a number of non-limiting Examples.

EXTRACTION AND PREPARATION OF EACH COMPONENT.

1. Extraction of Turmeric Extract:

100 gms of turmeric dried rhizomes were pulverized into a coarse powder and extracted with 500ml of 95:5 ratio of ethanol and water (95 % ethanol). This extraction was repeated for 3 times at 60-70 □ with same volume and proportion of solvent. Combined extracts were concentrated to 75 ml at 50-60 □ under the vacuum of 100mm Hg.

The concentrated mass is subjected to crystallization for 160 hrs. at 25±5D temperature. The crystallized crude product was filtered and washed with 10 ml of 95 % ethanol, followed by recry stallization with 95% ethanol at 25 ± 5 □ temperature.

The crystallized pure product is filtered and dried in vacuum oven at 60 □ under vacuum pressure of 100 mm Hg, until the LOD (loss of drying) of the product is NMT ( not more than) 5%.

Yield: 3.5 gms. Of curcumin with 96% purity. Preparation of Tetrahydrocurcumin from extracted curcumin: white curcumin.

100 grams of above obtained curcumin was dissolved in 2 liters of acetone. To this 50 grams of activated raney nickel catalyst is added in a hydrogenator vessel at a temp of 40-50 □ for 3-4 hrs with continuous flow of hydrogen gas with 2kg pressure under continuous stirring. Once the reaction is completed, raney nickel (catalyst) is filtered using by Buchner funnel and the reaction mass was concentrated by using vacuum at 40-50D to get the crude Tetrahydrocurcumin. The crude Tetrahydrocurcumin is further recrystallized in ethyl acetate and acetone in a ratio of 40:60 at room temperature and the crystallized mass is filtered & dried.

Yield: 80.88 gms of TCH was obtained with 99% purity Curcumin/ THC solution in peppermint oil:

2.5%w/v (2.5gmsOfcurcumin/ Tetrahydrocurcumin is dissolved in 100 ml. Of peppermint oil) urcumin/ THC solution in propylene glycol.

The active ingredient curcumin or tetra hydro curcumin concentration in propylene glycol or peppermint oil is 2.5%w/v. i.e., 2.5 gms of curcumin / THC is dissolved in 100 mlof propylene glycol Manufacturing of Seamless capsule a. Shell composition

Mixed the sodium alginate, agar powder, gaur gum and dextrose with 10 gms of water and thenglycerin and food colour is added. The entire mixture is dissolved at a temperature 90 - 100D and checked the viscosity of mixture, which was obtained as 120 mpa. Then the mixture is defoamed and allowed to stand to give a capsule shell liquid.

The capsule was prepared by mixing the above liquid into the seamless capsules manufacturing equipment at controlled temperature 20 ± 2 □ near the multiplex nozzle and maintained capsule shell liquid with the temperature of 70 ± 2 □. The liquid is cooled with cooling oil with about 10 □temperature, then dried the capsule at 25 □ with 50% or less humidity to get the final seamless capsules.

The obtained capsules were translucent and yellowish in color. The capsule was taken and imposed the pressure by fingers to be ruptured easily with a snap, where the cracking sound feeling of the capsule could be felt. b. Core Liquid ( composition ) of the capsule can be i. Tetrahydrocurcumin in Propylene glycol, ii. Curcumin in Propylene glycol, iii. Tetrahydrocurcumin in peppermint oil & iv. Curcumin in peppermint oil.

Curcumin and THC either individually or in combination can be used as the core liquid.

The active ingredient curcumin or tetra hydro curcumin concentration in propylene glycol or peppermint oil is 2.5%w/v. c. The manufacturing of encapsulated capsule.

The manufacturing of the encapsulated capsule with inner core comprising the active ingredient / antioxidant herbal formulated protected by outer vegetative cell is manufactured in a specialized system. Core liquid stored in a vessel is introduced into a seamless capsule manufacturing system, through core liquid inlet pipe and the shell liquid is inserted in to the system by inlet pipe where it covered by the inletconical vessel containing core liquid, the nozzle unit is present at bottom of the conical vessel, where shell liquid covers the core liquid in sphere shape and the pump delivers the both the liquids. These are ejected into cooling liquid forming the seamless mini capsules. The size and weight of the capsules are adjusted by setting the vibrator, which is on top of the encapsulation.

Shell liquid is prepared mixed with water & glycerin heated with stirring under vacuum the outer layer of the stainless steel vessel is steam jacketed the molten mass is kept in steam jacketed storage vessel at constant temperature and pumped into the machine. The encapsulation process begins when the molten mass pumped into machine simultaneously. Shell material enters through inlet and surrounding the conical vessel. Vibrator produces appropriate pressure on both of the material towards nozzle.

Shell material covers the core material in spherical shape at nozzle point horrifies the nozzle and pressure produced by vibrator is adjusted according to required size and weight of the capsules.

Characteristics of the seamless capsule which is further

Size of the capsule: 3 mm Capsule weight: 15 mg Empty shell weight: 3 mg Core weight: 12mg

Insertion of Seamless capsules in Cigarette filters:

Once the seamless capsules are formed they can be inserted into cigarette specifically into the filter component.

The cigarette dimension include: A tobacco rod length is about 57 mm and the filter element length 27 mm. Total length of the cigarette with filter is about 84mm. The tobacco rod includes a charge of tobacco cut filter weighing about 0.65 grams contained in circumscribing cigarette paper wrap of the type Brown Preroll Smoking Papers, GSM: Less than 80.. The tipping material circumscribes the length of filter at about 30mm length of the tobacco rod. The filters are cut into a two up configuration a breakable capsule is manually placed into the opening of the cavity at one end of the each fiber segment. The breakable capsules are seated in desirable location in such a way that with light pressure applied from a coffee stirrer stick and the filters are the attached to the tobacco rods using tipping paper the microcapsules are embedded in the cigarette filter, at an average distance of 16mm in the filterThat is each tobacco rod and each filter element are aligned abutting end to end relationship and secure together using non permeable tipping paper. The tipping paper is adhesively secured to the filter element and the adjacent portion of the tobacco rod.

The present novel composition in cigarette works as an effective anti-inflammatory, anti- apoptotic and reduces the oxidative stress. However the primary intention is to specifically reduce is nicotine induced inflammation, apoptosis, and oxidative stress.

Cigarette smoke (CS) is a major risk factor in cancer development. Among the potential mediators of CS-induced alterations is nuclear factor-kappa B (NF-kB), whose activation has been implicated in chemical carcinogenesis and tumorigenesis. We have demonstrated that CS activates NF-kB in a wide variety of different cell types and that this activation is mediated through induction of IKK, leading to phosphorylation and degradation of IkBa One such agent, curcumin has been shown to be pharmacologically safe, to be involved in the suppression of carcinogenesis and to suppress effects of various inflammatory stimuli.

In the present study it is found that curcumin could block CS-induced NF-kB activation in human non-small cell lung carcinoma cells and whether it abrogates the expression of genes implicated in carcinogenesis. Curcumin, with a purity >95%, dissolved in dimethylsulfoxide (DMSO) as alOOmM stock solution and stored af20C. To determine the effect of curcumin on CS-induced IKK activation, IKK was assayed to examine the effect of curcumin on induction of NF-kB activation. It was identified that curcumin and its derivatives activates NF-kB in a dose-and time-dependent manner in lung cells.

EXPERIMENTATAL STUDIES

1. Antioxidant study: DPPH scavenging activity assay

Plant extract preparation

The curcumin and tetrahydrocurcumin extracted by the above process have been used for the antioxidant scavenging activity study and the biological studies below. Materials and method

Tetrahydrocurcumin obtained in the step-2 was used. The other chemicals used were EDTA, sodium nitropruside, sulphanilamide, potassium superoxide, O-phosphoric acid, napthyl ethylene diaminedihydrochloride, potassium chloride, ferrous sulphate, thiobarbituric acid (TBA), trichloro acetic acid (TCA), and butyrated hydroxyl toluene (BHT).

DPPH radical scavenging assay

To study the radical scavenging activity 1 ml of various concentrations of THC ( i.e., 1, 5 and 50 mM) were Added to 1 ml of 0.1 Mm of DPPH. The reactants were allowed to react in dark and the resultant absorbance was checked at 517nm using spectrophotometry machine. The experiment was performed in triplicates with ascorbic acid as a control

The effect of THC and curcumin on scavenging of the stable free radical of DPPH are mentioned below in Table 1 Table 1 It is observed that curcumin was most active followed by THC. At 10 mM curcumin showed 35.2 % scavenging compared to THC which showed 32.0% whereas at 50 mM curcumin showed 100 % scavenging compared to THC which showed 47.5%.

The DPPH scavenging activity principle is based on the reduction of alcoholic DPPH solution in the presence of a hydrogen-donating antioxidant due to the formation of the non radical 2,2-diphenyl- 1-picrylhydrazine (DPPH-H) . These results indicate that antioxidant activity contained in sample reduced l,l-diphenyl-2-picrylhydrazyl free radical to 1,1,- diphenyl-2-picrylhydrazin. The research showed that DPPH scavenging activities of curcumin and tetrahydrocurcumin, and the IC50values were 2.89mM and 7.85pg/ml, respectively. It showed that curcumin has higher DPPH scavenging activity.

2. Effect of Curcumin on Pulmonary Emphysema caused by Porcine Pancreatic Elastase (PPE) and Cigarette Smoke (CS).

The effects of curcumin and tetrahydrocurcumin against the induction of antioxidant gene expression using primary mouse alveolar macrophages in vitro was checked to ensure the bioactivity of curcumin and continued evaluation for the effects of oral administration of curcumin and tetrahydrocurcumin against the PPE+ and CS+ exposed mouse models.

A stock solution of curcumin and tetrahydrocurcumin of concentration of 3mM was prepared in ethanol and diluted to an appropriate concentration with culture medium. Mouse alveolar macrophages were exposed to curcumin and tetrahydrocurcumin (5pMor 10mM) or culture medium alone for 6h.

After incubation, the cells were harvested for RNA isolation and RNA is evaluated. RNA evaluation data revealed as follows.

Curcumin and THC combotreatment up regulated antioxidant gene expression in lung at 6h after porcine pancreatic elastase ((PPE) treatment. Values expressed for b2 microglobulin and the fold increase against the value of the control group HO-1 mRNA (2.64 fold), GCLC mRNA (1.55 fold) GCLM mRNA (1.60 fold) and GSR mRNA (1.33 fold) whereas for curcumin alone the results were as follows. Values expressed for b2 microglobulin and the fold increase against the value of the control group HO-1 mRNA (1.64 fold), GCLC mRNA (1.25 fold) GCLM mRNA (1.30 fold) and GSR mRNA (1.10 fold) and for tetrahydrocurcumin alone, the results were as follows Values expressed for b2 microglobulin and the fold increase against the value of the control group HO-1 mRNA (1.40 fold), GCLC mRNA (1.35 fold) GCLM mRNA (1.20 fold) and GSR mRNA (1.11 fold) as shown in table 2. Table 2

Curcumin exposure up regulated antioxidant gene expression in mouse alveolar macrophages in vitro PPE induced emphysema mouse model: five units of PPE dissolved in 50 mΐ of saline alone was then injected into trachea of nine-week-old mice.

Curcumin and tetrahydrocurcumin (lOOmg/ Kg) suspended in 200 mΐ of vehicle alone was administrated by oral gavage at 1 h and 24 h before intra-tracheal PPE or saline treatment and daily thereafter throughout a 21 days experimental period.

Cigarette smoke exposure of mice: Nine-week-old mice were exposed to CS generated from commercially available filtered cigarettes. Followed experimental settings we have used: 15 ml of stroke volume and 12 puffs/ min. to generate CS. The CS was diluted with compressed air; we chose a protocol of 60 min 5% CS exposure per day because the mice were able to tolerate it up to 10 days.

Initially we found that both curcumin and tetrahydrocurcumin treatment increased antioxidant gene expression at 6hr after oral PPE administration & it is known that curcumin and tetrahydrocurcumin facilitates the Nrf2 translocation and increased the expression of antioxidant genes (HO-1, GCLC, GCLM, and GSR) in a dose-dependent manner.lt was found that oral administration of curcumin and tetrahydrocurcumin significantly inhibited PPE-induced pulmonary inflammation and emphysema in association with the induction of antioxidant gene expression ( as said values expressed for b2 microglobulin and the fold increase against the value of the control group HO-1 mRNA (2.64 fold), GCLC mRNA (1.55 fold) GCLM mRNA (1.60 fold) and GSRmRNA (1.33 fold)Curcumin and THC combination seems to induce its anti-inflammatory effect through several different mechanisms.

The plasma curcumin concentration in mice was reported to be 60 nM at 30 min and 36 nM at 2h after oral administration of 100 mg/kg curcumin. In humans, it was reported that an oral dose of up to 8g/ day of curcumin for 3 months is safe without over toxicity.

The plasma tetrahydrocurcumin concentration in mice was reported to be 50 nM at 30 min and 32nM at 2h after oral administration of 100 mg/kg tetrahydrocurcumin.

It is observed that NF-E2 related factor 2 (Nrf2), which is a key antioxidant transcriptional factor, decreased in the lungs of patients effected with emphysema & which makes them highly receptacle to oxidative stress and reactive electrophiles. The study concludes that curcumin and tetrahydrocurcuminare potent inducer of Nrf2 related antioxidant enzymes and demonstrated that oral curcumin and tetrahydrocurcumin attenuates PPE and CS induced pulmonary emphysema in mice. Intratracheal PPE administration produced an air space enlargement compared with control group mice and Curcumin, THC and the combination group treatment have inhibited the increase of Lm values upon 21 days treatment.

The above extracted components are combined in a particular dosage of lOOmg / kg and can be delivered in by encapsulating the same in a capsule. The capsules are incorporated in the fdter elements of the cigarettes. By incorporating the active herbal extract material in one or more capsules, in a fdter, the herbal active component is pyrolyzed during the smoking of the cigarette. In addition, by positioning the one or more capsules containing the additive material in the fdter section, the activated carbon can maintain its ability to modify cigarette smoke, which includes removing volatile organic components, such as 1,3 -butadiene, acrolein, isoprene, etc., from mainstream smoke by triggering the anti-inflammatory effect.

A representative fdtered smoking article includes at least one breakable capsule containing natural Curcumin, white curcumin or its derivatives in suitable base incorporated into its fdter element. The seamless breakable capsule can be adapted to rupture in response to pressure applied by the smoker to the outside of the fdter element. Such a Smoking article can be smoked with or without breaking the capsule contained within its fdter elements.

While the foregoing written description of the invention enables one of ordinary skill to make and use what is considered presently to be the best mode thereof those of ordinary skill will understand and appreciate the existence of variations, combinations, and equivalents of the specific embodiment, method, and examples herein. The invention should therefore not be limited by the above described embodiment, method, and examples, but by all embodiments and methods within the scope and spirit of the invention as claimed.