JP5208151B2 - Composition for preventing influenza virus infection comprising nullde extract, air filter comprising the composition, and air purifier comprising the filter - Google Patents

Description

  The present invention relates to a composition for preventing influenza virus infection containing a nullde extract, an air filter containing the composition, and an air cleaner equipped with the filter.

  Influenza usually refers to an infectious disease called flu, and the pathogen, the influenza virus of the Orthomyxoviridae family. Common symptoms are chills, fever, sore throat, myalgia, headache, cough, helplessness and discomfort, with milder symptoms such as fever and cough, often causing fatal complications.

  Influenza continues to spread worldwide, causing thousands to tens of thousands of people to die every season, causing outbreaks, and in the case of global outbreaks for several years, usually about one million people died . In the 20th century, there were three global epidemics of flu caused by a new strain of influenza, killing tens of millions of people. Such varieties often occur when humans are infected from other animals and when a species whose primary host is a human receives a gene from a species whose primary host is another animal. The advent of H5N1 that appeared in Asia in 1990 caused great interest in the global flu epidemic, but the species at this time was not mutated to specialize in human-to-human infection. In April 2009, a variant of H1N1 emerged in Mexico and spread across countries.

  Strictly speaking, influenza is classified into three types, influenza A, influenza B, and influenza C, among five genera included in the Orthomyxoriridae family of RNА viruses.

  Influenza A contains only one species, influenza A virus. Wild aquatic birds act as natural hosts for several influenza A species. Type A causes a pandemic through interspecies transmission and is the most toxic of influenza. Type A is further divided into several serotypes (subtypes) depending on the presence or absence of antibody reaction. Subtypes known to cause flu in humans are arranged in descending order according to the number of deaths at the time of the epidemic. First, H1N1 caused a 1918 Spanish sensation and a 2009 global sensation. H2N2 was the subtype that caused the Asian fluency in 1957 and H3N2 was the Hong Kong fluency in 1968. In addition, there are H5N1, H7N7, H1N2, H9N2, H7N2, H7N3, and H10N7 as subtypes of influenza A.

  Among them, influenza virus type A H1N1 subtype (English: Influenza A virus subtype H1N1) or H1N1 is the most common type of influenza that affects humans as a subtype of influenza A. . In addition to human beings, it can also infect and develop swine and birds.

  H1N1 caused a flu, also called the Spanish flu, and recorded between 50 and 100 million deaths between 1918 and 1919. In 2005, the genome of H1N1 was published in science, and the virus of this genome caused a Spanish sensation in 1918. Compared to the current one, only about 25-30 of about 4400 amino acids were mutated. It was made clear. Low pathogenic H1N1 is now widely distributed in the world, and in 2006 it was reported that human infection accounted for almost half.

  However, the number of deaths due to the H1N1 virus has increased worldwide since March 2009, and this virus was called swine flu at the beginning of the outbreak. The name is abolished, and the Korean speech and government use the name of the new type of flu or the new type of influenza A (H1N1) virus. On June 11, the World Health Organization became a pandemic of H1N1 influenza contagious disease warning levels. I raised it.

  Currently, more than 10,000 people have been infected in Korea based on the announcement on September 16, 2009, 8 are severely hospitalized, and 14 have died from chronic complications or acute complications It is reported.

  On the other hand, influenza B includes one species that is an influenza B virus. Type B is only sensitive to humans and appears rarer than type A. Other animals suspected of being sensitive to type B are oops and ferrets. This species exhibits a mutation rate that is 2-3 times lower than type A, has poor genetic diversity, and there is only one subtype. Because of this low antigenic diversity, type B also has a small host range and causes little epidemic.

  Influenza C also includes one species that is an influenza C virus. Type C is sensitive to humans, dogs and pigs, but usually shows only mild symptoms in children. Appears less frequently than types A and B.

  Conventionally, drugs used for the treatment of influenza include oseltamivir (trade name: Tamiflu), zanamivir (product life: Relenza), peramivir, Amantadine and the like. Oseltamivir, called Tamiflu, is currently mainly used for the treatment of H1N1 influenza A. Tamiflu is the world's only exclusive avian influenza (AI) treatment. It is an antiviral agent that prevents the enzyme function for proliferating the virus and exerts a therapeutic effect, and has a great therapeutic effect if taken within 48 hours after onset. The main therapeutic effects are a decrease in exacerbation of flu symptoms, a decrease in the occurrence of secondary complications such as bronchitis and pneumonia, and a decrease in fluency incubation period. It is also used as a therapeutic agent for influenza A and B. Zanamivir is also called Zanamivir, and the trade name is Relenza. It acts with a neuraminidase inhibitor and is used to treat influenza A and B.

  However, oseltamivir has a side effect that shows a serious feeling of vomiting, and zanamivir has a high antiviral effect, but has a disadvantage that its bioavailability is low and its excretion in the kidney is fast.

  Most anti-influenza therapeutic agents developed to date show side effects. Therefore, the development of an effective anti-influenza composition for the prevention and treatment of influenza is urgent.

  On the other hand, influenza is usually infected by inhalation of virus-containing fumes that are released into the air by coughing or sneezing. In addition, although it can be transmitted from the excrement, saliva, runny nose, stool and blood of other individuals, most of the transmission can be said to be a droplet infection by inhalation of fumes. Such influenza can be inactivated by surfactants such as sunlight, disinfectant, soap and the like to prevent infection, but has a disadvantage that it is not useful for removing viruses contained in the air.

  Therefore, if such a virus in the air can be removed, influenza infection can be effectively prevented.

  The air filter is a breathable member that filters contaminant particles out of the gas that passes through the filter and purifies the gas that has passed through the filter. Such F filters are used in various places such as home appliances, vehicles, buildings, etc., for example, air purifiers, air conditioners, dryers, vacuum cleaners, humidifiers, dehumidifiers, vehicle air conditioners, center in buildings. Used in air conditioning systems.

  On the other hand, an air cleaner is a device that purifies contaminated air and converts it into fresh air, and a large number of filters are used in one air cleaner. Generally, a pretreatment filter that removes dust with large particles, a deodorization filter that removes harmful substances such as odors, volatile organic compounds, and formaldehyde, a HEPA filter that is a high-performance filter that removes fine dust, and a HEPA filter One or two or more intermediate filters that are installed on the front surface and protect the HEPA filter from dust may be used in one air cleaner.

  The more such filters are used, the better the air cleaning performance. However, in order to increase resistance to the entire air circulation, it is common to use 3 to 5 stage filters as a whole.

  Recently, functional filters that can selectively and more elaborately remove substances harmful to the human body using air purifiers or replace the harmful substances with substances beneficial to the human body have been studied. Examples that have been studied include a high-performance fine substance filter that can filter finer substances for infants, or a filter for exclusive use of the new illness group that can filter harmful substances due to the new symptom group.

  Nurde is a deciduous oak tree belonging to the family Anacardiaceae that grows naturally in Asia, and is also called a quintuplet tree. The height reaches 7 m, the leaves are shifted by 7 to 13 leaflets, there are cuts at the edge of the leaf, and there are wings between the leaflets. From August to September, pale yellow or white florets bloom in different sexes, or bloom conically together. In spring, young leaves are eaten half-boiled, but there are toxic ingredients in humans, so be careful. Melaphis chinensis, which parasitizes the mullet, makes a bag-like gallbladder, which is called a quintuplet.

  Here, in view of the above points, the present inventor is searching for an anti-influenza composition. Nurde extract has an excellent anti-influenza activity, and a composition containing this as an active ingredient prevents influenza virus infection. The present invention was completed by finding that the influenza virus in the air can be effectively removed by coating the filter with the composition.

  The objective of this invention is providing the composition for the prevention of influenza virus infection containing a nullde extract.

  Another object of the present invention is to provide an air filter comprising the composition.

  Another object of the present invention is to provide an air purifier including the filter.

  In order to achieve the above-described object, the present invention provides, as one aspect, a composition for preventing influenza virus infection comprising a nullde extract as an active ingredient.

  According to a preferred embodiment of the present invention, the composition may further comprise a ginkgo extract.

  According to a preferred embodiment of the present invention, the composition may be used for quasi-drugs.

  According to yet another aspect of the present invention, an air filter comprising the composition is provided.

  According to still another aspect of the present invention, an air purifier including the air filter is provided.

  The composition containing the nullde extract according to the present invention has excellent anti-influenza activity, and thus is used for the prevention of influenza virus infection, and the filter coated with the composition effectively removes influenza virus in the air. It can be removed and is useful for preventing influenza virus infection.

  Hereinafter, the present invention will be described in detail.

  The present invention provides a composition for preventing influenza virus infection comprising a nullde extract as an active ingredient.

  According to another embodiment of the present invention, the composition may further include a ginkgo biloba extract in addition to the nullde extract.

  In the context of the present invention, the term “prevention” means any action that suppresses influenza virus infection or delays the onset of disease by the use of the composition.

  In the present invention, the influenza virus may be an influenza A virus, an influenza B virus, or an influenza C virus, preferably an H1N1 influenza virus, more preferably an influenza A virus H1N1 variant. It may be a new influenza A (H1N1) virus called a novel flu.

  The H1N1 influenza virus is the influenza virus type A H1N1 subtype (Influenza A virus subtype H1N1), and is the most common type of influenza that affects humans. Infection and disease may occur in pigs and birds unexpectedly, and specific types include A / PR / 8 (H1N1), A / WSN / 33 (H1N1), A / Bervig-Mission / 1 / 18 (rvH1N1), A / Singapore / 6/86 (H1N1), and the like.

  The new influenza A (H1N1) virus is a virus that caused the 2009 influenza pandemic (A / California / O4 / 09, A / California / 7/2009, etc.).

  In the present invention, the influenza virus causes flu, cold, sore throat, bronchitis, and pneumonia, and particularly may cause avian flu, swine flu, or goat flu.

  In the present invention, the types of nulls include, but are not limited to, Rhus javanica and Rhus chinensis.

  In the present invention, as the nullde and ginkgo biloba, commercially available ones may be purchased, or ones collected or cultivated from nature may be used.

  In the present invention, as the site of the nullde, not only the site of leaves, fruits, bark, roots, etc., but also a pentaploid that is a gallbladder formed by the murdephimynensis (Melaphis chinensis) parasitic on the nulle can be used. Uses leaves, bark, or quintuplets, but is not limited to this.

  In the present invention, ginkgo may use parts such as leaves, fruits, bark, and roots, and preferably leaves, but is not limited thereto.

  In the present invention, the nullde or ginkgo biloba extract may be applied by any method that can extract from a simple extraction method to a fat-soluble component in the extraction method, and is pulverized to facilitate the extraction. It is obtained by extraction with an extraction solvent, filtration and concentration.

  Examples of the extraction solvent include water, ethanol, methanol, butanol, n-hexane, n-heptane, and DMSO. These two or more solvents may be mixed and used as an extraction solvent. However, a person skilled in the art can appropriately select from known methods according to the amount of raw material to be extracted, the extraction method, and the like. The extraction time and temperature can also be appropriately selected from known methods by those skilled in the art in consideration of extraction efficiency, extraction solvent, and the like. A preferable extraction time is 10 minutes to 1 hour, and a preferable extraction temperature is 40 to 100 ° C.

  Moreover, after extracting the extract extracted by the said extraction method with well-known filtration methods, such as vacuum filtration, it can concentrate by distillation etc. Such extraction, filtration, and concentration methods are widely known in the art, and thus those skilled in the art can appropriately select them to produce a null deer or ginkgo biloba extract.

  According to another embodiment of the present invention, the composition for preventing influenza virus infection of the present invention can be used as a non-pharmaceutical product.

  That is, the composition of the present invention may be added to quasi-drugs for the purpose of preventing influenza virus infection. When the composition of the present invention is used as a quasi-drug additive, the composition may be added as it is, or may be used together with other quasi-drugs or quasi-drug components, or may be used as appropriate by a conventional method. Good. The mixing amount of the active ingredient may be appropriately determined according to the purpose of use.

  Preferably, the quasi-drug composition may be used in the manufacture of a filter coating agent, hand washing, rinsing, disinfecting and cleaning agent, shower foam, wet tissue, soap, humidifier filler, mask, or fragrance. .

  The present invention also provides an air filter comprising the composition for preventing influenza virus infection.

  In the present invention, the term “air filter” refers to removing microorganisms and dust in the air, preventing inflow of dangerous substances or contaminants such as microorganisms and dust from the outside, and preventing secondary contamination by the filter. Means a filter for. Therefore, the air filter according to the present invention may be used as an air filter for automobiles, an air filter for household appliances, an air filter for an air conditioner, an air filter for a gas mask, an air filter for an air purifier, an air filter for a clean loam, It is preferably used for an air filter for an air purifier.

  Furthermore, this invention provides the manufacturing method of the air filter containing the composition for prevention of the said influenza virus infection, and demonstrates the manufacturing method of the air filter containing the composition for prevention of the influenza virus infection mentioned above below.

The method for producing an air filter containing the composition for preventing influenza virus infection of the present invention,
(A) producing a composition for preventing influenza virus infection;
(B) coating the filter substrate with the composition for preventing influenza virus infection;
(C) drying the coated filter;
In step (b),
After immersing a roller in the composition for preventing influenza virus infection, applying the composition to the filter substrate using the immersed roller;
Immersing a roller containing the filter substrate in the composition for preventing influenza virus infection;
It is performed by any one or more steps selected from the group consisting of spraying the composition for preventing influenza virus infection on the filter substrate.

  As the filter substrate, metal, plastic, non-woven fabric, film or the like may be used. As the air filter for the air purifier, a highly porous nonwoven fabric is preferably used, but is not limited thereto.

  Examples of the plastic material include polypropylene, polyethylene, polyurethane, acrylic, PVC, and polystyrene. However, the plastic material is not limited thereto, and is preferably polypropylene.

  Non-woven fabric means a sheet-like cloth using fibers or the like that are appropriately arranged with fibers or the like, or an adhesive or its own fusing force. Depending on the base material, a paper-based non-woven fabric, a fiber-based non-woven fabric, etc. is there. Nonwoven fabric materials include rayon, tencel, or polypropylene, but are not limited thereto, and any material known in the art can be used as an ordinary nonwoven fabric material. Polypropylene is preferable.

  The filter substrate of the present invention is a weaving form in which a filament yarn produced by melting and spinning a plastic resin at a temperature near the melting point is woven, or a web of fibers (lattice entangled) without weaving the fibers. Can be manufactured into a non-woven fiber filter to which it is bonded, or it can be manufactured into a porous foam filter with the addition of a blowing agent.

  In order to manufacture an air filter, first, a composition for preventing influenza virus infection is manufactured. The composition for preventing influenza virus infection may be preferably dissolved or diluted with a certain solvent and used in the production of an air filter in the form of a solution. Here, as the solvent, water, ethanol, methanol, butanol, or n-hexane, n-heptane, DMSO, or a mixed solvent thereof may be used.

  A filter substrate is coated with the composition for preventing influenza virus infection. The coating method includes a method in which a roller provided with a water-absorbing member such as a sponge is immersed in the composition, and then the roller is rolled on a filter substrate and dried to coat the filter base. There are a method in which a material is directly immersed in the composition and dried to coat, a method in which the composition is sprayed on the filter substrate and dried to coat.

  The coating process for producing the filter of the present invention can be applied to an already produced filter substrate, and can also be applied at the stage of fiber form before weaving the filter. In the case of the fiber form, the filter weaving fiber is first subjected to a coating process in which the composition of the present invention is immersed or sprayed, and after the coating process, the filter substrate is woven.

  When drying the filter coated with the composition of the present invention by dipping or spraying, the filter may be dried at room temperature or hot air, and any drying method known in the art can be used without limitation. By drying, the composition of the present invention is well adsorbed on the filter, and the filter can have antibacterial effect for a long time.

  In addition to the composition of the present invention, the filter of the present invention is an existing antibacterial substance, deodorant (eg, flavonoid, phytoncide, wood vinegar, plant extract, cyclodextrin, metal ion, titanium dioxide) or a dust collecting filter. These components may be separately applied together or separately for each component, and the order of application is not particularly limited.

  The present invention also provides an air purifier including the air filter.

  Referring to an air purifier as an example in which the air filter of the present invention is used, as described above, the air purifier includes a number of filters, but includes a composition for preventing influenza virus infection according to the present invention. The air filter may be used as a functional filter, and can be used as any filter included in other air purifiers.

  The air purifier to which the air filter of the present invention is applied is not limited, and examples thereof include an air purifier in a home or office room, a filter for automobile / indoor purifying, and the like. Ordinary components included in other air purifiers may be included. Preferably, the air purifier of the present invention is formed by disposing the air filter of the present invention between an air inlet and an outlet.

  The present invention also provides an air cleaning method using the air cleaner.

  Hereinafter, the present invention will be described in more detail with reference to examples and experimental examples. However, the following examples are merely illustrative of the present invention, and the content of the present invention is not limited to the following examples.

[Production Example 1: Production of Nurde extract]
The nullde used in this Production Example was used after collecting the leaf site, drying to obtain the extract of the present invention efficiently, and then pulverizing it into a powder. 5 l of water was added to 1 kg of nullde, extracted by boiling in a water bath at 80 ° C. for 12 hours, filtered through filter paper, and concentrated to obtain a nullde extract.

[Production Example 2: Production of Ginkgo biloba extract]
The ginkgo biloba used in this production example was used after collecting the leaf part, and then drying to obtain the extract of the present invention efficiently, and then pulverizing it into a powder. 5 g of water was added to 1 kg of ginkgo biloba, extracted by boiling in a water bath at 80 ° C. for 12 hours, filtered through filter paper, and concentrated to obtain a ginkgo biloba extract.

[Production Example 3: Production of Air Filter According to the Present Invention Coated with Nurde Extract]
A polypropylene filter substrate was spray coated with a solution obtained by diluting the nullde extract produced in Production Example 1 in water, and dried at 140 ° C. for 4 minutes to produce a filter.

[Production Example 4: Production of Air Filter According to the Present Invention Coated with Nurde Extract and Ginkgo Extract]
A polypropylene filter base material is spray-coated with a solution obtained by diluting a mixture of the same amount of the Nurde extract produced in Production Example 1 and the Ginkgo biloba extract produced in Production Example 2 into water, and 140 A filter was produced by drying at 4 ° C. for 4 minutes.

[Experimental Example 1: Measurement of killing activity of influenza virus by Nurde extract and Ginkgo biloba extract]
The influenza virus killing activity of the Nurde extract and Ginkgo biloba extract obtained in Production Examples 1 and 2 of the present invention was measured as follows.

  As an influenza virus, the WHO standard strain influenza virus H1N1 (A / California / O4 /) was introduced as the new influenza A (H1N1) virus declared the 2009 pandemic (World Health Organization, declared on June 11, 2009). 09) and A / PR / 8/34 (H1N1) and A / WSN / 33 (H1N1) were used as other H1N1 type viruses already known as H1N1 types.

  As a sample, the nullde extract obtained in Production Example 1 alone, the ginkgo extract obtained in Production Example 2 alone, and the nullde extract and ginkgo extract obtained in Production Examples 1 and 2 were mixed in the same amount. The resulting mixture was used. A more specific experimental method is as follows.

First, MDCK (Mardine Darvine Canine Kidney) cell line was inoculated at a concentration of 1.5 × 10 ⁶ cells / ml and then in a 6-well plate for 24 hours in an incubator at 37 ° C. and 5% CO ₂ concentration. Cultured. In addition, each single extract and mixture was diluted with water at 100 mg / ml (1 ×), and this was diluted 10 times to prepare a sample. 90 μl of each of the prepared samples was placed in a 96-well plate and treated with 10 μl of influenza virus. After reacting for 10 minutes, the solution was diluted 10-fold with PBS. Thereafter, the MDCK cell line, which became a full dish in the culture dish, was infected with 1 ml of the diluted solution for 1 hour. On the other hand, a control group of “infection + non-administration” (a group of cells to which no extract was administered after infection with influenza virus) was reacted under the same conditions, then inoculated into an MDCK cell line and cultured.

  Thereafter, the medium was removed, 2 × MEM medium containing 2 × agarose and 10 μg / ml trypsin was mixed at a weight ratio of 1: 1, and then 2 ml each was poured onto a plate containing the MDCK cell line. The MDCK cell line was cultured in an incubator at 37 ° C. for 2 days, and then fixed with 1 ml of 4% paraformaldehyde and washed with water to remove agarose. Thereafter, the number of plaques was measured by staining with crystal violet, and the number was calculated in terms of plaque forming units (pfu / ml). The results are shown in Tables 1-3.

Formula 1
Virus inhibition rate (%) = (1−extract treated group / control group) × 100
Table 1
The titer of new influenza A (H1N1) (A / California / O4 / 09) after each sample treatment

Table 2
Influenza A / WSN / 33 (H1N1) titer after each sample treatment

Table 3
Influenza A / PR / 8 (H1N1) titer after each sample treatment

  Through the results of Tables 1 to 3, the virus is completely killed when the Nurde extract and a mixture of Nurde extract and Ginkgo biloba in the same amount are treated as compared with the control group. It can be seen that the Nurde extract according to the invention and the mixture of Nurde extract and Ginkgo biloba extract have excellent anti-influenza virus killing activity.

[Experimental Example 2: Measurement of killing activity of influenza virus of air filter according to the present invention]
The influenza virus killing activity of the air filter treated with the nullde extract obtained in Production Examples 3 and 4 of the present invention and the air filter treated with the nullde extract and the ginkgo biloba extract was measured as follows.

  As an influenza virus, the WHO standard strain influenza virus H1N1 (A / California / O4 /) was introduced as the new influenza A (H1N1) virus declared the 2009 pandemic (World Health Organization, declared on June 11, 2009). 09) and A / PR / 8/34 (H1N1) and A / WSN / 33 (H1N1) were used as other H1N1 type viruses already known as H1N1 types.

  As a sample, the air filter obtained in Production Examples 3 and 4 was used. As a control group, an air filter which was not treated with any extract was used. A more specific experimental method is as follows.

  After collecting the filter medium to 2 × 2 cm, apply a certain amount of virus solution to the filter and absorb and react with the filter medium for 10 minutes, and then put a medium (1 ml) on the absorbed filter. Shake for a minute to wash out the virus. This was diluted to prepare a sample. 90 μl of each of the prepared samples was placed in a 96-well plate and treated with 10 μl of influenza virus. After reacting for 10 minutes, the solution was diluted 10-fold with PBS. Thereafter, the MDCK cell line, which became a full dish in the culture dish, was infected with 1 ml of the diluted solution for 1 hour. On the other hand, a control group of “infection + non-administration” (a cell group to which no extract was administered after infection with influenza virus) was reacted under the same conditions, then inoculated into an MDCK cell line and cultured.

  Thereafter, the medium was removed, 2 × MEM medium containing 2 × agarose and 10 μg / ml trypsin was mixed at a weight ratio of 1: 1, and then 2 ml each was poured onto a plate containing the MDCK cell line. The MDCK cell line was cultured in an incubator at 37 ° C. for 2 days, and then fixed with 1 ml of 4% paraformaldehyde and washed with water to remove agarose. Thereafter, the number of plaques was measured by staining with crystal violet, and the number was calculated in terms of plaque forming units (pfu / ml). The results are shown in Tables 4-6.

Table 4
The titer of new influenza A (H1N1) (A / California / O4 / 09) after each sample treatment

Table 5
Influenza A / WSN / 33 (H1N1) titer after each sample treatment

Table 6
Influenza A / PR / 8 (H1N1) titer after each sample treatment

  From the results in Tables 4 to 6, it can be seen that the nullde extract-treated air filter according to the present invention and the mixture-treated air filter of nullde extract and ginkgo biloba extract have excellent influenza virus killing activity.

Claims (8)

A composition for preventing influenza virus infection, comprising a nullde extract as an active ingredient,
The influenza virus is, Ri H1N1 influenza virus der,
The composition for preventing influenza virus infection, wherein the H1N1 influenza virus is A / California / 04/09 . The influenza virus according to claim 1, wherein the extract is extracted using water, ethanol, methanol, butanol, n-hexane, n-heptane, DMSO, or a mixed solvent thereof. A composition for preventing infection.   The composition for preventing influenza virus infection according to claim 1, wherein the nullde is a leaf, fruit, bark, root, or quintuplet site.   The composition for preventing influenza virus infection according to claim 1, wherein the disease caused by influenza virus infection is flu, cold, sore throat, bronchitis, pneumonia, avian flu, swine flu, or goat flu. object.   The composition for preventing influenza virus infection according to claim 1, wherein the composition is used for manufacturing a quasi-drug. Wherein non-pharmaceutical products, filter coating agent, hand washing, mouth rinse, antiseptic cleaner, and wherein the shower foam, a wet tissue, soap, humidifier fillers, masks or fragrances, claim 5 Composition for preventing influenza virus infection. Air filter comprising a composition for preventing influenza virus infection by claim 1. An air cleaner comprising an air filter according to claim 7 .