WO2003053179A1

WO2003053179A1 - Cigarette filters containing lipophile flavonoids and/or tocopherols and tocotrienols

Info

Publication number: WO2003053179A1
Application number: PCT/EP2002/011235
Authority: WO
Inventors: Pierantonio Bimbato; Adriano Tamanini
Original assignee: Filtrona Italia S.P.A.
Priority date: 2001-12-21
Filing date: 2002-10-08
Publication date: 2003-07-03
Also published as: AU2002350500A1; ITMI20012756A1

Abstract

Cigarette filters comprising antioxidant substances such as lipophile flavonoids and/or tocopherols and tocotrienols supported on an adsorbent material in which said antioxidant substances are obtained by means of extraction using the supercritical CO2 technique and added to the adsorbent material from CO2 in liquid state, thus avoiding the presence of traces of solvents in the filters.

Description

CIGARETTE FILTERS CONTAINING LIPOPHILE FLAVONOIDS AND/OR TOCOPHEROLS AND TOCOTRIENOLS FIELD OF THE INVENTION

The present invention relates to cigarette filters containing antioxidant substances. PRIOR ART

It is know that cigarette smoke has noxious effects on health. In particular, is has been widely proved in the last decades that smoking is implicated in the development of cardiovascular and pulmonary pathologies and various types of tumour, especially of the mouth, pharynx, oesophagus and lungs. Cigarette smoke is composed of two phases: a solid phase (tar) and a gas phase. Most of the noxious effects of smoke are linked to the presence in these two phases of highly reactive oxidizing species which, interacting with membrane lipids, proteins, cellular cytoskeletal structures and DNA, have a negative effect on cellular function. The solid phase contains stable oxidizing species, of which the semiquinones, in equilibrium with quinones and hydroquinones, are particularly important; these are capable of reducing oxygen to superoxide which dismutates to form highly toxic H₂0₂. The gas phase instead contains high concentrations of very reactive free radicals, with a half-life of about 2-3 seconds. Notwithstanding the short half-life, the concentration of free radicals in the gas phase is maintained at high levels for around 10 minutes, as they are continuously formed and consequently reach a steady state situation. The most important free radical contained in the gas phase of smoke is nitrogen oxide (NO). This radical is able to form complexes with metal protein, such as hemoglobulin, and participates in a series of reactions which lead to the formation of nitrogen dioxide and peroxide and alcoxy radicals, all highly toxic. In order to decrease the toxicity of smoke, cigarettes are provided with filters with the function of reducing as much as possible the toxic compounds contained therein.

Particularly efficacious filters are perforated filters, which allow the smoke to be diluted with air up to 50%, and composite filters. The latter are composed of two filtering portions, preferably in cellulose acetate, delimiting a central cavity which contains absorbent material such as activated charcoal and/or molecular sieves. In recent years numerous attempts have been made with the aim of eliminating specific toxic components, considered noxious for the health of the smoker, from smoke.

For example, patent application WO 96/00019 describes a filter capable of withholding the nitrogen oxide, carbon monoxide, free radicals, H₂0₂₎ aldehydes and nitrosocompounds from cigarette smoke. The aforesaid filter comprises biological substances containing iron, copper and/or magnesium complexed with a prophyrin ring and/or iron bound stereospecifically to protein molecules.

Patent application US 5,829,449 describes the introduction into the filter, the tobacco or the wrapping of a cigarette of a compound comprising L-glutathione and selenium with the aim of reducing the oxidizing species contained in cigarette smoke and at the same time protect the tissues from oxidation through in situ action.

The disadvantages and limitations of the patents and known techniques above all relate to the problem that, in order for the filter media to absorb the antioxidant substances, organic solvents are used. As a result toxic residues of organic solvents remain in the cigarette filters. In addition to toxicity caused by the presence of organic solvents, absorption may not be homogeneous as not all the granules of the media can receive the same quantity of extract. SUMMARY OF THE INVENTION It has now surprisingly been found that when lipophile flavonoids and/or tocopherols and tocotrienols are added to traditional cigarette filters, there is a considerable increase in the capacity of said filters to decrease the concentration of oxidizing species in smoke.

Filters in which said substances are extracted from relative vegetables or concentrates by C0₂ in supercritical state and incorporated in the support material of the filter by C0₂ in liquid state have proved to be particularly advantageous.

Therefore, the present invention relates to cigarette filters comprising an adsorbent material, characterized in that they comprise antioxidant substances selected from lipophile flavonoids and/or tocopherols and tocotrienols supported on said adsorbent material, in which said antioxidant substances are obtained by means of extraction with the supercritical C0₂ technique and added to said absorbent material by C0₂ in liquid state. In addition to a high level of efficacy, the filters of the present invention have the great advantage of being totally free from organic solvents. DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to cigarette filters comprising antioxidant substances such as lipophile flavonoids and/or tocopherols and tocotrienols. The basic structure of the filters of the present invention is preferably the same as known composite filters, which may be of two types: a) Triple solid filters composed of: cellulose acetate/activated charcoal absorbed on cellulose acetate/cellulose acetate or cellulose acetate/activated charcoal absorbed on paper /cellulose acetate b) Triple cavity filters, in which two filtering segments are separated by a segment filled with activated charcoal.

The invention consists in the fact that the activated charcoal is replaced by an adsorbent material containing in adsorbed form lipophile flavonoids and/or tocopherols and tocotrienols obtained by extraction with the supercritical C0₂ technique and added to said adsorbent material by C0₂ in liquid state and therefore without using organic solvents. At the end of the process the unit is decompressed and the adsorbent material charged with the adsorbed antioxidant substances is recovered from the vessel of the separator. The cigarette filters of the present invention contain said adsorbent material in a quantity ranging from 18 to 78% and said antioxidant substances in a quantity ranging from 45 to 17,300 μg and, preferably, from 225 to 4,800 μg for each filter. According to a preferred application, the aforesaid antioxidant substances are of vegetable origin, although they may also be of synthetic origin. Particularly preferred for the purposes of the present invention are lipophile flavonoids and/or tocopherols and tocotrienols extracted from Gingko biloba, Orthosiphon spicatus, Scutellaria baicalensis and barley bran. The aforesaid antioxidant substances are obtained by means of extraction with C0₂ in supercritical state from vegetable products or from raw extracts. This type of extraction has the advantage of being performed at a temperature close to ambient temperature, in particular at a temperature ranging from 35 to 45°C, and therefore the properties of the extracted substances remain intact. The lipophile flavonoids and/or tocopherols and tocotrienols extracted are then adsorbed on an adsorbent material.

The adsorbent material is selected from the group comprising activated charcoal, sepiolite and molecular sieves.

Preferably, the extraction of antioxidant substances and their adsorption are effected through a process characterized by the following stages: a) the vegetable material comprising lipophile flavonoids and/or tocopherols and tocotrienols or a raw extract of this mixed in advance with supporting material is placed in the extraction vessel of a unit for treatment with C0₂ in supercritical state; b) the adsorbent material is placed in the separation vessel of the same unit; c) extraction in a current of CO₂ in supercritical state is performed; d) the extracted substances are adsorbed on the adsorbent material by C0₂ in liquid state; e) the unit is decompressed to obtain total elimination of the C0₂. The unit utilized in the aforesaid process comprises an extractor and a separator in stainless steel, cylindrical in shape and equipped with an external heat regulation sleeve wound by insulating material. The heating circuits for the extractor and separator are separate.

A suitable quantity of material to be subjected to extraction is placed in the extraction vessel inserted inside the extractor. This is a cylindrical stainless steel container with a capacity of 200 ml, closed at the ends by two porous septa which prevent discharge of fine particles or other impurities which could contaminate the extract or the unit itself.

The adsorbent material, closed in a polypropylene net, is placed in the separation vessel with a capacity of 350 ml, inside the separator. A quantity of adsorbent material ranging from 15 g to 25 g is normally utilized.

The temperature and pressure conditions at which extraction is to be performed are set and the unit is pressurized.

A condenser liquefies the C0₂, which is compressed by a pump and heated until reaching the temperature and pressure conditions required and thus reaches the extractor in supercritical state, at a temperature ranging from 35 to 45°C and a pressure ranging from 71 to 400 bar.

In the extractor the supercritical fluid flows through the material extracting the aforesaid antioxidant substances. The mixture obtained then passes to the separator where the decrease in pressure to 50 bar and temperature to 25°C causes the dissolution capacity of the C0₂ to decrease and the extracted substances are adsorbed on the adsorbent material. The temperature of the separator is regulated from the outside by the operator, usually at 25°C, acting so that the liquid C0₂ is at a level that covers the adsorbent material. It is important that the adsorbent material is always covered by liquid C0₂ and a constant flow of liquid C0₂ is maintained. This can be done by regulating the temperature of the separator appropriately, or by eliminating small quantities of C0₂ through a specific valve. The level of the liquid can be viewed through two diametrally opposite inspection windows, located halfway up the separator. Extraction is carried out for a time preferably ranging from 30 to 60 minutes and absorption for a time ranging from 30 to 60 minutes. Extraction and adsorption are preferably performed in parallel, even if, in this case, not all the material extracted is adsorbed. At the end of this period the unit is decompressed and the adsorbent material charged with the adsorbed antioxidant substances is recovered from the separator vessel. At the end of the process the recovered material is utilized to prepare the filters of the present invention using techniques known in the art.

The use of this process is particularly advantageous as it is possible to work at temperatures close to ambient temperature, thus maintaining intact the antioxidant properties of the active substances. Moreover, this process avoids the use of organic solvents and the consequent need to purify the final product and also prevents traces of organic solvents in filters.

As shall be described in detail in the examples below, which should be considered as non-limiting examples of the present invention, the filters of the present invention are capable of considerably reducing the concentration of free radicals in cigarette smoke. EXAMPLE 1

Preparing filters comprising lipophile flavonoids from Gingko biloba a) 5 g of unrefined methanol extract of Gingko biloba with a lipophile flavonoid content of 148 mg, obtained through extraction of the dried and chopped leaves at ambient temperature, is mixed thoroughly with 50 g of sepiolite in powder form acting as a support.

The mixture obtained is placed in the extraction vessel of the supercritical C0₂ unit, purchased from the Swiss Company Sieber Sitec Eng.A.G., model

01/200, while 20 g of sepiolite is placed in the separation vessel of the same unit to act as adsorbent material. Extraction in supercritical C0₂ is performed at a temperature of 40°C and a pressure of 180 bar for 30 minutes.

The C0₂ with the extracted substances is transferred to the separation vessel and maintained at a pressure of 50 bar and a temperature of 25°C for 30 minutes.

The sepiolite charged with the flavonoids is recovered from the extraction vessel.

The quantity of flavonoids absorbed on the 20 g of sepiolite proved to be 148 mg.

The sepiolite charged with flavonoids is employed to prepare triple cavity filters, each of which is filled with about 0.1 g of sepiolite charged with flavonoids, in place of the activated charcoal. b) The preparation described above was repeated with an extraction pressure of 230 bar. c) The preparation described above was repeated employing activated charcoal as adsorbent material in place of the sepiolite, with an extraction pressure of 230 bar.

The efficiency of the filters was analysed using the following methods: 1 ) Spectrophotometric methods (Methods in Enzymology, 1994, Vol 234, p. 279). 2) Method using EPR spectroscopy (Electron Paramagnetic Resonance). The EPR spectra were recorded with a Varian spectrometer model E-109 (E-Line Century series EPR spectrometer system). The method employed uses the DPPH radical:

1a. R = H - b. R = (CH₃)3CCH2C(CH₃)2- The DPPH radical is very steady in time and is used as a standard in EPR spectroscopy. It presents a signal in 5-fold solution, due to the two non-equivalent nitrogen atoms. The calibration line of the DPPH in toluene was predetermined, with a concentration interval between 1.0 x 10 ^~5 and 2.0 x 10 ^"^ M. The smoke from 5 cigarettes, each with a filter prepared as described above, was made to bubble directly into a solution of DPPH, at known concentration. The decrease in the EPR signal of the DPPH is then assessed after smoking obtained by comparing the filters containing activated charcoal as is (white) with those with activated charcoal treated with Gingko biloba (example 1 ) or with Orthosiphon spicatus (example 2). These operating modes are based on methods indicated in literature such as: Nature 1958, n. 4617, pages 1199-1200 Biochimica et Biophysica Acta 1427, 13-23 (1999) Biochimica et Biophysica Acta 1472, 643-650 (1999) Anticancer Research 19: 5457-5462 (1999) The results obtained with method 1 ) are shown in Table 1. Table 1

Gingko biloba Activated 230 30 40 8.00 - 9.10 charcoal

The last sample in table 1 was also tested with method 2). The reduction in free radicals detected with EPR spectroscopy was 7.4%.

EXAMPLE 2

Preparing filters comprising lipophile flavonoids from Orthosiphon spicatus a) 30 g of dried and chopped Orthosiphon spicatus leaves are placed in the extraction vessel of the unit of example 1.

20 g of activated charcoal are placed in the separation vessel.

Extraction is performed at 40°C and 100 bar for 30 minutes.

Adsorption of the extracted flavonoids on the activated charcoal is performed as in example 1.

At the end of the process the adsorbent material charged with the 45 mg of lipophile flavonoids is recovered from the separation vessel and then used to prepare triple cavity filters employing about 0.1 g of the material obtained for each filter. b) The example 2a) was repeated using 20 g of sepiolite as adsorbent material. c) The example 2b) was repeated utilising an extraction time of 60 minutes. The results obtained with method 1) are shown in Table 2.

Table 2

The last samp e in table 2 was also tested with method 2). The reduction in free radicals detected with EPR spectroscopy was 13.6%. EXAMPLE 3 Preparing filters comprising lipophile flavonoids from Scutellaria baicalensis a) 40 g of Scutellaria baicalensis roots are placed in the extraction vessel of the unit of example 1.

20 g of activated charcoal are placed in the separation vessel.

Extraction is performed at 40°C and 200 bar for 30 minutes.

Adsorption of the flavonoids extracted on the activated carbon is performed as in example 1.

At the end of the process the adsorbent material charged with 108 mg of lipophile flavonoids is recovered from the separation vessel and then utilized to prepare triple cavity filters employing about 0.1 g of the material obtained for each filter. b) Example 3a) is repeated utilizing as the material to extract 40 g of doped Scutellaria baicalensis roots, that is conditioned for 24 hours in an environment saturated with vapours from a methanol-water solution. c) Example 3a) is repeated utilizing as absorbent material 20 g of sepiolite. d) Example 3b) is repeated utilizing as absorbent material 20 g of sepiolite. The efficiency of the filters is assessed as described in example 1.

The results obtained with method 1) are shown in the table 3. Table 3

EXAMPLE 4 Preparing filters comprising tocopherols and tocotrienols from bran a) 50 g of barley bran, obtained during pearling selecting the central fraction with the most tocopherols and tocotrienols, is loaded in the extraction vessel of the unit of example 1.

20 g of activated charcoal is placed in the separation vessel.

Extraction is performed at 40°C and 140 bar for 30 minutes.

Adsorption is performed as in example 1.

At the end of the process the adsorbent material charged with 960 g of tocopherols and tocotrienols is recovered from the separation vessel and then utilized to prepare triple cavity filters employing about 0.1 g of the material obtained for each filter. b) The example 4a) is repeated utilizing as adsorbent material 20 g of sepiolite. The efficiency of the filters is assessed as in example 1.

The results obtained with method 1) are shown in table 4. Table 4

Claims

1. Cigarette filters comprising an adsorbent material, characterized in that they contain antioxidant substances selected from lipophile flavonoids and/or tocopherols and tocotrienols adsorbed on said adsorbent material, in which said antioxidant substances are extracted from the relative vegetable species with the supercritical CO₂ technique and then adsorbed on said adsorbent material by C0₂ in liquid state.

2. Filters as claimed in claim 1 characterized in that said adsorbent material is selected from the group comprising activated charcoal, sepiolite and molecular sieves.

3. Filters as claimed in claim 1 , characterized in that said lipophile flavonoids and/or tocopherols and tocotrienols are contained in quantities ranging from 45 to 17,300 μg for each filter.

4. Filters as claimed in claim 1 , characterized in that said lipophile flavonoids and/or tocopherols and tocotrienols are contained in quantities ranging from

225 to 4,800 μg for each filter.

5. Filters as claimed in claim 1 , characterized in that said lipophile flavonoids are extracted from Gingko biloba, Orthosiphon spicatus or Scutellaria baicalensis.

6. Filters as claimed in claim 1 , characterized in that said tocopherols and tocotrienols are extracted from barley bran, or are of synthetic origin.

7. Filters as claimed in claim 1 , characterized in that extraction and adsorption of said antioxidant substances are performed through the following stages: a) the vegetable material comprising lipophiles flavonoids and/or tocopherols and tocotrienols or an unrefined extract of this previously mixed with a support material in powder form is placed in the extraction vessel of a unit for treating with CO2 in supercritical state; b) the adsorbent material is placed in the separation vessel of the same unit; c) an extraction in C0₂ in supercritical state is performed; d) adsorption by C0₂ in liquid state is performed; e) the unit is decompressed to totally eliminate the C0₂.

8. Filters as claimed in claim 7 characterized in that said extraction is performed at a temperature ranging from 35 to 45°C and a pressure ranging from 71 to 400 bar and in that said absorption is performed at a temperature of 25°C and a pressure of 50 bar.