FR2811573A1 - Extracts of ginger Zingiber officinale comprise anti-inflammatory, anti-platelet aggregation and antifungal activity - Google Patents

Abstract

Process for the preparation of pharmaceutically-active material from rhizomes of Zingiber officinale (ginger). Process for the preparation of pharmaceutically-active active material from rhizomes of Zingiber officinale (ginger) comprising: (a) preparing a crude liquid from the ginger rhizomes; (b) introducing this into an inverse phase chromatography column and eluting successively with water, a first eluant and a second eluant in which the second eluant has a lower polarity than the first eluant, but is greater than chloroform, (c) evaporating the first eluate, and (d) evaporating the second eluate. The first stage in which the crude liquid is obtained from the rhizomes may be effected in either of two ways. In the first method, the fresh rhizomes are chopped and filtered, the filtrate is extracted with a first organic solvent and evaporated the residue is extracted with a second organic solvent and evaporated, and the two extracts are combined. In the second method of preparing the crude liquid, powdered dried rhizomes are extracted with an organic solvent or with steam and the crude liquid evaporated, or the extraction may be effected with supercritical CO2. Independent claims cover pharmaceutical compositions containing and extract as described.

Description

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PROCESS FOR PREPARING A POWERFUL EXTRACT OF MATERIAL
ACTIVITY ANTI-INFLAMMATORY, ANTI-AGGREGATION
PLAOUETTAIRE AND ANTIFONGIQUE FROM ZINGIBER
OFFICINAL AND PHARMACEUTICAL COMPOSITIONS CONTAINING
LEDIT EXTRACT
The present invention relates to a method for preparing a potent extract for anti-inflammatory, anti-platelet aggregation and antifungal activities from Zingiber officinale.

 Unrefined Chinese medicines or spices, such as Zingiber officinale, Eugenia caryophyllata, Allium sativum, have been used in medicine and to enhance the taste of food. Raw ginger is used as an antiemetic, expectorant, antitussive and accelerator of the digestive system. The semi-dried raw ginger is also used for stomach pains, chest pains, spinal pain, cough, common cold and cure for a form of edema called "water retention". The zingerone is the main component that explains the spicy nature of ginger; gingerol and shogaol are other pungent ingredients in ginger. Gingerol has a cardiotonic action, suppresses the contraction of isolated portal veins in mice, and modulates the contraction of eicosanoid-induced mouse and rat blood vessels. Shogaol has a response that increases blood pressure.

Gingerol and shogaol are mutagenic, while zinger and zingerone have been shown to have anti-mutagenic activity. Shogaol has inhibitory activity on paw edema induced by

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 carrageenan and platelet aggregation [US Patent No. 5,804,603, background of the invention].

 So far, numerous reports have shown that Zingiber officinale exhibits various physiological activities. Typical examples include a cancer metastasis suppressor, described in Japanese Patent Publication No. 7-258104; a synthetic enhancer for neurotropic factor, which is effective for degenerative neurological conditions such as Alzheimer's dementia or Parkinson's disease, described in Japanese Patent Publication No. 7-25777; an anti-rheumatic agent, described in Japanese Patent Publication No. 6,236,653, in U.S. Patent Nos. 5,494,668 and 5,683,698; an antimicrobial composition, described in Japanese Patent Publication No. 6-227931; and an analgesic composition described in Japanese Patent Publication No. 6-107556. Ginger contains 1 to 4% essential oil (oleoresin). Over the past 45 years, numerous chemical studies have been conducted on the constituents of the essential oil. In all, more than 200 different volatile substances have been identified in the essential oil, which is the site of the pharmacological activity. The essential oil contains a mixture of various terpenes as well as some other non-terpene compounds. Although this is essentially speculative, experimental data and observations suggest that ginger inhibits both cyclooxygenase and lipooxygenase products, that is, it may be a dual inhibitor of eicosanoids. In all, 56 patients (28 with rheumatoid arthritis, 18

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 osteoarthritis and muscle discomfort) used ginger powder for their affections. More than three-quarters of patients with arthritis had varying degrees of relief from pain and edema. All patients with muscle discomfort observed pain relief.

None of the patients reported side effects during the ginger consumption period, which ranged from 3 months to 2.5 years. (Srivastava and Mustafa, Medical Assumptions, 1992, 39 342-348).

 Nonsteroidal anti-inflammatory drugs have three major actions, all of which are related to the inhibition of cyclooxygenase leading to a decrease in prostanoid formation. First, an anti-inflammatory action achieved by decreased production of vasodilator prostaglandins (PGE2, PGI2), which means less vasodilatation and, indirectly, less edema. Secondly, an analgesic effect obtained by decreased production of prostaglandins (lower sensitization of nociceptive nerve endings relative to the inflammatory mediators bradykinin and serotonin). Third, an antipyretic effect probably due to a decrease in the PGE2 mediator generated in response to inflammatory pyrogens, much like interleukin 1. Since ginger inhibits the synthesis of prostanoids and also lipoxygenase products, its enhancement effects arthritis and muscle discomfort may be related to a decrease in the formation of prostanoids and leukotrienes. Because of such a possibility, a decrease in the formation of #dema induced by the

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 carrageenin in the paw of a rat after administration of 3 g of ginger extract has been demonstrated and the power of the extract in the acute inflammation test appears to be comparable to that shown by the acetylsalicylic acid mentioned in the same study (Mascolo N. et al., Journal of Ethnopharmacology 1989, 27, 129-140).

 Dermatophytes, particularly Trichophyton rubrum and Trichophyton mentagrophytes, are the common pathogens of onychomycosis and athlete's foot [Roberts DT., British Journal of Dermatology. 141 Suppl.

56: 1-4, Nov. 1999; Roldan YB et al., Mycoses, 43 (5): 181-3,2000]. Pityrosporum ovale (Malassezia furfur) is the etiological agent of pityriasis versicolor, folliculitis Pityrosporum and intertrigo Malassezia. Several studies indicate a strong association between Pityrosporum ovale and seborrheic dermatitis and dandruff, a less severe form of seborrheic dermatitis [Nenoff P. et al., Dermatology.

191 (4): 311-4, 1995; Bulmer AC. et al., Mycopathologia, 147 (2) 63-5, 1999].

 The present invention provides ginger rhizome extracts that demonstrate activity in an in vitro platelet anti-aggregation assay, carrageenan-induced paw edema inhibitory activity, and activity. antifungal against Trichophyton mentagrophytes and Pityrosporum ovale. The extracts are prepared by extraction of ginger rhizomes with an organic solvent (such as ethyl ether, acetone, methanol and ethanol) or with supercritical CO2, or by steam distillation of ginger rhizomes for get a raw liquid, and in

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 subjecting said crude liquid to reverse phase chromatography to obtain extracts containing shogaols, gingerols and / or dehydrogingerdione.

 As presented in the background of the invention, ginger has been used for its anti-inflammatory effect and in the relief of pain.

 The present invention is to provide an effective method of preparing a potent product for anti-inflammatory, platelet anti-aggregation and antifungal activities from ginger rhizomes. The strong product prepared according to the process of the present invention has a substantially constant composition, so that the pharmacological effects thereof are defined.

 The effective method of preparing a potent product for anti-inflammatory, platelet anti-aggregation and antifungal activities from ginger rhizomes in accordance with the present invention comprises the following steps: a) preparing a raw liquid from rhizomes of ginger; b) introducing the crude liquid into a reverse phase chromatography column, and eluting the column with water, with a first eluent and with a second eluent successively, said second eluent having a lower polarity than that of the first eluent but stronger than that of chloroform, so that a first eluate resulting from elution with the first eluent and a second eluate resulting from elution with the second eluent are obtained; c) removing the first eluent from the first eluate by evaporation, so that a first concentrated eluate is

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obtained and can be used as a powerful product; and d) removing the second eluent from the second eluate by evaporation so that a second concentrated eluate is obtained and can be used as a strong product; wherein step a) comprises steps i) to iv), or comprises step I), step I '), or step I "), said steps i) to iv) consisting of: i ) chop fresh rhizomes of ginger and filter the resulting mixture to obtain a filtrate and a residue, ii) extract the filtrate with a first organic solvent, recover the extraction solution with the first organic solvent resulting, and evaporate the first organic solvent of the extraction solution to obtain a first concentrated extraction solution, iii) extracting the residue with a second organic solvent, recovering the extraction solution with the second resulting organic solvent, and evaporating the second organic solvent from the extraction solution to obtain a second concentrated extraction solution; and iv) combining the first concentrated extraction solution and the second concentrated extraction solution to obtain the raw liquid; said step I) consis to:
I) extracting powder of dried ginger rhizomes with the second organic solvent, recovering the extraction solution with the second organic solvent

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resulting and evaporating the second organic solvent from the extraction solution to obtain the crude liquid; said step II) consisting of:
(I) steaming the powder of dried ginger rhizomes and concentrating the resulting distillate by evaporation to obtain the crude liquid; and said step I ") consisting of:
I ") extract dried ginger rhizome powder with supercritical CO2, recover the resulting supercritical CO2 extraction solution and evaporate the CO2 from the extraction solution to obtain the crude liquid.

 The potent product for antiinflammatory, platelet anti-aggregation and antifungal activities prepared according to the process of the present invention preferably comprises 0 to 10 mg of 6-shogaol per gram of the product, 1 to 150 mg of 6gingerol per gram. of the product and from 0 to 40 mg of dehydrogingerdione per gram of the product.

 The present invention also provides a potent pharmaceutical composition for anti-inflammatory, platelet anti-aggregation and antifungal activity comprising a therapeutically effective amount of said crude liquid prepared in step a) of the method of the present invention as a principle. active, in admixture with a pharmaceutically acceptable carrier or diluent, for the active ingredient.

 The present invention also provides a powerful pharmaceutical composition for anti-inflammatory, anti-aggregation activity

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 platelet and antifungal composition comprising a therapeutically effective amount of said product prepared according to the process of the present invention, as active ingredient, in admixture with a pharmaceutically acceptable carrier or diluent, for the active ingredient. Preferably, said product prepared according to the process of the present invention is the first concentrated eluate prepared in step c). Alternatively, said product prepared according to the process of the present invention is the second concentrated eluate prepared in step d).

 Preferably, said first eluent is methanol and said second eluent is acetone.

 Preferably, step a) of the process of the present invention comprises steps i) to iv).

 Preferably, said first organic solvent is ethyl ether.

 Preferably, said second organic solvent is acetone, methanol, ethanol or a combination thereof. More preferably, said second organic solvent is acetone.

 Preferably, step a) of the process of the present invention comprises step I).

 Preferably, step a) of the process of the present invention comprises step I ').

 Preferably, step a) of the process of the present invention comprises step I ").

 A reverse phase chromatography column suitable for use in the process of the present invention includes (but is not limited to) a reverse phase chromatography column packed with porous resin, for example Diaion HP-20 (Mitsubishi

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 Co.), Sephadex LH-20 (Pharmicia Co.) and RP-18 (Nacalai tesque Co.).

 The powerful pharmaceutical composition of the antifungal activity of the present invention is preferably applied topically, for example in the form of a shampoo, a bath gel, a soap, a body lotion. , a body cream and a detergent. Preferably, the strong anti-fungal activity pharmaceutical composition of the present invention is used in the treatment of diseases associated with Trichophyton mentagrophytes or Pityrosporum ovale, including but not limited to athlete's foot (tinea pedis), ringworm microscopic teat (tinea capitis), jock itch (tinea cruris), dermatophytosis of the glabrous skin (tinea glabrosa), onychomycosis, pityriasis capitis simplex, pityriasis versicolor, folliculitis pityrosporum, seborrheic dermatitis and dandruff. In particular, the antifungal pharmaceutical composition of the present invention is in the form of a shampoo for use in the treatment of dandruff.

 It is believed that the above description provides the means to realize the present invention adequately. The following specific examples should therefore be interpreted as being illustrative only, and not limiting in any way as to the rest of the description.

Determination of active ingredients
In the following examples, high performance liquid chromatography (abbreviated as HPLC) was used to determine the active ingredients of

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 products prepared herein. The HPLC spectra were recorded on an HPLC instrument (Shimadzu LC-10AT HPLC, Japan) using a Cosmosil 5C-18 column (250 mm x 4.6 mm, filled with 5 m diameter particles) by a DSC process. elution. A sample of HPLC was prepared by diluting a suitable amount of a product with a mobile phase solution (hydrogen cyanide: water = 65:35, v / v) to 25 ml, and followed by filtration on a membrane of 0.25 lira. The filtrate was introduced into the HPLC column and eluted with the mobile phase solution. A UV detector (Shimadzu SPD-6AV, Japan) was used to detect the absorption of the eluate at 230 nm.

Example 1
2100 g of fresh ginger rhizomes were chopped and filtered to obtain a filtrate and a residue. 500 ml of the filtrate were extracted with 500 ml of ethyl ether three times, the layers of organic phase were separated from the aqueous phase layers, before combination.

The ethyl ether of the combined extraction solution was evaporated in vacuo to give a concentrated ethyl ether (I-OE) extract. The ginger residue was extracted with 3000 ml of acetone three times, the extraction solutions were recovered by filtration and combined. Acetone was evaporated from the combined extraction solution in vacuo to give a concentrated acetone (I-O) extract (14.5 g). In a 300 mm x 30 mm reverse phase chromatography column packed with 180 g of Diaion HP-20 resin having a diameter of 500 m to 800 m, 7 g of a mixture of the concentrated ethyl ether extraction product. (I-OE) and the product

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 extract with concentrated acetone (1-0) were injected. 1500 ml of water, 2500 ml of methanol, 2000 ml of acetone and 2000 ml of chloroform were used to elute. The aqueous eluate, the methanol eluate, the acetone eluate and the chloroform eluate were collected separately and concentrated in vacuo to give 0.27 g of concentrated aqueous eluate (I-OW), 1.45 g of concentrated methanolic eluate (I-OM), 2.68 g of concentrated acetone eluate (I-OA) and 0.83 g of concentrated chloroform eluate (I-OC). The amounts (mg) of 6-shogaol, 6-gingerol and 6-dehydrogingerdione per gram of I-O, I-OM and I-OA determined by HPLC are listed in Table 1.

Table 1

<Tb>
<tb> Content
<tb> (mg / g) <SEP> IO <SEP> I-OM <SEP> I-OA
<tb> 6-shogaol <SEP> 1.10 <SEP> 0.14 <SEP> 1.15 <SEP> 0.0-
<tb> 6-gingerol <SEP> 59.98 <SEP> 0.99 <SEP> 103.37 <SEP> 8.57 <SEP> 2.51 <SEP> 0.89 <SEP>
<tb> 6-dehydrogingerdione <SEP> 7.68 <SEP> 0.42 <SEP> 8.94 <SEP> 0.41-
<Tb>
Example 2
500 g of shade-dried ginger rhizomes were reduced to powder and the resulting powder was extracted with 30 L of acetone three times (with 10 1 each). The three extraction solutions were combined together after filtration and then concentrated in vacuo to give 24 g of concentrated acetone extract (II-O). In a reverse phase chromatography column packed with 600 g of Diaion HP-20 resin, 20 g of the concentrated acetone extractant (II-O) were injected, which

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 then eluted with 4 l of water, 6.5 l of methanol, 15 l of acetone and 5 l of chloroform successively. The aqueous eluate, the methanol eluate, the acetone eluate and the chloroform eluate were collected separately and concentrated in vacuo to give 2.5 g of concentrated aqueous eluate (II-OW), 7.1 g of concentrated methanolic eluate (II-OM), 6.9 g of concentrated acetone eluate (II-OA) and 3.5 g of concentrated chloroform eluate (II-OC). The amounts (mg) of 6-shogaol, 6-gingerol and 6-dehydrogingerdione per gram of II-O, II-OM and II-OA determined by HPLC are listed in Table 2.

Table 2

<Tb>
<tb> Content
<tb> (mg / g) <SEP> II-O <SEP> II-OM <SEP> II-OA
<tb> 6-shogaol <SEP> 1, <SEP> 98 <SEP> 0.00 <SEP> 4, <SEP> 96 <SEP> ~ <SEP> 0.00 <SEP> -
<tb> 6 <SEP> -gingerol <SEP> 43.06 <SEP> 0.84 <SEP> 70.87 <SEP> 1.85 <SEP> 2, <SEP> 54 <SEP> 0, <SEP> 00 <SEP>
<tb> 6-dehydrogingerdione <SEP> 9.33 <SEP> 0.85 <SEP> 19.15 <SEP> 4.57 <SEP> 2.35 <SEP> 0.28
<Tb>
Example 3
10 kg of shade-dried ginger rhizomes were reduced to powder and the resulting powder was steamed for 5 hours. The distillate was concentrated in vacuo to give 410 g of concentrated distillate (III-0). In a reversed phase chromatography column packed with 600 g of Diaion HP-20.20 g resin of the concentrated distillate (III-O) was injected, which was then eluted with 4.5 l of water, 4.5 1 of methanol, 3 l of acetone and 5 l of chloroform successively. The aqueous eluate, the methanolic eluate, the acetone eluate and the chloroform eluate were collected separately and

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 concentrated in vacuo to obtain 0.03 g of concentrated aqueous eluate (III-OW), 14.5 g of concentrated methanolic eluate (III-OM), 0.85 g of concentrated acetone eluate (III-OA) and 0.2 g of concentrated chloroform eluate (III-OC). The concentrated distillate (III-0) does not contain 6-shogaol, 6-gingerol and 6-dehydrogingerdione as determined by HPLC.

Example 4
10 g of powder of ginger rhizomes dried in the shade were extracted with 1000 ml of acetone at 50 C for two hours. The extraction solution was separated and concentrated under vacuum (40 C, 75 mmHg) to obtain a concentrated acetone extractant (IV-O). The color and viscosity of the product (IV-O), as well as its yield, are shown in Table 3.

Example 5
10 g of shaded ginger rhizome powder were steam stripped and the oily distillate, after being separated from the aqueous distillate, was lyophilized to obtain an oily extract (VO). The color and viscosity of the oily extract (V-0), as well as its yield, are shown in Table 3.

Example 6
To 10 g of ginger rhizomes powder dried in the shade in a 250 ml extraction chamber, CO2 was introduced at a rate of 45 l / min. The pressure of the chamber was set at an absolute pressure of 17225.103 to 27560.103 Pa with a high pressure pump (Model N EK-1, LEWA Co, USA) and the chamber temperature was maintained at 35-60. C with a heat exchanger (Model N H-2410, HOTEC Co., USA) and an external circulation system. The extraction was stopped when the volume

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 of CO2 introduced reached 300 1, and a supercritical CO 2 extraction product (VI-O) was obtained after evaporation of CO2. The color and viscosity of the product (VI-O), as well as its yield, are shown in Table 3. The contents of the pungent components determined by HPLC are shown in Table 4.

Table 3

<Tb>
<tb> IV-O <SEP> VO <SEP> VI-O
<tb> L * <SEP> 87.6 <SEP> 80.4 <SEP> 96.3
<tb> A * <SEP> -9.1-0.1 <SEP> -9.6
<tb> B * <SEP> 31.1 <SEP> 9.6 <SEP> 22.0
<tb> Viscosity <SEP> (cps) <SEP> 15.6 <SEP> 11.8 <SEP> 12.1
<tb> Yield <SEP> (%) <SEP> 3.8 <SEP> 2.2 <SEP> 3.9
<Tb>
*: the values of L, A and B were determined using a colorimetric system # 90 (Nippon
Denshoku Inc., Co, Ltd., Japan), wherein L is intensity, unlike red / green and B is the yellow / blue difference.

Table 4

<Tb>
<tb> Content <SEP> (mg / g) <SEP> VI-O
<tb> 6-shogaol <SEP> 17.30 <SEP> 0.00 <SEP>
<tb> 6-gingerol <SEP> 26.29 <SEP> 0, <SEP> 00 <SEP>
<tb> 6-dehydrogingerdione <SEP> 19.20 <SEP> 1.19
<Tb>
Example 7: Antiplatelet test
Blood, collected from the marginal vein of the rabbit ear was mixed with EDTA (100 mM) in a volume ratio of 14: 1, and centrifuged at 90 g for 10 minutes at room temperature. to obtain platelet rich plasma. The latter was further centrifuged at 500 g for 10 minutes, the plasma-rich upper layer was removed from it, and the remaining lower layer was suspended with Tyrode's liquid containing 2 mM EDTA and not

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 of calcium. The suspension was further centrifuged at 500 g for 10 minutes and the platelets were suspended in tyrode liquid without EDTA. After centrifugation under the same conditions, the platelets were suspended in Tyrode's liquid having the following compositions (mM): NaCl (136.8), KCl (2.8), NaNCO3 (11.9), MgCl2 ( 1.1), NaH2PO4 (0.33), CaCl2 (1.0), glucose (11.2) and serum albumin (0.35%).

Platelet counts were determined with a Coulter counter (Model ZM) and adjusted to 4.5 x 108 platelets / ml.

Table 5. Inhibitory Effects of Ginger Extracts on Platelet Aggregation Induced by Arachidonic Acid and Collagen

<Tb>
<tb> Concentration <SEP> for <SEP> a <SEP> effect <SEP> inhibitor <SEP> of
<tb> 50 <SEP>% <SEP> (g / ml)
<tb> Extract <SEP> of <SEP> gingerb) <SEP><SEP> Acid arachidonic <SEP> Collagen
<tb> IO <SEP> 3.8 <SEP> 0.8 <SEP> 5.5 <SEP> 0.4
<tb> I-OM <SEP> 1.7 <SEP> 0.3 <SEP> 2.7 <SEP> 0.4 <SEP>
<tb> II-O <SEP> 3.1 <SEP> 0.5 <SEP> 6.5 <SEP> 1.2
<tb> II-OA <SEP> 10.9 <SEP> 3.2 <SEP> 21.8 <SEP> 2.2 <SEP>
<tb> II-OC <SEP> 6.9 <SEP> 0.7 <SEP> 16.6 <SEP> 4.3 <SEP>
<tb> II-OM <SEP> 2.0 <SEP> 0.2 <SEP> 6.9 <SEP> 2.4
<Tb>

The platelets were incubated with ginger extracts or with 0.5% DMSO (control) at 37 ° C for 3 minutes, then arachidonic acid (100 M) or collagen (10 g / ml) was added to trigger aggregation. Aspirin and indomethacin are positive controls. The percentage of inhibitory effect is calculated as follows: [(degree of inhibition of the control) - (degree of inhibition of the ginger extract) (degree of inhibition of the control) x 100%

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Values are presented as mean standard deviation, n = 3-6. b) Prepared in Examples 1 and 2 Example 8: Evaluation of the inhibitory activity on edema of the paw induced by carrageenan.

 A study on inhibitory activity on carrageenan induced paw edema was performed according to the method mentioned by Winter, C.A. et al.

(Winter, C.A. et al., Proc.Soc.Exc., Biol.Med 111: 544-547, 1962). Wistar male mice weighing 150 g were injected, without feeding overnight, at the left hind paws thereof, 0.1 ml of a 1% carrageenan suspension, and then samples of test or a vehicle as a control were rubbed on the left hind paws evenly (10 mg / paw). Three hours later, the volumes of the hind legs were determined using a volumetric scanner (Cat # 7150, UGO Basil, Italy), and the difference between the left hind paw and the right hind paw was used as an indication of the edema of the paw induced by carrageenan.

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Table 6. Inhibitory activity of ginger extracts on the edema of the paw induced by carrageeninea

<Tb>
<tb> Treatmentb) <SEP> Dosage <SEP> Activity
<tb> (mg / paw) <SEP> inhibitory <SEP> on
<tb> 1 <SEP>'<SEP> edema <SEP> of <SEP> la
<tb> paw <SEP> induced <SEP> by
<tb><SEP> carrageenan <SEP> (%)
<tb> IO <SEP> 10 <SEP> 18 <SEP>
<tb> I-OE <SEP> 10 <SEP> 19
<tb> I-OM <SEP> 10 <SEP> 29
<tb> I-OA <SEP> 10 <SEP> 25
<tb> II-O <SEP> 10 <SEP> 18 <SEP>
<tb> II-OW <SEP> 10 <SEP> 0
<tb> II-OM <SEP> 10 <SEP> 26
<tb> II-OA <SEP> 10 <SEP> 25
<tb> II-OC <SEP> 10 <SEP> 8
<tb> III-O <SEP> 10 <SEP> 0 <SEP>
<tb> III-OM <SEP> 10 <SEP> 11
<tb> III-OA <SEP> 10 <SEP> 15
<tb> [6] -dehydrogingerdione <SEP> 5 <SEP> 2 <SEP> 6 <SEP>
<Tb>
a) Inhibitory activity on # carrageenan-induced paw edema (%) was calculated as: [(mean degree of #dema of control group mice) - (mean degree of # demy of the test group mice) / (average degree of #dema of control group mice)] x 100%

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Values are presented as mean standard deviation, n = 3-6. b) Prepared in Examples 1,2 and 3 Example 9: Evaluation of the inhibitory activity on Trichophyton mentagrophytes or on Pityrosporum ovale.

Anti-Trichophyton mentagrophytes test
The minimum inhibitory concentration (MIC) of ginger extract was determined according to the method previously described by Edwards, JR et al.

(Edwards, J.R. et al., Antimicrobial Agents Chemotherapy 33: 215-222, 1989). The analyte was dissolved and serially diluted in a solvent (100% DMSO) to the desired base concentrations. For each concentration tested, an aliquot of 0.01 ml was added to a 48-well plate containing 0.99 ml of potato dextrose broth (Potato Dextrose Broth) (DIFCO, USA) with 100 ml. 104 CFU / ml Trichophyton mentagrophytes (American National Collection of Microorganism Culture ATCC 9533). The plates were incubated at 28 ° C for 72 hours and then examined visually and scored. A control vehicle was used as a blank control. Each concentration was evaluated in duplicate. The results are shown in Table 7.

Oval anti-Pityrosporum test
The minimum inhibitory concentration (MIC) of ginger extract was determined using the method mentioned above (Edwards, JR et al., Antimicrobial Agents Chemotherapy 33: 1989). The analyte was dissolved and diluted serially in a solvent (100% DMSO) to

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 desired base. For each concentration tested, an aliquot of 0.01 ml was added to a 48-well plate containing 0.99 ml of Sabouraud Medium fluid (Fluid Sabouraud Medium) (DIFCO, USA) with 103-104 CFU / ml. ml of Pityrosporum ovale (American National Collection of ATCC 38593 Microorganism Culture). The plates were incubated at 37 ° C for 48 hours and then examined visually and scored. A control vehicle was used as a blank control.

Each concentration was evaluated in duplicate. The results are shown in Table 7.

Table 7. Inhibitory Effects of Ginger Extracts on Pityrosporum ovale (Po) and Trichophyton mentagrophytes (Tm)

<Tb>
<tb> MIC <SEP> (g / ml)
<tb> Extract <SEP> of <SEP> ginger) <SEP> Po <SEP> Tm
<tb> IO <SEP> 100 <SEP> 30
<tb> I-OM <SEP> 100 <SEP> 30
<tb> II-O <SEP> 500 <SEP> 30
<tb> II-OC <SEP> 100 <SEP> 30
<tb> II-OM <SEP> 100 <SEP> 30
<tb> III-O <SEP> 500 <SEP> 100
<tb> Witness <SEP> to <SEP> White <SEP> b) <SEP> b) <SEP> ----]
<Tb>
') Prepared in Examples 1,2 and 3 (b) No inhibitory effect on development

Claims (32)

 1. A process for preparing a potent product for anti-inflammatory, platelet anti-aggregation and antifungal activities, from rhizomes of Zingiber officinale, comprising the following steps of: a) preparing a raw liquid from rhizomes of Zingiber officinale; b) introducing the crude liquid into a reverse phase chromatography column, and eluting the column with water, with a first eluent and with a second eluent successively, said second eluent having a lower polarity than that of the first eluent but stronger than that of chloroform, so that a first eluate resulting from elution with the first eluent and a second eluate resulting from elution with the second eluent are obtained; c) removing the first eluent from the first eluate by evaporation so that a first concentrated eluate is obtained and can be used as a strong product; and d) removing the second eluent from the second eluate by evaporation so that a second concentrated eluate is obtained and can be used as a strong product; wherein step a) comprises steps i) to iv), or comprises step I), step I '), or step I "), said steps i) to iv) consisting of: i ) chop fresh rhizomes of Zingiber officinale and filter the resulting mixture to obtain a filtrate and a residue; <Desc / Clms Page number 21>  ii) extracting the filtrate with a first organic solvent, recovering the extraction solution with the first resulting organic solvent, and evaporating the first organic solvent from the extraction solution to obtain a first concentrated extraction solution; iii) extracting the residue with a second organic solvent, recovering the extraction solution with the second resulting organic solvent, and evaporating the second organic solvent from the extraction solution to obtain a second concentrated extraction solution; and iv) combining the first concentrated extraction solution and the second concentrated extraction solution to obtain the raw liquid; said step I) consisting of: I) extracting dried rhizome powder from Zingiber officinale with the second organic solvent, recovering the extraction solution with the second resulting organic solvent, and evaporating the second organic solvent from the extraction solution to obtain the crude liquid; said step II) consisting of: (I) steaming the dried rhizome powder of Zingiber officinale, and concentrating the resulting distillate by evaporation to obtain the crude liquid; and said step I ") consisting of: I ") extract dried Zingiber officinalis rhizome powder with supercritical CO2, recover the extraction solution with supercritical CO 2 <Desc / Clms Page number 22>  resulting and evaporate the CO 2 from the extraction solution to obtain the crude liquid. The method of claim 1, wherein the potent anti-inflammation or anti-platelet aggregation product comprises from 0 to 10 mg of 6-shogaol per gram of the product, from 1 to 150 mg of 6gingerol per tablet. gram of the product and from 0 to 40 mg of dehydrogingerdione per gram of the product. The process according to claim 1 or 2, wherein said first eluent is methanol and said second eluent is acetone. The method of claim 1, 2 or 3, wherein step a) comprises steps i) to iv). The process of claim 4, wherein said first organic solvent is ethyl ether. The method of claim 4, wherein said second organic solvent is acetone, methanol, ethanol or a combination thereof. The process of claim 6, wherein said second organic solvent is acetone. The method of claim 1, 2 or 3, wherein step a) comprises step I). <Desc / Clms Page number 23>  The process of claim 8, wherein said second organic solvent is acetone, methanol, ethanol, or a combination thereof. The process of claim 9, wherein said second organic solvent is acetone. The method of claim 1, 2 or 3, wherein step a) comprises step I '). The method of claim 1, 2 or 3, wherein step a) comprises step I "). The process of claim 1, wherein said reverse phase chromatography column is packed with a porous resin. An anti-inflammatory pharmaceutical composition comprising a therapeutically effective amount of said crude liquid prepared in step a) of the process according to claim 1, as active ingredient, in admixture with a pharmaceutically acceptable carrier or diluent for the active ingredient. A pharmaceutical composition for inhibiting platelet aggregation, which comprises a therapeutically effective amount of said crude liquid prepared in step a) of the process according to claim 1, as active ingredient, in admixture with a carrier or a carrier. pharmaceutically acceptable diluent for the active ingredient. <Desc / Clms Page number 24> An antifungal pharmaceutical composition comprising a therapeutically effective amount of said raw liquid prepared in step a) of the process according to claim 1, as active ingredient, in admixture with a pharmaceutically acceptable carrier or diluent for the active ingredient. The pharmaceutical composition of claim 14, 15 or 16, wherein step a) comprises steps i) to iv). The pharmaceutical composition of claim 14, 15 or 16, wherein step a) comprises step I). The pharmaceutical composition of claim 17, wherein said first organic solvent is ethyl ether. 20. The pharmaceutical composition according to claim 17, 18 or 19, wherein said second organic solvent is acetone, methanol, ethanol or a combination thereof. The pharmaceutical composition of claim 20, wherein said second organic solvent is acetone. 22. The pharmaceutical composition of claim 14, 15 or 16, wherein step a) comprises step I '). 23. The pharmaceutical composition of claim 14, 15 or 16, wherein step a) comprises step I "). <Desc / Clms Page number 25> 24. An anti-inflammatory pharmaceutical composition comprising a therapeutically effective amount of the product prepared according to the process of any one of claims 1 to 13, as active ingredient, in admixture with a vehicle or a pharmaceutically acceptable diluent for the active ingredient. A pharmaceutical composition for inhibiting platelet aggregation, which comprises a therapeutically effective amount of the product prepared according to the process of any one of claims 1 to 13, as an active ingredient, in admixture with a carrier or a carrier. pharmaceutically acceptable diluent for the active ingredient. An antifungal pharmaceutical composition comprising a therapeutically effective amount of the product prepared according to the process of any one of claims 1 to 13, as an active ingredient, in admixture with a pharmaceutically acceptable carrier or diluent for the active ingredient. 27. The pharmaceutical composition according to claim 24, 25 or 26, wherein said product is the first concentrated eluate prepared in step c). The pharmaceutical composition of claim 24, 25 or 26, wherein said product is the second concentrated eluate prepared in step d). <Desc / Clms Page number 26> 29. A pharmaceutical composition according to claim 16 or 26 which is used in the treatment of Trichophyton mentagrophytes or Pityrosporum ovale associated disease. 30. The pharmaceutical composition according to claim 29, wherein said disease is selected from the group consisting of athlete's foot, microscopic moth, jock itch, dermatophytosis of the glabrous skin, onychomycosis, pityriasis capitis simplex. , pityriasis versicolor, folliculitis pityrosporum, seborrheic dermatitis and dandruff. 31. The pharmaceutical composition according to claim 29, which is in the form of a shampoo, a bath gel, a soap, a body lotion, a body cream or a body wash. detergent. 32. The pharmaceutical composition according to claim 31, which is in the form of a shampoo for use in the treatment of dandruff.