O-Quinone Compounds as Agents Neutralising Nitric Oxide

Abstract

The invention relates to O-quinone compounds of general formula (I) as agents neutralising nitric oxide, and to the therapeutic or cosmetic use thereof.

##STR00001##

Claims

1. A compound of formula (I): ##STR00009## for preventing and/or treating diseases and/or disorders resulting from an excess of NO, wherein: R.sup.1 is selected from the group consisting of —CH═CH—COOR.sup.2 and —Z—O—R.sup.3; is a group of formula —Y—O—X-(3,4)diphenol (II); is a pyranose ring, substituted with —CH.sub.2—OH, at least one hydroxyl function and a rhamnose; and Z are —CH.sub.2—CH.sub.2—; R.sup.3 is a group of formula —W—O—CO—CH═CH-(3,4)diphenol (III); and W is a pyranose ring, substituted with —CH.sub.2—OH, at least one hydroxyl function and a rhamnose;

2. The compound according to claim 1 selected from the group consisting of a first form of the quinone of the verbascoside and a second form of the quinone of the verbascoside.

3. An extract of Plantago lanceolata enriched with a quinone from verbascoside.

4. An extract of Plantago lanceolata enriched with a quinone from verbascoside, where the extract is obtained from aerial parts of the plant by alcoholic maceration.

5. The compound according to claim 1 for preventing and/or treating inflammatory diseases and/or disorders.

6. The compound according to claim 1 for preventing and/or treating skin inflammatory diseases and/or disorders selected from the group consisting of psoriasis, atopic dermatitis, contact dermatitis, skin irritation, contact hypersensitivity reaction, skin allergic signs, excessive vasodilation, rosacea, solar erythema and acne.

7. The compound according to claim 1 for preventing and/or treating joint inflammatory diseases and/or disorders selected from the group consisting of arthritis, arthrosis, rheumatoid polyarthritis, ankylosing spondylarthritis, erythematous lupus and chondritis.

8. The compound according to claim 1 for preventing and/or treating coughs.

9. The compound according to claim 1 for preventing and/or treating anaphylactic shocks or septic shocks.

10. The compound according to claim 1 for combatting intrinsic or extrinsic ageing of the skin; for combatting signs of skin ageing; for combatting skin neurogenic inflammatory processes; for improving the comfort of sensitive skins; for reinforcing the barrier function of the skin; for stimulating hydration of the skin; for improving the comfort of dry skins; for controlling sweating; for stimulating lipolysis; and for inhibiting hair loss.

11. A pharmaceutical or cosmetic composition containing at least one compound of formula (I) according to claim 1 and a physiologically acceptable carrier.

12. The composition according to claim 11, for application on skin or for oral administration.

13. The composition according to claim 12, wherein the composition is a cream, a gel, an oil, a lotion or a milk.

14. The composition according to claim 12, wherein the composition is a syrup, a lozenge to be sucked, a gelatin capsule or a tablet.

15. A method for preventing and/or treating diseases and/or disorders resulting from an excess of NO, comprising administering to a mammal in need thereof, a therapeutic amount of the compound according to claim 1.

16. The method according to claim 15, wherein the compound is applied to skin or administered orally.

17. A method for preventing and/or treating diseases and/or disorders resulting from an excess of NO, comprising administering to a mammal in need thereof, a therapeutic amount of the composition according to claim 11.

18. The method according to claim 17, wherein the composition is applied to skin or administered orally.

Description

FIGURE

[0102] FIG. 1: Apparatus allowing application of the specific method for evaluating the neutralization of nitric oxide as described in example 1.

EXAMPLES

Example 1. Method for Evaluating the Neutralization of Nitric Oxide

[0103] Nitric oxide is prepared extemporaneously in a container A, obturated with a flexible polymeric plug through which pass an entering tubing and an exiting tubing, with an inner diameter comprised between 50 μm and 300 μm, but particularly of 100 μm (cf. FIG. 1), with a volume comprised between 10 ml and 100 ml, but particularly of 30 ml, by adding a volume comprised between 0.1 and 2 ml, but particularly of 0.5 ml of an aqueous solution of sodium nitrite (NaNO.sub.2, ref. Sigma-Aldrich 237213) at a concentration comprised between 0.10% and 2.00% but particularly of 0.70%, at a volume comprised between 5 ml and 80 ml of the following aqueous solution:

TABLE-US-00001 Sulfuric acid (H.sub.2SO.sub.4, ref. Sigma-Aldrich 32,050-1):  1 ml Ferrous sulfate (FeSO.sub.4, 7H.sub.2O, ref. Sigma-Aldrich 215422): 5.5 g  Deionized water: 200 ml

[0104] The container B in which is measured the nitric oxide is a container opaque to light, with a double jacket allowing circulation of water maintaining the temperature at 25° C., obturated with a plug through which pass the tubing stemming from the container A and an exhaust tubing (cf. FIG. 1) with a volume comprised between 10 ml and 100 ml, preferably 30 ml which receives a volume comprised between 5 ml and 80 ml, preferably 20 ml, of an aqueous solution of phosphate buffer at pH 7.4.

[0105] Once they are mixed, the solutions are stirred for the whole period of the measurement with a magnetic stirrer, in container A and in container B.

[0106] Nitrogen gas is admitted at a flow rate comprised between 20 ml/min and 120 ml/min, preferably 80 ml/min in the bottom of the container A. The nitrogen thereby carries away the nitric oxide through the exiting tubing of container A as far as into container B where it is measured with a device provided for this purpose equipped with an amperometric probe provided with a specific membrane (apparatus of ref. TBR 1025 from World Precision Instruments, probe ISO-NOP). The probe thereby delivers a variable current between 10 and 1,000 nA proportional to the concentration of nitric oxide in the tested solution, of the order of magnitude of μM. The electric current produced by the probe under the influence of nitric oxide is recorded with a suitable device.

[0107] Before any measurement, the probe is calibrated by measuring the current produced by releasing known increasing amounts of nitric oxide. It is thus checked that there exists a linear relationship between the electric current produced by the probe and the molar concentration of nitric oxide and the correspondence between this electric current and the molar concentration of nitric oxide is determined. For this, 20 ml of the following aqueous solution is placed in the container B disconnected from the container A during the calibration period:

TABLE-US-00002 Sulfuric acid (H.sub.2SO.sub.4, ref. Sigma-Aldrich 32,050-1):  1 ml Potassium iodide (KI, ref. Sigma-Aldrich 12636): 3.31 g   Deionized water: 200 ml

[0108] 50 μl, 100 μl, 150 μl and 200 μl of a solution of sodium nitrite (NaNO.sub.2, ref. Sigma-Aldrich 237213) are then successively added to a concentration of 360 μM.

[0109] The curve giving the correspondence between the nitric oxide concentration in the reaction medium and the electric intensity produced by the probe expressed in nA is plotted for each addition of the nitric oxide solution.

Evaluation of the Neutralization Effect on a Molecule to be Tested

[0110] During a typical measurement, 20 ml of the ferrous sulfate solution are placed in the container A and 20 ml of phosphate buffer at pH 7.4 are placed in the container B. In the container A, nitrogen gas is admitted for 10 mins, which passes in transit towards the container B, in order to remove the dissolved oxygen present in the solutions of the containers A and B. After these 10 mins, the recording of the measurement of nitric oxide is launched.

[0111] At time T.sub.0, the sodium nitrite solution is injected into the container A through the flexible polymeric plug by means of a syringe equipped with a needle. The recording of the nitric oxide shows a positive evolution which will stabilize shortly after about 3 mins, before slowly decreasing in order to return to the baseline in about 20 mins. At time T.sub.1=T.sub.0+4 mins, 400 μl of a hydro-ethanol solution with an alcoholic titer comprised between 0% and 100% according to the solubility of the product to be tested, are injected, containing an amount P of product from 4 to 8 mg, according to the efficiency of the product to be tested, through the flexible plug of the container B by means of a syringe equipped with a needle.

[0112] A control is produced with 400 μl of solvent in which is dissolved the product to be tested.

[0113] The value E of the neutralization effect of nitric oxide by the product to be tested, expressed in μM of nitric oxide neutralized per second and per mg of product to be tested is obtained by the following expression:

[00001]$\begin{array}{cc}E=\frac{\Delta .Math..Math.H_{\mathrm{test}}-\Delta .Math..Math.H_{\mathrm{tem}}}{30.Math..Math.P}& \left(1\right)\end{array}$

With: ΔH.sub.test=difference in concentration in nitric oxide test, expressed in pM of nitric acid between T.sub.1 and T.sub.2=T.sub.1+30 s, obtained with the solution of product to be tested. ΔH.sub.tem=difference in concentration of control nitric oxide, expressed in pM of nitric oxide between T.sub.1 and T.sub.2=T, +30 s, obtained with the solvent of the product to be tested.
P=amount of product to be tested in mg.

Example 2. Measurement of the Neutralization Effect of Nitric Oxide and Effects of Diverse Molecules

[0114] The test described in example 1 was applied for testing the following molecules:

Rosmarinic acid (Aldrich 536954)
p-coumaric acid (Sigma C9008)
(+) catechin hydrate (Sigma C1251)
Caffeic acid (Sigma-Aldrich C0625)
Chlorogenic acid (Sigma-Aldrich C3878)
Quercetin dihydrate (Sigma Q0125)
Porcine hemoglobin (Sigma H4131)

Verbascoside (HWI Analytik GMGH 0082-05-80)

Luteolin (Sigma L9283).

[0115] After having calibrated the probe for measurement of the concentration of nitric oxide of the apparatus described in example 1, in the container A, as described in example 1 and illustrated in FIG. 1, 20 ml of an aqueous solution is placed:

TABLE-US-00003 of sulfuric acid (H.sub.2SO.sub.4, ref. Sigma-Aldrich 32,050-1):  1 ml of ferrous sulfate (FeSO.sub.4, 7H.sub.2O, ref. Sigma-Aldrich 215422): 5.5 g  of deionized water: 200 ml

[0116] In the container B, as described above and in FIG. 1, 20 ml of a phosphate buffer solution at a pH of 7.4 is placed.

[0117] Nitrogen gas is admitted into the apparatus at a flow rate of 80 ml/min for 10 mins, and then the recording of the value of the intensity of the electric current produced by the probe for detection of nitric oxide is launched and (at time T.sub.0) it is injected into the container A, a 0.5 ml of an aqueous solution of sodium nitrite (NaNO.sub.2, ref. Sigma-Aldrich 237213) at a concentration of 0.70%.

[0118] At time T.sub.1=T.sub.0+4 mins, 400 μl of a hydro-ethanol solution with an alcoholic titre comprised between 0% and 100% depending on the solubility of the product to be tested, containing an amount P of product to be tested expressed in mg, adjusted according to the efficiency of the tested product.

[0119] The values of the concentrations of nitric oxide are noted at time T.sub.0, T.sub.1=T.sub.0+4 mins and T.sub.2=T.sub.1+30 s. The values of E calculated according to formula (1) appears in the table 1 below.

TABLE-US-00004 TABLE 1 Tested molecules E (μM .Math. s.sup.−1.Math. mg.sup.−1) Hemoglobin (control+) 2.27 Caffeic acid 0.957 Rosmarinic acid 0.043 p-coumaric acid 0.13 Catechin 0.13 Chlorogenic acid 0.043 Luteolin 0.174 Quercetin 0.087 Verbascoside 0.058

Example 3. Preparation and Measurement of the Effect of o-Quinone of Caffeic Acid

[0120] The ortho-quinone of caffeic acid:

##STR00008##

may be obtained crystallized, by oxidation of caffeic acid by the o-chloranil in solution in a mixture of ether and of tetrahydrofurane (4/1) at −70° C., according to a method described in the prior art (20, 21).

[0121] According to the method for measuring the neutralization effect of nitric oxide as described in example 1, the caffeoyl-quinone produces an effect E of 24.36 μM.Math.s.sup.−1.Math.mg.sup.−1.

[0122] A high value of E is therefore observed for this molecule comparatively with the effects produced by molecules already known as capable of neutralizing nitric oxide.

Example 4. Preparation of an Extract of Plantain and Evaluation of its Neutralization Activity of Nitric Oxide

[0123] In order to determine the method for treating the plant and the extraction method leading to better neutralization efficiency of nitric oxide, it was proceeded with the extraction of a dry plant and of a fresh plant with hydro-alcoholic solutions with titers varying from 10 to 10 ranging from 0% to 100% of ethanol, either by maceration at room temperature for 48 h, in solvent volumes corresponding to 10 times the weight of the plant (v/w), or by extraction upon boiling at reflux for 30 mins with solvent volumes also corresponding to 10 times the weight of the plant (v/w). Subsequently to these preliminary tests, the following method was selected, since it produces the most active extract:

[0124] Aerial parts of Plantago lanceolate are harvested and rapidly frozen at −18° C. in order to ensure their preservation. 1 kg of this batch is coarsely milled and put to ripen in 10 liters of aqueous ethanol at 60% for 48 h. This alcoholic titer is defined by taking into account the water content of the plant, determined beforehand by drying on a representative sample. At the end of the maceration, the plant is removed by sifting, and then the extracted solution is filtered on a filter with 0.45 μM of porosity for removing the major portion of the germs present. The solution is then concentrated under reduced pressure so as to obtain a total volume of 1 l, totally without any ethanol.

[0125] This solution is then dried by freeze-drying in order to obtain a final weight of 43 g.

[0126] The efficiency E of neutralization of nitric oxide is 0.16 μM.Math.s.sup.−1.Math.mg.sup.−1.

Example 5. Preparation of an Extract of Aerial Parts of Plantago lanceolate Enriched with o-Quinones from Verbascoside

[0127] Aerial parts of Plantago lanceolate are harvested and rapidly frozen to −18° C. in order to ensure their preservation. 1 kg of this batch is coarsely milled and set to ripen in 10 liters of aqueous ethanol at 60% for 48 h. This alcoholic titer is defined by taking into account the water content of the plant, determined beforehand by drying on a representative sample. At the end of the maceration, the plant is removed by sifting, and then the extracted solution is filtered on a filter with 0.45 μM of porosity for removing the major portion of the germs present. The solution is then concentrated under reduced pressure so as to obtain a total volume of 1 l, totally without any ethanol.

[0128] This concentrated solution is then extracted with a counter current with 5 times 200 ml of ethyl acetate. The organic solutions are collected, dried on anhydrous sodium sulfate, filtered and then dry evaporated under reduced pressure. 1.12 g of dry extract is thereby obtained which is taken up with 25 ml of pure ethanol.

[0129] This ethanol solution is subject to flash chromatography (apparatus Grace Reveleris X2) on a silica gel column of 40 g (Grace Reveleris Silica Cartridge 40 g) with a dichloromethane-methanol gradient like in table 2 below:

TABLE-US-00005 TABLE 2 Time (mins) Dichloromethane % Methanol % 0 100 0 15 80 20 18 80 20 18.5 50 50 23.5 50 50

[0130] The fractions of 2 ml which are subject to the neutralization test of nitric oxide are collected. The most active fractions are retained which correspond to the elution volumes of 568 ml to 576 ml, which provide the whole 17.7 mg of dry material after evaporation of the solvent under reduced pressure, and which has a neutralization efficiency E of nitric oxide of 0.96 μM.Math.s.sup.−1.Math.mg.sup.−1.

[0131] The presence of the o-quinone of the verbascoside is then checked in high resolution mass spectrometry in tandem, with electrospray ionization in a negative mode, by the presence of a doubly charged ion at m/z 311, corresponding to the relevant quinone and having the characteristic fragmentation of phenylhydantoic heterosides, in this case related to that of the verbascoside.

Example 6. Galenic Formula of a Cream Containing an Extract of Plantain, Intended for Treating Skin Inflammations (Notably Dermatitises, Eczema, Psoriasis) Adapted to Body Use

[0132] A cream is prepared having the following weight formula, according to a method and with standard materials as known from the prior art:

TABLE-US-00006 Purified water: 67.28%    Glycerol (Glycerine, AMI Chimie): 20%  Cetostearyl alcohol (Lanette O, AMI Chimie): 7% Palmitostearic acid (Stearin, Stearinerie Dubois): 3% Extract of fresh aerial parts 1% of Plantago lanceolata as described in example 5: Benzyl alcohol (Geogard 221, Lonza): 0.87%   Cetostearyl sodium sulfate (Lanette E, AMI Chimie): 0.7%   Dehydroacetic acid (Geogard 221, Lonza): 0.09%   Sodium hydroxide (Sigma-Aldrich): 0.058%   

Example 7. Galenic Formula of a Cream Containing a Plantain Extract, Intended for Treating Skin Inflammations (Notably Dermatitises, Eczema, Psoriasis) Adapted for Use on the Face

[0133]

TABLE-US-00007 Purified water: 67.28%    Glycerol (Glycerine, AMI Chimie): 10%  Cetostearyl alcohol (Lanette O, AMI Chimie): 7% Isopropyl myristate (Isopropyl Myristate, Interchimie): 3% Extract of fresh aerial parts 1% of Plantago lanceolata as described in example 5: Benzyl alcohol (Geogard, Lonza): 0.87%   Cetostearyl sodium sulfate (Lanette E, AMI Chimie): 0.7%   Dehydroacetic acid (Geogard, Lonza): 0.09%   Sodium hydroxide (Sigma-Aldrich): 0.058%   

Example 8. Galenic Formula of a Washing Gel Containing a Plantain Extract, Intended for Cleaning Hair, the Face and the Body of Subjects Having Skin Inflammations (Notably Dermatitises, Eczema, Psoriasis)

[0134]

TABLE-US-00008 Mixture of fatty alcohol ethersulfate (Texapon ASV50, AMI   50% Chimie): Purified water: 34.03%  Glycerol (Glycerine, AMI Chimie):   10% Sodium chloride (sodium chloride, VWR):   4% Extract of fresh aerial parts   1% of Plantago lanceolata as described in example 5: Benzyl alcohol (Geogard 221, Lonza): 0.87% Dehydroacetic acid (Geogard 221, Lonza): 0.09% Sodium hydroxide (Sigma-Aldrich): 0.01%

Example 9. Galenic Formula of a Gel Intended for Treating Skin Inflammations (Notably Dermatitises, Eczema, Psoriasis) Adapted to Body Use and to Use on the Face

[0135]

TABLE-US-00009 Purified water: 95.69%  Carbomer (Carbopol 980, Lubrizol):   2% Extract of fresh aerial parts   1% of Plantago lanceolata as described in example 5: Benzyl alcohol (Geogard 221, Lonza): 0.87% Dehydroacetic acid (Geogard 221, Lonza): 0.09% Sodium hydroxide (Sigma-Aldrich): 0.35%

Example 10. Galenic Formula of a Cream Containing a Plantain Extract, Intended for Treating Inflammations Relative to Acne

[0136]

TABLE-US-00010 Purified water: 71.04%  Cetostearyl alcohol (Lanette O, AMI Chimie):   10% Essential oil of lemon (Myrtea):  7.5% Essential oil of sweet orange (Myrtea):  7.5% Extract of fresh aerial parts   1% of Plantago lanceolata as described in example 5: Cetostearyl sodium sulfate (Lanette E, AMI Chimie):   2% Benzyl alcohol (Geogard 221, Lonza): 0.87% Dehydroacetic acid (Geogard 221, Lonza): 0.09%

Example 11. Galenic Formula of a Syrup for Coughs

[0137]

TABLE-US-00011 Purified water: 44.23%    Saccharose: 44.22%    Glycerol (Glycerine, AMI Chimie): 5% Extract of fresh aerial parts 4% of Plantago lanceolata as described in example 5: Natural raspberry aroma: 1.5%   Citric acid monohydrate: 0.75%   Sodium benzoate: 0.3%  

Example 12. Galenic Formula of Gelatin Capsules for Treating Inflammatory Signs of Arthrosis

[0138] For a gelatin capsule no. 0 (Capsugel):

TABLE-US-00012 Extract of fresh aerial parts 0.350 g of Plantago lanceolata as described in example 5: Maltodextrin: 0.045 g

REFERENCES

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