HK1085997B - Depigmentation agents - Google Patents

Description

Decolorizing agent

Technical Field

The present invention relates to 4-cyclopentyl resorcinol monohydrate (4-cyclopentyl resorcinol monohydrate) and its form I polymorph.

Background

In humans, skin color results from a complex series of cellular processes that are carried out within a population of cells called melanocytes. Melanocytes are located in the lower part of the epidermis and their function is to synthesize the pigment, melanin, which protects the body from the damaging effects of ultraviolet radiation.

The mechanism of skin pigmentation formation, melanogenesis, comprises the following major steps: tyrosine → L-Dopa (Dopa) → dopaquinone dopachrome → melanin. The first two reactions in the series are catalyzed by the enzyme tyrosinase. Tyrosinase activity is promoted by the action of alpha-melanocyte stimulating hormone and UV rays.

Typically, melanogenesis results in darker skin tone (i.e., tan). However, melanogenesis can also lead to undesirable pigmentation patterns. Examples of such undesirable pigmentation include age spots, freckles (liver spots), melasma, hyperpigmentation, and the like. This has led to research to find compounds that inhibit melanogenesis. One goal of this study is tyrosinase, an enzyme that catalyzes an initial step in melanin production.

U.S. patent 6132740 discloses a class of tyrosinase inhibitors. These compounds are 4-cycloalkyl resorcinols. One compound disclosed in the' 740 patent is 4-cyclopentyl resorcinol. Example 2 of the' 740 patent describes the preparation of 4-cyclopentyl resorcinol. The synthesis described in example 2 can result in the production of oil. Although this oil is a potent tyrosinase inhibitor, it cannot be readily produced in the required amounts to support clinical development.

The synthesis of example 2 produced a large number of various positional isomers of 4-cyclopentyl resorcinol. Examples of such isomers include 2-cyclopentyl resorcinol, 4, 6-dicyclopentyl resorcinol, 2, 4-dicyclopentyl resorcinol, and the like. It is difficult to separate 4-cyclopentylresorcinol from its positional isomers, especially when all compounds are present as oils. Accordingly, there is a need in the art for solid forms of 4-cyclopentyl resorcinol that can be more readily produced than the anhydrates of the prior art.

Summary of The Invention

According to the present invention, a novel solid form of 4-cyclopentyl resorcinol has been discovered. The solid form is 4-cyclopentyl resorcinol monohydrate. 4-cyclopentyl resorcinol monohydrate can be represented by the following formula:

formula 1

Another aspect of the present invention relates to specific crystalline polymorphs of 4-cyclopentyl resorcinol monohydrate. This polymorph is referred to as polymorph form I. It has a characteristic powder X-ray diffraction pattern (XRPD) as described hereinafter. The single crystal structure of polymorph form I is also determined and reported below.

The monohydrate of 4-cyclopentyl resorcinol and its form I polymorph can be used to lighten skin (i.e., as a depigmenting agent). In a more specific embodiment, the compound is incorporated into a topical dosage form that the patient can apply directly to the area of skin in need of lightening.

In yet another embodiment, the invention relates to an article of manufacture containing a monohydrate or form I polymorph thereof packaged for retail sale with instructions advising the consumer how to use the product to lighten the skin.

Figure 1 depicts an X-ray powder diffraction pattern of form I polymorph over the range of 2.5 ° 2 Θ to 40 ° 2 Θ.

Figure 2 shows an X-ray powder diffraction pattern of form I polymorph over a 7 ° 2 θ to 28 ° 2 θ shrinkage range.

Figure 3 shows X-ray powder diffraction patterns of 4 different batches of polymorph form I.

Detailed Description

A.Characterization method

1) Experimental X-ray powder diffraction

These X-ray powder diffraction (XPRD) analyses of lot numbers 2, 3 and 4 shown in FIG. 3 were performed using a Shimadzu XRD-6000X-ray powder diffractometer using Cu Ka radiation. The instrument is equipped with a fine focus X-ray tube. The tube voltage and current intensity were set to 40kV and 40mA, respectively. The divergence slit and the anti-divergence slit are set at 1 deg., and the receiving slit is set at 0.15 mm. Diffracted radiation was detected with a NaI scintillation detector. Continuous scans from 2.5-40 deg. 2 theta using theta-2 theta of 3 deg./min (0.4s/0.02 deg. steps). Silicon standards were analyzed to check instrument calibration (instrument alignment). Data were collected and analyzed using Shimadzu XRD-6000v.4.1 software. Samples for analysis were prepared by placing the samples on a silicon sample holder and aligning with a frosted glass slide.

The X-ray powder diffraction (XPRD) analyses shown in fig. 1, 2 and 3 (batch #1) were performed using a Cu ka radiation using an Inel XRG-3000 diffractometer equipped with a bend position-sensitive detector. Data were collected in real time at a resolution of 0.03 ° over a 2 θ range of 120 °. The tube voltage and current strength were 40kV and 30mA, respectively. Samples were packed in aluminum racks with silicon plugs and analyzed. Silicon standards were analyzed daily to check the instrument calibration. Data were collected using INEL windlot v3.11 software and analyzed using Shimadzu XRD-6000v.4.1 software.

As will be apparent to those skilled in the art, the results of any X-ray powder diffraction may vary. This variation may be attributed to the test sample preparation, the particular model of X-ray diffractometer used, the skill of the operator, and the like. If the term "about" is used in defining the position of a characteristic peak in an X-ray powder diffraction pattern, the term is defined as a 2 θ value ± 0.2 ° 2 θ.

2) X-ray single crystal structure determination

The type I single crystal is laid in a random orientation. Preliminary examination and data collection was performed using Mo K α radiation on a Bruker SAMRT IK CCD diffractometer available from Bruker AXS, inc. 6145 diffraction points were collected over a set angle of 3 ° < θ < 28 °, and the cell constants and orientation matrix were obtained by minimum two-fold refinement of the collected data. The structure is solved by a direct method. By making a pair F²The full matrix method least square fine structure. The crystal structure was determined at 160K, space group P2₁Solved in/c and accurate to a final R of 0.04 (F)²＞2σ)。

B) 4-cyclopentyl resorcinol monohydrate

As described above, novel forms of 4-cyclopentyl resorcinol have been discovered. This novel form is the monohydrate of 4-cyclopentyl resorcinol. Such materials may be represented by the following formula:

the term "monohydrate" is generally used to describe a substance in which 1 water molecule is associated with one molecule (i.e., 1: 1 molar ratio) of a given compound. The term "monohydrate" as used herein should not be considered in this limiting sense. The inventors have found that the molar ratio of water to 4-cyclopentyl resorcinol can vary. The term "4-cyclopentyl resorcinol monohydrate" as used herein refers to a material comprising from about 0.7 moles of water to about 1.4 moles of water per mole of 4-cyclopentyl resorcinol. In a more specific embodiment, the monohydrate has from about 0.8 to about 1.2 moles of water per mole of 4-cyclopentyl resorcinol. In a more specific embodiment, the monohydrate has from about 0.9 to about 1.2 moles of water per mole of 4-cyclopentyl resorcinol.

This new form has many advantages over prior art forms. One major advantage is the simplicity of separating 4-cyclopentyl resorcinol from the above positional isomers such as 2-cyclopentyl resorcinol, 4, 6-dicyclopentyl resorcinol, 2, 4-dicyclopentyl resorcinol. The reader will note that examples 1-3 illustrate this simplified recovery. The hydrate of 4-cyclopentyl resorcinol will crystallize from the reaction mixture. It can be separated from the positional isomers by filtration rather than by distillation or column chromatography as required in prior art forms.

C) Polymorph form I of 4-cyclopentyl resorcinol

4-cyclopentyl resorcinol monohydrate can exist as a crystalline polymorph. A crystalline polymorph has been identified to date. For simplicity, hereinafter will be referred to as "form I polymorph".

Form I polymorph can be identified by its characteristic X-ray powder diffraction pattern. Examination of figures I-III shows that form I polymorph exhibits three characteristic peaks. The characteristic peaks are those with significant relative intensity in the powdered XPRD pattern.

One peak occurs at about 8.1 ° 2 θ. The second peak occurs at about 23.8 ° 2 θ. The third peak occurs at about 16.2 ° 2 θ. Any of these peaks can be used alone or in combination to determine form I polymorph.

In addition to these characteristic peaks, examination of FIGS. I-III shows that additional secondary peaks can be identified. The intensity of these additional peaks varies with the particular orientation of the polymorph sample. These additional peaks can be used to confirm the presence of the form I polymorph, but their absence does not apply to determining that a particular material is not a form I polymorph. These secondary peaks include: 13.9, 14.3, 18.4, 19.3, 20.1, 21.2, 25.8 and 26.54 (expressed in ° 2 θ, ± 0.2 ° 2 θ).

The structure of the single crystal of polymorph form I was also determined. The unit cell parameters are shown in table I below. Table II describes the atomic coordinates and isotropic displacement parameters. Table III describes the hydrogen atom coordinates and isotropic displacement parameters.

TABLE I

Space group and unit cell parameters for polymorph form I

TABLE II

Atomic coordinates and isotropic displacement parameters of 4-cyclopentyl resorcinol monohydrate

U_eqDefined as orthogonalizing U^ijOne third of the tensor trace.

TABLE III

Hydrogen atom coordinates and isotropic displacement parameters of 4-cyclopentyl resorcinol monohydrate

U_eqDefined as orthogonalizing U^ijOne third of the tensor trace.

C) Preparation method

Reaction scheme I

4-cyclopentyl resorcinol monohydrate and its form I polymorph can be prepared by analogous methods known to those skilled in the art, and these methods are described in reaction scheme I.

The initial step was to prepare 4-cyclopentyl resorcinol. This can be accomplished as described in U.S. patent 6132740, which is hereby incorporated by reference. A Friedel-Crafts reaction is carried out in which resorcinol is contacted with an excess of cyclopentanol in the presence of a catalyst such as polyphosphoric acid and the mixture is heated until the reaction is complete. The 4-cyclopentyl resorcinol can be recovered by extraction. Evaporation of the organic phase of the extract yields a mixture of 4-cyclopentylresorcinol and its positional isomers.

To obtain the monohydrate of 4-cyclopentyl resorcinol, or its form I polymorph, the above isolation and recovery process must be modified. This can be achieved using methods similar to those known in the art to recrystallize the anhydrate in the presence of sufficient water. The anhydrate is dissolved in the recrystallization solvent and cooled to precipitate the desired monohydrate from solution as polymorph form I. One suitable recrystallization solvent is a mixture of water and toluene. The ratio of toluene to water can vary widely. The form I polymorph can be isolated by filtration or evaporation, as is known in the art.

It was found that separate extraction and recrystallization were not necessary. The form I polymorph can be directly recovered by using a mixture of toluene and water as the extraction solvent. On cooling, polymorph form I crystallizes from solution. Likewise, other recrystallization solvents may also be used. Polymorph form I has been produced from mixtures of ethanol/water, methanol/water and isopropanol/water. The reader will note that this recrystallization is described in more detail in examples 1-3.

D. Pharmacology and dosage

As mentioned above, U.S. Pat. No. 6132740 describes the pharmacology of 4-cyclopentyl resorcinol. It is a tyrosinase inhibitor. It can be used for inhibiting melanin production by melanocyte (i.e. inhibiting melanogenesis). 4-cyclopentyl resorcinol monohydrate and its form I polymorph are also tyrosinase inhibitors (hereinafter "compound"). They can be used to inhibit melanogenesis in the same manner as described in the' 740 patent. Thus, the compounds are useful for lightening improperly pigmented skin areas.

Examples of such inappropriate pigmentation include sunlight and simple blotches (including age spots/freckles), melasma/chloasma and post-inflammatory hyperpigmentation. The compounds may also be used to reduce the amount of non-diseased skin melanin in order to induce a lighter skin tone when used as desired by the user, or to prevent the accumulation of melanin in skin exposed to UV radiation. They can also be used in combination with desquamating agents (including glycolic acid or trichloroacetic acid dehulline) to lighten skin tone and prevent re-pigmentation.

The compounds may also be used in combination with sunscreens (UVA or UVB blockers) to prevent re-pigmentation, to combat sun or UV induced darkening of the skin or to enhance their ability to reduce skin melanin and their skin depigmenting effects. The compounds used in the present invention may also be used in combination with 4-hydroxyanisole. The compounds used in the present invention may also be used in combination with ascorbic acid, its derivatives and ascorbic acid based products (such as magnesium ascorbate) or other products with anti-aging mechanisms (such as resveratrol) that promote or enhance their ability to reduce skin melanin and their skin depigmentation effects.

As a general guideline, the compounds will be administered topically. They will be applied directly to the area of skin where bleaching or lightening is desired. Topical formulations such as creams, lotions, ointments, gels, etc., containing about 0.1-10 w/w% of the compound are prepared. The compound is then applied to the affected area 1 to 4 times per day. If the compound is administered systemically, it is administered daily from about 0.1mg/kg to about 100mg/kg, optionally in divided doses.

E.Pharmaceutical preparation

If desired, the compounds can be administered directly without any carrier. However, for ease of administration, they are generally formulated into a pharmaceutical carrier. Also, most of them are generally formulated into dermatological or cosmetic carriers. In the present application, the terms "dermatological carrier" and "cosmetic carrier" are used interchangeably. They refer to formulations designed for direct administration to the skin or hair (i.e., topical formulations).

For oral administration, the compounds may be formulated as solid or liquid preparations such as capsules, pills, tablets, lozenges, melts, powders, suspensions or emulsions. Solid unit dosage forms may be conventional gel-type capsules containing, for example, surfactants, lubricants, and inert fillers such as lactose, sucrose, and corn starch, or they may be sustained release formulations.

In another embodiment, the compounds may be tableted using conventional tablet bases such as lactose, sucrose and corn starch in combination with binders such as acacia, corn starch or gelatin, disintegrating agents such as potato starch or alginic acid, and lubricating agents such as stearic acid or magnesium stearate. Liquid formulations may be prepared by dissolving the active ingredient in a water-soluble or non-water-soluble pharmaceutically acceptable solvent, which may also contain suspending agents, sweetening agents, flavoring agents, and preserving agents as are known in the art.

For parenteral administration, the compounds may be dissolved in a physiologically acceptable pharmaceutical carrier and administered as a solution or suspension. Exemplary suitable pharmaceutical carriers are water, saline, dextrose solution, fructose solution, ethanol or animal, vegetable or synthetic source oils. The pharmaceutical carrier may also contain preservatives, buffers and the like as are known in the art. When compounds are administered by intrathecal injection, they may also be dissolved in cerebrospinal fluid, as is known in the art.

Typically, however, the compounds will be incorporated into formulations suitable for topical administration. Any topical formulation known in the art may be used. Examples of such topical formulations include lotions, sprays, creams, ointments, salves, gels, and the like. The actual method of preparing the topical formulations is known and obvious to those skilled in the art and is described in detail in Remington's pharmaceutical science, 1990 (supra); and Pharmaceutical document Forms and Drug delivery systems, 6 th edition, Williams & Wilkins (1995).

In yet another embodiment, the above-described formulations can be packaged for retail sale (i.e., industrial kits or products). The package will contain instructions advising the patient how to use the product to lighten their skin. Such instructions may be printed on the cartridge, as individual loose sheets, or on the face of the container holding the formulation, etc.

F. Examples of the embodiments

The following examples are provided to further illustrate the invention. They should not be construed as limiting the invention in any way.

Example I

The following example describes one method of preparing 4-cyclopentyl resorcinol monohydrate form I polymorph.

Resorcinol (150g, 1.36mol), cyclopentanol (125ml, 1.38mol) and phosphoric acid (85% in water) were charged in a 500ml round bottom flask equipped with a stir bar. The flask was equipped with a reflux condenser, purged with nitrogen, and the mixture was heated at 120 ℃ (oil bath temperature) for 26 hours. After this time, TLC analysis indicated that resorcinol starting material was still present. Cyclopentanol (25ml, 0.25mol) was added to the reaction mixture and heating was continued for 2.5 hours. On cooling, the mixture was diluted with water (500ml) and ethyl acetate (600 ml). The organic layer was separated and the aqueous layer was extracted with ethyl acetate (3X 500 ml). The combined organic layers were neutralized by careful addition of an excess of saturated aqueous sodium bicarbonate, washed with brine (300ml), dried (magnesium sulfate) and concentrated. The residue was dissolved in toluene (500ml) and water (20ml, 1.11mol, 0.8 eq) was added. The solution was stirred for approximately 30s and cooled in an ice/water bath with periodic stirring. After 4 hours, the solid was filtered and left to air dry in the petri dish for 16 hours to give polymorph form I as a coloured crystal (118.22 g). Recrystallization from toluene gave polymorph form I as a white tablet (93g, 35%). Test values C67.44, H8.22%; c₁₁H₁₆O₃Theoretical C67.32, H8.22%. IR data (v)_max/cm^-1): 3199.2br, 2963.8s, 2863.5s, 1624.2m, 1604.7s, 1528.3s, 1457.3s, 1395.3s, 1349.7w, 1287.4m, 1265.2s, 1228.0s, 1179.4m, 1166.9m, 1108.1s, 977.8s, 826.5s, 749.1m, 723.9m, 703.8m and 627.9 m.

An X-ray powder diffraction pattern was generated using a sample of 4-cyclopentyl resorcinol monohydrate produced as described above. The results of this test are depicted in fig. I, II and fig. III (batch # 1).

Example II

Single crystals of polymorph form I were obtained by recrystallization from isopropanol and water. The structure of this form I polymorph single crystal is reported in table I above.

Example III

The following protocol describes an alternative method for preparing 4-cyclopentyl resorcinol monohydrate. Toluene and water were used as extraction solvents.

To be full of N₂The pressure reactor of (2) was charged with resorcinol (44.0g, 0.40mol), cyclopentanol (44mL, 0.49mol) and 85% H₃PO₄Aqueous solution (55mL, 0.80 mol). Heating the slurry to 95-120 deg.C for 6-18 hr. The pink reaction mixture was cooled to-70 ℃ and diluted with water (50mL) and toluene (200 mL). The layers were separated at 60 + -5 deg.C. The bottom orange aqueous layer was discarded. The remaining pink organic layer was extracted with 2X 50mL of water at 60. + -. 5 ℃ and then stirred with carbon (5g) at 60. + -. 5 ℃ for 1-2 hours. The slurry was filtered hot through Supercel and the filter cake was rinsed with hot toluene (50 ml). The orange filtrate was diluted with 5mL of water and cooled to-30 ℃ at which time the product crystallized. The slurry was cooled to 0-5 deg.C and the product was collected, washed with cold toluene (40mL) and dried on a funnel by suction to give 41g of 4-cyclopentyl resorcinol monohydrate as a white to pale pink solid (. gtoreq.98% area, HPLC). If desired, the material can be recrystallized from hot toluene (5mL/g) and filtered (carbon (10% by weight) filtration) through Supercel and/or silica gel, carbon (10 wt%) to give 4-cyclopentyl resorcinol form I polymorph as a white solid (. gtoreq.99.7%, HPLC).

The powder X-ray diffraction patterns of three (3) different batches of polymorph form I produced according to the protocol described above were determined. FIG. III depicts this data (lots # 2-4).

Example IV

The water content of the 4-cyclopentyl resorcinol monohydrate produced as in example 1 was determined by Karl Fischer analysis. These analyses were performed in the following manner:

i) coulomb Karl Fischer (KF) analysis

The analysis was performed using a Mettler Toledo DL39 Karl Fischer titrator. Approximately 15-20mg of sample was placed in a KF titration vessel containing Hydranal-Coulomat AD and mixed for 10 seconds to ensure dissolution. Then by means of electrochemical oxidation: 2I- ═ I₂+2e generator electrode (generator electrode) producing iodine titrates the sample. The assay was repeated three times to ensure reproducibility.

ii) volumetric Karl Fischer (KF) analysis for water determination

The analysis was performed using a Mettler Toledo DL38 Karl Fischer titrator. Approximately 10-20mg of the sample was placed in a KF titration vessel containing Hydranal absolute methanol and mixed for 10 seconds to ensure dissolution. The sample was then titrated with Hydranal Composite 5 to the appropriate endpoint. The assay was repeated twice to ensure reproducibility. Titrants were standardized using Hydranal Water Standard 10.0.

The following results were obtained:

TABLE II

Method of producing a composite material	Run 1% HO	Run 2% HO	Run 3% HO	Average% HO	Average number of moles HO
						Volumetric method	8.95	9.04	-	9.00	1.08
Volumetric method	9.22	9.80	-	9.51	1.15
						Coulomb method	8.12	8.21	8.58	8.30	0.99

The data in Table II illustrate that applicants have produced monohydrate 4-cyclopentyl resorcinol.

Claims (6)

1.4-cyclopentyl resorcinol monohydrate.

A crystalline polymorph of 4-cyclopentyl resorcinol monohydrate that exhibits a single crystal X-ray crystallographic analysis at 160K with crystal unit cell parameters equal to:

TABLE I

Space group and unit cell parameters for polymorph form I

3. Use of a compound according to claim 1 for the manufacture of a medicament for reducing pigmentation.

4. Use of the crystalline polymorph of claim 2 in the manufacture of a medicament for reducing pigmentation.

5. A pharmaceutical formulation comprising an effective amount of a compound of claim 1 in admixture with at least one pharmaceutically acceptable carrier.

6. A pharmaceutical formulation comprising an effective amount of the crystalline polymorph of claim 2 in admixture with at least one pharmaceutically acceptable carrier.