WO2009099410A1

WO2009099410A1 - Preparation and use of (r)-8-keto-methylnaltrexone

Info

Publication number: WO2009099410A1
Application number: PCT/US2008/001659
Authority: WO
Inventors: Alfred A. Avey; Julio Perez
Original assignee: Progenics Pharmaceuticals Inc
Current assignee: Progenics Pharmaceuticals Inc
Priority date: 2008-02-06
Filing date: 2008-02-06
Publication date: 2009-08-13
Anticipated expiration: 2010-08-06

Abstract

This invention relates to synthesis of (R)-8-keto-MNTX, as shown in Formula (I), and related methods and products.

Description

PREPARATION AND USE OF (R)-8-KETO-METHYLNALTREXONE

Field of the Invention

The present invention relates to the preparation of (i?)-8-keto-methylnaltrexone and related methods and products.

Background of the Invention

Methylnaltrexone (MNTX) is a quaternary derivative of the opioid antagonist, naltrexone. MNTX exists as a salt, for example, a bromide salt. The bromide salt of MNTX is also known in the literature as: Methylnaltrexone bromide; N- Methylnaltrexone bromide; Naltrexone methobromide; Naltrexone methyl bromide; MRZ 2663BR. MNTX was first reported in the mid-70s by Goldberg et al. as described in U.S. Patent No. 4,176,186. It is believed that addition of the methyl group to the ring nitrogen of naltrexone forms a charged compound with greater polarity and less liposolubility than naltrexone, preventing MNTX from crossing the blood-brain barrier in humans. As a consequence, MNTX exerts its effects in the periphery rather than in the central nervous system with the advantage that it does not counteract the analgesic effects of opioids on the central nervous system.

MNTX is a chiral molecule, wherein the quaternary nitrogen can possess either the (R) or (S) configuration. For example, (iϊ)-MNTX refers to a molecule of MNTX having (R) stereochemistry at the quaternary nitrogen, while (S)-MNTX refers to a molecule of MNTX having (S) stereochemistry at the quaternary nitrogen. All of the reported functions of MNTX described in the literature indicate it is a peripheral opioid antagonist. Some of these antagonist functions are described in U.S. Patents 4,176,186, 4,719,215, 4,861,781, 5,102,887, 5,972,954, 6,274,591, 6,559,158, and 6,608,075, and in U.S. patent application publication numbers 2003/0022909A1, 20040266806, 20040259899 and 20050004155. These uses include reducing the side-effects of opioids without reducing the analgesic effect of opioids. Such side-effects include nausea, emesis, dysphoria, pruritus, urinary retention, bowel hypomotility, constipation, gastric hypomotility, delayed gastric emptying and immune suppression. The art discloses that MNTX not only reduces the side-effects stemming from opioid analgesic treatment but also reduces the side-effects mediated by endogenous opioids alone (or in conjunction with exogenous opioid treatment) such as gastrointestinal dysfunction including inhibition of gastric emptying, constipation, inhibition of gastrointestinal motility from any cause such as surgery, inflammation or excessive vagal stimulation and other such conditions including, but not limited to, those mentioned above. However, it is unclear from the art whether the MNTX used in these studies was a mixture of (R) and (S) stereoisomers or a single stereoisomer.

The art suggests that isolated stereoisomers of a compound sometimes may have contrasting physical and functional properties, although it is unpredictable in any particular circumstance. Quaternary narcotic antagonists exhibit such contrasting physical and functional properties, making it important to develop procedures to isolate and identify MNTX as pure (i?)-MNTX or (S)-MNTX. Goldberg, et al.'s U.S. Patent No. 4,176,186, and more recently Cantrell, et al.'s WO 2004/043964 A2 describe a protocol for the synthesis of MNTX involving the quaternization of a tertiary N- substituted morphinan alkaloid with a methylating agent. However, both Goldberg et al and Cantrell εt a! remain silent as to the sterεoisornεr(s) produced by the synthesis. Based on the method of synthesis described in each, it is unknown whether the MNTX so produced was either (R) or (S) or a mixture of both. Furthermore, (S)-N- methylnaltrexone ((.S)-MNTX), in pure form, and a method of making pure (S)-MNTX had not been described in the literature. Therefore, researchers would have been unable to definitively characterize and distinguish the stereoisomer(s) obtained by the Goldberg et al or Cantrell et al synthesis in the absence of pure (S)-MNTX as a standard.

In addition to the isolation and characterization of each stereoisomer of quaternary narcotic antagonists, it can be desirable to isolate the antagonists from any additional impurities in the manufacture of a pharmaceutical composition. Generally, pharmaceutical compositions also require a high level of purity to meet regulated standards for drug quality and purity. For example, in the synthesis of MNTX as described above, impurities are often formed, including degradants or by-products of manufacture, which may hinder the therapeutic effects of MNTX and/or may be toxic if present in high enough quantity. As such, it is desirable to have the ability to determine both the stereochemical configuration and the purity of MNTX. To do this, it is important to identify, isolate, and chemically characterize impurities, which can be used in chromatographic procedures as standards to confirm the purity of MNTX. Summary of the Invention

The present invention relates to the identification, purification, and synthesis of an impurity of (7?)-MNTX. It has been discovered that this compound can arise as an impurity that is a degradant when certain solutions of (7?)-MNTX are stored under certain conditions. The compound is (i?)-8-keto-methylnaltrexone. This compound is a salt, and will therefore have a counterion. This compound can also exist as a zwitterion. Accordingly, to one aspect of the invention, there is provided a compound of Formula (I),

I and tautomers thereof, wherein X is a counterion. In some embodiments, X can be halide, sulfate, phosphate, sulfonate, nitrate, carboxylate or an anionic-charged organic species. In a particular embodiment, X is bromide. In some embodiments, the compound is isolated with at least 0.5% purity, or at least 1% purity, or at least 5% purity, or at least 10% purity, or at least 15% purity, or at least 25% purity, or at least 50%, at least 75% purity, at least 95% purity, or, more preferably, at least 97% purity. In some embodiments, the compound is a solid. In other embodiments, the compound is packaged as a solution in a sealed bottle. In one embodiment, the compound is packaged as a solution in sealed bottle having a septum.

According to another aspect of the invention, there are provided kits including a compound of Formula (I) and indicia in or on the kit indicating that the compound is present in the kit. In some embodiments, the indicia indicates the purity of the compound. In some embodiments, the indicia indicates the chemical structure of the compound by providing a chemical formula or a structural drawing. In some embodiments, the kit includes a second compound that is (7?)-methylnaltrexone, (S)- methy lnaltrexone . Another aspect of the present invention provides methods for synthesizing (i?)-8- keto-methylnaltrexone comprising treating methylnaltrexone with an aqueous solution comprising a salt to form a compound of Formula (I), or tautomers thereof, and isolating the compound. In some embodiments, the treating comprises addition of a solution comprising a salt to a solution of methylnaltrexone, wherein the salt is, for example, sodium chloride. In some embodiments, the compound is isolated as a solid. In some embodiments, the method further comprises isolating the compound as a solid. In some embodiments, the compound is isolated with at least 0.5% purity, or at least 1% purity, or at least 5% purity, or at least 10% purity, or at least 15% purity, or at least 25% purity, or at least 50%, at least 75% purity, or, more preferably, at least 90% purity.

The present invention also provides methods for determining an impurity, comprising injecting a reference solution comprising a compound of Formula (I) into an HPLC column under a set of conditions to obtain a first HPLC chromatogram, wherein the amount and/or chemical identity of the compound present in the reference solution is known; injecting a sample solution comprising methylnaltrexone into the HPLC column under said set of conditions to obtain a second HPLC chromatogram; and determining the presence and/or the amount of the compound in the sample solution. In some embodiments, the reference solution is injected multiple times. In some embodiments, the determining comprises comparing retention times of peaks in the first HPLC chromatogram and peaks in the second HPLC chromatogram to determine the presence of the compound in the sample solution. In other embodiments, the determining comprises quantifying peak areas of the sample solution and peak areas of the reference solution on the HPLC chromatograms and estimating from these the amount of the compound in the sample solution. The sample solution can comprise (R)- methylnaltrexone and/or (5)-methylnaltrexone, or a mixture of (7?)-methylnaltrexone and (5)-methylnaltrexone. In some embodiments, the HPLC column is a reverse phase column and the column is eluted using a mobile phase comprising water, methanol, trifluoroacetic acid, or mixtures thereof.

The present invention also provides methods for determining an impurity in a material consisting essentially of methylnaltrexone, comprising injecting into an HPLC column, in a single or series of injections, a sample solution containing the material and spiked with a reference compound having a known chemical structure of Formula (I); obtaining an HPLC chromatogram; and determining the presence and/or the amount of the compound in the material. The sample solution can comprise (7?)-methylnaltrexone, or a mixture of (Λ)-methylnaltrexone and (5)-methylnaltrexone. In some embodiments, the HPLC column is a reverse phase column and the column is eluted using a mobile phase comprising water, methanol, trifluoroacetic acid, or mixtures thereof. The method may further comprise documenting in a written form the chemical identity of the compound and the amount of the compound as an impurity.

The present invention also provides methods for determining an impurity in a material consisting essentially of methylnaltrexone, comprising injecting, in a single or series of injections, a solution in which the material is dissolved into an HPLC column and obtaining an HPLC chromatogram; determining the amount in the material of a compound known to have the structure of Formula (I); and documenting in a written form the chemical identity of the compound and the amount of the compound as an impurity in the material. In some cases, the amount in the material of the compound is determined by (i) identifying a peak on the chromatogram that corresponds to a peak on a control chromatogram of a compound known to have the structure of Formula (I), (ii) identifying a peak on the chromatogram that corresponds to a relative retention time of a compound known to have the structure of Formula (I), and/or (iii) identifying a peak on the chromatogram that corresponds that corresponds to a known amount of a spike of the compound known to have the structure of Formula (I). The sample solution can comprise (i?)-methylnaltrexone, or a mixture of (R)- methylnaltrexone and (5)-methylnaltrexone. In some embodiments, the HPLC column is a reverse phase column and the column is eluted using a mobile phase comprising water, methanol, trifluoroacetic acid, or mixtures thereof.

Brief Description of the Drawings

FIG. 1 shows the synthesis of (i?)-8-keto-MNTX, according to one embodiment of the invention.

FIG. 2 shows an HPLC chromatogram of (a) the diketone tautomer of (i?)-8-keto- MNTX, (b) the enol tautomer of (7?)-8-keto-MNTX, and (c) (Λ)-MNTX, upon reverse phase chromatography.

FIG. 3 shows an HPLC chromatogram of (a) the diketone tautomer of (Λ)-8-keto- MNTX, (b) the enol tautomer of (/?)-8-keto-MNTX, and (c) (Λ)-MNTX, upon ion exchange chromatography. FIG. 4 shows an HPLC chromatogram of (a) the diketone tautomer of (/?)-8-keto- MNTX, (b) the enol tautomer of (J?)-8-keto-MNTX, and (c) CK)-MNTX, upon recrystallization.

FIG. 5 A shows the UVspectrum of the keto tautomer of (7?)-8-keto-MNTX.

FIG. 5B shows the UV spectrum of the enol tautomer of (i?)-8-keto-MNTX.

FIG. 6 shows the UV spectrum of a 1 :1 solution of 1,3 cyclohexadione and naltrexone.

FIG. 7 shows the proton NMR of (7?)-8-keto-MNTX.

FIG. 8 shows the proton NMR of a sample of (i?)-8-keto-MNTX that has been stored for four days.

FIG. 9 shows the IR spectrum of (i?)-8-keto-MNTX after four days.

FIG. 10 shows the MS peaks observed for (7?)-8-keto-MNTX.

Detailed Description

The present invention provides synthetic routes for the synthesis of (i?)-8-keto- methylnaltrexone ((/?)-8-keto-MNTX), and related products and methods. (Λ)-8-Keto-MNTX has the structure in Formula (I):

I or tautomers thereof, wherein X is a counterion. The counterion can be, for example, a halide (e.g., iodide, bromide, chloride or fluoride), sulfate, phosphate, nitrate, or an anionic-charged organic species (e.g., sulfonates, such as mesylate, besylate, tosylate, triflate, and the like, carboxylates, such as formate, acetate, citrate, fumarate, and the like). In one embodiment, the counterion is a halide, such as bromide.

(Λ)-8-keto-MNTX may exist in two different tautomeric forms, diketone and enol, as shown in Equation (1).

diketone

The present invention provides methods for synthesizing (7?)-8-keto-MNTX. FIG. 1 shows the synthesis of (i?)-8-keto-MNTX, according to one embodiment of the invention, wherein (i?)-MNTX is treated with an aqueous solution comprising a salt to convert (.K)-MNTX to (7?)-8-keto-MNTX. The salt may be any salt known to those of ordinary skill in the art, including alkali salts such as NaCl or KCl, other inorganic salts, organic salts, and the like. In one embodiment, the salt is NaCl. The aqueous solution may contain 50%, 40%, 30%, more preferably, 10% salt.

Because 8-keto-MNTX contains a chiral quaternary amine, it may possess two different stereochemical configurations (e.g., (i?)-8-keto-MNTX or (5)-8-keto-MNTX) based on the stereochemistry at the quaternary amine of the MNTX molecule utilized as starting material. Methods described herein utilize MNTX with at least 99.5% in the (R) configuration as the starting material, such that the (/?)-8-keto-MNTX isomer is the major isomer formed. While (S)-8-keto-MNTX may possibly be formed, it would be present in very low or negligible quantities. The synthesis of pure (Λ)-MNTX is described in U.S. Patent Application No. 11/441,395, filed May 25, 2006, Publication Number US 2007-0099946 Al. The stereochemical configuration of (^)-MNTX may be determined by using pure (S)-MNTX as a reference standard. The synthesis of pure (S)- MNTX is described in U.S. Patent Application No. 11/441,452, filed May 25, 2006, Publication Number US 2007-0265293 Al. (Λ)-MNTX has been found to be an opioid antagonist, while (S)-MNTX is a partial opioid agonist.

The method can further involve purification of (i?)-8-keto-MNTX using at least one purification technique, such as chromatography or crystallization. The chromatography can be reverse-phase chromatography, regular-phase chromatography, and/or ion exchange chromatography. In some embodiments, the regular-phase chromatography may involve use of an alumina or silica gel column. In some cases, methods of the invention may involve use of a reverse phase HPLC column followed by ion exchange chromatography. For reverse phase chromatography, the HPLC column may be eluted using a mobile phase comprising water, methanol, trifluoroacetic acid, or mixtures thereof. The crystallization solvent can be an organic solvent, a mixture of organic solvents, or a mixture of organic solvent(s) plus water. A preferred solvent can be an alcohol, such as methanol. Methods for both chromatography and crystallization are known to those of ordinary skill in the art.

In some cases, the HPLC column is a reverse phase column and the column is eluted using a mobile phase comprising water, methanol, trifluoroacetic acid, or mixtures thereof.

In some embodiments, multiple crystallizations are performed in order to isolate (/?)-8-keto-MNTX. For example, a sample containing a mixture of (/J)-MNTX and (R)- 8-keto-MNTX may be treated (e.g., concentrated in solution) such that crystallization of (/?)-MNTX occurs. The crystallized (iϊ)-MNTX may be removed, resulting in a mother liquor having a higher concentration of (i?)-8-keto-MNTX. This crystallization process may be repeated multiple times to afford a mother liquor that is enriched in (i?)-8-keto- MNTX. In one embodiment, (J?)-8-keto-MNTX is obtained in 90% purity. It should be understood that one of ordinary skill in the art can optimize the methodologies described herein to obtain higher purity and/or higher yield of (i?)-8-keto-MNTX, in which the crystallized product is greater than 99%, or even greater than 99.9%, pure. The relative retention times for (Λ)-8-keto-MNTX, (R)-MNTX and (S)-MNTX are 0.49, 1.00, and 0.89, respectively, as determined by HPLC Method B, described herein.

Where purity of a solid is concerned, purity is the weight percent of the (R)-S- keto-MNTX versus that of other compounds in the solid. Where purity of a solution is concerned, purity is the weight percent of the (/?)-8-keto-MNTX versus that of other compounds in the solution. Purity may be determined by HPLC, wherein the percent purity of a compound refers to the weight percent of the compound present in solution relative to the weight percent of other compounds present in the same solution.

The present invention may advantageously provide (i?)-8-keto-MNTX in sufficient purity in order to enable its use as a reference or standard in various analytical methods (e.g., HPLC), as described more fully below. In some embodiments, (R)-S- keto-MNTX may be isolated with at least 0.5% purity, at least 1% purity, at least 5% purity, at least 10% purity, at least 15% purity, at least 25% purity, at least 50% purity, at least 75% purity, or, more preferably, with at least 90% purity. In some embodiments, (/?)-8-keto-MNTX may be isolated and/or packaged as a solid. In some embodiments, (/?)-8-keto-MNTX can be packaged as a solution in a container such as a sealed bottle or vial, or a sealed bottle comprising a septum.

In another aspect, the present invention provides methods for determining the presence and/or amount of (/?)-8-keto-MNTX in a sample (e.g., a sample comprising (R)- MNTX). For example, (i?)-8-keto-MNTX may be formed as an impurity during the synthesis of (i?)-MNTX. As used herein, the term "impurity" may refer to degradants which arise during storage of (i?)-MNTX and/or by-products formed in a chemical reaction for manufacturing of (i?)-MNTX. In one embodiment, the method comprises injecting a reference solution comprising (i?)-8-keto-MNTX into an HPLC column under a set of conditions to obtain a first HPLC chromatogram wherein the amount and/or chemical identity of (7?)-8-keto-MNTX present in the reference solution is known, injecting a sample solution comprising MNTX into the HPLC column under the same set of conditions to obtain a second HPLC chromatogram, and comparing the first HPLC chromatogram with the second HPLC chromatogram to determine the presence and/or amount of the impurity. The reference solution may be formed by dissolving a sample (e.g., solid sample) of (i?)-8-keto-MNTX in a first solvent, and the sample solution may be formed by dissolving a solid sample in a second solvent. In some embodiments, the reference solution may contain an additional compound(s), wherein the amount and/or identity of the additional compound(s) is also known. In one embodiment, the sample (e.g., sample solution) may comprise (i?)-MNTX. It should be understood that the invention may encompass other samples suspected of containing (i?)-8-keto-MNTX.

In one embodiment, the presence of (i?)-8-keto-MNTX in the sample solution may be determined by comparing retention times of peaks in the first HPLC chromatogram with the retention times of peaks in the second HPLC chromatogram. For example, the standard solution comprising (Λ)-8-keto-MNTX may produce a chromatogram with a peak corresponding to (/?)-8-keto-MNTX and having a particular retention time. A sample solution may then be injected into the HPLC column under the same conditions as the standard solution, and the resulting chromatogram may be studied to determine if a peak exists at the same retention time as the peak corresponding to (R)- 8-keto-MNTX in the HPLC chromatogram of the standard solution. The existence of such a peak can indicate that (i?)-8-keto-MNTX is present in the sample. In another embodiment, the amount of (i?)-8-keto-MNTX in the sample solution may be determined by comparing the area of peaks in the first HPLC chromatogram with the area of peaks in the second HPLC chromatogram, and calculating from these the content of (7?)-8-keto- MNTX in the sample solution.

In some embodiments, the present invention provides methods for determining an impurity in a material consisting essentially of methylnaltrexone, where a sample solution containing the material and spiked with a reference compound having a known chemical structure of Formula (I), as described herein, is injected into an HPLC column and an HPLC chromatogram is obtained to determine the presence and/or the amount of the compound in the material.

Methods of the invention may further comprise, documenting in a written form the chemical identity of the compound and the amount of the compound as an impurity in the material.

In other embodiments, the present invention provides methods for determining an impurity in a material consisting essentially of methylnaltrexone, wherein a solution in which the material is dissolved is injected into an HPLC column and an HPLC chromatogram is obtained to determine the amount in the material of a compound known to have the structure of Formula (I), as described herein. The chemical identity of the compound and the amount of the compound as an impurity in the material may then be documented, in a written form. The amount in the material of the compound may be determined by (i) identifying a peak on the chromatogram that corresponds to a peak on a control chromatogram, (ii) identifying a peak on the chromatogram that corresponds to a relative retention time of a compound known to have the structure of Formula (I), and/or (iii) identifying a peak on the chromatogram that corresponds that corresponds to a known amount of a spike of the compound known to have the structure of Formula (I). Some embodiments of the invention may be useful in determining the amount and/or presence of (i?)-8-keto-MNTX in a sample comprising (.K)-MNTX. The sample may be a sample of freshly manufactured material or the sample may be one stored for a defined period of time. In one embodiment, a sample of (Λ)-MNTX may be stored and periodically analyzed using methods described herein to determine the presence and/or amount of (7?)-8-keto-MNTX in the sample which may have been formed by, for example, degradation of (K)-MNTX. In some cases, the sample may be placed in conditions to intentionally promote degradation (.K)-MNTX, wherein the sample is periodically analyzed using methods described herein to determine the presence and/or amount of (R),(R)- 8-keto-MNTX in the sample. The sample solution may also comprise (/?)-MNTX, (S)-MNTX, or mixtures thereof. In one embodiment, the sample solution comprises (R)- MNTX and/or (S)- MNTX. In another embodiment, the sample solution comprises a mixture of (iϊ)-MNTX and (S)-MNTX.

In another aspect of the invention, kits are provided containing (i?)-8-keto- MNTX. For example, a kit including (i?)-8-keto-MNTX may be provided for the purpose of producing standards for analytical methods, such as HPLC. In one embodiment, the kit may comprise (i?)-8-keto-MNTX and indicia in or on the kit indicating that (i?)-8-keto-MNTX is present in the kit. In some cases, the indicia indicates the purity of (i?)-8-keto-MNTX. The indicia may also indicate the chemical structure of the compound by providing a chemical formula or a structural drawing. It should be understood that the kit may also include additional compounds, similarly identified by the indicia.

In some cases, the kit comprises a solution of (7?)-8-keto-MNTX. In some cases, the kit comprises a solid sample of (/?)-8-keto-MNTX. A "kit," as used herein, typically defines a package or an assembly including one or more of the compositions of the invention, and/or other materials associated with the invention, such as solvents. Each of the compositions of the kit may be provided in liquid form (e.g., in solution), or in solid form. In certain cases, some of the compositions may be constitutable or otherwise processable, for example, by the addition of a suitable solvent or other species, which may or may not be provided with the kit. Examples of other compositions or components associated with the invention include, but are not limited to, solvents, surfactants, diluents, salts, buffers, emulsifiers, chelating agents, antioxidants, drying agents, needles, syringes, packaging materials, tubes, bottles, flasks, beakers, dishes, frits, filters, rings, clamps, wraps, patches, containers, and the like, for example, for using, modifying, assembling, storing, packaging, preparing, mixing, diluting, and/or preserving the compositions components for a particular use, for example, to a sample.

A kit of the invention may, in some cases, include instructions in any form that are provided in connection with the compositions of the invention in such a manner that one of ordinary skill in the art would recognize that the instructions are to be associated with the compositions of the invention. For instance, the instructions may include instructions for the use, modification, mixing, diluting, preserving, assembly, storage, packaging, and/or preparation of the compositions and/or other compositions associated with the kit. In some cases, the instructions may also include instructions for the delivery of the compositions, for example, for a particular use, e.g., to a sample. The instructions may be provided in any form recognizable by one of ordinary skill in the art as a suitable vehicle for containing such instructions, for example, written or published, verbal, audible (e.g., telephonic), digital, optical, visual (e.g., videotape, DVD, etc.) or electronic communications (including Internet or web-based communications), provided in any manner.

As used herein, "instructions" can define a component of instructional utility (e.g., directions, guides, warnings, labels, notes, FAQs or "frequently asked questions," etc.), and typically involve written instructions on or associated with the invention and/or with the packaging of the compounds of the invention. Instructions can also include instructional communications in any form (e.g., oral, electronic, audible, digital, optical, visual, etc.), provided in any manner such that a user will clearly recognize that the instructions are to be associated with compounds of the invention.

Examples Example 1 : Synthesis of IRV8-Keto-MNTX

(i?)-8-Keto-MNTX was synthesized by forced degradation of (Λ)-MNTX, as shown in FIG. 1. The forced degradation was carried out by stirring a solution of (R)- MNTX (20 g) in a 1% solution of sodium chloride (4 g) in water (400 mL) at 50 °C for 11 days under a room air atmosphere. The reaction was then cooled to room temperature and filtered. The concentration of the (i?)-8-keto-MNTX increased to about 1% after 4 days and was not observed to increase substantially after that time.

Example 2: Purification of (/?)- 8-Keto-MNTX

To isolate (Λ)-δ-keto-MNTX, MNTX was recrystallized by concentrating the mother liquor by rotary evaporation until crystallization of (i?)-MNTX commenced, and then allowing the remaining (/J)-MNTX to crystallize overnight. The (/?)-MNTX crystals were filtered, resulting in a mother liquor having a higher concentration of (i?)-8- keto-MNTX. This concentration/crystallization process was repeated two more times to afford approximately 20 mL of mother liquor enriched to approximately 6% (i?)-8-keto- MNTX. Isolation of (i?)-8-keto-MNTX was then accomplished by reverse phase chromatography. The enriched mother liquor was loaded directly onto a 40 M flash Cl 8 reverse phase Biotage column. The column was eluted with 1.2 L of a linear gradient consisting of water:methanol:TFA (100:0:0.1 to 80:20:0.1), and the eluant was collected in 21 mL fractions. The fractions were analyzed by HPLC, and the fractions containing (/?)-8-keto-MNTX were combined. The solvent was removed by rotary evaporation in a 34 °C bath. The resulting residue was reloaded on to the Cl 8 column and purified using 4 L of a linear gradient of water:methanol:TFA (100:0:0.1 to 90:10:0.1) as eluant. Fractions containing (i?)-8-keto-MNTX were combined and the solvent was removed using rotary evaporation to afford ca. 100 mg residue, which contained 90% pure (R)-S- keto-MNTX, as shown by the HPLC chromatogram in FIG. 2. Peak A corresponds to the diketone tautomer of (i?)-8-keto-MNTX (RRT = 2.2); peak B corresponds to the enol tautomer of (/?)-8-keto-MNTX (RRT = 2.6); peak C corresponds to the (Λ)-MNTX (RRT = 3.0), as determined by HPLC Method A, described herein.

The residue was submitted to ion exchange chromatography on a 6 g Bio-Rad AGX8 resin to convert the product to the bromide salt. The ion exchange column was prepared by eluting 100 mL of IN HBr and then distilled water until the pH of the eluant was 6-7. The (i?)-8-keto-MNTX was dissolved in water and loaded onto the ion exchange column and eluted with distilled water. The product was eluted in the first 100 mL, and the water was removed by rotary evaporation at 34 °C to give 70 mg of an amber glass. FIG. 3 shows the HPLC chromatogram of the product after ion exchange chromatography, wherein peaks A, B, and C correspond to the diketone tautomer of (R)- 8-keto-MNTX, enol tautomer of (/?)-8-keto-MNTX, and (tf)-MNTX, respectively. The residue was recrystallized from 0.5 mL of methanol to afford (/?)-8-keto-MNTX as ivory colored crystals (15 mg of approx. 90% pure enol form), as shown by the HPLC chromatogram in FIG. 4, where peaks A, B, and C again correspond to the diketone tautomer of (i?)-8-keto-MNTX, enol tautomer of (i?)-8-keto-MNTX, and (R)-MNTX, respectively.

Example 3: Characterization of (i?)-8-keto-MNTX

As described herein, (/?)-8-keto-MNTX is an unsymmetrical 1,3-diketone which exists in two tautomeric forms, the enol form and diketone form, as shown in Equation

(I)- The product was isolated from crystallization as the enol tautomer while the diketone tautomer was more stable in solution. The crystalline enol tautomer was dissolved in water and isomerized to an equilibrium mixture of the enol and diketone forms, as observed by HPLC. In aqueous solution, the diketone appeared to be favored by a ratio of 6:1, as observed by the absorption spectra. The molecular spectra, NMR, IR, UV -vis, and high resolution mass spectra of the isolated compound were consistent with the enol form of (Λ)-8-keto MNTX.

FIG. 5 A shows the UV spectrum of the Peak A, which exhibits an absorbance peak at about 285 nm, consistent with a dihydromorphinone type structure. In contrast, FIG. 5B shows the UV spectrum of the Peak B, which exhibits an absorbance peak at about 266 nm, consistent with enol form of a 1,3 diketone. Thus, Peak A corresponds to the keto tautomer of (7?)-8-keto-MNTX and Peak B corresponds to the enol tautomer of (i?)-8-keto-MNTX. To further illustrate this correlation, the absorption spectrum of a solution of 1,3 cyclohexadione and naltrexone (1 :1) was measured, as shown by in FIG. 6. In aqueous solution, the diketone appeared to be favored by a ratio of 6:1, as observed by the absorption spectra. Peak C corresponds to (i?)-MNTX.

The NMR (in D₂O) of the isolated 2.5 min peak (enol) shows the characteristic resonance of a vinylic proton at 6.2 ppm, in addition to the C-5 hydrogen for both the diketone at 5.1 and enol at 4.9 PPM. (FIG. 7) These protons washed out over a period of several days, as shown in FIG. 8.

The IR spectrum in a KBr pellet shows absorbances consistent with a mixture of the diketone and enol forms of the 8-keto MNTX. (FIG. 9)

MS shows a parent ion at 370 amu and molecular formula Of C₂₁H₂₄NO₅. (FIG. 10)

HPLC analysis was performed on a Hewlett Packard 1100 series using HPLC Method A, as described herein.

HPLC Method A:

Column: Phenomonex Synergi hydro RP column (C 18, 5 μ, 150 X 4.6 mm)

Flow rate: 1.0 mL/min.

Column temperature: 40 °C.

Detector: diode array detector monitoring @ 220 and 210nm. Elution: isocratic. 60% water, 30% buffer (700 ml of water, 300 mL methanol, 3 mL triethylamine and sufficient phosphoric acid to give a pH of 3.4), 10% methanol.

HPLC Method B:

HPLC Method B is described as "HPLC Method II" in Publication Number US 2007-0099946 Al.

What is claimed:

Claims

1. A compound of Formula (I),

I or tautomers thereof, wherein X is a counterion.

2. The compound of claim 1, wherein X is halide, sulfate, phosphate, sulfonate, nitrate, carboxylate, or an anionic-charged organic species.

3. The compound of claim 1, wherein X is bromide.

4. The compound of claim 1, wherein the compound is isolated with at least 0.5% purity.

5. The compound of claim 1, wherein the compound is isolated with at least 1% purity.

6. The compound of claim 1 , wherein the compound is isolated with at least 5% purity.

7. The compound of claim 1, wherein the compound is isolated with at least 10% purity.

8. The compound of claim 1, wherein the compound is isolated with at least 15% purity.

9. The compound of claim 1 , wherein the compound is isolated with at least 25% purity.

10. The compound of claim 1, wherein the compound is isolated with at least 50% purity.

11. The compound of claim 1 , wherein the compound is isolated with at least 75% purity.

12. The compound of claim 1, wherein the compound is isolated with at least 90% purity.

13. The compound of claims 1-12, wherein the compound is a solid.

14. The compound of claim 1, wherein the compound is packaged as a solution in a sealed bottle.

15. The compound of claim 1 , wherein the compound is packaged as a solution in a sealed bottle comprising a septum.

16. A kit, comprising: a compound of Formula (I),

or tautomers thereof, wherein X is a counterion, and indicia in or on the kit indicating that the compound is present in the kit.

17. The kit of claim 16, wherein X is halide, sulfate, phosphate, sulfonate, nitrate, carboxylate or an anionic-charged organic species.

18. The kit of claim 16, wherein X is bromide.

19. The kit of claim 16, wherein the indicia indicates the purity of the compound.

20. The kit of claim 16, wherein the indicia indicates the chemical structure of the compound by providing a chemical formula or a structural drawing.

21. A method of synthesizing (i?)-8-keto-methylnaltrexone, comprising: treating methylnaltrexone with an aqueous solution comprising a salt to form a compound of Formula (I),

I or tautomers thereof, wherein X is a counterion; and isolating the compound.

22. The method of claim 21, wherein X is halide, sulfate, phosphate, sulfonate, nitrate, carboxylate, or an anionic-charged organic species.

23. The method of claim 21 , wherein X is bromide.

24. The method of claim 21 , wherein the treating comprises addition of a solution comprising a salt to a solution of methylnaltrexone.

25. The method of claim 24, wherein the salt is sodium chloride.

26. The method of claim 21, wherein the compound is isolated as a solid.

27. The method of claim 21, wherein the compound is isolated with at least 0.5% purity.

28. The method of claim 21, wherein the compound is isolated with at least 1% purity.

29. The method of claim 21, wherein the compound is isolated with at least 5% purity.

30. The method of claim 21 , wherein the compound is isolated with at least 10% purity.

31. The method of claim 21 , wherein the compound is isolated with at least 15% purity.

32. The method of claim 21, wherein the compound is isolated with at least 25% purity.

33. The method of claim 21, wherein the compound is isolated with at least 50% purity.

34. The method of claim 21, wherein the compound is isolated with at least 75% purity.

35. The method of claim 21, wherein the compound is isolated with at least 90% purity.

36. A method for determining an impurity, comprising: injecting a reference solution comprising a compound of Formula (I),

I or tautomers thereof, wherein X is a counterion, into an HPLC column under a set of conditions to obtain a first HPLC chromatogram, wherein the amount and/or chemical identity of compound present in the reference solution is known; injecting a sample solution comprising methylnaltrexone into the HPLC column under said set of conditions to obtain a second HPLC chromatogram; and determining the presence and/or the amount of the compound in the sample solution.

37. The method of claim 36, wherein X is halide, sulfate, phosphate, sulfonate, nitrate, carboxylate, or an anionic-charged organic species.

38. The method of claim 36, wherein X is bromide.

39. The method of claim 36, wherein the determining comprises comparing retention times of peaks in the first HPLC chromatogram and peaks in the second HPLC chromatogram to determine the presence of the compound in the sample solution.

40. The method of claim 36, wherein the determining comprises quantifying peak areas of the sample solution and peak areas of the reference solution on the HPLC chromatograms and estimating from these the amount of the compound in the sample solution.

41. The method of claim 36, wherein the sample solution comprises (R)- methylnaltrexone and/or (5)-methylnaltrexone.

42. The method of claim 36, wherein the sample solution comprises a mixture of (R)- methylnaltrexone and (iS)-methylnaltrexone.

43. The method of claim 36, wherein the HPLC column is a reverse phase column and the column is eluted using a mobile phase comprising water, methanol, trifluoroacetic acid, or mixtures thereof.

44. The method of claim 36, further comprising documenting in a written form the chemical identity of the compound and the amount of the compound as an impurity.

45. A method for determining an impurity in a material consisting essentially of methylnaltrexone, comprising: injecting into an HPLC column a sample solution containing the material and spiked with a reference compound having a known chemical structure of Formula (I),

I wherein X is a counterion; obtaining an HPLC chromatogram; and determining the presence and/or the amount of the compound in the material.

46. The method of claim 45, wherein X is halide, sulfate, phosphate, sulfonate, nitrate, carboxylate, or an anionic-charged organic species.

47. The method of claim 45, wherein X is bromide.

48. The method of claim 45, wherein the sample solution comprises (R)- methy lnaltrexone .

49. The method of claim 45, wherein the sample solution comprises a mixture of (R)- methylnaltrexone and (5)-methylnaltrexone.

50. The method of claim 45, wherein the HPLC column is a reverse phase column and the column is eluted using a mobile phase comprising water, methanol, trifluoroacetic acid, or mixtures thereof.

51. The method of claim 45, further comprising documenting in a written form the chemical identity of the compound and the amount of the compound as an impurity.

52. A method for determining an impurity in a material consisting essentially of methylnaltrexone, comprising: injecting a solution in which the material is dissolved into an HPLC column and obtaining an HPLC chromatogram; determining the amount in the material of a compound known to have the structure of Formula (I),

I wherein X is a counterion, and documenting in a written form the chemical identity of the compound and the amount of the compound as an impurity in the material.

53. The method of claim 52, wherein the amount in the material of the compound is determined by (i) identifying a peak on the chromatogram that corresponds to a peak on a control chromatogram, (ii) identifying a peak on the chromatogram that corresponds to a relative retention time of a compound known to have the structure of Formula (I), and/or (iii) identifying a peak on the chromatogram that corresponds that corresponds to a known amount of a spike of the compound known to have the structure of Formula (I).

54. The method of claim 52, wherein X is halide, sulfate, phosphate, sulfonate, nitrate, carboxylate, or an anionic-charged organic species.

55. The method of claim 52, wherein X is bromide.

56. The method of claim 52, wherein the sample solution comprises (R)- methylnaltrexone .

57. The method of claim 52, wherein the sample solution comprises a mixture of (R)- methylnaltrexone and (S)-methylnaltrexone.

58. The method of claim 52, wherein the HPLC column is a reverse phase column and the column is eluted using a mobile phase comprising water, methanol, trifluoroacetic acid, or mixtures thereof.