Purification of rifabutin formulations

Abstract

The invention provides compositions and methods of creating a rifabutin powder suitable for IV formulation preparation.

Description

[0039] Solvent selection

[0040] Using the same silica and ratio determined to be most suitable in the previous experiment (90 A pore size, 75-315pm diameter and a rifabutin to silica ratio of 1:4) binding of rifabutin was also tested with other alkane solvents. As shown in FIG. 5, binding with n-hexane and n-heptane worked similar to n-pentane, while cyclohexane is less suitable as rifabutin does not completely bind to the silica.

[0041] Method 2 lab scale (FIG. 6)

[0042] Rifabutin is mixed with perlite in a 1:2 ratio and packed into a glass column. A second column is packed with silica gel, using a silica to rifabutin ratio of about 1:5. Both columns contain a glass frit onto which a layer of sand and perlite is added prior to packing, in order to retain the undissolved rifabutin or silica gel respectivley. An alkane solvent is added to column 1. While running through the column, the alkane solvent dissolves the rifabutin. The alkane solvent containing rifabutin is collected and transferrred to the second column. While the solution runs through column 2 rifabutin adsorbs to the silica. The alkane solvent depleted of rifabutin is subssequently added back to column 1. These steps are repeated until all rifabutin is adsorbed to the silica in column 2 and only insoluble impurities remain in column 1. Rifabutin is then eluted from column 2 using acetone. In a last step the solvent is evaporated to obtain purified (IV-grade) rifabutin.

[0043] Method 2 large scale (FIG. 7)

[0044] Rifabutin and an alkane solvent are added into a reactor (A) and mixed with a mechanical stirrer (B) to dissolve the rifabutin. The solution flows through a filter in the bottom of the reactor (C), retaining undissolved rifabutin, and an additional filter D. Using a pump (E) the solution is pumped through a column (F) filled with silica gel. Rifabutin is adsorbed on the silica gel in the column. The alkane solvent depleted of rifabutin flows through tubing (G) back to the reactor (A). The alkane solvent is circulated through the system until all rifabutin is adsorbed to the silica gel in the column (F) and only insoluble impurities suspended in the alkane solvent remain in the reactor (A). After the extraction phase, the rifabutin is eluted from the coulmn (F) using acetone. In a last step the solvent is evaporated at reduced pressure to obtain purified (IV-grade) rifabutin. Example 2

[0045] 10g of rifabutin and 20g of perlite were mixed and filled into a glass column (column 1). A Second column was packed with 55g of silica gel (90A, 75-315pm). The bottom of the glass columns contained a glass frit onto which sand and additional perlite was added to retain the undissolved rifabutin or silica gel respectivley.

[0046] Pentane was ran through column 1 to dissolve rifabutin and collected in a beaker. After approx. 200mL of pentane were collected the rifabutin in pentane solution was ran through column 2. Rifbautin is adsorbed on the silica gel and the almost colorless pentane was collected in a beaker before then again being transferred to column 1 to dissolve additional rifabutin. These steps were repeated about 20 times, until all rifbutin was tranferred from column 1 to column 2 where it was adsorbed to the silica gel and only insoluble impurities remained in column 1.

[0047] Rifabutin was then eluted from column 2 using approximately 150mL of acetone. The solvent was evaporated under vacuum to obtain purified (IV-grade) rifabutin with a yield of 97.5%.

[0048] HPLC analysis shows an overall increase in purity of about 2% after purification (Table 4). As shown by the results from the sub-visible particle test (Table 5), sub-visible particle forming impurities are in the acceptable range. Table 4: HPLC results, Method 2, Example 2 Peak RRT % Area Impurity original purified 1 0.47 0.1 - 16-desacetylrifabutin 2 0.49 0.1 0.08 3 0.55 0.16 0.15 Hydroxy rifabutin 4 0.62 0.05 - 5 0.72 0.06 - 6 0.78 0.12 - 7 0.84 0.12 0.06 8 0.89 0.13 0.07 9 0.92 0.44 0.39 Rifabutin 14 R epimer 10 1.00 96.43 98.49 RBT 11 1.19 0.45 - Rifabutin N-oxide 12 1.58 0.22 0.2 Didehydrorifabutin 13 1.70 0.09 0.09 14 1.89 0.1 - 15 1.98 0.11 - 16 2.00 0.12 - 17 2.03 0.06 - 18 2.06 0.14 - 19 2.08 0.06 - 20 2.10 0.1 - Table 5: Sub-visible particles test results obtained with purified drug substance (Method 2) particles / mL Particle size original purified > 10 pm 1804.0 2.6 > 25 pm 98.7 0.0 Method 3

[0049] Besides using continuous extraction processes, as described in method 1 and method 2, purification can also be achieved by simply dissolving standard grade rifabutin in a sufficient amount of solvent and remove the undissolved impurities by filtration.

[0050] Rifabutin is mixed with one of the alkane solvents in a ratio that allows complete dissolution of rifabutin. The solution is stirred until rifabutin is fully dissolved and subsequently filtered. After filtration the solvent is evaporated to obtain purified (IV-grade) rifabutin powder. Example 3

[0051] 1.5g rifabutin were mixed with 250mL of n-heptane and heated at 50°C under stirring for 1 hour. The solution was filtered through perlite to remove any undissolved impurities. The heptane was removed using a rotary evaporator to obtain purified (IV-grade) rifabutin powder. The yield was 95.3%.

[0052] HPLC analysis shows an overall increase in purity of about 2% after purification (Table 6). As shown by the results from the sub-visible particle test (Table 7), sub-visible particle forming impurities are in the acceptable range. Table 6: HPLC results, Method 3, Example 3 Peak RRT % Area Impurity original purified 1 0.32 0.06 - 2 0.46 0.1 - 16-desacetylrifabutin 3 0.49 0.11 0.09 4 0.55 0.18 0.15 Hydroxy rifabutin 5 0.62 0.07 - 6 0.68 0.07 - 7 0.72 0.07 - 8 0.78 0.16 0.08 9 0.84 0.1 0.08 10 0.88 0.14 0.11 11 0.93 0.46 0.44 Rifabutin 14 R epimer 12 1.00 95.94 98.02 RBT 13 1.19 0.52 0.12 Rifabutin N-oxide 14 1.49 0.25 0.22 15 1.55 0.13 0.10 Didehydrorifabutin 16 1.57 0.05 - 17 1.66 0.14 - 18 1.74 0.11 - 19 1.75 0.14 - 20 1.78 0.08 - 21 1.81 0.14 0.07 22 1.82 0.07 - 23 1.84 0.1 - Table 7: Sub-visible particles test results obtained with purified drug substance (Method 3) Particle size i        > 10 pm i        > 25 pm particles / mL original                         purified 1804.0                             7.1 98.7                              0.2 Testing methods HPLC analysis

[0053] Weigh approx. 10 mg of rifabutin into a clean glass vial. Dissolve in 1 ml acetonitrile and dilute to 1 mg / mL using diluent. Chromatographic system: HPLC Waters Alliance equipped with UV detector or equivalent Software: Empower 3 System of equivalent Column: Intersil ODS-3V 250 x 4.6 mm 5 pm Flow: 1.0 mL / min Diluent: 0.1% Triethylamine / Acetonitrile 8 / 2 pH6.0 with trifluoracetic acid (TFA) Column temperature: 50 °C Sample concentration: 1 mg / mL Dissolution phase: Acetonitrile / diluent 1 / 9 Injection volume: 20 pl Detector UV: 254 nm Analysis time: 90 min Acquisition time: 90 min Mobile phase: Phase A: 0.1% Triethylamine / Acetonitrile 8 / 2 pH2.5 with TFA Phase B: 0.1% Triethylamine / Acetonitrile 1 / 9 pH2.5 with TFA Gradient: Time [min] Phase A [%] Phase B [%] 0 75 25 35 75 25 45 45 55 50 40 60 60 30 70 70 30 70 71 0 100 80 0 100 81 75 25 91 75 25

[0054] As particle forming impurities are not detected by current pharmacopeial analytical methods, a new method was developed to test the drug substance regarding its suitability for the preparation of intravenous formulations. Sub-visible particle test

[0055] About 400mg of rifabutin is dissolved in 2mL of a mixture of isosorbide dimethyl ether (DMI) and acetic acid in water (40% DMI, 2.4% acetic acid). The obtained solution is filtered through a 0.2pm Nylon filter and 1.5mL are added to 23.5mL of 0.9% saline solution to a final concentration of 10 mg / mL. Solutions are stirred for 15min on a roller mixer to allow sub-visible particle formation.

[0056] Sub-visible particles are determined using a suitable instrument (e.g. PAMAS SVSS particle counter). Particle numbers in the lOmg / mL solution should be not more than 12.5 particles / mL greater 10pm and not more than 1.5 particles / mL greater 25pm to pass the test. INCORPORATION BY REFERENCE

[0057] References and citations to other documents, such as patents, patent applications, patent publications, journals, books, papers, web contents, publicly accessible databases, have been made throughout this disclosure. All such documents are hereby incorporated herein by reference in their entirety for all purposes. EQUIVALENTS

[0058] Various modifications of the invention and many further embodiments thereof, in addition to those shown and described herein, will become apparent to those skilled in the art from the full contents of this document, including references to the scientific and patent literature cited herein. The subject matter herein contains important information, exemplification, and guidance that can be adapted to the practice of this invention in its various embodiments and equivalents thereof.

Claims

1. A purified rifabutin powder from which sub-visible particles have been removed.

2. The rifabutin powder of claim 1, wherein a lOmg / mL aqueous solution comprising the rifabutin powder has no more than about 12.5 particles / mL that are greater I Opm in diameter and no more than about 1.5 particles / mL greater than 25pm in diameter.

3. The rifabutin powder of claim 2, wherein the aqueous solution is suitable for intravenous administration.

4. The rifabutin powder of claim 1, wherein the sub-visible particles forming impurities are insoluble in an alkane solvent.

5. The rifabutin powder of claim 4, wherein the alkane solvent is selected from the group consisting of n-pentane, n-hexane, n-heptane, and cyclohexane.

6. The alkane solvent of claim 5, wherein the alkane solvent is a class 3 solvent.

7. A pharmaceutical composition comprising the rifabutin powder of any one of claims 1-6, and a pharmaceutically acceptable solvent and / or excipient.

8. The pharmaceutical composition of claim 7, wherein the pharmaceutical composition is suitable for intravenous administration.

9. A method of producing a rifabutin for intravenous administration, the method comprising: combining a rifabutin powder with an alkane solvent to create a mixture; exposing the mixture to a solid-liquid extraction process; and extracting purified rifabutin from the mixture.

10. The method of claim 9, wherein the extracting step comprises distilling the solvent away from the rifabutin powder.

11. The method of claim 9, wherein the extracting step comprises trapping the rifabutin using a silica gel.

12. The method of claim 9, wherein the solid-liquid extraction process is repeated.

13. The method of claim 9, wherein an amount of rifabutin is completely dissolved in analkane solvent.

14. The method of claim 9, wherein insoluble impurities are removed with one or more filters.

15. The method of claim 9, wherein the mixture is heated to or above about 20 °C.

16. The method of claim 9, wherein the mixture is heated at or lower than about 50 °C.

17. The method of claim 9, wherein the method is conducted in one or more reaction vesselsthat are connected by one or more connections.

18. The method of claim 9, wherein the alkane solvent is selected from the group consisting of n-pentane, n-hexane, n-heptane, and cyclohexane.

19. The method of claim 18, wherein the alkane solvent is a class 3 solvent.

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