Claims:1. A process for sterilizing an active pharmaceutical ingredient (API) comprising the steps of:
(a) providing an API,
(b) providing a controlled atmosphere by replacing oxygen from the controlled atmosphere of the API with an inert gas, wherein the inert gas selected is from carbon dioxide, neon, freon, or a mixture thereof, and
(c) exposing the API in the controlled atmosphere of step (b) to a gamma radiation.
2. The process as claimed in claim 1, wherein the controlled atmosphere is of carbon dioxide gas.
3. The process as claimed in claim 1, wherein the controlled atmosphere is of neon gas.
4. The process as claimed in claim 1, wherein the API is brinzolamide.
5. The process as claimed in claim 1 or 4, wherein the API is in a powder form.
6. The process as claimed in claim 1 or 4, wherein the API is in a micronized powder form.
7. The process as claimed in claim 1, wherein the API is exposed to gamma radiation at a temperature ranging from -50°C to -78°C.
8. The process as claimed in claim 1, wherein the API is exposed to gamma radiation at a dose in the range of 5-30 kGy.
9. The process as claimed in claim 1 or 8, wherein the API is exposed to gamma radiation at a dose of 7 kGy to 25 kGy.
10. The process as claimed in any one of claims 1, 8 and 9, wherein the API is exposed to gamma radiation at a dose of 25 kGy.
11. The process as claimed in claim 1, wherein the API is exposed to gamma radiation for a time period ranging from 8 hours to 10 hours.
12. The process as claimed in claim 1, wherein the API is packed in a package under the controlled atmosphere replacing oxygen with inert gas before it is exposed to gamma radiation.
13. The process as claimed in claim 1 or 12, wherein the API is packed firstly in a primary package under the controlled atmosphere replacing oxygen with inert gas and the primary package is then packed in a secondary package under the controlled atmosphere replacing oxygen with inert gas; optionally wherein the primary package is made of polyethylene and the secondary package is made of aluminium.
, Description:FIELD OF THE INVENTION
[001] The present invention pertains to sterilization of active pharmaceutical ingredients. In particular, the present invention pertains to terminal sterilization of active pharmaceutical ingredients.

BACKGROUND OF THE INVENTION
[002] According to WHO, the active pharmaceutical ingredient is “Any substance or combination of substances used in a finished pharmaceutical product (FPP), intended to furnish pharmacological activity or to otherwise have direct effect in the diagnosis, cure, mitigation, treatment or prevention of disease, or to have direct effect in restoring, correcting or modifying physiological functions in human beings”. Particularly, in many formulations like ophthalmic formulation, API needs to be sterilized before preparing FPP or sterilization done on FPP depending on nature of API and other components. A sterile topical ophthalmic suspension has typically been manufactured in the past in one of three ways: by bulk sterilization of a milled suspension, by aseptic addition of sterile micronized raw material into a sterile vehicle, or by aseptic addition of a sterile raw material to a sterile menstruum followed by ball milling and aseptic addition of the sterile concentrate into a sterile vehicle.
[003] A sterile active pharmaceutical ingredients (APIs) could be manufactured by aseptic processing, which is very costly for a drug substance considering the low value of the API compared with the final drug formulation. Alternatively, a terminal sterilization is more convenient and cost effective. The terminal sterilization is a process of sterilizing a sample in its final container. The US pharmacopoeia recognize five procedures under chapter <1211> 1. Steam Sterilization, 2. Dry Heat Sterilization, 3. Gas Sterilization (ethylene oxide), 4. Sterilization by Ionizing Radiation (gamma ray, electron beam), 5. Sterilization by Filtration.
[004] The API brinzolamide is a carbonic anhydrase inhibitor (specifically, carbonic anhydrase II) which is found primarily in erythrocytes and in other tissues specifically in the eye. An ophthalmic suspension containing brinzolamide is used for the treatment of open-angle glaucoma and raised intraocular pressure due to excess aqueous humor production. The IUPAC name of brinzolamide is (5R)-5-ethylamino-3-(3-methoxypropyl)-2,2-dioxo-2?6,9-dithia-3-azabicyclo [4.3.0]nona-7,10-diene-8-sulfonamide, having formula of C12H21N3O5S3, molecular weight of 383.50, and melting point of about 131 ºC. The PCT published document WO2012053011 describes preparing of a sterile ophthalmic suspension comprising brinzolamide by aseptically filtering the milled slurry of brinzolamide with surfactant which is filtered through 0.22 µm filter and then added other adjuvants or carriers to obtain the finalized sterile suspension. Another PCT published document WO2015068105 describes preparation of a sterile, ophthalmic pharmaceutical suspension comprising a sterile active ingredient (brinzolamide), at least one surfactant, polymer and other carriers or adjuvants.
[005] The European patent EP941094B1 describes that a steam sterilization of brinzolamide employing saturated steam under pressure is carried out in an autoclave at 120°C or dry heat sterilization where the unit is operating at not less than 250°C. US6071904 describes sterilization of suspension containing brinzolamide by autoclaving method. The PCT published document WO2014057499 describes sterilization of brinzolamide by dry heat sterilization at a temperature between 80- 120°C for 5-20 hours. Another PCT published document WO2015110993 describes preparation of aqueous sterile, ophthalmic pharmaceutical formulation comprising a) brinzolamide as carbonic anhydrase inhibitor, b) an amphiphilic polymer, and c) a surfactant, in which the sterile brinzolamide is obtained by heating at temperature around 70 ºC or by autoclaving method. All of these documents suffer from various disadvantages, for example, the steam sterilization recrystallizes brinzolamide at autoclaving temperatures forming large needle-type crystals in the FPP. Further, the powdered samples of brinzolamide may get degraded by absorbing the moisture or melted at higher dry temperature. In view of these, the dose of API brinzolamide would be highly uneven which affect the overall treatment. The large needle like crystal formation in the FPP can be cured by ball milling the formulation to reduce the size of brinzolamide, however, this ultimately increase the cost of process.
[006] Alternatively, another method of sterilization is gaseous sterilization preferably using ethylene oxide. The European Patent EP2394637 provide the sterilization of brinzolamide using pure ethylene oxide for 200 minutes under reduced pressure of 612 mBar in Sterimed sterilization device. The Indian Patent application IN421/CHE/2011 describes process of sterilization of API brinzolamide and FPP of ophthalmic suspension comprising brinzolamide using ethylene oxide. The US published Patent document US2015065493 provides a process for sterilizing an active pharmaceutical ingredient (brinzolamide) by contacting with ethylene oxide, under conditions that at most 0.5% by weight of degradation products are formed, wherein the concentration of ethylene oxide is in the range of 300-450 mg/1, and the contacting time is in the range of 20-100 minutes. However, the above-mentioned prior art processes suffer distinct disadvantages which render them technically and economically objectionable. For example, ethylene oxide is highly flammable unless mixed with suitable inert gases, its mutagenic properties, and the possibility of toxic residues in treated materials, particularly those containing chloride ions. Further, one of the principal limiting factor of the ethylene oxide sterilization process is their limited ability to diffuse to the innermost product areas that require sterilization. Furthermore, the sterilization with ethylene oxide is difficult to perform and it is hardly suitable for industrial manufacture of active pharmaceutical ingredients as required by pharmacopoeias and current GMP.
[007] Another sterilization method is done by filtration method. The Japanese patent published document JP2015212231A describes preparation of aqueous solution obtained by dissolving brinzolamide in an alkaline aqueous solution having a pH of 10.5 to 13 which was sterilized by filtration technique and then adjusting pH with an acid to precipitate the crystals of brinzolamide in the aqueous solution. However, this method requires first solubilisation of API for proper filtration which would be costly and tedious in case of intermediate and insoluble APIs like brinzolamide.
[008] Yet another sterilization method is by ionic irradiation which has its own advantages including low chemical reactivity, low measurable residues, and fewer variables to control. Also radiation sterilization is unique in that the basis of control is essentially that of the absorbed radiation dose, which can be precisely measured by using dosimeters. The European patent EP2394637 provide the sterilization of brinzolamide using gamma radiation at dose of 15 KGy for 3-4 seconds in dosimeter ECB. However, EP2394637 provides no details about the degradation of brinzolamide active pharmaceutical ingredients, only measuring enantiomeric purity.
[009] Previous research efforts and achievements notwithstanding, there is a continuing need to develop improved processes which enable quick and efficient sterilization of bulk powder of pharmaceutical active ingredients, without interfering with the active ingredients. The present invention satisfies the existing needs, as well as others, and generally overcomes the deficiencies found in the prior art.
[0010] All publications herein are incorporated by reference to the same extent as if each individual publication or patent application were specifically and individually indicated to be incorporated by reference. Where a definition or use of a term in an incorporated reference is inconsistent or contrary to the definition of that term provided herein, the definition of that term provided herein applies and the definition of that term in the reference does not apply.

SUMMARY OF THE INVENTION
[0011] The present invention provides a new and improved process for sterilization of bulk powder of active pharmaceutical ingredients. The present invention is based upon a surprising finding that exposing a bulk powder of an active pharmaceutical ingredient, such as brinzolamide powder, to gamma radiation in a controlled atmosphere of carbon dioxide, neon, freon, or a mixture thereof, can sterilize the active pharmaceutical ingredient without interfering with the active ingredient. The gamma-ray sterilization process disclosed herein provides high quality, pure sterile API substance complying with the pharmacopeial requirements.
[0012] Accordingly, in one aspect the present invention provides a process for sterilizing an active pharmaceutical ingredient (API) comprising the steps of:
(a) providing an API,
(b) providing a controlled atmosphere by replacing oxygen from the atmosphere of the API with an inert gas, wherein the inert gas selected is from carbon dioxide, neon, freon, or a mixture thereof, and
(c) exposing the API in the controlled atmosphere of step (b) to a gamma radiation.
[0013] In another aspect, the present invention provides an economical and eco-friendly process for sterilization of active pharmaceutical ingredients.
[0014] In the gamma-ray sterilization process disclosed herein, the API, such as brinzolamide, may be used in the form of micronized or non-micronized bulk powder, preferably in the form of micronized bulk powder. In certain embodiments, the API bulk powder may be packed in a package under the controlled atmosphere before it is exposed to gamma radiation. The controlled atmosphere refers to an atmosphere wherein ambient air, specifically oxygen is replaced with carbon dioxide, neon, freon, or a mixture thereof, and then sealing the package containing the API bulk powder. In a preferred embodiment, the API bulk powder is packed firstly in a primary package made of polyethylene, preferably LDPE, and then in a secondary package made of aluminium, preferably triple laminated aluminium.
[0015] In various embodiments, the gamma-ray sterilization disclosed herein can be carried out at sub-zero temperature ranging from -10°C to -78°C, preferably ranging from -50°C to -78°C.
[0016] In various embodiments, the sterilization by gamma irradiation disclosed herein can be performed employing any pharmaceutically acceptable dose of gamma irradiation, preferably at a dose in the range of 5-30 kGy, more preferably at a dose in the range of 20-25 kGy and even more preferably, at a dose of 25 kGy.
[0017] Various objects, features, aspects and advantages of the inventive subject matter will become more apparent from the following detailed description of preferred embodiments.

DETAILED DESCRIPTION OF THE INVENTION
[0018] The following is a detailed description of embodiments of the present invention. The embodiments are in such detail as to clearly communicate the invention. However, the amount of detail offered is not intended to limit the anticipated variations of embodiments; on the contrary, the intention is to cover all modifications, equivalents, and alternatives falling within the spirit and scope of the present invention as defined by the appended claims.
[0019] Unless the context requires otherwise, throughout the specification which follow, the word “comprise” and variations thereof, such as, “comprises” and “comprising” are to be construed in an open, inclusive sense that is as “including, but not limited to.”
[0020] Reference throughout this specification to “one embodiment” or “an embodiment” means that a particular feature, structure or characteristic described in connection with the embodiment is included in at least one embodiment. Thus, the appearances of the phrases “in one embodiment” or “in an embodiment” in various places throughout this specification are not necessarily all referring to the same embodiment. Furthermore, the particular features, structures, or characteristics may be combined in any suitable manner in one or more embodiments.
[0021] As used in the description herein and throughout the claims that follow, the meaning of “a,” “an,” and “the” includes plural reference unless the context clearly dictates otherwise. Also, as used in the description herein, the meaning of “in” includes “in” and “on” unless the context clearly dictates otherwise.
[0022] In some embodiments, the numbers expressing quantities of ingredients, properties such as concentration, process conditions, and so forth, used to describe and claim certain embodiments of the invention are to be understood as being modified in some instances by the term “about.” Accordingly, in some embodiments, the numerical parameters set forth in the written description are approximations that can vary depending upon the desired properties sought to be obtained by a particular embodiment. In some embodiments, the numerical parameters should be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. Notwithstanding that the numerical ranges and parameters setting forth the broad scope of some embodiments of the invention are approximations, the numerical values set forth in the specific examples are reported as precisely as practicable.
[0023] The recitation of ranges of values herein is merely intended to serve as a shorthand method of referring individually to each separate value falling within the range. Unless otherwise indicated herein, each individual value is incorporated into the specification as if it were individually recited herein.
[0024] All methods described herein can be performed in suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g. “such as”) provided with respect to certain embodiments herein is intended merely to better illuminate the invention and does not pose a limitation on the scope of the invention otherwise claimed. No language in the specification should be construed as indicating any non-claimed element essential to the practice of the invention.
[0025] The headings and abstract of the invention provided herein are for convenience only and do not interpret the scope or meaning of the embodiments.
[0026] Various terms are used herein. To the extent a term used in a claim is not defined below, it should be given the broadest definition persons in the pertinent art have given that term as reflected in printed publications and issued patents at the time of filing.
[0027] The present invention provides a new and improved process for sterilization of bulk powder of active pharmaceutical ingredients. The present invention is based upon a surprising finding that exposing a bulk powder of an active pharmaceutical ingredient, such as brinzolamide powder, to gamma radiations in a controlled atmosphere of inert gas(es) such as carbon dioxide (CO2), neon, freon, or a mixture thereof, which can sterilize the active pharmaceutical ingredient without interfering with the active ingredient. The gamma-ray sterilization process disclosed herein provides high quality, pure sterile API substance complying with pharmacopeial requirements.
[0028] Thus, one aspect of the present invention provides a process for sterilizing an active pharmaceutical ingredient (API) comprising the steps of:
(a) providing an API,
(b) providing a controlled atmosphere by replacing oxygen from the atmosphere of the API with an inert gas, wherein the inert gas selected is from carbon dioxide, neon, freon, or a mixture thereof, and
(c) exposing the API in the controlled atmosphere of step (b) to a gamma radiation.
[0029] In a preferred embodiment, a bulk powder of an API can be sterilized by providing a controlled atmosphere by replacing oxygen from the atmosphere of the bulk powder of API with carbon dioxide and then exposing the bulk powder to gamma radiation in the controlled atmosphere of carbon dioxide (CO2).
[0030] In another preferred embodiment, a bulk powder of an API can be sterilized by providing a controlled atmosphere by replacing oxygen from the atmosphere of the bulk powder of API with neon and then exposing the bulk powder to gamma radiation in the controlled atmosphere of neon.
[0031] In another preferred embodiment, a bulk powder of an API can be sterilized by providing a controlled atmosphere by replacing oxygen from the atmosphere of the bulk powder of API with freon and then exposing the bulk powder to gamma radiation in the controlled atmosphere of freon.
[0032] In the gamma-ray sterilization process disclosed herein, the API, such as brinzolamide, may be used in the form of micronized or non-micronized bulk powder, preferably in the form of micronized bulk powder. In certain preferred embodiments, the API is used in the form of micronized dry powder, e.g., micronized brinzolamide dry powder, wherein at least 90% of the particles (d90) are of a size of not more than 10µm and at least 50% of the particles (d50) are of a size of not more than 5µm, complying with the current USP monograph as well as ICH Q3A.
[0033] In certain embodiments, the API bulk powder may be packed in a package under the controlled atmosphere before it is exposed to gamma radiation. The controlled atmosphere refers to replacement of ambient air, specifically oxygen with carbon dioxide, neon, freon, or a mixture thereof, and then sealing the package containing the API bulk powder.
[0034] In an embodiment, the package is selected from but not limiting to a pouch, bag, container or any other suitable pharmaceutical package, to protect the API from sunlight.
[0035] In another embodiment, the API bulk powder is packed under the controlled atmosphere firstly in a primary package made of polyethylene and this primary package is then packed under the controlled atmosphere in a secondary package made of aluminium. The interior of the primary package and the secondary package can be filled or flushed with the said inter gas(es) to replace and exclude oxygen and/or moisture that may cause degradation of API during gamma irradiation. The controlled atmosphere provided in the primary package and the secondary package may be the same or different from one another. In one exemplary embodiment, the primary package is made of low-density polyethylene (LDPE). In further exemplary embodiment, the secondary package is a polylaminated aluminium package, for example a triple laminated aluminium bag or triple laminated aluminium pouch.
[0036] As disclosed herein, the term “primary package” relates to the material that first envelops or protect a product and hold it, as in present case, the API.
[0037] As disclosed herein, the term “secondary package” relates to the material that surrounds or is outside to the first package.
[0038] In various embodiments, the gamma-ray sterilization disclosed herein can be carried out at a sub-zero temperature ranging from -10°C to -78°C, preferably ranging from -50°C to -78°C.
[0039] In various embodiments, the sterilization by gamma irradiation disclosed herein can be performed employing any pharmaceutically acceptable dose of gamma irradiation, preferably at a dose in the range of 5-30 kGy. In certain embodiments, bulk powder of an API can be exposed to gamma radiation at a dose ranging between 7 kGy to 25 kGy or preferably at a dose in the range of 20-25 kGy, or more preferably at a dose of 25 kGy. In certain embodiments, bulk powder of an API can be exposed to gamma radiation at a dose of 7 kGy, 10 kGy, 15 kGy, 20 kGy, 25 kGy or 30 kGy.
[0040] In various embodiments, the gamma ray irradiation can be carried out for a time period to sterilize the API bulk powder effectively, for example ranging from 8 hours to 10 hours.
[0041] In one or more embodiments, the gamma ray irradiation can be carried out on API bulk powder for a time period ranging from about 8 hours to about 10 hours to receive gamma rays dose of not less than 20 kGy or 25 kGy.
[0042] While any API may be employed in the process according to the present invention, preferably, the API is brinzolamide.
[0043] While any suitable gamma radiation source may be employed in the process according to the present invention, preferably, the source is Cobalt-60.
[0044] As employed herein, the terms “sterilization”, “sterilize” and “sterilizing” refer to elimination of microorganisms such as bacteria, spores, viruses and moulds in an active pharmaceutical ingredient (API). When an API is “sterilized” or “sterile” there are no living organisms in or on an API.
[0045] In various embodiments, the gamma-ray sterilization process according to the present invention can produce sterile API with a purity of at least about 99% by RS-HPLC (Rapid Separation High Performance Liquid Chromatography). The specified controlled atmosphere can prevent or minimize API degradations, which lead to by products (known and unknown impurities) and impact product quality, due to gamma irradiation.
[0046] In an exemplary embodiment, the disclosed sterilization process can produce sterile brinzolamide with a purity of at least about 99% by RS-HPLC with all known and unknown impurities within the United States Pharmacopeia (USP) specification.
[0047] In another exemplary embodiment, the disclosed sterilization process can produce sterile brinzolamide in a purity of at least about 99% by RS-HPLC, with about 0.1% to about 0.2% by RS-HPLC of impurity B.
[0048] In another exemplary embodiment, the disclosed sterilization process can produce sterile brinzolamide in a purity of at least about 99% by RS-HPLC, with about 0.02% to about 0.1% by RS-HPLC of impurity D.
[0049] As used herein, unless otherwise indicated, the term " As used herein, unless otherwise indicated, the term "impurity B" refers to (R)-4-amino-3,4-dihydro-2-(3-methoxypropyl)-2H-thieno[3,2-e]-1,2-thiazine-6-sulfonamide 1,1-dioxide. As used herein, unless otherwise indicated, the term "impurity D" refers to 4-oxo-3,4-dihydro-2-(3-methoxypropyl)-2H-thieno[3,2-e]-1,2-thiazine-6-sulfonamide 1,1-dioxide.
[0050] In various embodiments, a sterile API produced according to the gamma-ray sterilization process disclosed herein may be combined with one or more pharmaceutically acceptable excipients to obtain a pharmaceutical composition.
[0051] In an exemplary embodiment, a sterile brinzolamide produced according to the gamma-ray sterilization process disclosed herein may be combined with one or more pharmaceutically acceptable excipients to obtain a pharmaceutical composition for the treatment of open angle glaucoma and/or increased intraocular pressure.
[0052] While the foregoing description discloses various embodiments of the invention, other and further embodiments of the invention may be devised without departing from the basic scope of the disclosure. The invention is not limited to the described embodiments, versions or examples, which are included to enable a person having ordinary skill in the art to make and use the invention when combined with information and knowledge available to the person having ordinary skill in the art.
EXAMPLES
[0053] The present invention is further explained in the form of following examples. However, it is to be understood that the foregoing examples are merely illustrative and are not to be taken as limitations upon the scope of the invention. Various changes and modifications to the disclosed embodiments will be apparent to those skilled in the art. Such changes and modifications may be made without departing from the scope of the invention.
Example 1
Sterilization of API brinzolamide
[0054] 500 mg samples of micronized brinzolamide USP grade powder were packed separately under a controlled atmosphere of inert gas CO2 or Neon or Freon respectively in LDPE bag. Each LDPE bag was placed into a triple laminated aluminium bag, which was then sealed under controlled atmosphere of CO2 or neon or freon respectively. Each sealed aluminium bag was placed in dry ice to cool it to about -50°C to -78°C and was then exposed to gamma radiation of 25 kGy. The irradiated API powder was tested according to USP monograph. The details about purity profile and degradation products (impurities) detected by Reversed-Phase High-Performance Liquid Chromatography (RS-HPLC) method are given in Table 1.
Table 1: Purity & Impurity Profile by RS- HPLC of Control and Sterilized API
Gas used Sterilized API % Purity Impurity B Impurity D
Air 97.95 0.03 0.58
CO2 98.75 0.19 0.09
Neon 98.9 0.19 0.06
Freon 98.88 0.19 0.07

[0055] Gamma-ray irradiation of brinzolamide under CO2, neon or freon controlled atmosphere provides sterile brinzolamide complying with pharmacopeial requirements. The data also show that neon or CO2 or Freon controlled atmosphere is more effective in reducing brinzolamide degradation due to gamma irradiation dose of 25 kGy at lower temperatures of -50°C to -78 °C
[0056] It is submitted that the air stability data (without controlled atmosphere) is failing immediately at 0 month (initial stage as provided in table 1), hence, no further studies at 1 month, 2 months, 3 months and 6 months were done.
Example 2
Intermediate stability study of API brinzolamide sterilized with CO2 and gamma radiation
[0057] The stability study of the micronized brinzolamide sterilized with CO2 and gamma radiation in accordance with the present invention as per Example 1 was carried out at intermediate conditions by storing samples at 30 ± 2°C / 65±5% RH (relative humidity) for three months and six months, and their purity profile and degradation products (impurities) were detected by RS-HPLC. Details of the purity and impurity profile are given in Table 2.
Table 2: Purity & Impurity Profile by RS- HPLC of API Sterilized with CO2 and gamma radiation under intermediate stability conditions
Parameters Initial 3 months 6 months
Appearance White powder White to off-white White to off-white
Sterilized API % Purity 99.24 99.15 99.11
Impurity B (%) 0.17 0.21 0.24
Impurity D (%) 0.06 0.09 0.09

Example 3:
[0058] The stability study of the micronized brinzolamide sterilized with neon gas and gamma radiation in accordance with the present invention as per Example 1 was carried out at intermediate conditions by storing samples at 30 ± 2°C / 65±5% RH (relative humidity) for three months and six months, and their purity profile and degradation products (impurities) were detected by RS-HPLC. Details of the purity and impurity profile are given in Table 3.

Table 3: Purity & Impurity Profile by RS- HPLC of API Sterilized with neon and gamma radiation under intermediate stability conditions
Parameters Initial 3 months 6 months
Appearance White powder White to off-white White to off-white
Sterilized API % Purity 99.24 99.18 99.11
Impurity B (%) 0.16 0.22 0.22
Impurity D (%) 0.06 0.09 0.09

Example 4:
[0059] The accelerated stability study of the micronized brinzolamide sterilized with CO2 and gamma radiation in accordance with the present invention as per Example 1 was carried out by storing samples at 30 ± 2°C / 65±5% RH (relative humidity) for one, two, three months and six months, and their purity profile and degradation products (impurities) were detected by RS-HPLC. Details of the purity and impurity profile are given in Table 4.
Table 4: Purity & Impurity Profile by RS- HPLC of API Sterilized with CO2 and gamma radiation under accelerated stability conditions
Parameters Initial 1 month 2 months 3 months 6 months
Appearance White powder White powder White to off-white White to off-white White to off-white
Sterilized API % Purity 99.24 99.17 99.14 99.10 99.06
Impurity B (%) 0.17 0.21 0.21 0.22 0.23
Impurity D (%) 0.06 0.08 0.08 0.08 0.09

Example 5:
[0060] The accelerated stability study of the micronized brinzolamide sterilized with neon and gamma radiation in accordance with the present invention as per Example 1 was carried out by storing samples at 30 ± 2°C / 65±5% RH (relative humidity) for one, two, three months and six months, and their purity profile and degradation products (impurities) were detected by RS-HPLC. Details of the purity and impurity profile are given in Table 5.
Table 5: Purity & Impurity Profile by RS- HPLC of API Sterilized with neon and gamma radiation under accelerated stability conditions
Parameters Initial 1 month 2 months 3 months 6 months
Appearance White powder White powder White to off-white White to off-white White to off-white
Sterilized API % Purity 99.24 99.19 99.16 98.93 98.90
Impurity B (%) 0.16 0.21 0.22 0.23 0.25
Impurity D (%) 0.06 0.07 0.08 0.08 0.09