FIELD OF THE INVENTION
The present invention relates to a novel polymorphic form I of rifaximin, a rifamycin class of antibiotics and process for preparation thereof.
BACKGROUND OF THE INVENTION
Rifaximin of formula I, is an antibiotic belonging to rifamycin class of antibiotics and is chemically known as (2S,16Z,18E,20S,21S,22R,23R,24R,25S,26S,27S, 28E)-5,6,21,23,25-pentahydroxy-27-methoxy-2,4,ll,16,20,22,24,26-octamethyl-2,7-(epoxypentadeca-[l,11,13]trienimino)benzofuro[4,5-e]pyrido[l,2-a]-benzimi dazole-l,15(2H)-dione, 25-acetate.
(Formula Removed)
Rifaximin is a pyrido-imidazo rifamycin which shows its broad antibacterial activity such as, in the gastrointestinal tract against localized gastrointestinal bacteria that cause infectious diarrhea, irritable bowel syndrome, small intestinal bacterial overgrowth, Crohn's disease, and/or pancreatic insufficiency. Rifaximin was first described and claimed in Italian patent IT 1154655. US Patent 4, 341,785 and 4,557,866 disclose a process for the preparation of rifaximin starting from rifamycin S or O. The above patents describe purification steps of rifaximin by performing crystallization of crude rifaximin from a 7:3 mixture of ethyl alcohol/water and drying under vacuum without describing exact experimental conditions required for crystallization and drying.
Recently published US Patent 7,045,620 discloses three polymorphic forms α, ß and  of rifaximin. Form α and ß show pure crystalline characteristics while the y form is poorly crystalline. These polymorphic forms are differentiated on the basis of water content and XRPD. This patent also discloses processes for preparation of these polymorphs which involve use of specific reaction conditions during crystallization like dissolving rifaximin in ethyl alcohol at 45 to 65 °C, precipitation by adding water to form a suspension, filtering suspension and washing the resulted solid with demineralized water, followed by drying at room temperature under vacuum for a period of time between 2 and 72 hours. The above process provided a general criterion to get three polymorphic forms α, ß and  of rifaximin. However, the crystallization conditions are specific in order to obtain a particular form. Purely crystalline forms α and ß are obtained by immediate filtration of suspension when temperature of reaction mixture is brought finally to 0 °C whereas in order to obtain the poorly crystalline form , the reaction mixture is stirred for 5-6 hours after temperature is set to 0 °C and then filtered the suspension. In addition to above specific conditions, it is mandatory for a particular form to have specific water content such as to obtain a form, water content should be lower than 4.5%, for ß form it should be higher than 4.5% and to obtain  form, water content should be below 2%. US Patent 7,709,634 discloses an amorphous form of rifaximin and the process for the preparation thereof. The amorphous form is prepared by dissolving rifaximin in solvents such as alkyl esters, alkanols and ketones and precipitating by addition of anti-solvents selected from hydrocarbons, ethers or mixtures thereof.
US patent publication 2009/0312357A1 discloses an amorphous form of rifaximin which is purely amorphous and method of converting amorphous form to crystalline form.
US patent publication 2009/0130201A1 discloses polymorphic forms 5 and s of rifaximin and methods of their preparation by dissolving rifaximin in ethyl
alcohol at 45 to 65 °C, precipitation by adding water to form a suspension, filtering the suspension and washing the resulted solid with demineralized water, followed by drying for a period of time between 2 and 72 hours until a water content in the range 2.5-6 % is obtained.
Another US patent publication 2010/0174064A1 discloses several polymorphic forms of rifaximin such as , form , form , form -dry, form -dry', form B, amorphous form and form 6 of rifaximin. Main differentiating point of these polymorphs is respective water content and XRPD.
It is evident from above, that rifaximin can exist in number of polymorphic forms, formation of these polymorphic forms depends upon specific reaction conditions applied during crystallization and drying. Nevertheless, there is continuing need of new polymorphic forms of rifaximin to meet challenges in the upcoming pharmaceutical market. It can be noted that in recent years, solid-state properties of drugs have received great attention in the pharmaceutical industry, as a major contributing factor to both bio-availability and formulation characteristics. The ability of some substances to exist in more than one crystalline form, called polymorphism, was accredited as one of the most important solid-state property of drugs. While polymorphs have the same chemical composition, they differ in packing and geometrical arrangement thereof and exhibit different physical properties such as melting point, shape, color, X-ray diffraction pattern, infrared absorption, and solid state NMR spectrum, density, hardness, deformability, stability, dissolution, and the like. Depending on their temperature-stability relationship, one crystalline form may give rise to thermal behavior different from that of another crystalline form. Thermal behavior can be measured in the laboratory by techniques such as capillary melting point, thermogravimetric analysis ("TGA"), and differential scanning calorimetry ("DSC"), which have been used to distinguish polymorphic forms.
One of the most important physical properties of pharmaceutical compounds is their solubility in aqueous solution, particularly their solubility in the gastric
juices of a patient. For example, where absorption through the gastrointestinal tract is slow, it is often desirable for a drug that is unstable to conditions in the patient's stomach or intestine to dissolve slowly, so that it does not accumulate in a deleterious environment. Different crystalline forms or polymorphs of the same pharmaceutical compounds can and reportedly do have different aqueous solubility or different dissolution rates (release profile) in-vivo. Therefore, discovery of a new polymorphic form of a pharmaceutically useful compound provides a new opportunity to improve performance characteristics of a pharmaceutical product. It enlarges repertoire of materials that a formulation scientist has available for designing, for example, a pharmaceutical dosage form of a drug with a targeted release profile or other desired characteristic. However, any failure to predict bioavailability of a drug may result in administration of either too small or too large undesired doses, which may be dangerous to patients and in extreme cases, lethal.
Other examples are known, where different crystalline forms behave differently during physical processing like milling and pressing. Many process-induced solid-solid transitions of substances are known, that lead to either other crystalline forms or an amorphous form of the substance. The solid-state experts are in a constant search for crystalline forms that can withstand physical stress and still retain their original properties.
Consequently, there is an ongoing search for a new polymorphic form of drug, which may provide improved performance thereof. A single molecule, such as rifaximin, may give rise to a variety of crystalline or amorphous form or a mixture of amorphous and crystalline form having distinct crystal structures and physical properties.
OBJECT OF THE INVENTION
It is principal object of present invention is to provide a new polymorphic of rifaximin.
Another object of present invention is to provide a process for preparation of a
new polymorph.
One another object of present invention is to provide an efficient process wherein
new polymorph results consistently.
One another object of present invention is to provide an efficient process wherein
new polymorph is free from other polymorphs.
Another object of the present invention is to provide processes for converting new
polymorphic form to crystalline form α, form ß and amorphous forms of rifaximin
and vice versa.
SUMMARY OF THE INVENTION
Accordingly, in one aspect, the present invention provides a new polymorphic form, named as Form I of rifaximin, which is highly pure and substantially free from other polymorphic form of rifaximin. Polymorphic form I may be characterized by X-ray powder diffraction peaks at least 5.2, 7.2, 8.4 and 8.7 expressed in degrees 20 ( + 0.2° 8) and any possible combination of peaks thereof. The polymorphic form I may be characterized by X-ray powder diffraction patterns as shown in Figures 1, IA and IB. The polymorphic form I may be further characterized by differential scanning calorimetry (DSC) substantially similar as shown in Figures 2 and 2A. Further, polymorphic form I may be characterized by water content of 2 to 4.5 %, preferably of 2 to 3 % as demonstrated by TGA curve shown in Figure 3.
In another aspect, the present invention provides a process for preparation of polymorphic form I which comprises steps:
a) providing a solution of crude rifaximin in ethyl alcohol at reflux temperature,
b) adding demineralized water in less than 10 minutes to reaction mixture at this temperature,
c) stirring the reaction mixture for 10-20 minutes at the same temperature,
d) cooling down the reaction solution at a temperature to 40 °C in 30 to 40 minutes,
e) optionally seeding the reaction mixture with rifaximin form I,
f) decreasing temperature of solution to 30 °C,
g) maintaining reaction mixture at 30 °C for 5 hours under stirring,
h) cooling reaction mixture to 0 °C slowly,
i) filtering the reaction mixture immediately
j) washing the resulting solid with a mixture of ethyl alcohol demineralized
water, k) drying the resulting solid at a temperature between 60 to 70°C under vacuum
for a time period between 8-14 hours, till water content is lower than 4.5%.
In one another aspect, the present invention provides a process for preparation of polymorphic form I which comprises steps:
a) providing a solution of rifaximin in a mixture of ethyl alcohol and demineralized water at reflux temperature,
b) cooling the reaction solution at temperature 25-40 °C in 30 to 60 minutes,
c) optionally seeding the reaction mixture with rifaximin form I;
d) decreasing temperature of solution to 30 °C,
e) stirring reaction mixture for 1-5 hours.
f) cooling the reaction mixture to 0 °C slowly,
g) filtering the reaction mixture within 6 hours,
h) washing the resulting solid with a mixture of ethyl alcohol and demineralized
water, i) drying the resulting solid at a temperature between 25 to 80 °C till water
content is lower than 4.5%. Another aspect of the present invention is to provide processes for inter-conversion of polymorphic form I to known forms like α, ß,  and amorphous forms of rifaximin or vice versa.

BRIEF DESCRIPTION OF THE DRAWINGS
Figures 1, 1A and 1B are exemplary XRPD patterns of rifaximin form I Figures 2 and 2 A are exemplary DSC of rifaximin form I Figure 3 is an exemplary TGA of rifaximin form I
Figure 4 is an exemplary graphical representation of intrinsic dissolution of rifaximin form I comparing with a -form of rifaximin.
DETAILED DESCRIPTION OF THE INVENTION
The present invention relates to a novel polymorphic form I of rifaximin, which is substantially free from other polymorphic forms of rifaximin. The polymorphic form I of the present invention exhibits an X-ray powder diffraction pattern, substantially similar as given in Figures 1, 1A and 1B.
The polymorphic form I is also characterized by X-ray powder diffraction peaks at least 5.2, 7.2, 8.4 and 8.7, expressed in degrees two-theta and any possible combination of peaks thereof. Further, the polymorph can be characterized by absence of any crystalline peak after 10 degrees two-theta.
According to present invention, the new polymorphic form I can be characterized by a distinct Differential Scanning Calorimetry ("DSC") thermogram substantially similar as shown in the Figures 2 and 2A. DSC in Figure 2 shows a broad endotherm in the range of 65 to 75 °C and an exothermic peak in the range of 215-220 °C using aluminum crucible with a lid having pin hole. However using same crucible having lid without pin hole give a different peak profile and represent rather better melting behavior. The DSC carried out without using pin hole method shows relatively sharp endotherm. The observed DSC peaks found in the range of 124-143 °C and variation in peak position depends on moisture content [2- 4.5%]. As moisture content increases, endothermic peak tends to appears at lower values, but when moisture content decreases endothermic peak tends to appears at moderately higher side, thus, supporting the fact that DSC thermogram is not so consistent in case of rifaximin. An exemplary

DSC thermogram showing endothermic peak at 137 °C as obtained by without pin
hole method is shown in Figure 2A.
According to present invention, polymorphic form I can be further characterized
by water content within the range of 2.0 to 4.5%, preferably 2-3%. This form
demonstrates weight loss by TGA as shown in Figure 3.
According to present invention, the novel polymorphic form I is quite stable and
after applying the physical techniques like pulverization, milling, micronization
the crystal structure does not distorted. The polymorphic form I after
pulverization, milling or micronization is also characterized by X-ray powder
diffraction and shows peaks at 5.2, 7.2, 8.4 and 8.7 expressed in degrees two-
theta.
According to one aspect of present invention the polymorphic form I can be
characterized by various parameters such as solubility, intrinsic dissolution, bulk
density, tapped density, particle size, morphology etc. Generically intrinsic
dissolution data of polymorphic form I is done and its comparison to a-form of
rifaximin is carried out as a form is most thermodynamically stable form
compared to other polymorphic form. The data is depicted in Table I and by
graphical representation in Figure 4.
Intrinsic Dissolution
Dissolution medium: 1000mL of 0.1M Sodium dihydrogen phosphate
monohydrate + 4.5g of sodium lauryl sulphate. (pH 7.4)
Temperature: 37±0.5 °C
Rotation speed: 100 rpm
Table-1
(Table Removed)
It is further observed from the comparative dissolution data as shown in Table 1 and graphical representation in Figure 4 that polymorphic form I has better dissolution profile than that of a form of rifaximin.
Polymorphic form I is further characterized by bulk density in the range of 0.25 to 0.35 g/ml and tapped density in the range of 0.45 to 0.55 g/ml. According to another aspect, present invention provides a process for preparation of novel polymorphic form I from rifaximin. The process involves providing a solution of rifaximin in ethyl alcohol at reflux temperature. Generally rifaximin is added to the ethyl alcohol at room temperature and heated at reflux temperature (80+2°C) until a clear solution is obtained. To the above solution demineralized water is added at the same temperature over time period of 1-15 minutes, preferably at 3-6 minutes. After the complete addition of water, it is advantageous to stir the reaction mixture at same temperature to obtain clear solution. Generally stirring can be performed for 1 hour or preferably for 10-20 minutes. The ratio of
ethyl alcohol and demineralized water can be in any proportion of 1: 0.35 to 1: 0.45 (V/V) in solution.
Alternatively, the preparation of rifaximin form I can also be carried out by dissolving rifaximin in a mixture of ethyl alcohol and water at reflux temperature. The volume of solvent used during recrystallisation can be 1-10 volumes to the weight of crude rifaximin. Preferably it may be 2-3 volumes to weight of rifaximin.
The solution obtained after addition of water is allowed to reflux for 10-20 minutes under stirring to obtain a clear solution and cooled down at a temperature of 35-40 °C. Cooling of reaction mass is carried out slowly may be in 60 minutes and preferably in 30-40 minutes. Thereafter, when temperature of reaction mixture reaches 40 °C, reaction mixture is optionally seeded with pure rifaximin form I and stirring is continued for further 10-15 minutes. Further, temperature of the reaction mixture is decreased to 30 oC and kept at same temperature for 1-5 hours under stirring and preferably for 2-4 hours. It is advantageous to perform normal stirring to obtain pure rifaximin form I consistently. Further, temperature of reaction mixture is decreased to 0 °C slowly in 60 minutes. As soon as temperature reaches 0 °C, reaction mixture is filtered immediately. In an alternate embodiment, reaction mixture can optionally be kept at 0 °C for sometime and filtered within 6 hours. The resulting solid is washed with a mixture of ethyl alcohol and demineralized water having same ratio of ethyl alcohol and demineralized water as used during crystallization. The solid, thus obtained, is dried at a temperature of above 20 °C. Generally, solid is dried at 25-80 °C and preferably drying is carried out between 35 to 70 °C under vacuum. Alternatively drying can be performed under atmospheric conditions at ambient temperature. Time period required for drying can vary from 8 to 18 hours as it depend upon water content. Preferably drying is carried out for about 12 hour until water content of compound reaches less than 4.5%, preferably between 2-3 %. The preferable temperature at which solid is dried is 35-70 °C. The term room

temperature used in the application refers to a temperature range between 25+2 °C.
The crucial steps of obtaining pure polymorphic form I depends on the conditions chosen for crystallization. In particular, composition of the solvent mixture from which crystallization is carried out i.e. 1:0.35 tol:0.45 (V/V) mixtures of ethyl alcohol and water, the temperature at which the reaction mixture is kept after crystallization and the period of time at which that temperature is maintained. In the present invention the reaction mass is first heated at reflux temperature, and then the temperature is brought to 40 °C in 30-40 minutes and decrease further to 30 °C and maintaining the solution at this temperature for 1-5 hours, finally decreasing the temperature to 0 °C and filtered immediately. Alternatively, pure polymorphic form I can be obtained by following steps like dissolving rifaximin in a mixture of ethyl alcohol and water [1:0.35 tol:0.45 (V/V)] at reflux temperature, and then temperature is brought to 40 °C in 30-40 minutes, seeding the reaction mixture with rifaximin form I and then temperature is decreased further to 30 °C and maintaining the solution at this temperature for 1-5 hours, finally decrease the temperature to 0 °C and filter immediately or after optional stirring up to 6 hours. More precisely, it is another crucial step of the preparation that solid obtained after filtration is washed from a mixture of 1:0.35 tol:0.45 (V/V) mixtures of ethyl alcohol and water.
Drying step plays an important role in obtaining pure polymorphic form I, including the temperature at which drying is carried out. In present invention, drying of wet rifaximin is carried out at a temperature between 25-80 °C and preferably drying is carried out between 35 to 70 °C under vacuum or at atmospheric conditions. Further, presence of water in polymorph is checked for instance by Karl Fischer method, in order to check the amount of remaining water present in the product under drying. Also, time period of drying is another valuable step in addition to above criteria in order to obtain pure polymorphic form I, substantially free from the other polymorphic forms of rifaximin. The

present invention includes the step of washing solid from a mixture of ethyl alcohol and water, and is further dried at a temperature between 25-80 °C and preferably drying is carried out between 35 to 70 °C for a time period between 8-14 hours.
According to one another aspect, the present invention provides a process for conversion of polymorphic form I into other known polymorphs such as purely crystalline forms like α and ß; ; or amorphous form. Particularly polymorphic form I is converted to a form of rifaximin by dissolving form I in acetone at room temperature and stirring until complete dissolution of solid. Thereafter demineralized water is added to above solution at room temperature stirred the reaction mixture for 30 minutes at the same temperature for complete crystallization and filtered the suspension. The obtained solid is washed with a mixture of acetone and demineralized water (3.75:6.25) and dried under vacuum at 40 °C until water content is in the range of 2-4%, preferably 2-3%. Another aspect of the present invention is to provide a process for conversion of polymorphic form I to ß form of rifaximin by dissolving polymorphic form I in acetone at room temperature and stirring the reaction mixture until complete dissolution of solid at room temperature thereafter demineralized water is added to the above solution in 10 minutes, and stirred for 30 minutes at 25 to 35 °C for complete crystallization. The suspension is filtered and the obtained solid is washed with a mixture of acetone and demineralized water (3.75:6.25) and dried under vacuum at 40 °C until water content is higher than 4.5%, preferably between 5-6%.
Further another aspect of the present invention is to provide a process for conversion of polymorphic form I to y form of rifaximin which comprises dissolving polymorphic form I in ethyl alcohol by heating at reflux temperature until complete dissolution of solid. Adding demineralized water to the reaction mixture at the above temperature and brought the temperature to 40 °C during 30-40 minutes. As crystallization starts, seeded with rifaximin and maintained at this

temperature until plentiful crystallization. The temperature of the suspension is brought to 30 °C and kept at this temperature for 5 hours under stirring and further the temperature of the suspension is decreased to 0 °C in 60 minutes and stirred for a period of 2 to 6 hours. The suspension is filtered and washed the obtained solid with a mixture of ethyl alcohol and demineralized water (70:30) and dried under vacuum at 65 °C until water content is lower than 2 %. Another aspect of the present invention is to provide a process for conversion of rifaximin form I to amorphous rifaximin which involves dissolving rifaximin form I in dimethyl formamide by heating at 40 °C (±2) and stirring till the complete dissolution of solid. Demineralized water is added to the above reaction mixture at a temperature of 40 °C (±2) during the 10 minutes. After the complete addition of water, the resulted suspension is stirred for 30 minutes at 40 °C (±2) for complete crystallization. The suspension is filtered and the obtained solid is washed with a mixture of (dimethylformamide:demineralized water) of (3.75:6.25) ratio and dried under vacuum at 40 °C for 10-12 hours to get amorphous rifaximin.
Yet another aspect of the present invention is to provide a process for preparation of polymorphic form I from polymorphic forms α, ß and y which comprises dissolving any crystalline polymorph, a mixture of crystalline and amorphous or pure amorphous in ethyl alcohol and heating at the suspension at reflux temperature till complete dissolution of the solid and then 30 ml of demineralized water is added at this temperature and the temperature is brought down to 40 °C during 30 to 40 minutes. As crystallization starts, seeded with rifaximin form I and is kept at this temperature till complete crystallization. The temperature of the suspension is then brought to 30 °C and is kept at this temperature for 5 hours under stirring. Further temperature is decreased to 0 °C in 60 minutes and the suspension is immediately filtered. The solid is washed with a mixture of ethyl alcohol/demineralized water and dried under vacuum at 65 °C till water content is in the range of 2.0 to 4.5%, preferably 2-3%.

The ordinary person skilled in the art understands that experimental differences may arise due to differences in instrumentation, sample preparation or other factors. In this case the measure variability of the instrument for X-ray powder diffraction peaks was about (+) 0.2 °9.
X-ray powder diffraction analysis of polymorphic form I was measured on a PAN analytical X'pert pro diffractometer with Cu Ka radiation. The X-ray powder diffractometer was equipped with a X'Celerator detector with a 20 range of 120 °C. Real time data were collected using Cu Ka radiation starting at approximately 4 °20 at a resolution of 0-0083556 °29. The tube voltage and amperage were set to 45 kV and 40mA, respectively. The monochromator slit was set at 5 mm by 160 µm. The pattern is displayed from 3-40 °20. Samples were mounted on sample holder and allowed to spin. The samples were analyzed for 10 minutes. Instrument calibration was performed using a silicon reference standard. DSC analysis was performed using a TA instruments differential scanning calorimeter Mettler Toledo 823e. The sample was placed in an aluminum DSC pan and the accurate weight was recorded. The pan was covered with a lid [with or without pin hole] and then crimped. The sample cell was equilibrated at 25 °C and heated under a nitrogen purge at a rate of 10 °C/min or 5 °C/min up to 250 °C. Indium metal was used as the calibration standard. The temperature reported was at the transition maxima.
The evaluation of the water content present in the analyzed samples has always been carried out by means of the Karl Fischer method.
TGA analysis was performed using a TA instruments METTLER TOLENDO TGA/SDTA-851e thermogravimetric analyzer. Each sample was placed in an aluminum sample pan and inserted into the TG furnace. The furnace was started directly from ambient temperature and heated up to a temperature of 350 °C under nitrogen at a rate of 10 °C/min. Nickel and Alumel™ were used as the calibration standards.

During the phase of preservation of product, particular care has to be taken so that ambient conditions do not change the water content of product, by preserving product in an ambient having controlled humidity or in closed containers that do not allow a significant exchange of water with exterior ambient. When intrinsic dissolution of new form I of rifaximin was carried out against the form, it was observed that rifaximin has better dissolution profile than a form. Intrinsic dissolution test was carried out as described in the monograph 1087 at pages 2512-2513 of the USP (U.S. Pharmacopoeia) 27 by taking samples of 100mg of each rifaximin polymorph.
Particularly 100 mg of rifaximin polymorphs were put into a die and compressed for 1 minute under a pressure of 5 tons by means of a punch in a hydraulic press. A compacted pellet was formed in the die with a single face of defined area exposed on the bottom of the die so that from 50% to 75% of the compacted pellet could dissolve in an appropriate dissolution medium. The holder containing the die was mounted on a laboratory stirring device, immersed in a glass vessel containing a dissolution medium and rotated at a rotation speed of 100 rpm by means of the stirring device, while keeping the temperature of the dissolution medium at 37±0.5 °C. The dissolution medium contained in the glass vessel consisted of 1000ml of 0.1 M aqueous phosphate buffer pH 7.4 containing 4.5g of sodium lauryl sulphate and was kept at 37±0.5 °C for the whole duration of the test. Further solubility study data was generated by taking 4.5g each of rifaximin form I and alpha form and transfer the weighed material into a 500mL round bottom flask containing 300mL of dissolution media and started the solubility study following the conditions like temperature of 37 "C ±0.5 °C and rotation speed of 100rpm (using paddle type stirrer). Dissolution media is prepared by dissolving 27.5g sodium dihydrogen phosphate monohydrate and 9.0g of sodium lauryl sulphate in 4000mL of water and adjusted the pH 7.41 with sodium hydroxide solution.

Crude rifaximin used for the preparation of polymorphic form I is prepared by the known process as reported in US patent 4,557,866. In particular, process comprises reaction of rifamycin S with 2-Amino-4-methyl pyridine in dichloromethane at room temperature under nitrogen atmosphere. Optionally reaction can also be carried out in the presence of other halogenated solvents and nitriles solvents or mixture thereof. Further, iodine dissolved in dichloromethane is added drop wise at room temperature and stirred. Further, suitable reducing agent such as L(-) ascorbic acid in water is added at room temperature and then cooled to 10-15 °C. The pH of resulting solution is adjusted tol.5-2.0 with slow addition of an acid such as 15% solution of dilute hydrochloric acid under stirring. The reaction mass is stirred for another 15-20 min and the layers are separated. The organic layer is washed with demineralized water, sodium thiosulfate solution (10%) and finally washed with demineralized water till pH of the organic layer is neutral. The solvent is evaporated under vacuum at 30-50 °C to get a residue which is purified by crystallizing from a mixture of ethyl alcohol and water and dried at 35-40 °C to get crude rifaximin having purity >98% by HPLC. In another aspect crude rifaximin can be purified by making use of adsorbing agents like neutral alumina or silica gel. Generally, crude rifaximin can be dissolved in a suitable solvent selected from halogenated solvents such as dichloromethane, chloroform; C3-C8 aliphatic esters such as methylacetate, ethylacetate, isopropylacetate; ethers such as tetrahydrofuran, 2-methyl tetrahydrofuran, diisopropylether, methyl tertiary butyl ether, hydrocarbons such as toluene; C1-C6 alcohols such as methanol, ethanol, n-propanol, isopropanol at suitable temperature and treated with suitable adsorbing agents for a sufficient time. Particularly in the solution of crude rifaxinin in dichloromethane, neutral alumina is added and stirred for lhour and filtered. The solvent is then distilled off and resulting residue is dissolved in ethyl acetate. Rifaximin can optionally be purified by crystallizing from a mixture of ethyl alcohol and water to obtain rifaximin having purity ≥99.5% by HPLC.

The schematic representation for preparation of crude rifaximin is as follows:
(Formula Removed)
Rifamycin S can be purified prior to use, if desired, by recrystallization or by heating slurry at reflux temperature of solvent. Suitable solvent for purification can be selected from C1-C6 alcohols such as methanol, ethanol, n-propanol, isopropanol. Particularly rifamycin S and ethanol are stirred and re fluxed for 1 hour. The reaction mixture is then cooled slowly to ambient temperature, stirred for 2 hours and filtered. Optionally the filtered product can be washed with a suitable solvent selected from methanol, ethanol, acetone and n-hexane, n-heptane, cyclohexane or mixture there of. The product is then dried to obtain pure rifamycin S showing des acetyl impurity below to 0.6%.
Having described the invention with reference to certain preferred aspects, other aspects of the invention will become apparent to one skilled in the art from consideration of the specification. The invention is further defined by reference to the following examples describing in detail the preparation of the compounds of this invention. It will be apparent to those skilled in the art that many modifications, both to materials and methods, may be practiced without departing from the scope of the invention.
Examples Example 1: Purification of Rifamycin S
Rifamycin S (500g) and Ethanol (1.5L) were stirred and refluxed for 1 hour. The reaction mixture was then cooled slowly to ambience, stirred at this temperature

for 2 hour and filtered. The product dried in vacuum oven at 40 °C to obtain 475g of pure Rifamycin S showing the des acetyl impurity below to 0.6%. Example 2: Preparation of rifaximin
Rifamycin S (300 g) was stirred in dichloromethane (900 ml) at room temperature for 15 minutes to get a clear solution and then 2-Amino-4-methyl pyridine (139.2g) was added at room temperature under nitrogen atmosphere. Iodine (57.0g) dissolved in dichloromethane (2100ml), was added drop wise in 30-45 minutes at room temperature. The reaction mass was stirred for 22-24 hours at 25-30 °C. After completion of the reaction, a 20% solution of L(-) ascorbic acid in water (300 ml) was added. The reaction mixture was stirred for 45-60 minutes at room temperature and then cooled to 10-15 oC. The pH of the resulting solution was adjusted to 1.5-2.0 with slow addition of dilute hydrochloric acid under stirring. The reaction mass was stirred for 15-20 minutes and layers were separated. The organic layer was washed with demineralized water (1500 ml), 10% sodium thiosulfate solution (1500 ml) and with demineralized water till pH was neutral. The solvent was distilled off under vacuum at 40-45 °C to get a residue which was taken in cyclohexane (1500 ml) and stirred for 1 hour. The resulting solid was filtered, washed with cyclohexane (300 ml) crystallized from a mixture of ethyl alcohol and water (600ml; 420ml ethyl alcohol and 180 ml water) to get 240g of crude rifaximin having purity 99.3% by HPLC. Example 3: Preparation of rifaximin Step-1: Preparation of crude rifaximin
Rifamycin S (300 g) was stirred in dichloromethane (900 ml) at room temperature for 15 minutes to get a clear solution and then 2-amino-4-methyl pyridine (139.2g) was added at room temperature under nitrogen atmosphere. Iodine (57.0g) dissolved in dichloromethane (2100ml), was added drop wise in 30-45 minutes at room temperature and was stirred for 22-24 hours. After completion of the reaction, a 20% solution of L (-) ascorbic acid in water (300 ml) was added and stirred for 45-60 minutes. The reaction mass was cooled to 10-15 °C and pH

of the resulting solution was adjusted to 1.5-2.0 with slow addition of dilute hydrochloric acid under stirring. The reaction mass was stirred for 15-20 minutes and layers were separated and the organic layer was washed with demineralized water (1500 ml), with 10% sodium thiosulfate solution (1500 ml) and demineralized water till pH was neutral. The solvent was distilled off under vacuum at 40-45 °C to obtain a residue which was crystallized from a mixture of ethyl alcohol and water (378ml ethyl alcohol and 162 ml water) and dried at 35-40 °C to obtain 240g crude rifaximin having purity 98.8% by HPLC. Step-2: Purification of crude rifaximin
Crude rifaximin (240g) was stirred in dichloromethane (2400ml) at room temperature, a neutral alumina (240g) was added, stirred for 1 hour and filtered. The solvent was then distilled off and residue was treated with ethyl acetate (2400ml) and stirred to dissolution. The resulting residue was crystallized from a mixture of ethyl alcohol and water (302ml ethyl alcohol and 130ml water) and dried at 35-40 °C to obtain 192g of rifaximin having purity 99.8% by HPLC. Example 4: Preparation of polymorphic form I.
To crude rifaximin (50g) ethyl alcohol (70 ml) was added at room temperature. The reaction mixture was heated to reflux temperature until complete dissolution. To the above solution 30 ml of demineralized water was added slowly at same temperature and the reaction mixture was further stirred for 15 minutes. After complete addition of water, the temperature was brought down to 30 °C during 40 to 50 minutes and then kept at this temperature for 5 hours under stirring till complete crystallization. The reaction mixture was cooled to 0 °C in 45-60 minutes and immediately filtered. The resulting solid was washed with a mixture of ethyl alcohol and demineralized water (75 ml; 1:0.43) and dried under vacuum at 65 °C for 12 hours, m/c -2.5%. Example 5: Preparation of polymorphic form I.
Crude rifaximin (50g) was taken in a mixture of ethyl alcohol (70 ml) and demineralized water (30 ml) at room temperature and suspension was heated to

80±2 °C. After complete dissolution, the temperature of reaction solution was brought down to 30 °C during 40 to 50 minutes. The reaction mixture was stirred for 5 hours maintaining same temperature till complete crystallization. Further the temperature of reaction mixture was cooled to 0 °C during 60 minutes and immediately filtered. The resulting solid was washed with a solution of mixture of ethyl alcohol and demineralized water (75 ml) and dried under vacuum at 65 °C for 12 hours.
Example 6: Preparation of polymorphic form I.
To crude rifaximin (50g) ethyl alcohol (70 ml) was added at room temperature. The reaction mixture was heated to reflux temperature until complete dissolution. To the above solution, 30 ml of demineralized water was added slowly at same temperature and reaction mixture was further stirred for 15 minutes. Thereafter, temperature of reaction mixture was brought down to 40 °C during 30 to 40 minutes. The reaction mixture was seeded with rifaximin form I and stirred for 10-15 minutes at the same temperature. The temperature is decreased to 30 °C and maintained at this temperature for 5 hours under stirring to get complete crystallization. Then reaction mixture was cooled to 0 °C during 45-60 minutes and immediately filtered. The solid was washed with a solution of a mixture of ethyl alcohol and demineralized water (75 ml) and dried under vacuum at 65 °C for 12 hours.
Example 7: Preparation of polymorphic form I.
To crude rifaximin (50g) ethyl alcohol (70 ml) was added at room temperature. The reaction mixture was heated to reflux temperature until complete dissolution. To the above solution 30 ml of demineralized water was added slowly at the same temperature and the reaction mixture was further stirred for 15 minutes. Thereafter, temperature of reaction mixture was brought down to 30 °C during 40 to 50 minutes and maintained at this temperature for 5 hours under stirring till complete crystallization. The reaction mixture was cooled to 0 °C during 45-60 minutes and immediately filtered. The solid was washed with a solution of a

mixture of ethyl alcohol and demineralized water (75 ml) and dried under vacuum
at 65 oC for 14 hours.
Example 8: Preparation of polymorphic form I.
A slurry of rifaximin (50g) and ethyl alcohol (105ml) was heated to reflux until complete dissolution. Water (45ml) was added slowly at same temperature to the above clear solution and further stirred for 15 minutes. The reaction mixture was cooled to 30 °C over a period of 1 hour and stirred for 3 hours. The reaction mixture was further cooled to 0 °C in 1 hour and filtered. The resulting solid was washed with a mixture of ethyl alcohol and water (1:0.43) and dried under vacuum at 35-40 °C for 13 hours, m/c-2.3% Example 9: Preparation of polymorphic form I.
A stirred slurry of rifaximin (50g) and ethyl alcohol (105ml) was heated to reflux until complete dissolution. Water (45ml) was added slowly at same temperature to the above clear solution and stirred for 15 minutes. The reaction mixture was cooled to 30 °C in over a period of 1 hour and stirred for 3 hours. The reaction mixture was further cooled to 0 °C in 1 hour and filtered. The resulting solid was washed with a mixture of ethyl alcohol and water (1:0.43) and dried under atmospheric conditions at room temperature for 14 hours, m/c-2.8% Example 10: Preparation of polymorphic form I from crystalline a form Crystalline rifaximin a (10g) was added in ethyl alcohol (14 ml) at room temperature. The reaction mixture was heated at reflux temperature until complete dissolution. To the above solution, demineralized water (6 ml) was added at the same temperature. After complete addition of water, the temperature was brought to 40 °C during 30 to 40 minutes. The reaction mixture was seeded with rifaximin and kept at this temperature until plentiful crystallization. The temperature of the suspension was then brought to 30 °C and maintained at this temperature for 5 hours under stirring. Further the temperature was decreased to 0 °C in 60 minutes and the suspension was immediately filtered. The solid was washed with a

mixture of ethyl alcohol/demineralized water (15 ml) and dried under vacuum at
65 °C for 12 hours.
Example 11: Preparation of crystalline a form from polymorphic form I
Rifaximin form I (20g) was added in acetone (14 ml) at room temperature. The reaction mixture was stirred until complete dissolution at the same temperature. To the above solution, demineralized water (100 ml) was added in 10 minutes. Increasing the temperature to 25-30 °C and stirred for 30 minutes at the same temperature for complete crystallization. The suspension is filtered and the solid obtained is washed with a mixture of acetone and demineralized water (40 ml) and dried under vacuum at 40 °C for 9 hours.
Example 12: Preparation of crystalline ß -form from polymorphic form I Rifaximin form I (20g) was added in acetone (14 ml) at room temperature. The reaction mixture was stirred until complete dissolution at the same temperature. To the above solution, demineralized water (100 ml) was added in 10 minutes. Increasing the temperature to 25-30 °C and stirred for 30 minutes at the same temperature for complete crystallization. The suspension is filtered and the solid obtained is washed with a mixture of acetone and demineralized water (40 ml) and dried under vacuum at 40 °C for 6 hours.

We Claim:
1. Polymorphic form I of rifaximin.
2. Polymorphic form I of rifaximin characterized by the following:
a.) powder x-ray diffraction pattern comprising peaks at least at 29,
5.2,7.2, 8.4 and 8.7 degrees ±0.2, b.) thermogravemetric analysis which shows water content within the
range of 2.0 to 4.5%.
3. Polymorphic form I of rifaximin characterized by the following:
a.) powder x-ray diffraction pattern comprising peaks at least at 29,
5.2,7.2, 8.4 and 8.7 degrees ±0.2, b.) thermogravemetric analysis which shows water content within the
range of 2.0 to 4.5%, c.) DSC peaks showing endotherm in the range of 124-143 °C.
4. A process for the preparation of polymorphic form I of rifaximin comprising
the steps of:
a.) preparing a solution of rifaximin in a mixture of ethyl alcohol and
demineralized water at reflux temperature; b.) cooling the reaction solution at temperature 40 °C; c.) optionally seeding the reaction mixture with rifaximin form I; d.) lowering temperature of solution to 30 °C; e.) stirring reaction mixture for 1-5 hours; f.) cooling the reaction mixture to 0 °C slowly; g.) filtering the reaction mixture within 6 hours; h.) washing the resulting solid with a mixture of ethyl alcohol and
demineralized water; i.) drying the resulting solid at a temperature between 25 to 80 °C for
sufficient time till water content is lower than 4.5%.
5. The process according to claim 4, wherein in step a) the ratio of ethyl alcohol and demineralized water is in proportion of 1: 0.35 to 1: 0.45 (V/V).

6. The process according to claim 4, wherein in step g) reaction mixture is
filtered immediately.
7. The process according to claim 4, wherein in step h) the mixture of ethyl alcohol and demineralized water used for washing, is in the ratio of 1:0.43.
8. The process according to claim 4, wherein in step i) drying is carried out under vacuum.
9. The process according to claim 4, wherein in step i) drying is carried out under atmospheric conditions.
10. The process according to claim 4, wherein in step i) drying is carried out till water content reaches within the range of 2.0-4.5%.