New Drug Approvals

Home » Posts tagged 'New molecules'

Tag Archives: New molecules

DRUG APPROVALS BY DR ANTHONY MELVIN CRASTO .....FOR BLOG HOME CLICK HERE

Blog Stats

  • 2,932,999 hits

Flag and hits

Flag Counter

Enter your email address to follow this blog and receive notifications of new posts by email.

Join 2,504 other followers

Follow New Drug Approvals on WordPress.com

Archives

Categories

Flag Counter

ORGANIC SPECTROSCOPY

Read all about Organic Spectroscopy on ORGANIC SPECTROSCOPY INTERNATIONAL 

Enter your email address to follow this blog and receive notifications of new posts by email.

Join 2,504 other followers

DR ANTHONY MELVIN CRASTO Ph.D

DR ANTHONY MELVIN CRASTO Ph.D

DR ANTHONY MELVIN CRASTO, Born in Mumbai in 1964 and graduated from Mumbai University, Completed his Ph.D from ICT, 1991,Matunga, Mumbai, India, in Organic Chemistry, The thesis topic was Synthesis of Novel Pyrethroid Analogues, Currently he is working with GLENMARK PHARMACEUTICALS LTD, Research Centre as Principal Scientist, Process Research (bulk actives) at Mahape, Navi Mumbai, India. Total Industry exp 30 plus yrs, Prior to joining Glenmark, he has worked with major multinationals like Hoechst Marion Roussel, now Sanofi, Searle India Ltd, now RPG lifesciences, etc. He has worked with notable scientists like Dr K Nagarajan, Dr Ralph Stapel, Prof S Seshadri, Dr T.V. Radhakrishnan and Dr B. K. Kulkarni, etc, He did custom synthesis for major multinationals in his career like BASF, Novartis, Sanofi, etc., He has worked in Discovery, Natural products, Bulk drugs, Generics, Intermediates, Fine chemicals, Neutraceuticals, GMP, Scaleups, etc, he is now helping millions, has 9 million plus hits on Google on all Organic chemistry websites. His friends call him Open superstar worlddrugtracker. His New Drug Approvals, Green Chemistry International, All about drugs, Eurekamoments, Organic spectroscopy international, etc in organic chemistry are some most read blogs He has hands on experience in initiation and developing novel routes for drug molecules and implementation them on commercial scale over a 30 year tenure till date Dec 2017, Around 35 plus products in his career. He has good knowledge of IPM, GMP, Regulatory aspects, he has several International patents published worldwide . He has good proficiency in Technology transfer, Spectroscopy, Stereochemistry, Synthesis, Polymorphism etc., He suffered a paralytic stroke/ Acute Transverse mylitis in Dec 2007 and is 90 %Paralysed, He is bound to a wheelchair, this seems to have injected feul in him to help chemists all around the world, he is more active than before and is pushing boundaries, He has 9 million plus hits on Google, 2.5 lakh plus connections on all networking sites, 50 Lakh plus views on dozen plus blogs, He makes himself available to all, contact him on +91 9323115463, email amcrasto@gmail.com, Twitter, @amcrasto , He lives and will die for his family, 90% paralysis cannot kill his soul., Notably he has 19 lakh plus views on New Drug Approvals Blog in 216 countries......https://newdrugapprovals.wordpress.com/ , He appreciates the help he gets from one and all, Friends, Family, Glenmark, Readers, Wellwishers, Doctors, Drug authorities, His Contacts, Physiotherapist, etc

Personal Links

Verified Services

View Full Profile →

Archives

Categories

Flag Counter

WCK ? New molecules from Wochkardt to treat bacterial infections


(2S, 5R)-7-OXO-N-[(3S)-PYRROLIDIN-3-YLOXY]-6-(SULFOOXY)-1,6-DIAZABICYCLO [3.2.1]OCTANE-2-CARBOXAMIDE

  • (2S,5R)-7-Oxo-N-((3S)-pyrrolidin-3-yloxy)-6-(sulfooxy)-1,6-diazabicyclo[3.2.1]octane-2-carboxamide
  • C11 H18 N4 O7 S, 350.35
  • Sulfuric acid, mono[(1R,​2S,​5R)​-​7-​oxo-​2-​[[[(3S)​-​3-​pyrrolidinyloxy]​amino]​carbonyl]​-​1,​6-​diazabicyclo[3.2.1]​oct-​6-​yl] ester

CAS 1452458-72-8

KEEP WATCHING THIS POST

 

SODIUM SALT CAS 1629221-44-8

Sulfuric acid, mono[(1R,​2S,​5R)​-​7-​oxo-​2-​[[[(3S)​-​3-​pyrrolidinyloxy]​amino]​carbonyl]​-​1,​6-​diazabicyclo[3.2.1]​oct-​6-​yl] ester, sodium salt (1:1)

Patent

WO 2015110886

http://www.google.com/patents/WO2015110886A1?cl=en

Formula (II) Formula (III) Formula (IV)

Hydrogenolysis

Formula (I)

Scheme – 1

 

 

Formula (VII) Formula (VIII)

Hydrazine hydrate

Formula I

Scheme – 2

 

Example 1

Synthesis of tert-butyl (3S)-2-(aminooxy)pyrrolidine-l-carboxylate (III):

Step 1; Preparation of 3-(R)-hydroxypyrrolidine hydrochloride (VIII):

To a stirred suspension of commercially available (25, 4i?)-4-hydroxy-2-pyrrolidinecarboxylic acid (L-hydroxyproline) (VII) (100 g, 0.762 mol) in anhydrous cyclohexanol (500 ml), was added 2-cyclohexen-l-one (5 ml). The resulting mixture was heated under reflux at about 154°C for about 48 hour. The obtained clear solution was allowed to cool to room temperature and then was cooled further to 10°C. To this, about 15 % solution of hydrochloric acid in ethanol (234 ml) was added and then stirred for 30 minutes. The separated solid was filtered under suction and washed with ethyl acetate (2 x 100 ml). The solid was dried under reduced pressure to obtain 47.5 g of 3-(R)-hydroxypyrrolidine hydrochloride (VIII) in 51 % yield. The solid was used without further purification in the next step.

Analysis:

Mass: 87.8 (M+l) as free base; for Molecular weight of 123.57 and Molecular Formula of C4Hi0ClNO; and

1H NMR (400MHz, DMSO): 5 9.58 – 9.32 (brd, 2H), 5.36 (brs, 1H), 4.36 – 3.39 (brs, 1H), 3.17 (brs, 2H), 3.11-2.96 (dd, 2H), 1.90 – 1.81 (m, 2H).

Step 2: Preparation of (3R)-l-(tert-butoxycarbonyl)-3-hydroxypyrrolidine (IX):

To a stirred suspension of 3-(i?)-hydroxypyrrolidine hydrochloride (VIII) (110 g, 0.9 mol) in dichloromethane (1100 ml), triethylamine (273 g, 2.7 mol) was added at 0-5°C. After 5 minute of stirring di-feri-butyldicarbonate [(Boc)20] (245 g, 1.125 mol) was added to the reaction mixture in small portions, followed by 4-dimethylaminopyridine (10.99 g, 0.09 mol). The reaction mixture was stirred for 2 hour and then poured in to water (1100 ml). The organic layer was separated and washed with saturated ammonium chloride solution (1×1100 ml) and water (1100 ml). The organic layer was dried over anhydrous sodium sulphate and the solvent evaporated under reduced pressure. The residue was purified by silica gel (60-120 mesh) column chromatography using 1-5% mixtures of acetone: hexane as an eluent. The combined fractions were evaporated, to obtain the 118 g of (3i?)-l-(ieri-butoxycarbonyl)-3-hydroxypyrrolidine (IX), as a white solid, in 71 % yield.

Analysis:

Melting point: 55 – 58°C;

Mass: 188 (M+l); for Molecular Weight of 187.24 and Molecular Formula of C9H17N03; and

1H NMR (400MHz, CDC13): 54.428 – 4.424 (s, 1H), 3.46 – 3.43 (m, 2H), 3.37 -3.28 (m, 2H), 2.36 – 2.30 (d, 1H), 2.00 – 1.86 (m, 2H), 1.44 (s, 9H).

Step 3: Preparation of (5)-3-[(l,3-dihydro-l,3-dioxo-isoindol-2-yl)oxy]pyrrolidine-l-carbox lic acid tert- butyl ester (X):

To a stirred solution of di-isopropyl azodicarboxylate (97.17 g, 0.481 mol) in tetrahydrofuran (1200 ml), a solution triphenyl phosphine (125.9 g, 0.481 mol) in tetrahydrofuran (300 ml) was added at temperature below -10°C. The resulting reaction mixture was stirred for further 45 minute at the same condition and a solution of (3i?)-l-(ieri-butoxycarbonyl)-3-hydroxypyrrolidine (IX) (60 g, 0.3204 mol) in tetrahydrofuran (300 ml) was added over a period of 15 minute. After another 45 minute of stirring, N-hydroxy phthalimide (52.4 g, 0.3204mol) was added in one portion to the reaction mass. The reaction mixture was allowed to warm to room temperature and stirred for 16 hour.

The completion of the reaction was monitored by thin layer chromatography. After completion of reaction, the solvent was evaporated under reduced pressure. The residue thus obtained was stirred with di-isopropyl ether (600 ml). The precipitate formed was filtered under suction. The filtrate was concentrated under reduced pressure and the residual mass was purified by silica gel (60-120 mesh) column chromatography using 1-5 % mixtures of acetone: hexane as an eluent. The solvent from the combined fractions was evaporated to obtain 63 g of (5)-3-[(l,3-dihydro-l,3-dioxo-isoindol-2-yl)oxy]pyrrolidine-1-carboxylic acid tert-buty\ ester (X), as a white solid, in 59% yield.

Analysis:

Melting point: 112-115°C;

Mass: 333.2 (M+l); for Molecular Weight of 332.36 and Molecular Formula of

1H NMR (400 MHz, CDC13): 57.86-7.83 (m, 2H), 7.78-7.75 (m, 2H), 4.99 – 4.94 (d, 1H), 3.80 – 3.68 (m, 2H), 3.60 – 3.53 (m, 2H), 2.28-2.25 (m, 1H), 2.02 (m, 1H), 1.48 (s, 9H).

Step 4: Preparation of tert-butyl (35)-2-(aminooxy)pyrrolidine-l-carboxylate (III):

To a stirred suspension of the (5)-3-[(l,3-dihydro-l,3-dioxo-isoindol-2-yl) oxy]pyrrolidine-l-carboxylic acid tert-buty\ ester (X) (12.68 g, 0.0381 mol) in dichloromethane (200 ml) was added 99% hydrazine hydrate (3.81 g, 0.0762 mol) drop-wise over a period of 30 minutes, at 25°C. After 2 hour of stirring, the separated solid was filtered and washed with dichloromethane (2 x 50 ml). The filtrate and washings were combined and washed with water (2 x 65 ml) and finally with brine (1 x 65 ml). The organic layer was dried over anhydrous sodium sulphate and the solvent was evaporated under reduced pressure to obtain 7.71 g of tert-buty\ (3S)-2-(aminooxy pyrrolidine- 1-carboxylate (III) as pale yellow oil.

Analysis:

Mass: 203 (M+l); for Molecular Weight of 202.26 and Molecular Formula of C9H18N203.

Example 2

Synthesis of (25, 5R)-7-oxo-N-r(35)-pyrrolidin-3-yl-oxyl-6-(sulfooxy)-l,6-diaza bicyclor3.2. lloctane-2-carboxamide (I) :

Step 1: Preparation of fert-butyl-(35)-3-[({[25, 5R)-6-(benzyloxy)-7-oxo-l,6-diazabicylco[3.2.1]oct-2-yl]carbonyl}amino)oxy]pyrrolidine-l-carboxylate (IV):

To a clear, stirred solution of sodium (25, 5i?)-6-(benzyloxy)-7-oxo-l,6-diazabicyclo[3.2.1]octane-2-carboxylate (II) (11.38 g, 0.0382 mol) in water (114 ml), was added EDC.HC1 (18.24 g, 0.0955 mol) at 15°C, in small portions. After 10 minutes, a solution of feri-butyl-(35)-3-(aminooxy) pyrrolidine- 1-carboxylate (III, 7.72 g, 0.0382 mol), prepared as per the literature procedure: US5233053, Chemistry Letters, 893-896, (1986) and depicted in scheme 2), in dimethylformamide (24 ml) was added drop wise, to the above stirred solution, at about 10°C. The reaction mass was allowed to warm to 25°C and HOBt (5.15 g, 0.0382 mol) was added in small portions over a period of 15 minutes and the reaction mixture was stirred further at room temperature for 16 hour. After completion of the reaction (monitored by thin layer chromatography using solvent system acetone: hexane (35:65)) the resulting mixture was filtered and the residue was washed with water (120 ml). The residual white solid was suspended in fresh water (120 ml) and the mixture stirred at 50°C, for 3 hour. The resulting suspension was filtered and the residual solid dried under reduced pressure to obtain 16.1 g of tert-buty\ (35)-3-[({ [25,5i?)-6-(benzyloxy)-7-oxo-l,6-diazabicylco[3.2.1]oct-2-yl]carbonyl}amino) oxy]pyrrolidine- 1-carboxylate (IV) as off white solid in 92% yield.

Analysis:

Mass: 461.3 (M+l); for Molecular weight of 460.53 and Molecular formula of

1H NMR (400MHz, CDC13): δ 9.08-9.03 (d, 1H), 7.43-7.36 (m, 5H), 5.06-4.88 (dd, 2H), 4.63-4.57 (d, 1H), 3.97-.396 (d, 1H), 3.64-3.53 (m, 2H), 3.47-3.37 (m, 2H), 3.31 (s, 1H), 3.02-2.99 (d, 1H), 2.75-2.73 (d, 1H), 2.29(m, 2H), 2.18-2.15 (m, 1H), 2.01-1.90 (m, 3H), 1.66 (m, 1H), 1.46 (s, 9H).

Step 2: Preparation of tert-butyl-(35)-3-[({[25,5R)-6-hydroxy-7-oxo-l,6-diazabicylco

[3.2.1]oct-2-yl]carbonyl}amino)oxy]pyrrolidine-l-carboxylate (V):

ieri-Butyl-(35)-3-[({ [25,5R)-6-(benzyloxy)-7-oxo-l,6-diazabicylco[3.2.1]oct-2-yl]carbonyl}amino)oxy]pyrrolidine-l-carboxylate (IV) (10 g, 0.02171 mol) was dissolved in a mixture of dimethylformamide and dichloromethane ( 1 : 1 , 50 ml : 50 ml) to obtain a clear solution. To this solution, was added 10% palladium on carbon (2.5 g, 50% wet) catalyst. The suspension was stirred for 4 hour, at 50 psi hydrogen atmosphere, at 25°C. After completion of the reaction (monitored by thin layer chromatography), the resulting mixture was filtered through a celite pad. The residue was washed with dichloromethane (50 ml). The solvent from the combined filtrate was evaporated under reduced pressure to obtain 8.04 g of ieri-butyl(35)-3-[({ [25,5i?)-6-hydroxy-7-oxo-l,6-diazabicylco[3.2.1]oct-2-yl]carbonyl

amino)oxy]pyrrolidine-l-carboxylate (V) as oil. This was used as such for the next reaction without further purification.

Analysis:

Mass: 371.2 (M+l); for Molecular Weight of 370.4 and Molecular Formula of

Step 3: Preparation of tert-butyl-(35)-3-[({[25,5R)-6-(sulfooxy)-7-oxo-l,6-diazabicylco[3.2.1]oct-2-yl]carbonyl}amino)oxy]pyrrolidine-l-carboxylate, tetrabutyl ammonium salt (VI):

To a stirred solution of ieri-butyl(35)-3-[({ [25,5i?)-6-hydroxy-7-oxo-l,6-diazabicylco[3.2.1]oct-2-yl]carbonyl}amino)oxy]pyrrolidine-l-carboxylate (V) (8.04 g, 0.0217 mol) in dimethylformamide (50 ml), was added sulfur trioxide dimethyl formamide complex (3.98 g, 0.0260 mol) in one portion, at about 10°C. The stirring was continued further for 30 minute and then the reaction mixture was allowed to warm to room temperature. After 2 hour, a solution of tetrabutylammonium acetate (7.83 g, 0.0260 mol) in water (25.8 ml) was added to the resulting reaction mass under stirring. After additional 2 hour of stirring, the solvent from the reaction mixture was evaporated under reduced pressure to obtain an oily residue. The oily mass was co-evaporated with xylene (2 x 20 ml) to obtain thick mass. This mass was partitioned between dichloromethane (100 ml) and water (100 ml). The organic layer was separated and the aqueous layer re-extracted with dichloromethane (50 ml). The combined organic extracts were washed with water (3 x 50 ml), dried over anhydrous sodium sulphate and the solvent evaporated under reduced pressure. The residual oily mass was triturated with ether (3 x 50 ml), each time the ether layer was decanted and finally the residue was concentrated under reduced pressure to obtain 11.3 g of tert-butyl(3S)-3-[({ [2S,5R)-6-(sulfooxy)-7-oxo-l,6-diazabicylco[3.2.1]oct-2-yl]carbonyl}amino)oxy] pyrrolidine- 1-carboxylate, tetrabutylammonium salt (VI), as a white foam, in 75 % yield.

Analysis:

Mass: 449.3 (M-l, without TBA); for Molecular weight of 691.94 and Molecular formula of C32H61N5O9S; and

1H NMR (400MHz, CDC13): 59.14-9.10 (d, 1H), 4.63 (s, 1H), 4.35 (s, 1H), 3.94-3.92 (d, 1H), 3.66-3.35 (m, 5H), 3.29-3.27 (m, 8H), 2.83-2.80 (d, 1H), 2.35-2.17 (m, 3H), 1.98-1.87 (m, 2H), 1.73 (m, 1H), 1.70-1.62 (m, 8H), 1.49-1.40 (m, 17H), 1.02-0.99 (t, 12H).

Step 4: Preparation of (25,5R)-7-oxo-iV-[(35)-pyrrolidin-2-yl-oxy]-6-(sulfooxy)-l,6-diazabicyclo [3.2.1]octane-2-carboxamide (I):

To a stirred solution of ieri-butyl(35)-3-[({ [25,5i?)-6-(sulfooxy)-7-oxo-l,6-diazabicylco[3.2.1]oct-2-yl]carbonyl}amino)oxy]pyrrolidine-l-carboxylate tetrabutyl ammonium salt (VI) (11 g, 0.0158 mol) in dichloromethane (55 ml), was added trifluoroacetic acid (55 ml) drop wise at about -10 °C over a period of 1 hour. After 1 hour of stirring, the resulting mixture was poured into hexane (550 ml), stirred well for 30 minute and the separated oily layer was collected. This procedure was repeated one more time and finally the combined oily layer was added to diethyl ether (110 ml) under vigorous stirring, at about 25 °C. The ether layer was removed by decantation from the precipitated solid. This procedure was repeated twice again with diethyl ether (2 x 110 ml). The solid thus obtained was stirred with fresh dichloromethane (110 ml) for 30 minutes and filtered. The residual solid was dried at about 45 °C under reduced pressure to obtain 5.7 g of (25,5i?)-7-oxo-N-[(35)-pyrrolidin-2-yl-oxy]-6-(sulfo-oxy)- l,6-diaza bicyclo[3.2.1] octane-2-carboxamide (I), as a white amorphous solid having XRPD as shown in Figure 1.

Analysis:

Mass: 349.2 (M-l); for Molecular Weight of 350.35 and Molecular Formula of

1H NMR (400MHz, DMSO-D6): δ 11.44 (brs, 1H), 8.80 (brs, 2H), 4.64-4.63 (m, 1H), 4.00 (s, 1H), 3.78-3.77 (d, 1H), 3.38-3.23 (m, 4H), 3.03-2.93 (dd, 2H), 2.48-2.11 (m, 1H), 2.00- 1.94 (m, 2H), 1.88- 1.86 (m, 1H), 1.71-1.65 (m, 2H).

Example 3

Preparation of Crystalline Form I of (25,5R)-7-oxo-jV-r(35)-pyrrolidin-2-yl-oxyl-6-(sulfooxy)-l,6-diaza bicyclor3.2.11 octane-2-carboxamide:

The solid (5 g) obtained in Step 4 of Example 2 was dissolved in water (30 ml) with stirring. To this solution, Isopropanol (210 ml) was slowly added at 25 °C and stirred for 12 hours. The separated solid was filtered and washed with additional isopropanol ( 10 ml) and dried under reduced pressure to obtain 3.9 g of (25,5i?)-7-oxo-N-[(35)-pyrrolidin-2-yl-oxy]-6-(sulfo-oxy)-l,6-diazabicyclo[3.2.1]octane-2-carboxamide as crystalline Form I, having XRPD as shown in Figure 2, in 78 % yield.

Analysis:

Purity as determined by HPLC: 95.56 %; and

X-ray powder diffraction pattern comprising peak at (2 Theta Values): 10.57 (± 0.2), 12.01 (± 0.2), 13.61 (± 0.2), 15.47 (± 0.2), 17.86 (± 0.2), 18.34 (± 0.2), 19.09 (± 0.2), 19.81 (± 0.2), 22.69 (± 0.2), 24.79 (± 0.2), 27.22 (± 0.2) and 33.41 (± 0.2)

//////

WCK ? , WCK Series by Wockhardt for treating the bacterial infection


 

(2S,5R)-7-0X0-N-[(2S)-PYRROLLIDIN-2-YL-METHYLOXY]-6-(SULFOOXY)-1,6-DIAZABICYCLO[3.2.1 ]OCTANE-2-CARBOXAMIDE

(2S,5R)-7-Oxo-N-((2S)-pyrrolidin-2-ylmethyloxy)-6-(sulfooxy)-1,6-diazabicyclo[3.2.1]octane-2-carboxamide

Sulfuric acid, mono[(1R,​2S,​5R)​-​7-​oxo-​2-​[[[(2S)​-​2-​pyrrolidinylmethoxy]​amino]​carbonyl]​-​1,​6-​diazabicyclo[3.2.1]​oct-​6-​yl] ester

KEEP WATCHING THIS POST

MW 364.37, C12 H20 N4 O7 S

CAS 1452459-04-9 FREE FORM

CAS Na SALT 1572988-44-3

Sulfuric acid, mono[(1R,​2S,​5R)​-​7-​oxo-​2-​[[[(2S)​-​2-​pyrrolidinylmethoxy]​amino]​carbonyl]​-​1,​6-​diazabicyclo[3.2.1]​oct-​6-​yl] ester, sodium salt (1:1)

 

PATENTS, WO 2015079329, WO 2015079389 , WO 2015063714, US 20130225554

Emergence of bacterial resistance to known antibacterial agents is becoming a major challenge in treating bacterial infections. One way forward to treat bacterial infections, and especially those caused by resistant bacteria, is to develop newer antibacterial agents that can overcome the bacterial resistant. Coates et al. (Br. J. Pharmacol. 2007; 152(8), 1147-1154.) have reviewed novel approaches to developing new antibiotics. However, the development of new antibacterial agents is a challenging task. For example, Gwynn et al. (Annals of the New York Academy of Sciences, 2010, 1213: 5-19) have reviewed the challenges in discovery of antibacterial agents.

Several compounds have been described in the prior art for use in treatment of bacterial infections (for example, see Patent Application Nos. PCT/IB2012/054296, PCT/IB2012/054290, US20130225554, PCT/US2010/060923, PCT/EP2010/067647, PCT/US2010/052109, PCT/US2010/048109, PCT/GB2009/050609, PCT/EP2009/056178, PCT/US2009/041200, PCT/US2013/034562, PCT/US2013/034589, PCT/IB2013/053092 and PCT/IB2012054706). However, there remains a need for potent antibacterial agents for preventing and/or treating bacterial infections, including those caused by bacteria that are resistant to known antibacterial agents.

PATENT

WO 2015079329

https://encrypted.google.com/patents/WO2015079329A2?cl=en

Formula (I)

Scheme -1

Formula (VII) Formula (VIII)

Formula (III) Formula (X) Formula (IX)

Scheme 2

 

Example 1

Synthesis of fert-butyl (25)-2-r(aminooxy)methyllpyrrolidine-l-carboxylate

Step 1: Synthesis of l-(tert-butoxycarbonyl)-(25)-pyrrolidine-2-carboxylic acid (VII):

To a stirred suspension of (2S)-pyrrolidine-2-carboxylic acid (L-proline) (200 g, 1.73 mol) in 1,4-dioxan and water mixture (1: 1, 1000 ml : 1000 ml) was added a solution of sodium hydroxide (138.97 g, 3.47 mol in 740 ml water) over a period of 20 minutes at 0 °C. Bi-feri-butyl dicarbonate (415.3 ml, 1.9 mol in 400 ml 1,4-dioxan) was added to the resulting clear solution over a period of 30 minutes, at temperature of about 0-5 °C. The reaction mixture was allowed to warm to room temperature and stirred for 16 hours. After completion of the reaction (monitored by thin layer chromatography), the reaction mixture was concentrated to 40 % of the initial volume under reduced pressure at 40-50 °C. The pH of the residual mixture was adjusted to 2 – 2.5 using 30 % aqueous potassium hydrogen sulphate at 15 °C under continuous stirring. The separated solid was filtered under suction and washed with water (2×400 ml) and dried under reduced pressure (4 mm Hg), to obtain 370 g of l-(ieri-butoxycarbonyl)-(25)-pyrrolidine-2-carboxylic acid (VII) as white solid.

Analysis:

Mass: 216 (M+l), for Molecular Weight: 215.24 and Molecular Formula:

1H NMR (400 MHz, CDC13): δ 10.60 (s, 1H), 4.35-4.24 (dd, 1H), 3.54-.3.34 (M, 2H), 2.27-1.91 (unresolved, 4H), 1.47-1.41 (d, 9H);

Purity as determined by HPLC: 99.92 %.

Step 2: Synthesis of tert-iutyl-(25)-2-(hydroxymethyl)-pyrrolidine-l-carboxylate (IX):

N-Methylmorpholine (113 ml, 1.114 mol) was added to the suspension of \-{tert-butoxycarbonyl)-(25)-pyrrolidine-2-carboxylic acid (VII, 30 g, 139 mmol) in tetrahydrofuran (2000 ml) under stirring at temperature of about 0 °C. Ethyl chloroformate (106 ml, 1.114 mol) was added drop- wise to the above obtained clear solution over a period of 30 minutes. After stirring for 1 hour, the resulting suspension was filtered over celite and the residue was washed with tetrahydrofuran (2×200 ml). To the combined filtrate was added dropwise a solution of sodium borohydride (42.1 g, 1.114 mol) in 210 ml water, containing a catalytic amount of sodium hydroxide, at temperature of about -10 °C over a period of 1-2 hours under stirring. The reaction mixture was allowed to warm to room temperature and stirred further for an hour. The reaction mixture was filtered through celite bed and the filtrate concentrated under reduced pressure to yield 180 g of ieri-butyl(25)-2-(hydroxymethyl)-pyrrolidene-l-carboxylate (IX) as colorless oil.

Analysis:

Mass: 202 (M+l), for Molecular Weight: 201.2 and Molecular Formula: C10H19NO3;

1H NMR (400 MHz, CDC13): δ 3.94-.3.92 (m, 1H), 3.80 (board, 1H), 3.63-3.54 (m, 2H), 3.45-3.40 (m, 1H), 3.32-3.28 (m, 1H), 2.01-1.96 (m, 1H), 1.84-1.75 (m, 2H), 1.63 (m, 1H), 1.45 (s, 9H);

Purity as determined by HPLC: 87.7 %.

Step 3: Synthesis of fert-butyl (25)-2-[[(l,3-dihydro-l,3-dioxo-2H-isoindol-2-yl)oxy] methyl] -pyrrolidine-1 -carboxylate (X) :

Triphenylphosphine (328.4 g, 1.253 mol) in tetrahydrofuran (1260 ml) was added to solution of Diisopropyl azodicarboxylate (253.3 g, 1.253 mol) in tetrahydrofuran at temperature of -15 °C under stirring. After stirring for an hour, N-feri-butoxylcarbonyl-L-prolinol (IX) (180 g, 0.895 mol) in tetrahydrofuran (540 ml) was added to the resulting mixture over a period of 15 minutes. After stirring the mixture for 45 minutes, N-Hydroxy phthalimide (146 g, 0.895 mol) was added and the mixture was allowed to warm to room temperature and stirred further for 16 hours. The solvent was evaporated under reduced pressure and residual oil was dissolved in dichloromethane (5000 ml) and washed with an aqueous 5 % sodium hydrogen carbonate solution (2×300 ml). The organic layer was dried over anhydrous sodium sulfate and the solvent evaporated under reduced pressure to obtain viscous oil. Diisopropyl ether (720 ml) was added to the oil, the mixture was stirred well and separated solid was filtered under suction. The filtrate was concentrated under reduced pressure and the residue was further purified by chromatography over a silica gel column (60 -120 mesh) and eluted with mixtures of ethyl acetate and hexane. Upon concentration of the combined eluted fractions, 230 g of teri-butyl (25)-2-[[( l,3-dihydro- l,3-dioxo-2H-isoindol-2-yl)oxy]methyl]-pyrrolidine- l-carboxylate (X) was obtained as yellow oil.

Analysis:

Mass: 347.3 (M+l), for Molecular Weight: 346.39 and Molecular Formula:

1H NMR (400 MHz, CDCI3): δ 7.80-7.78 (m, 2H), 7.72-7.70 (m, 2H), 4.32 (brs, 1H), 4.05 (brs, 2H), 3.36-3.31 (m, 2H), 2.27-2.25 (m, 1H), 2.08(m, 1H), 1.88-1.87 (m, 2H), 1.43 (s, 9H).

Step 4: Synthesis of fert-butyl (25)-2-[(aminooxy)methyl]pyrrolidine-l-carboxylate (HI):

To a stirred solution of the compound of Formula (X) ( 100 g, 0.288 mol) in dichloromethane (2000 ml) was added 99 % hydrazine hydrate (28.9 g, 0.577 mol) drop-wise over a period of 30 minutes at temperature of about 25 °C. The stirring was continued further for a period of 3 hours. The separated solid was filtered and the solid washed with additional dichloromethane (2 x 500 ml). The combined organic layer was collected and washed with water (2 x 500 ml). The organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure to obtain 62.4 g of tert-butyl (25)-2-[(aminooxy)methyl]pyrrolidine-l-carboxylate (III) as a colorless oil. This was used as such for the next reaction without further purification.

Analysis:

Mass: 215.1 (M- l), for Molecular Weight: 216.2 and Molecular Formula:

Example 2

Synthesis of (25,5R)-7-oxo-N-r(25)-pyrrolidin-2-yl-methyloxyl-6-(sulfooxy)-l,6- diazabicvclor3.2.11octane-2-carboxamide (I)

Step 1: Synthesis of tert-butyl (25)-2-{[({[25,5R)-6-(benzyloxy)-7-oxo-l,6-diazabicylco[3.2.1]oct-2-yl]carbonyl}amino)oxy]methyl}pyyrolidine-l-carboxylate

(IV):

Sodium(25,5i?)-6-(benzyloxy)-7-oxo-l,6-diazabicyclo[3.2.1]octane-2-carboxylate (II, 77.4 g, 0.259 mol; prepared according to the procedure disclosed in Indian patent application No. 699/MUM/2013) was dissolved in water (774 ml) to obtain a clear solution. To the clear solution was added EDC.HC1 (120.8 g, 0.632 mol) at temperature of about 15°C and after 10 minutes a solution of tert-buty\ (25)-2-[(aminooxy)methyl]pyrrolidine-l-carboxylate (III, 62.4 g, 0.288 moles prepared as per the literature procedure depicted in scheme 2) in dimethylformamide (125 ml) was added drop wise under continuous stirring at temperature of about 10 °C. The reaction mass was allowed to warm to temperature of about 25°C and then HOBt (38.96 g, 0.288 mol) was added in small portions over a period of 15 minutes and the resulting mixture was further stirred at room temperature for 16 hours. The reaction progress was monitored using thin layer chromatography using mixture of acetone and hexane (35: 65) as solvent system. The resulting suspension was filtered and the residue was washed with water (200 ml). The residual white solid was suspended in water (200 ml) and the mixture stirred with heating at temperatyre of about 50 °C for 3 hours. The resulting suspension was filtered, the residue dried at atmospheric temperature and then dried under vacuum to obtain 105 g of ierr-Butyl(25)-2- { [( { [25,5R)-6-(benzyloxy)-7-oxo- l,6-diazabicylco[3.2. l]oct-2-yl]carbonyl} amino)oxy]methyl}pyyrolidine-l-carboxylate (IV) as off white solid.

Analysis:

Mass: 475.4 (M+l), for Molecular Weight of 474.56 and Molecular Formula of

1H NMR (400 MHz, CDCI3): δ 10.16 (br s, 1H), 7.43-7.35 (m, 5H), 5.06-4.88 (dd, 2H), 4.12 (s, 1H), 3.94-.393 (d, 2H), 3.83 (unresolved s, 1H), 3.75-3.73 (m, 1H), 3.37-3.28 (dt, 2H), 3.02-2.86 (dd, 2H), 2.31-2.26 (m, 1H), 2.02-1.84 (m, 6H), 1.71-1.68 (m, 1H), 1.45 (s, 9H).

Step 2: Synthesis of tert-butyl(25)-2-{[({[25,5R)-6-hydroxy-7-oxo-l,6-diazabicylco

[3.2.1]oct-2-yl]carbonyl}amino)oxy]methyl}pyyrolidine-l-carboxylate (V):

tert-butyl(25)-2-{ [({ [25,5R)-6-(benzyloxy)-7-oxo-l,6-diazabicylco[3.2.1]oct-2-yl] carbonyl}amino)oxy]methyl}pyyrolidine-l-carboxylate (IV, 85 g, 0.179 mol) was dissolved in a mixture of dimethylformamide and dichloro methane (1: 1, 425 ml : 425 ml) to obtain a clear solution. To this solution was added 10 % Pd-C (17 g, 50 % wet) catalyst. The suspension was stirred for 4 hours under 7 psi hydrogen atmosphere at temperature of about 25 °C. The resulting mixture was filtered through celite under suction. The residue was washed with dichloromethane (170 ml). The solvent from the filtrate was evaporated under reduced pressure to furnish 68.8 g of tert-buty\(2S)-2-{ [( { [25,5i?)-6-hydroxy-7-oxo- l,6-diazabicylco[3.2. l]oct-2-yl]carbonyl} amino)oxy] methyl}pyyrolidine-l-carboxylate (V) as oil. The obtained product was used as such for the next reaction without further purification.

Analysis:

Mass: 385.4 (M+l), for Molecular Weight of 384.4 and Molecular Formula of C17H28N406.

Step 3: Synthesis of tert-butyl(25)-2-{[({[25,5R)-6-(sulfooxy)-7-oxo-l,6-diazabicylco[3.2.1]oct-2-yl]carbonyl}amino)oxy]methyl}pyyrolidine-l-carboxylate, tetra butyl ammonium salt (VI):

To solution of ieri-butyl(25)-2-{ [({ [25,5i?)-6-hydroxy7-oxo-l,6-diazabicylco [3.2.1]oct-2-yl]carbonyl}amino)oxy]methyl}pyyrolidine-l-carboxylate (V, 68.8 g, 0.178 mol) in dimethylformamide, (345 ml) was added sulfur trioxide dimethylformamide complex (30 g, 0.196 mol) under stirring at temperature of about 10 °C. The reaction mass was stirred at the same temperature for 30 minutes and then allowed to warm to room temperature. After 2 hours solution of tetra butyl ammonium acetate (59.09 g, 0.196 mol) in water (178 ml) was added to the reaction mixture under stirring. After 2 hours, the solvent from the reaction mixture was evaporated under reduced pressure to obtain an oily residue. The oily mass was co-evaporated with xylene (2×140 ml) to obtain thick mass. This mass was partitioned between dichloromethane (690 ml) and water (690 ml). The organic layer was separated and the aqueous layer re-extracted with dichloromethane (345 ml). The combined organic extracts were washed with water (3×345 ml) and dried over anhydrous sodium sulfate. The solvent was evaporated under reduced pressure and the resulting oily mass was triturated with ether (3×140 ml), each time the ether layer was decanted and finally the residue was concentrated under reduced pressure to obtain 102 g of ieri-butyl(25)-2- { [({ [25,5i?)-6-(sulfooxy)-7-oxo- l,6-diazabicylco[3.2.1]oct-2-yl]carbonyl}amino)oxy]methyl}pyyrolidine- l-carboxylate, tetrabutyl ammonium salt (VI) as fluffy material.

Analysis:

Mass: 463.4 (M- l without TBA), for Molecular Weight of 705.96 and Molecular Formula of C33H63N5O9 S;

1H NMR (400 MHz, CDCI3): δ 10.2 (s, 1H), 4.35 (s, 1H), 4.14 (s, 1H), 3.91 -3.92 (d, 2H), 3.74 (m, 1H), 3.36-3.27 (m, 10H), 2.96-2.88 (dd, 2H), 2.31-2.26 (m, 2H), 2.19-1.98 (m, 2H), 1.95-1.70 (m, 4H), 1.68- 1.62 (p, 8H), 1.49- 1.40 (m, 17H), 1.02-0.98 (t, 12H).

Step 4: (25,5R)-7-oxo-N-[(25)-pyrrolidin-2-yl-methyloxy]-6-(sulfooxy)-l,6-diaza bicyclo [3.2.1]octane-2-carboxamide (I):

feri-butyl(25)-2-{ [( { [25,5i?)-6-(sulfooxy)-7-oxo-l ,6-diazabicylco[3.2.1]oct-2-yl] carbonyl}amino)oxy]methyl}pyyrolidine- l-carboxylate, tetrabutylammonium salt (VI) (50 g, 0.0708 mol) was dissolved in dichloromethane (250 ml) and to the clear solution was slowly added trifluoroacetic acid (250 ml) at temperature of about -10 °C over a period of 1 hour under stirring. After stirring for an hour, the resulting mixture was poured into hexane (2500 ml) and the oily layer was separated. This procedure was repeated one more time and finally the separated oily layer was added to diethyl ether (500 ml) under vigorous stirring at temperature of about 25 °C. The ether layer was removed by decantation from the precipitated solid. This procedure was repeated twice again with diethyl ether (2x500ml). The solid thus obtained was stirred with fresh dichloromethane (500 ml) for 30 minutes and filtered. The residual solid was dried at temperature of about 45 °C under reduced pressure to yield 25 g of (25,5i?)-7-Oxo-N-[(25)-pyrrolidin-2-yl-methyloxy]-6-(sulfooxy)- l,6-diazabicyclo[3.2.1]octane-2-carboxamide (I) in amorphous form. The XRD of the obtained amorphous form is shown in Figure 1.

Analysis:

Mass: 363.2 (M- l), for Molecular Weight: 364.37 and Molecular Formula: C12H2oN407S;

1H NMR (400 MHz, DMSO-D6): δ 1 1.73 (s, 1H), 8.62-8.83 (d, 2H), 3.88-4.00 (m, 3H), 3.74-3.81 (m, 2H), 3.19 (t, 2H), 2.94-3.04 (dd, 2H), 1.96-2.03 (m, 2H), 1.80-1.92 (m, 3H), 1.54- 1.73 (m, 3H);

Purity as determined by HPLC: 90.30 %.

Example 3

Preparation of Crystalline Form I of (25,5R)-7-oxo-N-r(25)-pyrrolidin-2-yl- methyloxyl-6-(sulfooxy)-l,6-diaza bicvclor3.2.11octane-2-carboxamide

The amorphous solid obtained in the Step 4 of Example 2 was dissolved in water (75 ml) and to this solution isopropanol (200 ml) was slowly added at temperature of about 25 °C. The solution was further stirred for 12 hours. The separated solid thus obtained was filtered and washed with additional isopropanol (25 ml) and dried under reduced pressure to obtain 19 g of (25,5i?)-7-Oxo-N-[(25)-pyrrolidin-2-yl-methyloxy]-6-(sulfooxy)-l ,6-diazabicyclo[3.2.1]octane-2-carboxamide as crystalline Form I. The XRD of the obtained crystalline Form I is shown in Figure 2.

X-ray powder diffraction pattern comprising peak at (2 Theta Values): 8.08 (± 0.2), 1 1.45 (± 0.2), 16.26 (± 0.2), 17.89 (± 0.2), 18.15 (± 0.2), 19.66 (± 0.2), 21.15 (± 0.2), 23.55 (± 0.2), 24.23 (± 0.2), 24.94 (± 0.2), 25.66 (± 0.2) and 29.41 (± 0.2).

Typical X-ray analysis was performed as follows. Pass the test substance through sieve #100 BSS or gently grind it with a mortar and pestle. Place the test substance uniformly on a sample holder having cavity surface on one side, press the sample and cut into thin uniform film using a glass slide in such a way that the surface of the sample should be smooth and even. Record the X-ray diffractogram using the following instrument parameters:

Instrument : X-Ray Diffractometer

(PANalytical, Model X’Pert Pro

MPD)

Target source : CuK(a)

Antiscattering slit (Incident beam) : 1°

Programmable Divergent slit : 10 mm (fixed)

Anti- scattering slit (Diffracted beam) : 5.5 mm

Step width : 0.02°

Voltage : 40 kV

Current : 40 mA

Time per step : 30 seconds

Scan range : 3 to 40°

Example 4

Preparation of Pure (25,5R)-7-oxo-N-r(25)-pyrrolidin-2-yl-methyloxyl-6-(sulfooxy)- l,6-diazabicyclor3.2.11octane-2-carboxamide

(25,5i?)-7-Oxo-N-[(25)-pyrrolidin-2-yl-methyloxy]-6-(sulfooxy)- l,6-diazabicyclo [3.2.1] octane-2-carboxamide (5 g) was slowly dissolved in water (50 ml) under stirring until clear solution appears. To this clear solution 350 ml of isopropanol was added drop wise under stirring over the period of 2 hours. Formation of fine white precipitates was observed after the completion of the addition of isopropanol. The resulted fine suspension was stirred at temperature of about 25 °C for 20 hours. The formed white precipitates were filtered and vacuum dried at temperature of about 30-40 °C, under reduced pressure (2 mm Hg) to get 4.4 g of (2S,5i?)-7-oxo-N-[(2S)-pyrroMin-2-yl-methyloxy]-6-(sulfooxy)-l,6-diazabicyclo [3.2.1] octane-2-carboxamide.

The above obtained (25,5i?)-7-oxo-N-[(25)-pyrrolidin-2-yl-methyloxy]-6-(sulfooxy)-l,6-diazabicyclo[3.2.1]octane-2-carboxamide (3.4 gm) was dissolved in 34 ml of water to get clear solution. To the obtained clear solution 170 ml of isopropanol was added drop wise over a period of 1 hour. Formation of fine oily globules was observed and allowed to stand still for 15 minutes. The upper clear water and isopropanol layer was decanted from the oily mass. The clear decanted solution was allowed to stand at temperature of about 25 °C for 48 hours. Formation of crystals was observed and were collected by filtration. The collected crystals were dried at temperature of about 30-40 °C, under reduced pressure (2 mm Hg) to get 2 g of (2S,5i?)-7-oxo-N-[(2S)-pyrrolidin-2-yl-methyloxy]-6-(sulfooxy)- 1 ,6-diazabicyclo[3.2.1 ]octane-2-carboxamide which was analyzed for content of various components using HPLC and the results are described in Table 1.

The relative % content of (25,5i?)-7-oxo-N-[(25)-pyrrolidin-2-yl-methyloxy]-6-(sulfooxy)-l,6-diazabicyclo[3.2.1]octane-2-carboxamide with other substances (Table 1) was determined using HPLC (Agilent 1100 or equivalent). The HPLC column having 250 mm length and 4.6 mm ID packed with 5 μ particles of octa-decyl silane (ODS) was used. Mobile phase A used was a mixture of buffer (0.02 M potassium dihydrogen phosphate in HPLC grade water, pH adjusted to 2.5 with orthophosphoric acid and again readjusted to 7.0 with dilute ammonia), HPLC grade water and acetonitrile in a ratio of 40 : 60 : 0.2; v/v/v. Mobile phase B was mixture of buffer and acetonitrile in a ratio of 40 : 60; v/v. Mobile phase was run in gradient mode. Initially mobile phase A and B was run at 100 : 0 for 15 minutes, slowly ratio of mobile phase B was raised to 100 % in 10 minutes, held for 10 minutes at this concentration and brought back to initial condition in next 5 minutes and held for 10 minutes before next run. Flow rate of mobile phase was maintained at 1.0 ml/min. Column temperature was maintained at temperature of about 30°C. Detection was carried out using UV detector at wavelength 225 nm. Test solutions were prepared in mobile phase A. The method is capable of resolving diastereomers (Table 1, Sr. No. 1 and 2) with resolution of not less than 2.0.

%d bloggers like this: