Storage-stable ready-to-use formulations of tigecycline

Abstract

Liquid parenteral formulations are provided that include tigecycline and at least one or more pharmaceutically acceptable excipient or adjuvant, where the formulation is ready to use without additional steps for reconstitution at the time of administration.

Claims

1. A liquid ready to use parenteral formulation, comprising tigecycline, calcium chloride, sodium chloride, sodium bisulfite, and optionally an amino acid selected from L-Histidine, L-Cysteine HCl or L-arginine; wherein concentration of tigecycline in the ready to use parenteral formulation is from 1 mg/ml to about 5 mg/ml and pH of the ready to use parenteral formulation is from 5.5 to 6.5.

2. The formulation of claim 1, provided as a 100 ml, nitrogen flushed solution.

3. The formulation of claim 1, provided as a 5 ml, nitrogen flushed solution.

Description

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

(1) The present application now will be described more fully hereinafter with reference to the accompanying examples and experiments, in which illustrative embodiments of the application are shown. This application may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein; rather, these embodiments are provided so that this disclosure will be thorough and complete, and will fully convey the scope of the application to those skilled in the art.

(2) The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the application. As used herein, the singular forms “a”, “an” and “the” are intended to include the plural forms as well, unless the context clearly indicates otherwise. Unless otherwise defined, all terms (including technical and scientific terms) used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. It will be further understood that terms, such as those defined in commonly used dictionaries, should be interpreted as having a meaning that is consistent with their meaning in the context of the relevant art and will not be interpreted in an idealized or overly formal sense unless expressly so defined herein.

(3) As used herein, “tigecycline” refers to tigecycline and the pharmaceutically acceptable salts, solvates, hydrates and anhydrous forms thereof.

(4) As used here in “ready-to-use” when used in connection with a tigecycline formulation refers to a formulation that includes tigecycline in dissolved or solubilized form and/or is intended to be used as such or upon further dilution in intravenous diluents.

(5) As used herein, and unless otherwise specified, the term “storage-stable” refers to any tigecycline-containing composition or formulation having sufficient physical and chemical stability to allow storage at a convenient temperature, such as between about 0° C. and about 50° C., for a commercially reasonable period of time. The phrase “physical stability” refers to maintenance of colour or colourless state, dissolved oxygen level, head space oxygen level and particulate matter and the phrase “chemical stability” relates to formation of drug-related impurities in terms of total impurities, single maximum individual impurity, or maximum individual unknown impurity. For pharmaceutical products, stability is required for commercially relevant times after manufacturing, such as for about 6, 12, 18, 24, or 36 months, during which time a product is kept in its original packaging under specified storage conditions.

(6) As used herein, and unless otherwise specified, the term “about” means an acceptable error for a particular value as determined by one of ordinary skill in the art, which depends in part on how the value is measured or determined. In certain embodiments, the term about means within 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.1%, or 0.05% of a given value or range.

(7) In still further embodiments provided are ready-to-use liquid parenteral formulations including tigecycline and at least one or more pharmaceutically acceptable excipient or adjuvant.

(8) In one or more further embodiment of present application pharmaceutically acceptable excipients or adjuvants include but are not limited to one or more preservatives, polymers, pH adjusting agents, isotonicity adjusting agents, surfactants, chelating agents and antioxidants.

(9) Pharmaceutically acceptable excipients are not limited to complex forming agent, oxidizing agents and/or pH adjusting agents.

(10) Pharmaceutically acceptable excipients or adjuvants include but are not limited to one or more preservatives, complex forming agents, pH adjusting agents, surfactants and antioxidants.

(11) Examples of pharmaceutically acceptable preservatives include but are not limited to chlorobutanol, benzalkonium chloride, methyl paraben, propyl paraben, benzoic acid, sodium benzoate, sorbic acid, benzethonium chloride, cetyl pyridinium chloride, benzyl bromide, benzyl alcohol, phenylmercury nitrate, phenylmercury acetate, thiomersal, merthiolate, chlorhexidine, phenylethyl alcohol, quaternary ammonium chloride, sodium benzoate, etc. and combinations thereof.

(12) Examples of pharmaceutically acceptable complex forming agents include but are not limited to sodium chloride, potassium chloride, calcium chloride and magnesium chloride, Arginine, glucose, glycerol, etc. and combinations thereof.

(13) Examples of pharmaceutically acceptable antioxidants include but are not limited to butylated hydroxytoluene (BHT), butylated hydroxyanisole (BHA), sodium sulfite, Sodium bisulfite, propyl gallate (PG), monothioglycerol, ascorbic acid, sodium ascorbate, erythorbic acid, potassium metabisulfite, sodium metabisulfite, propionic acid, sodium formaldehyde sulphoxylate, reduced glutathione, thiourea, cysteine, n-aceticysteine, methionine, alkyl gallate, vitamin E or other tocopherol analogs such as tocopherol acetate and TPGS, etc. and combinations thereof.

(14) Examples of pharmaceutically acceptable pH adjusting agents include but are not limited to sodium hydroxide, hydrochloric acid, meglumine, boric acid, citric acid, acetic acid, phosphoric acid, succinic acid, potassium hydroxide, ammonium hydroxide, magnesium oxide, calcium carbonate, magnesium carbonate, magnesium aluminum silicates, malic acid, potassium citrate, sodium phosphate, lactic acid, gluconic acid, tartaric acid, fumaric acid, diethanolamine, monoethanolamine, sodium carbonate, sodium bicarbonate, triethanolamine, etc. and combinations thereof.

(15) The formulations according to the present application may be in the form of clear injectable solution, suspension or emulsion.

(16) In some embodiments the storage-stable ready-to-use injectable formulation may have a concentration of tigecycline of less than 10 mg/ml. In other embodiments the injectable formulation may have a concentration of tigecycline of less than 7 mg/ml. In another embodiment the injectable formulation may have a concentration of tigecycline of less than 5 mg/ml. In other embodiments the injectable formulation may have a concentration of tigecycline of less than 3 mg/ml. In still other embodiments the concentration of tigecycline in the formulation may be about 1 mg/ml.

(17) The storage-stable, ready-to-use injectable tigecycline-containing formulations disclosed herein do not require any additional reconstitution step(s) at the time of administration.

(18) The formulations have a controlled impurity profile suitable for regulatory approval at various storage conditions. The storage-stable ready-to-use tigecycline formulations are stored at 2-8° C. The storage-stable, ready-to-use tigecycline formulations for injection may retain at least 94% of the potency of tigecycline after storage for six months at 2-8° C. temperature and 60% relative humidity.

(19) The storage stable, ready-to-use, injectable formulations may be formulated to provide single or multiple dosage administration. The single dosage formulation may be packaged in IV bag, an ampoule, a vial, or a syringe. Multiple dosage formulations may be packaged in a vial. Multiple dosage formulations may preferably include at least one preservative.

(20) The formulations have a pH value from about 3 to about 9. In some embodiments the pH range is from about 4 to about 8. In other embodiments the pH is about 5.5-6.5.

(21) Storage-stable ready-to-use, injectable formulations disclosed herein contain tigecycline having a purity of from about 80% to about 120%. In some embodiments the formulation contains tigecycline having a purity of from about 90% to about 110%. In some embodiments the formulation contains tigecycline having a purity of about 100%.

(22) Methods of treatment of such antibiotics are disclosed including administering to an individual in need thereof a therapeutically effective amount of a storage stable, ready-to-use, injectable formulation as disclosed herein.

(23) Formulations as disclosed herein are useful as an antibiotic for skin and skin structure infections caused by susceptible isolates of Escherichia coli, Enterococcus faecalis (vancomycin-susceptible isolates), Staphylococcus aureus (methicillin-susceptible and -resistant isolates), Streptococcus agalactiae, Streptococcus anginosus grp. (includes S. anginosus, S. intermedius, and S. constellatus), Streptococcus pyogenes, Enterobacter cloacae, Klebsiella pneumoniae, and Bacteroides fragilis.

(24) Prepared formulations are also used in patients 18 years of age and older for the treatment of complicated intra-abdominal infections caused by susceptible isolates of Citrobacter freundii, Enterobacter cloacae, Escherichia coli, Klebsiella oxytoca, Klebsiella pneumoniae, Enterococcus faecalis (vancomycin-susceptible isolates), Staphylococcus aureus (methicillinsusceptible and -resistant isolates), Streptococcus anginosus grp. (includes S. anginosus, S. intermedius, and S. constellatus), Bacteroides fragilis, Bacteroides thetaiotaomicron, Bacteroides uniformis, Bacteroides vulgatus, Clostridium perfringens, and Peptostreptococcus micros.

EXAMPLES

(25) The following examples are for the illustration only and are not intended in any way to limit the scope of the present application.

Example 1

(26) TABLE-US-00002 TABLE 1 Ingredients Qty/IV bag Tigecycline 100 mg Calcium chloride 60 mg Sodium bisulfite 2.5 mg Sodium chloride 900 mg Sterile Water for Injection (WFI) 100 ml Meglumine (for pH adjustment) q.s.

(27) In this example, 900 mg sodium chloride, 60 mg Calcium chloride and 2.5 mg Sodium bisulfite were dissolved in 45 ml of sterile water for injection. The pH of the solution was adjusted to 5.5-6.5 range using 0.1 M Meglumine solutions. The volume of the solution was made up to 100 ml with sterile water for injection. The solution was nitrogen purged until the dissolved oxygen reaches <0.1 mg/L level. The solution was divided into two parts—80 ml and 20 ml. 80 ml part was poured in an infusion bag (Solution A), 20 ml part was poured in plastic tube (Solution B).

(28) Solution A and Solution B were then further nitrogen purged for another 30 minutes and closed tightly after nitrogen flushing and covered with black cover. Solution A and solution B were stored in refrigerator (2-8° C.) for 02 hours. 100 mg tigecycline was charged in solution B followed by the addition of solution B to solution A while the solution A was kept at ice bath (e.g., about 0° C.). Now, the total 100 ml solution in the infusion bag was nitrogen purged for another 15 minutes and sealed after nitrogen flushing and stored at refrigerator (2-8° C.).

(29) Stability data is summarized in Table 1A. As can be seen, total impurities did not exceed 1.13 and purity was at least 98.88 after 62 days.

(30) TABLE-US-00003 TABLE 1A Stability Day 1 Day 62 Purity 98.46 98.88 Open Ring Tig. ND ND Oxo Impurity 0.09 0.03 Rel. Comp. B 0.13 0.05 Epimer Impurity 0.57 0.69 Quinone Analogue ND 0.02 Minocycline ND 0.02 Tricyclic Impurity 0.02 0.08 Max. Unk Imp. 0.25 0.06 Total impurities 1.44 1.13

Example 2

(31) TABLE-US-00004 TABLE 2 Ingredients Qty/IV bag Tigecycline 100 mg Calcium chloride 60 mg Sodium bisulfite 2.5 mg Sodium chloride 900 mg Sterile Water for Injection 100 ml

(32) In this example, 900 mg sodium chloride, 60 mg Calcium chloride and 2.5 mg Sodium bisulfite were dissolved in 45 ml of sterile water for injection. The volume of the solution was made up to 100 ml with sterile water for injection. The solution was nitrogen purged until the dissolved oxygen reaches <0.1 mg/L level. The solution was divided into two parts—80 ml and 20 ml. 80 ml part was poured in an infusion bag (Solution A), 20 ml part was poured in plastic tube (Solution B).

(33) Solution A and Solution B were then further nitrogen purged for another 30 minutes and closed tightly after nitrogen flushing and covered with black cover. Solution A and solution B were stored in refrigerator (2-8° C.) for 02 hours. 100 mg tigecycline was charged in solution B followed by the addition of solution B to solution A while the solution A was kept at ice bath (e.g., about 0° C.). Now, the total 100 ml solution in the infusion bag was nitrogen purged for another 15 minutes and sealed after nitrogen flushing and stored at refrigerator (2-8° C.). Stability data is summarized in Table 2A. As can be seen, total impurities did not exceed 1.13 and purity was at least 98.87 after 62 days.

(34) TABLE-US-00005 Stability Day 1 Day 42 Purity 99.15 98.87 Open Ring Tig. ND ND Oxo Impurity 0.03 0.21 Rel. Comp. B 0.02 0.05 Epimer Impurity 0.52 0.62 Quinone Analogue ND 0.01 Minocycline ND 0.03 Tricyclic Impurity 0.03 0.02 Max. Unk Imp. 0.05 0.04 Total impurities 0.85 1.13

Example 3 (Solution Stability at RT)

(35) TABLE-US-00006 TABLE 3 Ingredients Qty/IV bag Tigecycline 100 mg Calcium chloride 60 mg Sodium bisulfite 2.5 mg Sodium chloride 900 mg Sterile Water for Injection 100 ml Meglumine (for pH q.s. adjustment)

(36) In this example, 900 mg sodium chloride, 60 mg Calcium chloride and 2.5 mg Sodium bisulfite were similarly dissolved in 45 ml of sterile water for injection. The pH of the solution was adjusted to 5.5-6.5 range using 0.1 M Meglumine solutions. The volume of the solution was made up to 100 ml with sterile water for injection. The solution was nitrogen purged until the dissolved oxygen reaches <0.1 mg/L level. The solution was divided into two parts 80 ml and 20 ml. 80 ml part was poured in the infusion bag (Solution A), 20 ml part was poured in plastic tube (Solution B).

(37) Solution A and Solution B were then further nitrogen purged for another 30 minutes and closed tightly after nitrogen flushing and covered with black cover. Solution A and solution B were stored in refrigerator (2-8° C.) for 02 hours. 100 mg tigecycline was charged in solution B followed by the addition of solution B to solution A while the solution A was kept at ice bath (e.g., about 0° C.). Now, the total 100 ml solution in the infusion bag was nitrogen purged for another 15 minutes and sealed after nitrogen flushing and stored at RT and analysed on particular interval to check the stability. Stability data is summarized in Table 3A.

(38) TABLE-US-00007 TABLE 3A API Initial 1 hr 10 min 2 hr 20 min 3 hr 30 min 4 hr 40 min % Purity 99.22 99.11 99.14 99.14 99.11 99.11 Open Ring Tig. ND ND ND ND ND ND Oxo Impurity 0.02 0.02 0.02 0.03 0.02 0.03 Rel. Comp. B 0.01 0.02 0.02 0.02 0.02 0.02 Epimer Impurity 0.5 0.53 0.54 0.55 0.56 0.56 Quinone Analogue ND ND ND ND ND ND Minocycline ND 0.04 0.04 0.04 0.04 0.04 Tricyclic Impurity 0.03 0.08 0.08 0.08 0.09 0.08 Max. Unk Imp. 0.05 0.05 0.05 0.05 0.05 0.05

Example 4

(39) TABLE-US-00008 TABLE 4 Ingredients Qty/vial Tigecycline 50 mg Calcium chloride 60 mg 5% Dextrose Solution 5 ml Sodium bisulfite 2.5 mg

(40) Calcium chloride 60 mg and sodium bisulfite 2.5 mg were dissolved in 3 ml 5% dextrose for injection. The volume was made up to 5 ml with 5% dextrose solution. The solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber color vial and then the 5 ml of the above solution was poured in the vial under nitrogen. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 4A.

(41) TABLE-US-00009 TABLE 4A Stability API Day 24 Purity 99.3 98.17 Open Ring Tig. ND ND Oxo Impurity 0.02 0.28 Rel. Comp. B 0.02 0.32 Epimer Impurity 0.48 0.82 Quinone Analogue ND 0.03 Minocycline ND ND Tricyclic Impurity 0.04 0.06 Max. Unk Imp. 0.05 0.05

Example 5

(42) TABLE-US-00010 TABLE 5 Ingredients Qty/vial Tigecycline 50 mg Calcium chloride 60 mg 5% Dextrose Solution 5 ml Sodium bisulfite 2.5 mg Meglumine (for pH q.s. adjustment)

(43) Calcium chloride 60 mg and sodium bisulfite 2.5 mg were dissolved in 3 ml 5% dextrose. pH was adjusted to 5.5-6.0 pH range using 0.1 M meglumine solution, as required. The volume was made up to 5 ml with 5% dextrose solution. The solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber colour vial and then the 5 ml of the above solution was poured in the vial under nitrogen. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 5A.

(44) TABLE-US-00011 TABLE 5A Stability API Day 24 Purity 99.3 98.52 Open Ring Tig. ND ND Oxo Impurity 0.02 0.13 Rel. Comp. B 0.02 0.24 Epimer Impurity 0.48 0.83 Quinone Analogue ND 0.03 Minocycline ND ND Tricyclic Impurity 0.04 0.06 Max. Unk Imp. 0.05 0.05

Example 6

(45) TABLE-US-00012 TABLE 6 Ingredients Qty/vial Tigecycline 50 mg Calcium chloride 60 mg 5% Dextrose Solution 5 ml Sodium bisulfite 2.5 mg Meglumine (for pH q.s. adjustment)

(46) Calcium chloride 60 mg and sodium bisulfite 2.5 mg were dissolved in 3 ml 5% dextrose. pH was adjusted to 4.5-5.0 pH range using 0.1 M meglumine solution, as required. The volume was made up to 5 ml with 5% dextrose solution. The solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber colour vial and then the 5 ml of the above solution was poured in the vial under nitrogen. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 6A.

(47) TABLE-US-00013 TABLE 6A Stability API Day 24 Purity 99.3 98.24 Open Ring Tig. ND ND Oxo Impurity 0.02 0.26 Rel. Comp. B 0.02 0.36 Epimer Impurity 0.48 0.81 Quinone Analogue ND 0.03 Minocycline ND ND Tricyclic Impurity 0.04 0.06 Max. Unk Imp. 0.05 0.05

Example 7

(48) TABLE-US-00014 TABLE 7 Ingredients Qty/vial Tigecycline 50 mg Calcium chloride 60 mg 0.9% NaCl Solution 5 ml Sodium bisulfite 2.5 mg

(49) Calcium chloride 60 mg and sodium bisulfite 2.5 mg were dissolved in 3 ml 0.9% NaCl Solution. The volume was made up to 5 ml with 0.9% NaCl Solution. The solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber color vial and then the 5 ml of the above solution was poured in the vial under nitrogen. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 7A.

(50) TABLE-US-00015 TABLE 7A Stability API Day 64 Purity 99.3 97.82 Open Ring Tig. ND ND Oxo Impurity 0.02 0.43 Rel. Comp. B 0.02 0.54 Epimer Impurity 0.48 0.97 Quinone Analogue ND 0.02 Minocycline ND ND Tricyclic Impurity 0.04 0.05 Max. Unk Imp. 0.05 0.16

Example 8

(51) TABLE-US-00016 TABLE 8 Ingredients Qty/vial Tigecycline 50 mg Calcium chloride 60 mg 0.9% NaCl Solution 5 ml Sodium bisulfite 2.5 mg Meglumine (for pH q.s. adjustment)

(52) Calcium chloride 60 mg and sodium bisulfite 2.5 mg were dissolved in 3 ml 0.9% NaCl Solution. pH was adjusted to 5.5-6.0 pH range using 0.1 M meglumine solution, as required. The volume was made up to 5 ml with 0.9% NaCl Solution. The solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber colour vial and then the 5 ml of the above solution was poured in the vial under nitrogen. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 8A.

(53) TABLE-US-00017 TABLE 8A Stability API Day 64 Purity 99.3 98.03 Open Ring Tig. ND ND Oxo Impurity 0.02 0.23 Rel. Comp. B 0.02 0.5 Epimer Impurity 0.48 1.01 Quinone Analogue ND 0.03 Minocycline ND 0.03 Tricyclic Impurity 0.04 0.04 Max. Unk Imp. 0.05 0.14

Example 9

(54) TABLE-US-00018 TABLE 9 Ingredients Qty/vial Tigecycline 50 mg Calcium chloride 60 mg 0.9% NaCl Solution 5 ml Sodium bisulfite 2.5 mg Meglumine (for pH q.s. adjustment)

(55) Calcium chloride 60 mg and sodium bisulfite 2.5 mg were dissolved in 3 ml 0.9% NaCl Solution. pH was adjusted to 4.5-5.0 pH range using 0.1 M meglumine solution, as required. The volume was made up to 5 ml with 0.9% NaCl Solution. The solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber colour vial and then the 5 ml of the above solution was poured in the vial under nitrogen. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 9A.

(56) TABLE-US-00019 TABLE 9A Stability API Day 24 Purity 99.3 98.5 Open Ring Tig. ND ND Oxo Impurity 0.02 0.12 Rel. Comp. B 0.02 0.3 Epimer Impurity 0.48 0.78 Quinone Analogue ND 0.04 Minocycline ND ND Tricyclic Impurity 0.04 0.05 Max. Unk Imp. 0.05 0.04

Example 10

(57) TABLE-US-00020 TABLE 10 Ingredients Qty/vial Tigecycline 50 mg Calcium chloride 60 mg 0.9% NaCl Solution 5 ml Sodium bisulfite 2.5 mg pH NA Meglumine (for pH NA adjustment) Storage Refg.

(58) Calcium chloride and sodium bisulfite were dissolved in 3 ml 0.9% NaCl Solution. The volume was made up to 5 ml with sterile water for injection. The solution was then cooled. Cooled solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber colour vial and then the 5 ml of the above cooled solution was poured in the vial under nitrogen in cooling condition. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 10A.

(59) TABLE-US-00021 TABLE 10A Stability API 42 days % Purity 99.2 98.37 Open Ring Tig. ND ND Oxo Impurity 0.02 0.17 Related Compound B 0.01 0.41 Epimer Impurity 0.5 0.84 Quinone Analogue ND 0.02 Minocycline ND ND Tricyclic Impurity 0.03 0.04 Max. Unk Imp. 0.05 0.14

Example 11

(60) TABLE-US-00022 TABLE 11 Ingredients Qty/vial Tigecycline 50 mg Calcium chloride 60 mg 0.9% NaCl Solution 5 ml Sodium bisulfite 2.5 mg pH 5.5-6.0 Meglumine q.s. Storage Refg.

(61) Calcium chloride and sodium bisulfite were dissolved in 3 ml 0.9% NaCl Solution. pH was adjusted to required pH 5.5-6.0 using 0.1 M meglumine solution. The volume was made up to 5 ml with sterile water for injection. The solution was then cooled. Cooled solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber colour vial and then the 5 ml of the above cooled solution was poured in the vial under nitrogen in cooling condition. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 11A

(62) TABLE-US-00023 TABLE 11A Stability API 42 days % Purity 99.2 98.47 Open Ring Tig. ND ND Oxo Impurity 0.02 0.16 Related Compound B 0.01 0.31 Epimer Impurity 0.5 0.78 Quinone Analogue ND 0.02 Minocycline ND ND Tricyclic Impurity 0.03 0.04 Max. Unk Imp. 0.05 0.21

Example 12

(63) TABLE-US-00024 TABLE 12 Ingredients Qty/vial Tigecycline  50 mg Calcium chloride  60 mg 0.9% NaCl Solution   5 ml Sodium bisulfite 2.5 mg pH 6.5-7.0 Meglumine q.s. Storage Refg.

(64) Calcium chloride and sodium bisulfite were dissolved in 3 ml 0.9% NaCl Solution. pH was adjusted to required pH 6.5-7.0 using 0.1 M meglumine solution. The volume was made up to 5 ml with sterile water for injection. The solution was then cooled. Cooled solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber colour vial and then the 5 ml of the above cooled solution was poured in the vial under nitrogen in cooling condition. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 12A

(65) TABLE-US-00025 TABLE 12A Stability API 29 days % Purity 99.2 98.98 Open Ring Tig. ND ND Oxo Impurity 0.02 0.01 Related Compound B 0.01 0.07 Epimer Impurity 0.5 0.66 Quinone Analogue ND 0.03 Minocycline ND ND Tricyclic Impurity 0.05 0.04

Example 13

(66) TABLE-US-00026 TABLE 13 Ingredients Qty/vial Tigecycline 50 mg Calcium chloride 60 mg 0.9% NaCl Solution  5 ml Sodium bisulfite  5 mg pH NA Meglumine NA Storage Refg.

(67) Calcium chloride and sodium bisulfite were dissolved in 3 ml 0.9% NaCl Solution. The volume was made up to 5 ml with sterile water for injection. The solution was then cooled. Cooled solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber colour vial and then the 5 ml of the above cooled solution was poured in the vial under nitrogen in cooling condition. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 13A.

(68) TABLE-US-00027 TABLE 13A Stability API 29 days % Purity 99.2 98.49 Open Ring Tig. ND ND Oxo Impurity 0.02 0.11 Related Compound B 0.01 0.24 Epimer Impurity 0.5 0.85 Quinone Analogue ND 0.03 Minocycline ND 0.03

Example 14

(69) TABLE-US-00028 TABLE 14 Ingredients Qty/vial Tigecycline  50 mg Calcium chloride  30 mg 0.9% NaCl Solution  05 ml Sodium bisulfite 2.5 mg pH No pH adjst. Meglumine NA Storage Refg.

(70) Calcium chloride and sodium bisulfite were dissolved in 3 ml 0.9% NaCl Solution. The volume was made up to 5 ml with sterile water for injection. The solution was then cooled. Cooled solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber colour vial and then the 5 ml of the above cooled solution was poured in the vial under nitrogen in cooling condition. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 14A.

(71) TABLE-US-00029 TABLE 14A Stability API 18 days % Purity 99.22 98.49 Open Ring Tig. ND ND Oxo Impurity 0.02 0.18 Rel. Comp. B 0.01 0.33 Epimer Impurity 0.5 0.74 Quinone Analogue ND 0.02 Minocycline ND 0.03 Tricyclic Impurity 0.03 0.02 Max. Unk Imp. 0.05 0.18

Example 15

(72) TABLE-US-00030 TABLE 15 Ingredients Qty/vial Tigecycline  50 mg Calcium chloride  30 mg 0.9% NaCl Solution  05 ml Sodium bisulfite 2.5 mg pH 5.5-6.5 Meglumine pH adjst. Storage Refg.

(73) Calcium chloride and sodium bisulfite were dissolved in 3 ml 0.9% NaCl Solution. pH was adjusted to required pH 5.5-6.0 using 0.1 M meglumine solution. The volume was made up to 5 ml with sterile water for injection. The solution was then cooled. Cooled solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber colour vial and then the 5 ml of the above cooled solution was poured in the vial under nitrogen in cooling condition. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 15A

(74) TABLE-US-00031 TABLE 15A Stability API 18 days % Purity 99.22 98.77 Open Ring Tig. ND ND Oxo Impurity 0.02 0.11 Rel. Comp. B 0.01 0.18 Epimer Impurity 0.5 0.7 Quinone Analogue ND 0.03 Minocycline ND 0.04 Tricyclic Impurity 0.03 0.02 Max. Unk Imp. 0.05 0.14

Example 16

(75) TABLE-US-00032 TABLE 16 Ingredients Qty/vial Tigecycline   50 mg Calcium chloride   30 mg 0.9% NaCl Solution   05 ml Sodium bisulfite 1.25 mg pH No pH adjst. Meglumine NA Storage Refg.

(76) Calcium chloride and sodium bisulfite were dissolved in 3 ml 0.9% NaCl Solution. The volume was made up to 5 ml with sterile water for injection. The solution was then cooled. Cooled solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber colour vial and then the 5 ml of the above cooled solution was poured in the vial under nitrogen in cooling condition. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 16A

(77) TABLE-US-00033 TABLE 16A Stability API 02 days % Purity 99.22 99.07 Open Ring Tig. ND ND Oxo Impurity 0.02 0.05 Rel. Comp. B 0.01 0.06 Epimer Impurity 0.5 0.55 Quinone Analogue ND ND Minocycline ND 0.04 Tricyclic Impurity 0.03 0.05 Max. Unk Imp. 0.05 0.05

Example 17

(78) TABLE-US-00034 TABLE 17 Ingredients Qty/vial Tigecycline   50 mg Calcium chloride   30 mg 0.9% NaCl Solution   05 ml Sodium bisulfite 1.25 mg pH 5.5-6.0 Meglumine pH adjst. Storage Refg.

(79) Calcium chloride and sodium bisulfite were dissolved in 3 ml 0.9% NaCl Solution. pH was adjusted to required pH 5.5-6.0 using 0.1 M meglumine solution. The volume was made up to 5 ml with sterile water for injection. The solution was then cooled. Cooled solution was then nitrogen purged until dissolved oxygen reached <0.1 mg/L. 50 mg tigecycline was transferred into an amber colour vial and then the 5 ml of the above cooled solution was poured in the vial under nitrogen in cooling condition. The headspace of the vial was flushed through with nitrogen and packed. The vials were stored at refrigeration condition. Stability data is summarized in Table 17A

(80) TABLE-US-00035 TABLE 17A Stability API 02 days % Purity 99.22 99.09 Open Ring Tig. ND ND Oxo Impurity 0.02 0.04 Rel. Comp. B 0.01 0.06 Epimer Impurity 0.5 0.55 Quinone Analogue ND ND Minocycline ND 0.04 Tricyclic Impurity 0.03 0.05 Max. Unk Imp. 0.05 0.05

Example 18

(81) TABLE-US-00036 TABLE 18 Ingredients Qty/IV bag Tigecycline  100 mg Calcium chloride   60 mg Sodium bisulfite 2.50 mg Sodium chloride  900 mg L-Cysteine HCl   30 mg WFI  100 ml pH 5.0-6.0 NaOH (for pH adjustment) q.s. HCl (for pH adjustment) q.s.

(82) 900 mg sodium chloride, 60 mg Calcium chloride, 2.5 mg sodium bisulfite, 30 mg cysteine HCl were dissolved in 70 ml water. pH was adjusted to 5.0-6.0 using NaOH & HCl solution. The solution was then nitrogen purged for an hour while kept on cooling. 100 mg tigecycline was dissolved in 10 ml water (nitrogen purged) and added to the above solution. The volume was made up to 100 ml with nitrogen purged cold water. The solution was further nitrogen purged for 15 minutes and sealed under nitrogen in the infusion bag. The above bag was packed in Aluminium pouch under vacuum. The formulation was stored at 2-8° C. Stability data is summarized in Table 18A.

(83) TABLE-US-00037 TABLE 18A Stability Day 32 % purity 98.35 Openring Tigecycline 0.01 Oxo Impurity 0.03 Rel. Comp. B 0.33 Epimer Impurity 0.81 Quinone Analogue 0.01 Minocycline ND Tricyclic Impurity 0.05 Max. Unk. impurity 0.15 Total Impurity 1.65

Example 19

(84) TABLE-US-00038 TABLE 19 Ingredients Qty/IV bag Tigecycline  100 mg Calcium chloride   60 mg Sodium bisulfite 2.50 mg Sodium chloride  900 mg L-Histidine   90 mg WFI  100 ml pH 5.0-6.0 NaOH (for pH adjustment) q.s. HCl (for pH adjustment) q.s.

(85) 900 mg sodium chloride, 60 mg Calcium chloride, 2.5 mg sodium bisulfite, 90 mg L-Histidine were dissolved in 70 ml water. pH was adjusted to 5.0-6.0 using NaOH & HCl solution. The solution was then nitrogen purged for an hour while kept on cooling. 100 mg tigecycline was dissolved in 10 ml water (nitrogen purged) and added to the above solution. The volume was made up to 100 ml with nitrogen purged cold water. The solution was further nitrogen purged for 15 minutes and sealed under nitrogen in the infusion bag. The above bag was packed in Aluminium pouch under vacuum. The formulation was stored at 2-8° C. Stability data is summarized in Table 19A.

(86) TABLE-US-00039 TABLE 19A Stability Day 32 % purity 98.28 Openring Tigecycline ND Oxo Impurity 0.19 Rel. Comp. B 0.07 Epimer Impurity 0.81 Quinone Analogue 0.01 Minocycline 0.01 Tricyclic Impurity 0.01 Max. Unk. impurity 0.17 Total Impurity 1.72

Example 20

(87) TABLE-US-00040 TABLE 20 Ingredients Qty/IV bag Tigecycline  100 mg Calcium chloride   60 mg Sodium bisulfite 2.50 mg Sodium chloride  900 mg Methionine  160 mg WFI  100 ml pH 5.0-6.0 NaOH (for pH adjustment) q.s. HCl (for pH adjustment) q.s.

(88) 900 mg sodium chloride, 60 mg Calcium chloride, 2.5 mg sodium bisulfite, 90 mg methionine were dissolved in 70 ml water. pH was adjusted to 5.0-6.0 using NaOH & HCl solution. The solution was then nitrogen purged for an hour while kept of cooling. 100 mg tigecycline was dissolved in 10 ml water (nitrogen purged) and added to the above solution. The volume was made up to 100 ml with nitrogen purged cold water. The solution was further nitrogen purged for 15 minutes and sealed under nitrogen in the infusion bag. The above bag was packed in Aluminium pouch under vacuum. The formulation was stored at 2-8° C. Stability data is summarized in Table 20A.

(89) TABLE-US-00041 TABLE 20A Stability Day 69 % purity 97.71 Openring Tigecycline ND Oxo Impurity 0.02 Rel. Comp. B 0.05 Epimer Impurity 1.36 Quinone Analogue 0.01 Minocycline ND Tricyclic Impurity 0.02 Max. Unk. impurity 0.14 Total Impurity 2.29

(90) Above example was kept in an accelerated stability study at various temperatures. Results for this study is summarized in Table no. 20B.

(91) TABLE-US-00042 TABLE 20B Condition API 2-8° C. 25° C. 40° C. 60° C. Days Day 1 Day 69 Day 7 Day 7 60 hr % purity 99.18 97.71 96.28 91.98 74.88 Openring Tigecycline ND ND ND ND ND Oxo Impurity 0.02 0.02 0.01 0.01 0.02 Rel. Comp. B 0.02 0.05 0.07 0.08 0.13 Epimer Impurity 0.49 1.36 2.58 6.73 20.64 Quinone Analogue ND 0.01 0.04 0.21 1.94 Minocycline ND ND 0.01 0.01 0.01 Tricyclic Impurity 0.05 0.02 0.04 0.11 0.6 Max. Unk. impurity 0.04 0.14 0.2 0.27 0.39 Total Impurity 0.82 2.29 3.72 8.02 25.12

Example 21

(92) TABLE-US-00043 TABLE 21 Ingredients Qty/IV bag Tigecycline  50 mg Calcium chloride  75 mg Sodium chloride 450 mg Methionine  40 mg L-arginine 175 mg WFI  50 ml pH 5.0-6.0 NaOH (for pH adjustment) q.s. HCl (for pH adjustment) q.s.

(93) 450 mg Sodium chloride, 175 mg L-arginine, 75 mg calcium chloride dihydrate, 40 mg methionine were dissolved in 40 ml sterile water for injection. pH was then adjusted to 5.0-5.5 using NaOH and HCl solution and volume was made up to 50 ml with water. The solution was then nitrogen purged for an hour (dissolved oxygen level<0.1 mg/L), the solution was kept on cooling. 50 mg tigecycline was added to the above solution while the solution was kept on cooling and protected from light. The above solution was further nitrogen purged for 15 minutes and sealed under nitrogen in the infusion bag. The solution was then stored at 2-8° C. The solution in the infusion bag was further packed in Aluminium pouch under vacuum. Stability data is summarized in Table 21A.

(94) TABLE-US-00044 TABLE 21A Stability Day 31 % purity 98.4 Openring Tigecycline ND Oxo Impurity 0.02 Rel. Comp. B 0.02 Epimer Impurity 0.7 Quinone Analogue 0.03 Minocycline ND Tricyclic Impurity 0.1 Max. Unk. impurity 0.07

Example 22

(95) TABLE-US-00045 TABLE 22 Ingredients Qty/IV bag Tigecycline  50 mg Calcium chloride 250 mg Sodium chloride 450 mg Methionine  40 mg WFI  50 ml pH 5.0-6.0 NaOH (for pH adjustment) q.s. HCl (for pH adjustment) q.s.

(96) 450 mg Sodium chloride, 250 mg calcium chloride dihydrate, 40 mg methionine were dissolved in 40 ml sterile water for injection. pH was then adjusted to 5.0-5.5 using NaOH and HCl solution and volume was made up to 50 ml with water. The solution was then nitrogen purged for an hour (dissolved oxygen level<0.1 mg/L) while the solution was kept on cooling. 50 mg tigecycline was added to the above solution while the solution was kept on cooling and protected from light. The above solution was further nitrogen purged for 15 minutes and sealed under nitrogen in the infusion bag. The solution was then stored at 2-8° C. The solution in the infusion bag was further packed in Aluminium pouch under vacuum. Stability data is summarized in Table 22A.

(97) TABLE-US-00046 TABLE 22A Stability Day 31 % purity 98.67 Openring Tigecycline ND Oxo Impurity 0.02 Rel. Comp. B 0.02 Epimer Impurity 0.67 Quinone Analogue 0.01 Minocycline ND Tricyclic Impurity 0.06 Max. Unk. impurity 0.05

Example 23

(98) TABLE-US-00047 TABLE 23 Ingredients Qty/IV bag Tigecycline 100 mg Calcium chloride 500 mg Sodium chloride 900 mg Methionine  80 mg L-arginine 350 mg WFI 100 ml pH 5.0-5.5 NaOH (for pH adjustment) q.s. HCl (for pH adjustment) q.s.

(99) 900 mg Sodium chloride, 500 mg Calcium chloride dihydrate, 350 mg Arginine and 80 mg methionine were dissolved in 80 ml sterile water for injection. The pH was adjusted to 5.0 and the solution was nitrogen purged for an hour (dissolved oxygen level<0.1 mg/L) while the solution was kept on cooling. 100 mg tigecycline was added to and pH was adjusted to 5.2-5.3. The volume was made up to 100 ml with sterile water for injection. The sample was further nitrogen purged for 15 minutes while the solution was kept on cooling and protected from light and sealed under nitrogen in the infusion bag. The solution in the infusion bag was packed in Aluminium pouch under vacuum. The solution was then stored at 2-8° C. Stability data is summarized in Table 23A.

(100) TABLE-US-00048 TABLE 23A Stability Day 6 % purity 99.16 Openring Tigecycline ND Oxo Impurity 0.01 Rel. Comp. B 0.01 Epimer Impurity 0.46 Quinone Analogue ND Minocycline ND Tricyclic Impurity 0.07 Max. Unk. impurity 0.04

Example 24

(101) TABLE-US-00049 TABLE 24 Ingredients Qty/IV bag Tigecycline  100 mg Calcium chloride 1000 mg Sodium chloride  900 mg Methionine  80 mg Arginine  350 mg WFI  100 ml pH 5.0-5.5 NaOH (for pH adjustment) q.s. HCl (for pH adjustment) q.s.

(102) 900 mg Sodium chloride, 1000 mg Calcium chloride dihydrate, 350 mg Arginine and 80 mg methionine were dissolved in 80 ml sterile water for injection. The pH was adjusted to 5.0 and the solution was nitrogen purged for an hour (dissolved oxygen level<0.1 mg/L) while the solution was kept on cooling. After purging 50 mg tigecycline was added and pH was adjusted to 5.2-5.3. The volume was made up to 100 ml with sterile water for injection. The sample was further nitrogen purged for 15 minutes while the solution was kept on cooling and protected from light and sealed under nitrogen in the infusion bag. The solution in the infusion bag was further packed in Aluminium pouch under vacuum. The solution was then stored at 2-8° C. Stability data is summarized in Table 24A.

(103) TABLE-US-00050 TABLE 24A Stability Day 7 % purity 98.88 Openring Tigecycline ND Oxo Impurity 0.03 Rel. Comp. B ND Epimer Impurity 0.52 Quinone Analogue ND Minocycline ND Tricyclic Impurity 0.05 Max. Unk. impurity 0.05

Example 25

(104) TABLE-US-00051 TABLE 25 Ingredients Qty/IV bag Tigecycline  100 mg Calcium chloride 4000 mg Sodium chloride  900 mg Methionine  80 mg Arginine  350 mg WFI  100 ml pH 5.0-5.5 NaOH (for pH adjustment) q.s. HCl (for pH adjustment) q.s.

(105) 900 mg Sodium chloride, 4000 mg Calcium chloride dihydrate, 350 mg Arginine and 80 mg methionine were dissolved in 80 ml sterile water for injection. The pH was adjusted to 5.0-5.5 and the solution was nitrogen purged for an hour (dissolved oxygen level<0.1 mg/L) while the solution was kept on cooling. After purging 50 mg tigecycline was added and pH was adjusted to 5.2-5.3. The volume was made up to 100 ml with sterile water for injection. The sample was further nitrogen purged for 15 minutes while the solution was kept on cooling and protected from light and sealed under nitrogen in the infusion bag. The solution in the infusion bag was packed in Aluminium pouch under vacuum. The solution was then stored at 2-8° C. Stability data is summarized in Table 25A.

(106) TABLE-US-00052 TABLE 25A Stability Day 6 % purity 98.95 Openring Tigecycline ND Oxo Impurity 0.02 Rel. Comp. B 0.01 Epimer Impurity 0.52 Quinone Analogue ND Minocycline ND Tricyclic Impurity 0.05 Max. Unk. impurity 0.04