Doxycycline for Injection

This article is compiled based on the United States Pharmacopeia (USP) – 2025 Edition

Issued and maintained by the United States Pharmacopeial Convention (USP)

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1 DEFINITION

Doxycycline for injection contains an amount of Doxycycline Hyclate equivalent to NLT 90.0% and NMT 120.0% of the labeled amount of doxycycline (C₂₂H₂₄N₂O₈).

2 IDENTIFICATION

A. The UV spectrum of the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay.

B. The retention time of the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay

3 ASSAY

PROCEDURE

Protect solutions containing doxycycline from light.

Solution A: Transfer 3.1 g of monobasic potassium phosphate, 0.5 g of edetate disodium, and 0.5 ml of triethylamine to a 1000-ml volumetric flask. Add about 850 ml of water and mix. Dilute with water to volume and adjust with 1 N sodium hydroxide to a pH of 8.5 ± 0.1. Pass through a suitable filter of 0.22-um pore size.

Solution B: Methanol

Mobile phase: See Table 1

Diluent: 0.01 N hydrochloric acid

Standard solution: 0.12 mg/mL of USP Doxycycline Hyclate RS in Diluent. Sonicate as needed to dissolve.

Sample solution: Nominally 0.1 mg/mL of doxycycline prepared as follows. Dissolve the contents of one container of Doxycycline for Injection using 1% of the final volume of Diluent and transfer to a suitable volumetric flask. Dilute with Diluent to volume and mix

Chromatographic system

(See Chromatography (621). System Suitability)

Mode: LC

Detector: UV 270 nm. For Identification A, use a diode array detector in the range of 200-400 nm.

Column: 2.1mm * 5 - cm; 1.7-um packing LZ [NOTE-A 1.7-um guard column with packing 17 was used during method validation.

Column temperature: 60°

Flow rate: 0.6 mL/min

Injection volume: 5 µL

System suitability

Sample: Standard solution

Suitability requirements

Tailing factor: NMT 1.5

Relative standard deviation: NMT 2.0%

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of the labeled amount of doxycycline (C_{22}*H_{28}*N_{3}*O_{g}) in the portion of Doxycycline for Injection taken:

Result = (r_U/(r_S) × (C_S /C_U) × P x F x 100

r_U = peak response from the Sample solution

r_S = peak response from the Standard solution

C_S = concentration of USP Doxycycline Hyclate RS, in the Stardard solution (mg/ml)

C_U = nominal concentration of doxycycline in the Sample sulation (mg/ml)

P = potency of doxycycline in USP Doxycycline Hyclate RS (µg/mg)

F = conversion factor 0.001 mg/μg

Acceptance criteria: 90.0%-120.0%

4 IMPURITIES

ORGANIC IMPURITIES

Protect solutions containing doxycycline from light.

Mobile phase, Diluent, and Chromatographic system: Proceed as directed in the Assay

System suitability stock solution 1: 1 mg/mL each of USP Doxycycline Related Compound A. RS and USP Methacycline Hydrochloride RS in Diluent

System suitability stock solution 2: 1.2 mg/mL of USP Doxycycline Hyciate RS in Diluent

System suitability solution: Transfer 5 mL of Systern suitability stock solution 2 to a 25-ml volumetric flask, heat on a steam bath for 60 min, and evaporate to dryness on a hot plate, taking care not to char the residue. Dissolve the residue in Diluent, add 0.5 mL of System suitability stock solution 7, and dilute with Diluent to volume. Pass the solution through a suitable filter and use the filtrate. This solution contains a mixture of 4-epidoxycycline, doxycycline related compound A, methacycline, and doxycycline. [NOTE-The solution is stable up to 14 days when stored in a refrigerator.)

Standard solution: 2.3 µg/mL of USP Doxycycline Hyclate RS in Diluent

Sample solution: Nominally 2.0 mg/ml of doxycycline prepared as follows. Dissolve the contents of one container of Doxycycline for Injection using 20% of the final volume of Diluent and transfer to a suitable volumetric flask. Dilute with Diluent to volume and mix.

System suitability

Samples: System suitability solution and Standard solution

Suitability requirements

Resolution: NLT 1.5 between methacycline and 4-epidoxycycline, NLT 1.5 between 4-epidoxycycline and doxycycline related compound A NLT 2.0 between doxycycline related compound A and doxycycline, System suitability solution

Relative standard deviation: NMT 5.0% for the doxycycline peak, Standard solution

Analysis

Samples: Standard solution and Sample solution

Calculate the percentage of each impurity in the portion of Doxycycline for Injection taken:

Result = (r_U/(r_S) × (C_S /C_U) × P x F x 100

r_U = peak response of each impurity from the Sample solution

r_S = peak response of doxycycline from the Standard solution

C_S = concentration of USP Doxycycline Hyclate RS, in the Stardard solution (mg/ml)

C_U = nominal concentration of doxycycline in the Sample sulation (mg/ml)

P = potency of doxycycline in USP Doxycycline Hyclate RS (µg/mg)

F = conversion factor 0.001 mg/μg

Acceptance criteria: See Table 2. Disregard peaks less than 0.1%.

^a (4S,4aR,55,5aR, 12aS)-4-(Dimethylamino)-1,4,4,5,58,6,11,12a-octahydro-3,5,10,12,12a-pentahydroxy-6-methylene-1,11-dioxo-2-naphthacenecarboxamide.

^b (4R,4aR,55,5aR,6,12aS)-4-(Dimethylamino)-1,4,4,5,5a,6,11,12a-octahydro-3,5,10,12,12a-pentahydroxy-6-methyl-1,11-dioxo-2-naphthacenecarboxamide.

^c(4R,4aR,5S,5aR,6R,12aS)-4-(Dimethylamino)-1,4,4a,5,5a,6,11,12a-octahydro-3,5,10,12,12a-pentahydroxy-6-methyl-1,11-dioxo-2- naphthacenecarboxamide.

^d (4S,4aR,5S,5aR,6S,12aS)-4-(Dimethylamino)-1,4,4a,5,5a,6,11,12a-octahydro-3,5,10,12,12a-pentahydroxy-6-methyl-1,11-dioxo-2- naphthacenecarboxamide.

5 SPECIFIC TESTS

STERILITY TESTS (21): Meets the requirements. if the membrane filtration test is used, use Fluid D instead of Fluid A.

PH (791)

Sample solution: Constitute as directed in the labeling.

Acceptance criteria: 1.8-3.3

LOSS ON DRYING (731)

Analysis: Dry 100 mg in a capillary-stoppered bottle under vacuum at a pressure not exceeding 5 mm of mercury at 60 deg for 3 h.

Acceptance criteria: NMT 2.0% for the article containing added substances; NMT 4.0% for the article containing no added substances

PARTICULATE MATTER IN INJECTIONS (288): Meets the requirements for small-volume injections

BACTERIAL ENDOTOXINS TEST (85): Contains NMT 1.14 USP Endotoxin Units/mg of doxycycline

CONSTITUTED SOLUTION: At the time of use, it meets the requirements in Injections and Implanted Drug Products (1), Product Quality Tests Common to Parenteral Dosage Forms. Specific Tests. Completeness and Clarity of Solutions.

6 ADDITIONAL REQUIREMENTS

PACKAGING AND STORAGE: Preserve as described in Packaging and Storage Requirements (659), Injection Packaging, protected from light.