Tetracycline Capsules

General Notices

Action and use

Tetracycline antibacterial.

Definition

Tetracycline Capsules contain Tetracycline Hydrochloride.

The capsules comply with the requirements stated under Capsules and with the following requirements.

Content of tetracycline hydrochloride, C22H24N2O8,HCl

95.0 to 105.0% of the stated amount.

Identification

A. Carry out the method for thin-layer chromatography, Appendix III A, using silica gel H as the coating substance. Adjust the pH of a 10% w/v solution of disodium edetate to 8.0 with 10m sodium hydroxide and spray the solution evenly onto the plate (about 10 mL for a plate 100 mm × 200 mm). Allow the plate to dry in a horizontal position at room temperature for at least 1 hour. Before use, dry the plate in an oven at 110° for 1 hour. Use a mixture of 6 volumes of water, 35 volumes of methanol and 59 volumes of dichloromethane as the mobile phase. Apply separately to the plate 1 µL of each of the following solutions. For solution (1) extract a quantity of the contents of the capsules containing 10 mg of Tetracycline Hydrochloride with 20 mL of methanol and centrifuge. Solution (2) contains 0.05% w/v of tetracycline hydrochloride BPCRS in methanol. Solution (3) contains 0.05% w/v of each of tetracycline hydrochloride BPCRS, chlortetracycline hydrochloride BPCRS and doxycycline hyclate BPCRS in methanol. Dry the plate in a current of air and examine under ultraviolet light (365 nm). The principal spot in the chromatogram obtained with solution (1) is similar in position, colour and size to the principal spot in the chromatogram obtained with solution (2). The test is not valid unless the chromatogram obtained with solution (3) shows three clearly separated spots.
B. To a quantity of the contents of the capsules containing 10 mg of Tetracycline Hydrochloride add 20 mL of warm ethanol (96%), allow to stand for 20 minutes, filter and evaporate the filtrate to dryness on a water bath. To 0.5 mg of the residue add 2 mL of sulfuric acid; a deep crimson colour is produced. Add 1 mL of water; the colour changes to deep yellow.
C. The residue obtained in test B yields the reactions characteristic of chlorides, Appendix VI.

Tests

Dissolution

Comply with the requirements for Monographs of the British Pharmacopoeia in the dissolution test for tablets and capsules, Appendix XII B1, using as the medium 900 mL of 0.1m hydrochloric acid and rotating the basket at 100 revolutions per minute. Withdraw a sample of 10 mL of the medium. Measure the absorbance of the filtered sample, suitably diluted if necessary, at the maximum at 353 nm, Appendix II B. Calculate the total content of tetracycline hydrochloride, C22H24N2O8,HCl, in the medium taking 310 as the value of A(1%, 1 cm) at the maximum at 353 nm.

4-Epitetracycline hydrochloride

Carry out the method for liquid chromatography, Appendix III D, using the following solutions. For solution (1) shake a quantity of the mixed contents of 20 capsules containing 25 mg of Tetracycline Hydrochloride in 80 mL of 0.01m methanolic hydrochloric acid for 10 minutes, dilute to 100 mL with the same solvent, mix and filter if necessary. Solution (2) contains 0.0020% w/v of 4-epitetracycline hydrochloride EPCRS in 0.01m methanolic hydrochloric acid. Solution (3) contains 0.0015% w/v each of 4-epitetracycline hydrochloride EPCRS and tetracycline hydrochloride BPCRS in 0.01m methanolic hydrochloric acid.

The chromatographic procedure may be carried out using (a) a stainless steel column (20 cm × 4.6 mm) packed with end-capped octadecylsilyl silica gel for chromatography (10 µm) (Nucleosil C18 is suitable) and maintained at 40°, (b) as the mobile phase with a flow rate of 2 mL per minute a mixture of 5 volumes of dimethylformamide and 95 volumes of 0.1m oxalic acid the pH of which has been adjusted to 3.9 with triethylamine and (c) a detection wavelength of 280 nm.

The test is not valid unless the resolution factor between the two principal peaks in the chromatogram obtained with solution (3) is at least 2.0.

In the chromatogram obtained with solution (1) the area of any peak corresponding to 4-epitetracycline is not greater than the area of the principal peak in the chromatogram obtained with solution (2).

Anhydrotetracycline hydrochloride and 4-epi-anhydrotetracycline hydrochloride

Carry out the method for liquid chromatography, Appendix III D, using the following solutions. For solution (1) shake a quantity of the mixed contents of 20 capsules containing 25 mg of Tetracycline Hydrochloride in 20 mL of 0.01m methanolic hydrochloric acid for 10 minutes, dilute to 25 mL with the same solvent, mix and filter if necessary. Solution (2) contains 0.0010% w/v each of anhydrotetracycline hydrochloride EPCRS and 4-epianhydrotetracycline hydrochloride EPCRS in 0.01m methanolic hydrochloric acid. Solution (3) contains 0.0010% w/v each of 4-epianhydrotetracycline hydrochloride EPCRS and tetracycline hydrochloride BPCRS.

The chromatographic conditions described under Epitetracycline hydrochloride may be used but use as the mobile phase a mixture of 10 volumes of dimethylformamide, 12 volumes of acetonitrile and 78 volumes of 0.1m oxalic acid the pH of which has been adjusted to 3.9 with triethylamine. The order of emergence of the peaks is tetracycline, 4-epianhydrotetracycline and anhydrotetracycline.

The test is not valid unless the resolution factor between the two principal peaks in the chromatogram obtained with solution (3) is at least 6.0.

In the chromatogram obtained with solution (1) the area of any peaks corresponding to anhydrotetracycline hydrochloride and 4-epianhydrotetracycline hydrochloride is not greater than the area of the respective principal peak in the chromatogram obtained with solution (2).

Loss on drying

When dried at 60° at a pressure not exceeding 0.7 kPa for 3 hours, the contents of the capsules lose not more than 3.0% of their weight. Use 1 g.

Assay

Carry out the method for liquid chromatography, Appendix III D, using the following solutions. For solution (1) shake a quantity of the mixed contents of 20 capsules containing 25 mg of Tetracycline Hydrochloride in 80 mL of 0.01m methanolic hydrochloric acid for 10 minutes, dilute to 100 mL with the same solvent, mix and filter if necessary. Solution (2) contains 0.025% w/v of tetracycline hydrochloride BPCRS in 0.01m methanolic hydrochloric acid. Solution (3) contains 0.025% w/v each of 4-epitetracycline hydrochloride EPCRS and tetracycline hydrochloride BPCRS in 0.01m methanolic hydrochloric acid.

The chromatographic conditions described under 4-Epitetracycline hydrochloride may be used.

The assay is not valid unless the resolution factor between the two principal peaks in the chromatogram obtained with solution (3) is at least 2.0.

Calculate the content of C22H24N2O8,HCl in the capsules using the declared content of C22H24N2O8,HCl in tetracycline hydrochloride BPCRS.