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Category  Antibacterial.

      Erythromycin consists primarily of erythromycin A (C₃₇H₆₇NO₁₃).  The sum of the percentages of erythromycin A, erythromycin B, and erythromycin C is not
less than 85.0 per cent and not more than 100.5 per cent, calculated on the anhydrous basis.

Description  White or slightly yellow powder or colourless or slightly yellow crystals. 

Solubility  Slightly soluble in water (the solubility decreases as the temperature rises); freely soluble in ethanol; soluble in methanol. 

Stability  It is slightly hygroscopic but relatively stable in dry state.  In aqueous solutions, it retains its activity during prolonged storage under refrigeration or when frozen, but slowly loses activity over several days at room temperature or higher. 

Contra-indication  It is contra-indicated in patients with acute porphyria and in those with hypersensitive to erythromycin and other macrolides. 

Warning

      1. It should be used with extreme caution in patients with a history of cardiac arrhythmias or QT prolongation interval especially in high doses of erythromycin or in those with hepatic function impairment or in elderly patients with reversible hearing loss. 

      2. Caution should be exercised when it is to be used concomitantly with hepatotoxic medications, alfentanil, astemizole, carbamazepine, chloramphenicol, clindamycin, cisapride, dispyramide, lincomycin, cyclosporine, warfarin, xanthine derivatives (e.g., aminophylline, theophylline, caffeine, oxtriphylline (except dyphylline)), pimozide, probenecid or alkalinizing agents (e.g., sodium bicarbonate, acetazolamide).

      3. It may cause abdominal pain and cramping, nausea, vomiting and diarrhea. 

      4. Superinfection with resistant organisms and pseudo-membranous colitis associated with erythromycin use may occur. 

      5. Oral candidiasis may occur following prolonged or repeated erythromycin therapy. 

      6.  Drug-induced rhabdomyolysis should be considered in patients receiving erythromycin concomitantly with lovastatin or another 3-hydroxy-3-methylglutarylcoenzyme A (HMG-CoA) reductase inhibitor. 

      7. Risk-benefit should be considered if it is to be used in pregnant or nursing women. 

Precaution

      1. Hepatic function should be determined periodically. 

      2. Group A beta-hemolytic streptococcal infections should be treated for at least 10 days to prevent the development of acute rheumatic fever or acute glomerulonephritis. 

Packaging and storage  Erythromycin shall be kept in tightly closed containers, protected from light, and stored at a temperature not exceeding 30º. 

Labelling  The label on the container states (1) storage condition; (2) parenteral or non-parenteral grade. 

Identification 

      A. The infrared absorption spectrum is concordant with the spectrum obtained from Erythromycin RS (Appendix 2.1) or with the reference spectrum of Erythromycin. 

      B. To 5 mg add 2 ml of sulfuric acid and shake gently:  a reddish brown colour is produced. 

      C. Dissolve 3 mg in 2 ml of acetone and add 2 ml of hydrochloric acid :  an orange colour is produced, which changes to red and then to deep purplish red.  Add 2 ml of chloroform and shake:  the chloroform layer becomes purple. 

      D. To 5 mg add 5 ml of a 0.02 per cent w/v solution of xanthydrol  in a mixture of 1 volume of hydrochloric acid and 99 volumes of 5 M acetic acid and heat on a water-bath:  a red colour is produced. 

Crystallinity  It is crystalline (Appendix 4.14). 

Specific rotation  –71º to –78º, calculated on the anhydrous basis, determined in a 2.0 per cent w/v solution in ethanol.  Measure the angle of rotation 30 minutes after preparing the solution (Appendix 4.8). 

Water  Not more than 10.0 per cent w/w (Karl Fischer Method, Appendix 4.12).  Use 20 ml of methanol containing 10 per cent of imidazole in place of methanol in the titration vessel. 

Sulfated ash  Not more than 0.2 per cent w/w (Appendix 5.3). 

Thiocyanate  Not more than 0.3 per cent w/v.  (Note Prepare all solutions in low-actinic volumetric flasks and use within 30 minutes.)  Carry out the determination as described in the “Ultraviolet and Visible Spectrophotometry” (Appendix 2.2). 

Standard solutions  Transfer about 100 mg of potassium thiocyanate, previously dried at 105º for 1 hour, cooled, and accurately weighed, to each of two 50-ml volumetric flasks.  Add about 20 ml of methanol  to each flask, swirl to dissolve, dilute with methanol  to volume, and mix.  Transfer 5.0 ml of each of these stock solutions to separate 50-ml volumetric flasks, dilute with methanol  to volume, and mix.  Transfer 5.0 ml of each of these intermediate solutions to separate 50-ml volumetric flasks.  To each flask add 1.0 ml of iron(III) chloride TS, dilute with methanol  to volume, and mix.

Test solution  Transfer about 100 mg of the test substance, accurately weighed, to a 50-ml volumetric flask.  Add 20 ml of methanol, and swirl to dissolve. Add 1.0 ml of iron(III) chloride TS, dilute with methanol  to volume, and mix. 

Blank solution Transfer 1.0 ml of iron(III) chloride TS to a 50-ml volumetric flask, dilute with methanol to volume, and mix.

Procedure  Measure the absorbances of each Standard solution and Test solution at the maximum at about 492 nm, using Blank solution as the blank. 

Calculation  Calculate the suitability value, S, by the expression: 

 (A₁/W₁)(W₂/A₂), 

in which A₁ and A₂ are the absorbance values obtained from the respective Standard solutions, and W₁ and W₂ are the weights, in mg, of the potassium thiocyanate taken to prepare the corresponding Standard solutions. In a suitable determination, the value, S, is not less than 0.985 and not more than 1.015.  Calculate the percentage of thiocyanate in the Erythromycin taken by the expression: 

(58.08/97.18)(AU/WU)(0.5)[(W₁/A₁) + (W₂/A₂)], 

in which 58.08 and 97.18 are the molecular weights of the thiocyanate moiety and of potassium thiocyanate, respectively, A_U is the absorbance of the Test solution, W_U is the weight, in mg, of Erythromycin taken to prepare the Test solution, and the other terms are as defined above. 

Related substances  Carry out the test as described in the “High-pressure Liquid Chromatography” (Appendix 3.5) under Assay, using, the chromatograms of the Assay preparation and the Diluted standard preparation obtained in the Assay, calculate the percentage of any individual related substance observed having the greatest response, other than erythromycin A, erythromycin B, erythromycin C, and erythromycin A enol ether, in the Erythromycin taken by the expression: 

25(CP/W)(r_i/r_S), 

in which C is the concentration, in mg per ml, of Erythromycin RS in Diluted standard preparation; P is the designated percentage of erythromycin A in the Erythromycin RS; W is the weight, in mg, of Erythromycin taken to prepare the Assay preparation; r_i is the peak response of an individual related substance, other than erythromycin A, erythromycin B, erythromycin C, or erythromycin A enol ether, observed in the chromato gram obtained from Assay preparation; and r_S is the erythromycin A peak response in the chromatogram obtained from Diluted standard preparation:  not more than 3.0 per cent w/v of any individual related substance is found.  Calculate the percentage of erythromycin A enol ether in the Erythromycin taken by the expression: 

(25/11)(CP/W)(r_E/r_S),

in which 11 is the response factor for erythromycin A enol ether in relation to that of erythromycin A; r_E is the peak response of the erythromycin A enol ether peak observed in the chromatogram obtained from Assay preparation; and the other terms are as defined above: not more than 3.0 per cent w/v of erythromycin A enol ether is found.  The percentage of erythromycin B obtained in the Assay is not more than 12.0 per cent w/v; and the percentage of erythromycin C obtained in the Assay is not more than 5.0 per cent w/v. 

Assay  Carry out the determination as described in the “High-pressure Liquid Chromatography” (Appendix 3.5). 

      Solution A  Dissolve 1.75 g of dipotassium hydrogenphosphate in 50 ml of water, adjust with diluted phosphoric acid (1 in 10) or 0.2 M sodium hydroxide to a pH of 9.0, add 400 ml of water, 165 ml of 2-methyl-2-propanol, and 30 ml of acetonitrile.  Dilute with water to 1000 ml, and mix. 

      pH 3.5 buffer  Adjust 20 ml of phosphate buffer pH 7.0 with phosphoric acid to a pH of 3.5.

      Mobile phase  Prepare a mixture of 5 volumes of Solution A, 2 volumes of acetonitrile, and 1 volume of water.  Make adjustments if necessary.

      Diluent  Prepare a mixture of 15 volumes of phosphate buffer pH 7.0 and 1 volume of methanol. 

(Note  Use the following solutions promptly, or within 1 day if stored in a refrigerator.) 

      Standard preparation  Transfer about 100 mg of Erythromycin RS, accurately weighed, to a 25-ml volumetric flask, add 5 ml of methanol, swirl to dissolve, dilute with Diluent to volume, and mix. 

      Diluted standard preparation  Transfer 3.0 ml of Standard preparation to a 100-ml volumetric flask, dilute with Diluent  to volume, and mix.  This solution contains about 0.12 mg of Erythromycin RS per ml. 

      Erythromycins B and C standard preparation Transfer about 5 mg each of Erythromycin B RS and Erythromycin C RS, both accurately weighed, to a 25-ml volumetric flask, add 5 ml of methanol, swirl to dissolve, dilute with Diluent to volume, and mix.

      Resolution solution Transfer about 2 mg of NDemethylerythromycin A RS to a 10-ml volumetric flask, add 0.4 ml of Standard preparation, dilute with Erythromycin B and C standard preparation to volume, and mix

      Erythromycin A enol ether retention time solution Dissolve about 10 mg of Erythromycin RS in 2 ml of methanol.  Add 10 ml of pH 3.5 buffer, mix, and allow to stand for about 30 minutes. 

      Assay preparation  Transfer about 100 mg of Erythromycin, accurately weighed, to a 25-ml volumetric flask, add 5 ml of methanol, and swirl to dissolve.  Dilute with Diluent to volume, and mix. 

      Chromatographic system  The chromatographic procedure may be carried out using (a) a stainless steel column (25 cm × 4.6 mm) packed with a rigid, spherical styrene-divinylbenzene copolymer (1000 Å), maintained at a constant temperature of about 65º, (b) Mobile phase at a flow rate of about 2 ml per minute, and (c) an ultraviolet photometer set at 215 nm. 

To determine the suitability of the chromatographic system, chromatograph Resolution solution, and record the peak responses as directed under Procedure:  the relative retention times are about 0.56 for N-demethylerythromycin A, 0.61 for erythromycin C, 1.0 for erythromycin A, and 1.6 for erythromycin B, and resolution factor between N-demethylerythromycin A and erythromycin C is not less than 0.8 and between erythromycin related compound N and erythromycin A not less than 5.5.  Chromatograph Erythromycin A enol ether retention time solution, and record the peak responses as directed under Procedure:  the retention time of the erythromycin A enol ether peak is about 3.2 relative to that of the erythromycin A peak as observed in the chromatogram obtained from Resolution solution.  Chromatograph Standard preparation and record the peak responses as directed under Procedure:  the relative standard deviation for replicate injections is not more than 2.0 per cent. 

      Procedure  Separately inject equal volumes (about 100 μl) of Standard preparation, Diluted Standard preparation, Erythromycins B and C standard solution and Assay preparation into the chromatograph, record the chromatograms for a period of time that is adequate to include the erythromycin A enol ether peak, if present, and measure the responses for the major peaks. 

      Calculation  Calculate the content of erythromycin A in the portion of Erythromycin taken by the expression: 

5(C_AP/W)(r_u/r_A),

in which C_A is the concentration, in mg per ml, of Erythromycin RS in Standard preparation, P is the designated percentage of erythromycin A in Erythromycin RS, W is the quantity, in mg, of Erythromycin taken to prepare Assay preparation, and r_u and r_A are the erythromycin A peak responses in the chromatograms obtained from Assay preparation and Standard preparation, respectively.  Calculate the content of erythromycin B and erythromycin C in the portion of Erythromycin  taken by the expression: 

25 (C_sP/W)(r_u/r_s), 

in which C_s is the concentration, in mg per ml, of the relevant Reference Substance in Erythromycins B and C standard solution, P is the designated percentage of erythromycin B or erythromycin C in the relevant Reference Substance, W is the quantity, in mg, ofErythromycin taken to prepare Assay preparation, and r_u and r_s are the peak responses of the relevant analyte in the chromatograms obtained from Assay preparation and Erythromycins B and C standard solution, respectively.