Erythromycin
(e rith'' roe mye' sin).
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733.94Erythromycin.
(3R*,4S*,5S*,6R*,7R*,9R*,11R*,12R*,13S*,14R*)-4-[(2,6-Dideoxy-3-C-methyl-3-O-methyl-
-l-ribo-hexopyranosyl)oxy]-14-ethyl-7,12,13-trihydroxy-3,5,7,9,11,13-hexamethyl-6-[[3,4,6-trideoxy-3-(dimethylamino)-
-d-xylo-hexopyranosyl]oxy]oxacyclotetradecane-2,10-dione
[114-07-8].» 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 percent and not more than 100.5 percent, calculated on the anhydrous basis.
Packaging and storage—
Preserve in tight containers.
USP Reference standards 
11
—
11—
USP Erythromycin RS 
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USP Erythromycin B RS
USP Erythromycin C RS
USP Erythromycin Related Compound N RS
N-Demethylerythromycin A.
C36H65NO13 719.91
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N-Demethylerythromycin A.
C36H65NO13 719.91
Identification—
The IR absorption spectrum of a solution of it containing 50 mg per mL, previously dried at a pressure not exceeding 5 mm of mercury at 60
for 3 hours, in chloroform, determined in a 0.1-mm cell, exhibits maxima only at the same wavelengths as that of a similar preparation of USP Erythromycin RS, except in the region between 1980 cm
1 and 2050 cm1.
for 3 hours, in chloroform, determined in a 0.1-mm cell, exhibits maxima only at the same wavelengths as that of a similar preparation of USP Erythromycin RS, except in the region between 1980 cm1 and 2050 cm1.
Specific rotation 781S:
between 71 and 78, determined after standing for 30 minutes.
781S:
between 71 and 78, determined after standing for 30 minutes.
Test solution:
20 mg per mL, in dehydrated alcohol.
Crystallinity 695:
meets the requirements.
695:
meets the requirements.
Water, Method I 921:
not more than 10.0%, 20 mL of methanol containing 10% of imidazole being used in place of methanol in the titration vessel.
921:
not more than 10.0%, 20 mL of methanol containing 10% of imidazole being used in place of methanol in the titration vessel.
Residue on ignition 281:
not more than 0.2%.
281:
not more than 0.2%.
Limit of thiocyanate—
Standard solutions—
Transfer about 100 mg of potassium thiocyanate, previously dried at 105 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 low-actinic volumetric flasks. To each flask add 1.0 mL of ferric chloride TS, dilute with methanol to volume, and mix. [note—Use these Standard solutions within 30 minutes. ]
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 low-actinic volumetric flasks. To each flask add 1.0 mL of ferric chloride TS, dilute with methanol to volume, and mix. [note—Use these Standard solutions within 30 minutes. ]
Test solution—
Transfer about 100 mg of Erythromycin, accurately weighed, to a 50-mL low-actinic volumetric flask. Add 20 mL of methanol, and swirl to dissolve. Add 1.0 mL of ferric chloride TS, dilute with methanol to volume, and mix. [note—Use this Test solution within 30 minutes. ]
Blank solution—
Transfer 1.0 mL of ferric chloride TS to a 50-mL low-actinic volumetric flask, dilute with methanol to volume, and mix. [note—Use this Blank solution within 30 minutes. ]
Procedure—
Determine the absorbances of each Standard solution and the Test solution at the wavelength of maximum absorbance at about 492 nm with a spectrophotometer, using the Blank solution to zero the instrument. Calculate the suitability value, S, by the formula:
(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 formula:
(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; AU is the absorbance of the Test solution; WU is the weight, in mg, of Erythromycin taken to prepare the Test solution; and the other terms are as defined above: not more than 0.3% is found.
Limit of related substances—
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 formula:
25(CP / W)(ri / rS)
in which C is the concentration, in mg per mL, of USP Erythromycin RS in the Diluted standard preparation; P is the designated percentage of erythromycin A in the USP Erythromycin RS; W is the weight, in mg, of Erythromycin taken to prepare the Assay preparation; ri 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 chromatogram obtained from the Assay preparation; and rS is the erythromycin A peak response in the chromatogram obtained from the Diluted standard preparation: not more than 3.0% of any individual related substance is found. Calculate the percentage of erythromycin A enol ether in the Erythromycin taken by the formula:
(25 / 11)(CP / W)(rE / rS)
in which 11 is the response factor for erythromycin A enol ether in relation to that of erythromycin A; rE is the peak response of the erythromycin A enol ether peak observed in the chromatogram obtained from the Assay preparation; and the other terms are as defined above: not more than 3.0% of erythromycin A enol ether is found. The percentage of erythromycin B obtained in the Assay is not more than 12.0%; and the percentage of erythromycin C obtained in the Assay is not more than 5.0%.
Assay—
Solution A—
Dissolve 1.75 g of dibasic potassium phosphate in 50 mL of water, adjust with diluted phosphoric acid (1 in 10) or 0.2 N sodium hydroxide to a pH of 9.0, add 400 mL of water, 165 mL of tertiary butyl alcohol, and 30 mL of acetonitrile. Dilute with water to 1000 mL, and mix.
Mobile phase—
Prepare a mixture of Solution A, acetonitrile, and water (5:2:1). Make any necessary adjustments (see System Suitability under Chromatography 621).
621).
Diluent—
Prepare a mixture of pH 7.0 buffer (see under Buffer Solutions in the section Reagents, Indicators, and Solutions) and methanol (15:1).
pH 3.5 Buffer—
Adjust 20 mL of pH 7.0 buffer (see under Buffer Solutions in the section Reagents, Indicators, and Solutions) with phosphoric acid to a pH of 3.5.
[note—Use the following solutions promptly, or within 1 day if stored in a refrigerator. ]
Standard preparation—
Transfer about 100 mg of USP 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 the Standard preparation to a 100-mL volumetric flask, dilute with Diluent to volume, and mix. This solution contains about 0.12 mg of USP Erythromycin RS per mL.
Erythromycins B and C standard solution—
Transfer about 5 mg each of USP Erythromycin B RS and USP 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 USP Erythromycin Related Compound N RS to a 10-mL volumetric flask, add 0.4 mL of Standard preparation, dilute with Erythromycins B and C standard solution to volume, and mix.
Erythromycin A enol ether retention time solution—
Dissolve about 10 mg of USP 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
(see Chromatography 621)—The liquid chromatograph is equipped with a 215-nm detector and a 4.6-mm × 25-cm column that contains packing L21 (1000 
) and is maintained at a constant temperature of about 65. The flow rate is about 2 mL per minute. Chromatograph the Resolution solution, and record the peak responses as directed for Procedure: the relative retention times are about 0.56 for erythromycin related compound N (N-demethylerythromycin A), 0.61 for erythromycin C, 1.0 for erythromycin A, and 1.6 for erythromycin B; and the resolution, R, between erythromycin related compound N and erythromycin C is not less than 0.8, and between erythromycin related compound N and erythromycin A not less than 5.5. Chromatograph the Erythromycin A enol ether retention time solution, and record the peak responses as directed for 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 the Resolution solution. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the relative standard deviation for replicate injections is not more than 2.0%.
621)—The liquid chromatograph is equipped with a 215-nm detector and a 4.6-mm × 25-cm column that contains packing L21 (1000 ) and is maintained at a constant temperature of about 65. The flow rate is about 2 mL per minute. Chromatograph the Resolution solution, and record the peak responses as directed for Procedure: the relative retention times are about 0.56 for erythromycin related compound N (N-demethylerythromycin A), 0.61 for erythromycin C, 1.0 for erythromycin A, and 1.6 for erythromycin B; and the resolution, R, between erythromycin related compound N and erythromycin C is not less than 0.8, and between erythromycin related compound N and erythromycin A not less than 5.5. Chromatograph the Erythromycin A enol ether retention time solution, and record the peak responses as directed for 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 the Resolution solution. Chromatograph the Standard preparation, and record the peak responses as directed for Procedure: the relative standard deviation for replicate injections is not more than 2.0%.
Procedure—
Separately inject equal volumes (about 100 µL) of the Standard preparation, the Diluted standard preparation, the Erythromycins B and C standard solution, and the 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, as determined in the chromatogram obtained from the Erythromycin A enol ether retention time solution (about five times the retention time of the main erythromycin A peak). Measure the areas of the peak responses. Calculate the percentage of erythromycin A in the portion of Erythromycin taken by the formula:
25(CAP / W)(rU / rA)
in which CA is the concentration, in mg per mL, of USP Erythromycin RS in the Standard preparation; P is the designated percentage of erythromycin A in USP Erythromycin RS; W is the quantity, in mg, of Erythromycin taken to prepare the Assay preparation; and rU and rA are the erythromycin A peak responses in the chromatograms obtained from the Assay preparation and the Standard preparation, respectively. Calculate the percentages of erythromycin B and erythromycin C in the portion of Erythromycin taken by the formula:
25(CSP / W)(rU / rS)
in which CS is the concentration, in mg per mL, of the relevant USP Reference Standard in the Erythromycins B and C standard solution; P is the designated percentage of erythromycin B or erythromycin C in the relevant USP Reference Standard; W is the quantity, in mg, of Erythromycin taken to prepare the Assay preparation; and rU and rS are the peak responses of the relevant analyte in the chromatograms obtained from the Assay preparation and the Erythromycins B and C standard solution, respectively.
Auxiliary Information—
Please check for your question in the FAQs before contacting USP.
| Topic/Question | Contact | Expert Committee |
|---|---|---|
| Monograph | Ahalya Wise, M.S.
Senior Scientific Liaison 1-301-816-8161 |
(SM12010) Monographs - Small Molecules 1 |
| Reference Standards | RS Technical Services 1-301-816-8129 rstech@usp.org |
USP35–NF30 Page 3078