Vancomycin

(van'' koe mye' sin).

C66H75Cl2N9O24 1449.25
Vancomycin
(Sa)-(3S,6R,7R,22R,23S,26S,36R,38aR)-44-[[2-O-(3-Amino-2,3,6-trideoxy-3-C-methyl-

-L-lyxo-hexopyranosyl)-

-d-glucopyranosyl]oxy]-3-(carbamoylmethyl)-10,19-dichloro-2,3,4,5,6,7,23,24,25,26,36,37,38,38a-tetradecahydro-7,22,28,30,32-pentahydroxy-6-[(2R)-4-methyl-2-(methylamino)valeramido]-2,5,24,38,39-pentaoxo-22H

8,11:18,21-dietheno-23,36-(iminomethano)-13,16:31,35-dimetheno-1H,16H-[1,6,9]oxadiazacyclohexadecino[4,5-m][10,2,16]-benzoxadiazacyclotetracosine-26-carboxylic acid;
[3S-[3R*,6S*(S*),7S*,22S*,23R*,26R*,36S*,38aS*]]-3-(2-Amino-2-oxoethyl)-44-[[2-O-(3-amino-2,3,6-trideoxy-3-C-methyl-

-L-lyxo-hexopyranosyl)-

-d-glucopyranosyl]oxy]-10,19-dichloro-2,3,4,5,6,7,23,24,25,26,36,37,38,38a-tetra-decahydro-7,22,28,30,32-pentahydroxy-6-[[4-methyl-2-(methylamino)-1-oxopentyl]amino]-2,5,24,38,39-pentaoxo-22H

8,11:18,21-dietheno-23,36-(iminomethano)-13,16:31,35-dimetheno-1H, 16H-[1,6,9]oxadiazacyclohexadecino[4,5-m][10,2,16]-benzoxadiazacyclotetraine-26-carboxylic acid

[1404-90-6].

DEFINITION

Vancomycin has a potency equivalent to NLT 950 µg/mg of C66H75Cl2N9O24, calculated on the anhydrous basis.

IDENTIFICATION

• A. Infrared Absorption

197K

• B. Identification Tests—General, Chloride

191

: It does not meet the requirements of the test.

ASSAY

• Antibiotics—Microbial Assays

Sample solution: Transfer 100 mg of Vancomycin to a 100-mL volumetric flask. Add 50 mL of water and 1 mL of 0.1 N hydrochloric acid, and swirl to dissolve, using sonication if necessary. Dilute with water to volume. Dilute a volume of this solution with Buffer No. 4 to yield a Test Dilution having a concentration assumed to be equal to that of the median dose of the standard.

Analysis: Proceed as directed for Vancomycin in the chapter.

Acceptance criteria: NLT 950 µg/mg of vancomycin on the anhydrous basis

IMPURITIES

• Heavy Metals, Method II

231

: NMT 30 ppm

SPECIFIC TESTS

• Composition of Vancomycin

Buffer: Triethylamine and water (1:500). Adjust with phosphoric acid to a pH of 3.2.

Solution A: Acetonitrile, tetrahydrofuran, and Buffer (7:1:92)

Solution B: Acetonitrile, tetrahydrofuran, and Buffer (29:1:70)

Mobile phase: See Table 1. Make adjustments if necessary, changing the acetonitrile proportion in Solution A to obtain a retention time of 7.5–10.5 min for the main vancomycin peak.

Table 1

Time (min)	Solution A (%)	Solution B (%)
0	100	0
12	100	0
20	0	100
22	0	100
23	100	0
30	100	0

System suitability solution: 0.5 mg/mL of USP Vancomycin Hydrochloride RS in water. Heat at 65

for 48 h, and allow to cool.

Sample stock solution: Transfer 250 mg of Vancomycin to a 25-mL volumetric flask. Add 5 mL of Solution A, then add 0.1 N hydrochloric acid dropwise with swirling until dissolution is achieved. Dilute with Solution A to volume.

Sample solution: Dilute 2.0 mL of the Sample stock solution to 50 mL with Solution A.

Chromatographic system

(See Chromatography

621

, System Suitability.)

Mode: LC

Detector: UV 280 nm

Column: 4.6-mm × 25-cm; 5-µm packing L1

Flow rate: 2 mL/min

Injection size: 20 µL

System suitability

Sample: System suitability solution

[Note—The elution order is compound 1, vancomycin B, and compound 2. Compound 2 elutes 3–6 min after the start of the period when the percentage of Solution B is increasing from 0% to 100%. ]

Suitability requirements

Resolution: NLT 3.0 between compound 1 and vancomycin B

Column efficiency: NLT 1500 theoretical plates for the vancomycin B peak

Analysis

Samples: Sample stock solution and Sample solution

Where baseline separation is not achieved, peak areas are defined by vertical lines extended from the valleys between peaks to the baseline. The main component peak may include a fronting shoulder, which is attributed to monodechlorovancomycin. This shoulder should not be integrated separately.

Correct any peak observed in the chromatograms obtained from the Sample stock solution and Sample solution by subtracting the area response of any peak observed in the chromatogram of Solution A at the corresponding retention time.

Calculate the percentage of vancomycin B in the portion of Vancomycin taken:

Result = {(D × rB)/[(D × rB) + rA]} × 100

D	=	= dilution factor, Sample stock solution to Sample solution, 25
rB	=	= corrected peak area response of the main peak from the Sample solution
rA	=	= sum of the corrected peak area responses of all the peaks, other than the main peak, from the Sample stock solution

Calculate the percentage of any individual peak, other than the main peak, in the portion of Vancomycin taken:

Result = {ri/[(D × rB) + rA]} × 100

ri	=	= corrected peak area response of any individual peak, other than the main peak, from the Sample stock solution
D	=	= dilution factor, Sample stock solution to Sample solution, 25
rB	=	= corrected peak area response of the main peak from the Sample solution
rA	=	= sum of the corrected peak area responses of all the peaks, other than the main peak, from the Sample stock solution

Acceptance criteria: NLT 92% of vancomycin B; NMT 3% of any individual peak other than the main peak

• Water Determination, Method I

921

: NMT 20%

ADDITIONAL REQUIREMENTS

• Packaging and Storage: Preserve in tight containers.

• USP Reference Standards

USP Vancomycin Hydrochloride RS

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 5000

Pharmacopeial Forum: Volume No. 30(6) Page 2055