Cephalexin Hydrochloride
(sef'' a lex' in hye'' droe klor' ide).

C16H17N3O4S·HCl·H2O 401.87
5-Thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid, 7-[(aminophenylacetyl)amino]-3-methyl-8-oxo-, monohydrochloride, monohydrate, [6R-[6,7 (R*)]]-;    
(6R,7R)-7-[(2R)-2-Amino-2-phenylacetamido]-3-methyl-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylic acid, monohydrochloride, monohydrate;    
7-(d-2-Amino-2-phenylacetamido)-3-methyl-3-cephem-4-carboxylic acid hydrochloride monohydrate     [105879-42-3].
DEFINITION
Cephalexin Hydrochloride contains the equivalent of NLT 800 µg/mg and NMT 880 µg/mg of cephalexin (C16H17N3O4S).
IDENTIFICATION
•  A. The retention time of the major peak of the Sample solution corresponds to that of the Standard solution, as obtained in the Assay.
•  B. Identification Tests—General, Chloride 191: 10 mg/mL meets the requirements
ASSAY
•  Procedure
Mobile phase:  0.985 g/L of sodium 1-pentanesulfonate in a mixture of acetonitrile, methanol, triethylamine, and water (20:10:3:170), adjusted with phosphoric acid to a pH of 3.0 ± 0.1
Standard stock solution:  1 mg/mL of USP Cephalexin RS in water
Standard solution:  0.4 mg/mL of cephalexin in Mobile phase from Standard stock solution
Sample stock solution:  1.15 mg/mL of Cephalexin Hydrochloride in water
Sample solution:  0.4 mg/mL of cephalexin in Mobile phase from Sample stock solution
Chromatographic system 
Mode:  LC
Detector:  UV 254 nm
Column:  4.6-mm × 25-cm; packing L1 of low acidity
Flow rate:  1.5 mL/min
Injection size:  20 µL
System suitability 
Sample:  Standard solution
Suitability requirements 
Relative standard deviation:  NMT 2.0%
Analysis 
Samples:  Standard solution and Sample solution
Calculate the quantity, in µg, of cephalexin (C16H17N3O4S) in each mg of Cephalexin Hydrochloride taken:
Result = (rU/rS) × (CS/CU) × P
rU== peak response from the Sample solution
rS== peak response from the Standard solution
CS== concentration of USP Cephalexin RS in the Standard stock solution (mg/mL)
CU== concentration of Cephalexin Hydrochloride from the Sample stock solution (mg/mL)
P== potency of cephalexin in USP Cephalexin RS (µg/mg)
Acceptance criteria:  800–880 µg/mg
IMPURITIES
Organic Impurities 
•  Procedure 1
Solution A:  1 g of sodium 1-pentanesulfonate in a mixture of 1000 mL of water and 15 mL of triethylamine. Adjust with phosphoric acid to a pH of 2.5 ± 0.1.
Solution B:  1 g of sodium 1-pentanesulfonate in a mixture of 300 mL of water and 15 mL of triethylamine. Adjust with phosphoric acid to a pH of 2.5 ± 0.1, and add 350 mL of acetonitrile and 350 mL of methanol.
Mobile phase:  See the gradient table below.
Time
(min)
Solution A
(%)
Solution B
(%)
0 100 0
1 100 0
33.3 0 100
34.3 0 100
Diluent:  18 mg/mL of monobasic potassium phosphate in water
Standard solutions:  0.08 mg/mL and 0.16 mg/mL of cephalexin (C16H17N3O4S) from USP Cephalexin RS in Diluent, taking into account the stated potency of the USP Cephalexin RS
Sample solution:  6 mg/mL of Cephalexin Hydrochloride in Diluent
Chromatographic system 
Mode:  LC
Detector:  UV 254 nm
Column:  4.6-mm × 25-cm; packing L1 of low acidity
Flow rate:  1 mL/min
Injection size:  20 µL
Analysis 
Samples:  Standard solutions and Sample solution
Plot the responses of the cephalexin peaks of the Standard solutions versus their concentrations, calculated on the anhydrous basis, in mg/mL, and draw a straight line through the two points and zero. From the line and the peak responses of the Sample solution, determine the concentration, I, in mg/mL, of each cephalexin-related substance from the Sample solution other than the cephalexin peak.
Calculate the percentage of each cephalexin-related substance represented by each peak of the Sample solution, other than the cephalexin peak.
Result = (I/C) × 100
I== concentration of each cephalexin-related substance other than cephalexin in the Sample solution (mg/mL)
C== concentration of cephalexin from the Sample solution (mg/mL)
Acceptance criteria 
Individual impurities:  NMT 1.0% of any individual cephalexin-related substance is found.
Total impurities:  NMT 5.0%
•  Procedure 2: Dimethylaniline 223: Meets the requirement
SPECIFIC TESTS
•  Crystallinity 695: Meets the requirements
•  pH 791: 1.5–3.0, in a solution containing 10 mg/mL
•  Water Determination, Method I 921: 3.0%–6.5%
ADDITIONAL REQUIREMENTS
•  Packaging and Storage: Preserve in tight containers.
•  USP Reference Standards 11
USP Cephalexin RS Click to View Structure
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 2585
Pharmacopeial Forum: Volume No. 36(1) Page 42