Packaging and storage—
Preserve in single-dose or in multiple-dose containers, preferably of Type I glass, protected from light.
Identification—
Dilute, if necessary, a volume of Injection, equivalent to about 25 mg of methotrexate, with water to obtain a solution having a concentration of about 2.5 mg per mL. Adjust with 0.1 N hydrochloric acid to a pH of 4.0. Place the slurry in a 50-mL centrifuge tube, and centrifuge. Decant the supernatant, add 25 mL of acetone, shake, and filter through a solvent-resistant, membrane filter of 0.45-µm pore size. Air-dry the filtered precipitate: the methotrexate so obtained responds to
Identification test
A under
Methotrexate.
Other requirements—
It meets the requirements under
Injections 
1
.
Assay—
pH 6.0 Buffer solution
,
Mobile phase,
System suitability preparation,
System suitability test, and
Standard preparation—Proceed as directed in the
Assay under
Methotrexate.
Assay preparation—
Transfer an accurately measured volume of Injection, equivalent to about 25 mg of methotrexate, to a 250-mL volumetric flask, dilute with Mobile phase to volume, and mix.
Procedure—
Proceed as directed for
Procedure in the
Assay under
Methotrexate. Calculate the quantity, in mg, of methotrexate (C
20H
22N
8O
5) in each mL of the Injection taken by the formula:
250(C/V)(PU/PS),
in which
C is the concentration, in mg per mL, of
USP Methotrexate RS in the
Standard preparation; V is the volume, in mL, of Injection taken; and
PU and
PS are the peak responses obtained from the
Assay preparation and the
Standard preparation, respectively.
