Methylene Blue Injection
»Methylene Blue Injection is a sterile solution of Methylene Blue in Water for Injection.It contains,in each mL,not less than 9.5mg and not more than 10.5mg of methylene blue (C16H18ClN3S·3H2O).
Packaging and storage— Preserve in single-dose containers,preferably of Type Iglass.
Identification—
A: The visible absorption spectrum of the solution employed for measurement of absorbance in the Assayexhibits maxima and minima at the same wavelengths as that of the Standard solution employed in the Assay,concomitantly measured.
B: Dilute a portion of the Injection with an equal volume of methanol.Dissolve 5mg of USP Methylene Blue RSin 1mLof a mixture of equal volumes of methanol and water.Apply 1µLof each solution to a thin-layer chromatographic plate,coated with a 0.25-mm layer of chromatographic silica gel,allow the spots to dry,and develop the chromatogram,using a mixture of water,alcohol,and acetic acid (4:3:3)as the solvent system,until the solvent front has moved about 10cm above the line of application.Remove the plate from the developing chamber,and allow the solvent to evaporate:the RFvalue of the principal spot obtained from the Methylene Blue corresponds to that obtained from the Reference Standard.
Bacterial endotoxins á85ñ— It contains not more than 2.5USP Endotoxin Units per mL.
pHá791ñ: between 3.0and 4.5.
Other requirements— It meets the requirements under Injections á1ñ.
Assay— Dilute an accurately measured volume of Injection,equivalent to about 100mg of methylene blue trihydrate to a 200-mLvolumetric flask,dissolve in and dilute with diluted alcohol to volume,and mix.Transfer 5.0mLof this solution to a 100-mLvolumetric flask,dilute with diluted alcohol to volume,and mix.Transfer 5.0mLof this solution to a 50-mLvolumetric flask,dilute with diluted alcohol to volume,and mix.This solution contains about 2.4µg of methylene blue trihydrate (2µg of anhydrous methylene blue)per mL.Prepare a Standard solution of USP Methylene Blue RSas directed in the Assayunder Methylene Blue.Concomitantly determine the absorbance of both solutions in 1-cm cells at the wavelength of maximum absorbance at about 663nm,with a suitable spectrophotometer,using diluted alcohol as the blank.Calculate the quantity,in mg,of methylene blue (C16H18ClN3S·3H2O)in each mLof the Injection taken by the formula:
(373.90/319.86)(40C/V)(AU/AS),
in which 373.90and 319.86are the molecular weights of methylene blue trihydrate and anhydrous methylene blue,respectively;Cis the concentration,in µg per mL,of USP Methylene Blue RSin the Standard solution;Vis the volume,in mL,of Injection taken;and AUand ASare the absorbances of the solution from the Injection and the Standard solution,respectively.
Auxiliary Information— Staff Liaison:Andrzej Wilk,Ph.D.,Senior Scientific Associate
Expert Committee:(PA5)Pharmaceutical Analysis 5
USP28–NF23Page 1264
Phone Number:1-301-816-8305