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 9.5 mg and not more than 10.5 mg of methylene blue (C16H18ClN3S·3H2O).
Packaging and storage Preserve in single-dose containers, preferably of Type I glass.
A: The visible absorption spectrum of the solution employed for measurement of absorbance in the Assay exhibits 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 5 mg of USP Methylene Blue RS in 1 mL of a mixture of equal volumes of methanol and water. Apply 1 µL of 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 10 cm above the line of application. Remove the plate from the developing chamber, and allow the solvent to evaporate: the RF value of the principal spot obtained from the Methylene Blue corresponds to that obtained from the Reference Standard.
Bacterial endotoxins 85 It contains not more than 2.5 USP Endotoxin Units per mL.
pH 791: between 3.0 and 4.5.
Other requirements It meets the requirements under Injections 1.
Assay Dilute an accurately measured volume of Injection, equivalent to about 100 mg of methylene blue trihydrate to a 200-mL volumetric flask, dissolve in and dilute with diluted alcohol to volume, and mix. Transfer 5.0 mL of this solution to a 100-mL volumetric flask, dilute with diluted alcohol to volume, and mix. Transfer 5.0 mL of this solution to a 50-mL volumetric flask, dilute with diluted alcohol to volume, and mix. This solution contains about 2.4 µg of methylene blue trihydrate (2 µg of anhydrous methylene blue) per mL. Prepare a Standard solution of USP Methylene Blue RS as directed in the Assay under Methylene Blue. Concomitantly determine the absorbance of both solutions in 1-cm cells at the wavelength of maximum absorbance at about 663 nm, with a suitable spectrophotometer, using diluted alcohol as the blank. Calculate the quantity, in mg, of methylene blue (C16H18ClN3S·3H2O) in each mL of the Injection taken by the formula:
(373.90 / 319.86)(40C / V)(AU / AS)in which 373.90 and 319.86 are the molecular weights of methylene blue trihydrate and anhydrous methylene blue, respectively; C is the concentration, in µg per mL, of USP Methylene Blue RS in the Standard solution; V is the volume, in mL, of Injection taken; and AU and AS are the absorbances of the solution from the Injection and the Standard solution, respectively.
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USP32NF27 Page 2946