» Reserpine Injection is a sterile solution of Reserpine in Water for Injection, prepared with the aid of a suitable acid. It contains not less than 90.0 percent and not more than 110.0 percent of the labeled amount of C33H40N2O9. It contains suitable antioxidants.
Packaging and storage Preserve in single-dose (or, if stabilizers are present, in multiple-dose), light-resistant containers, preferably of Type I glass.
Identification It responds to the Identification test under Reserpine Oral Solution.
Bacterial endotoxins 85 It contains not more than 71.5 USP Endotoxin Units per mg of reserpine.
pH 791: between 3.0 and 4.0.
Other alkaloids [noteConduct this test promptly after preparation of the test and standard solutions.] Pipet 10 mL each of the citric acid solution of the Injection, and of Solution 1 used in preparing the Standard preparation, respectively, obtained as directed in the Assay, into two separators. To the Injection solution add 100 mL of saturated sodium bicarbonate solution, and to Solution 1 add 10 mL of water, 10 drops of saturated sodium bicarbonate solution, and 90 mL of water, and extract both of the resulting solutions with 50 mL of ether. Transfer the aqueous phase to another separator, extract with a second 50-mL portion of ether, and discard the aqueous layers. Wash the ether layers in succession with two 25-mL portions of water, and discard the washings. Extract the combined ether layers with three 15-mL portions of 2 N sulfuric acid, collect the extracts in a 50-mL volumetric flask, add 2 N sulfuric acid to volume, and mix. The absorption spectrum of the solution from the Injection, in the range of 255 to 350 nm, measured in a 1-cm cell, 2 N sulfuric acid being used as the blank, exhibits maxima and minima only at the same wavelengths as that of the solution from the Standard preparation, concomitantly measured. Calculate the quantity, in mg, of total alkaloids in each mL of the Injection taken by the formula:
10(I / SV)in which I is the absorbance of the solution from the Injection at the wavelength of maximum absorbance at about 268 nm; S is that of the solution from the Standard preparation; and V is the volume, in mL, of Injection taken. The content of total alkaloids is not more than 114.0% of the labeled amount of C33H40N2O9, and does not differ by more than 10.0% from the amount of C33H40N2O9 determined in the Assay.
Other requirements It meets the requirements under Injections 1.
Standard preparation Dissolve 25.0 mg of USP Reserpine RS, accurately weighed, in 0.25 mL of chloroform, mix with about 30 mL of methanol previously warmed to 50, transfer the mixture to a 250-mL volumetric flask with the aid of warm methanol, cool the solution to room temperature, dilute with methanol to volume, and mix (Solution 1). Protect the solution from light. Just prior to use in the Assay, pipet 10 mL of Solution 1 into a 50-mL volumetric flask, add 36 mL of chloroform, and dilute with methanol to volume (Standard preparation).
Assay preparation Transfer to a 100-mL volumetric flask an accurately measured volume of Injection, equivalent to about 10 mg of reserpine, and dilute with citric acid solution (1 in 50) to volume.
Procedure Pipet 10 mL of the Assay preparation into a separator, or a suitably stoppered, 50-mL centrifuge tube, add 10 mL of chloroform, and shake for 2 minutes. Separate and withdraw the chloroform. Wash the citric acid solution with two 10-mL portions of chloroform, adding the washings to the main chloroform solution. To the combined chloroform solutions add 10 mL of sodium bicarbonate solution (1 in 100), shake for 2 minutes, and separate. Withdraw the chloroform, filtering it through a pledget of cotton, into a 50-mL volumetric flask containing 14.0 mL of methanol. Extract the aqueous bicarbonate layer in the extraction vessel with two 2-mL portions of chloroform, passing each portion successively through the filter into the volumetric flask. Add chloroform to volume, and mix.
Pipet duplicate 5-mL aliquots of the chloroform-methanol solution and of the Standard preparation into separate, 10-mL volumetric flasks. Add 2.0 mL of a 1 in 10 solution of hydrochloric acid in methanol to each flask. To one flask of each pair of duplicates (representing the Standard preparation and the extracted Assay preparation) add 1.0 mL of a 3 in 1000 solution of sodium nitrite in dilute methanol (1 in 2). To the second flask of each pair (constituting the blanks) add 1 mL of dilute methanol (1 in 2). Mix, and allow to stand for 30 minutes. Add 0.5 mL of ammonium sulfamate solution (1 in 20) to each flask, add methanol to volume, mix, and allow to stand for 10 minutes. Determine the absorbance of each solution in a 1-cm cell at the wavelength of maximum absorbance at about 390 nm, with a suitable spectrophotometer, using a mixture of methanol, chloroform, and water (5.4:3.6:1) as the blank.
Calculate the quantity, in mg, of C33H40N2O9 in each mL of the Injection taken by the formula:
10(A A0)U / V(A A0)S
in which V is the volume, in mL, of Injection taken, and the parenthetic expressions are the differences in absorbances of the nitrite-treated and blank solutions, respectively, from the Assay preparation (U) and the Standard preparation (S).
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USP32NF27 Page 3487