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C18H19ClN4 326.83
5H-Dibenzo[b,e][1,4]diazepine, 8-chloro-11-(4-methyl-1-piperazinyl)-.
8-Chloro-11-(4-methyl-1-piperazinyl)-5H-dibenzo[b,e][1,4]diazepine [5786-21-0].
» Clozapine contains not less than 98.0 percent and not more than 102.0 percent of C18H19ClN4, calculated on the dried basis.
Packaging and storage— Preserve in well-closed containers.
USP Reference standards 11
USP Clozapine RS Click to View Structure
USP Clozapine Resolution Mixture RS Click to View Structure
Contains the following components:
Impurity A: 8-chloro-5,10-dihydro-11H-dibenzo[b,e][1,4]diazepin-11-one.
Impurity B: 11,11¢-(piperazine-1,4-diyl)bis(8-chloro-5H-dibenzo[b,e][1,4]dizepine).
Impurity C: 8-chloro-11-(piperazin-1-yl)-5H-dibenzo[b,e][1,4]diazepine.
Impurity D: 4-chloro-N1-(2-[(4-methylpiperazin-1-yl)carbonyl]phenyl)benzene-1,2-diamine.
B: The retention time of the major peak in the chromatogram of the Test solution corresponds to the major peak in the chromatogram of the Resolution solution, as obtained in the test for Related compounds.
Melting range, Class I 741: between 182 and 186.
Loss on drying 731 Dry it at 105 for 4 hours: it loses not more than 0.5% of its weight.
Residue on ignition 281: not more than 0.1%.
Related compounds—
Diluent— Prepare a mixture of methanol and water (80:20).
Buffer— Dissolve about 2.0 g of monobasic potassium phosphate in 1000 mL of water, and adjust to a pH of 2.4 with phosphoric acid (85%). [note—The pH of this solution must not be below 2.4. ]
Solution A— Prepare a filtered and degassed mixture of Buffer, acetonitrile, and methanol (8:1:1).
Solution B— Prepare a filtered and degassed mixture of acetonitrile, methanol, and Buffer (2:2:1).
Mobile phase— Use variable mixtures of Solution A and Solution B as directed for Chromatographic system.
Resolution solution— Dissolve 4 mg of USP Clozapine Resolution Mixture RS in 4 mL of methanol, add 1 mL of water, and dilute with Diluent to 10 mL.
Standard stock solution— Dissolve an accurately weighed amount of USP Clozapine RS in Diluent to obtain a final solution having a known concentration of about 0.75 mg per mL of clozapine.
Standard solution— Dilute quantitatively a known volume of the Standard stock solution with Diluent to obtain a solution having a known concentration of about 0.75 µg per mL of clozapine.
Test solution— Dissolve 75.0 mg of the substance to be examined in 80 mL of methanol, and dilute with water to 100.0 mL.
Chromatographic system (see Chromatography 621) The liquid chromatograph is equipped with a 257-nm detector and a 4.6-mm × 15-cm column that contains 5-µm L1 packing. The flow rate is about 1.2 mL per minute. The chromatograph is programmed as follows.
Solution A (%) Solution B (%) Elution
0–4 100 0 isocratic
4–24 100®0 0®100 linear gradient
24–29 0 100 isocratic
29–40 0®100 100®0 linear gradient
Chromatograph the Resolution solution, and record the peak responses as directed for Procedure. Locate the clozapine peak and the peaks due to the specified impurities using the relative retention times provided in Table 1; the resolution, R, between impurity C and clozapine, is not less than 2.5. Chromatograph the Standard solution, and record the peak response as directed for Procedure: the relative standard deviation for replicate injections for clozapine is not more than 5.0%.
Procedure— Separately inject equal volumes (about 20 µL) of the Standard solution and the Test solution into the chromatograph, record the chromatograms, and measure the peak response for clozapine obtained from the Standard solution and the responses for all other peaks, except the clozapine peak, obtained from the Test solution. [note—Disregard any peak with an area less than 0.5 times the area of the clozapine peak in the chromatogram obtained from the Standard solution. ] Calculate the percentage of each related compound and any unknown impurity in the portion of Clozapine taken by the formula:
100(CS / CT)(1/F)(ri / rS)
in which CS and CT are the concentrations of clozapine, in mg per mL, in the Standard solution and the Test solution, respectively; F is the relative response factor for each impurity (given in Table 1); ri is the peak response for each individual peak obtained from the Test solution; and rS is the clozapine peak obtained from the Standard solution. The limits for the specified and unspecified impurities are given in Table 1.
Table 1
Peak Approximate
(w/w %)
Impurity C1 0.9 1.0 NMT 0.3
Clozapine 1.0
Impurity D2 1.1 0.35 NMT 0.2
Impurity A3 1.6 1.2 NMT 0.1
Impurity B4 1.7 1.0 NMT 0.2
Individual unspecified impurity 1.0 NMT 0.10
Total impurities NMT 0.6
1  8-Chloro-11-(piperazin-1-yl)-5H-dibenzo[b,e][1,4]diazepine.
2  4-Chloro-N1-(2-((4-methylpiperazin-1-yl)carbonyl)phenyl)benzene-1,2-diamine.
3  8-Chloro-5,10-dihydro-11H-dibenzo[b,e][1,4]diazepin-11-one.
4  11,11¢-(Piperazine-1,4-diyl)bis(8-chloro-5H-dibenzo[b,e][1,4]diazepine).
Assay— Transfer about 115 mg of Clozapine, accurately weighed, to a suitable container, dissolve in 70 mL of glacial acetic acid, and titrate with 0.1 N perchloric acid VS, determining the endpoint potentiometrically. Perform a blank determination, and make any necessary correction. Each mL of 0.1 N perchloric acid is equivalent to 16.34 mg of C18H19ClN4.
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