Related compounds—
Buffer solution, Solution A, Solution B, Mobile phase, and Standard preparation—
Proceed as directed in the
Assay under
Atracurium Besylate.
System suitability solution—
Heat a portion of the
Standard preparation at 90
for 30 minutes, and immediately chill to about 5
.
Diluted standard preparation—
Dilute a portion of the Standard preparation quantitatively, and stepwise if necessary, with Solution A to obtain a solution having a known concentration of about 0.02 mg per mL.
Test preparation—
Use the Assay preparation.
Chromatographic system—
Prepare as directed for Chromatographic system in the Assay. Chromatograph the System suitability solution and the Diluted standard preparation, record the chromatograms, and measure the responses for the degradation products by comparing the peak responses of the System suitability solution to those of the Diluted standard preparation as directed for Procedure: the retention times relative to the atracurium besylate cis-cis-isomer are about 0.22 for the acidic compound; 0.29 for laudanosine; 0.44 and 0.50 for the trans- and cis-isomers, respectively, of the hydroxy compound; and about 1.28 and 1.33 for the trans- and cis-isomers, respectively, of the monoacrylate.
Procedure—
Separately inject equal volumes (about 20 µL) of the
Diluted standard preparation and the
Test preparation into the chromatograph, record the chromatograms, and measure the peak responses, except the peak due to benzenesulfonic acid occurring at a retention time of about 0.08 relative to the atracurium besylate
cis-cis-isomer. Calculate the percentage of each impurity in the portion of
Test preparation taken by the formula:
100(C/M)(ri / rs),
in which
C is the concentration, in mg per mL, of
USP Atracurium Besylate RS in the
Diluted standard preparation; M is the concentration of atracurium besylate, in mg per mL, in the
Test preparation; ri is the peak response for each impurity obtained from the
Test preparation; and
rs is the sum of the responses of all the peaks obtained from the
Diluted standard preparation: not more than 6.0% of the acidic compound, not more than 6.0% of the combined
cis- and
trans-isomers of the hydroxy compound, not more than 3.0% of laudanosine, not more than 3.0% of the combined
cis- and
trans-isomers of the monoacrylate, and not more than 2.0% of other known synthetic impurities is found; not more than 0.1% of any other impurity is found; and not more than 15.0% of total impurities is found.
Assay—
Buffer solution, Solution A, Solution B, Mobile phase, and Standard preparation—
Proceed as directed in the
Assay under
Atracurium Besylate.
Assay preparation—
Transfer an accurately measured volume of Injection, equivalent to about 50 mg of atracurium besylate, to a 50-mL volumetric flask, dilute with Solution A to volume, and mix.
Chromatographic system (see Chromatography 
621
)—
The liquid chromatograph is equipped with a 280-nm detector and a 4.6-mm × 25-cm column that contains base-deactivated packing L1. The flow rate is about 1 mL per minute. The chromatograph is programmed as follows.
Time
(minutes) |
Solution A
(%) |
Solution B
(%) |
Elution |
| 0 |
80 |
20 |
equilibration |
| 0–5 |
80 |
20 |
isocratic |
| 5–15 |
80®40 |
20®60 |
linear gradient |
| 15–25 |
40 |
60 |
isocratic |
| 25–30 |
40®0 |
60®100 |
linear gradient |
Chromatograph replicate injections of the
Standard preparation, and record the peak responses as directed for
Procedure: the relative retention times are about 0.8 for the atracurium besylate
trans-trans-isomer, 0.9 for the
cis-trans-isomer, and 1.0 for the
cis-cis-isomer; the resolution,
R, between the atracurium besylate
trans-trans-isomer and the
cis-trans-isomer and between the atracurium besylate
cis-trans-isomer and the
cis-cis-isomer is not less than 2.0; and the relative standard deviation for replicate injections is not more than 2.0%.
Procedure—
Separately inject equal volumes (about 20 µL) of the
Standard preparation and the
Assay preparation into the chromatograph, record the chromatograms, and measure the responses for the three atracurium besylate isomer peaks. Calculate the quantity, in mg, of atracurium besylate (C
65H
82N
2O
18S
2) in each mL of the Injection taken by the formula:
50(C/V)(rU / rS),
in which
C is the concentration, in mg per mL, of
USP Atracurium Besylate RS in the
Standard preparation; V is the volume, in mL, of Injection taken for the
Assay preparation; and
rU and
rS are the sums of the peak responses of the atracurium besylate
trans-trans, trans-cis, and
cis-cis-isomers obtained from the
Assay preparation and the
Standard preparation, respectively.
