Packaging and storage—
Preserve in well-closed, light-resistant containers. Store at 25
, excursions permitted between 15
and 30
.
Identification—
B:
The retention times of the two principal peaks in the chromatogram of the Test solution correspond to those in the chromatogram of Standard solution 2, as obtained in Related compounds, Test 2. [NOTE—A third isomer may appear as a minor peak.]
C:
Heat 0.5 g in a crucible: violet vapors are evolved.
Microbial limits 
61
—
Proceed as directed for
Plate Method under
Total Aerobic Microbial Count: not more than 100 cfu per g.
Free iodine—
Transfer 2 g to a glass-stoppered tube, add about 20 mL of water, 5 mL of toluene, and 5 mL of 2 N sulfuric acid, shake vigorously, and allow the phases to separate: the toluene layer shows no red or pink color.
Limit of free iodide—
Transfer 5.0 g to a suitable container, add about 30 mL of water, and titrate with 0.001 N silver nitrate VS, determining the endpoint potentiometrically. Each mL of 0.001 N silver nitrate is equivalent to 126.9 µg of iodine. Not more than 0.39 mL of 0.001 N silver nitrate is required: not more than 10 µg of iodide per g is found.
Limit of free aromatic amine—
N-(1-Naphthyl)ethylenediamine dihydrochloride solution—
Prepare a fresh solution of N-(1-naphthyl)ethylenediamine dihydrochloride (3 in 1000) in a mixture of propylene glycol and water (70:30).
Blank solution—
Add 15 mL of water to a 25-mL volumetric flask.
Standard stock solution—
Dissolve an accurately weighed quantity of
USP Iohexol Related Compound B RS, and quantitatively dilute with water to obtain a solution having a known concentration of about 10 µg per mL.
Standard solution—
Transfer 10.0 mL of the Standard stock solution and 5 mL of water to a 25-mL volumetric flask.
Test solution—
Transfer about 200 mg of Iodixanol, accurately weighed, to a 25-mL volumetric flask, add 15 mL of water, and mix.
Procedure—
Treat the
Standard solution, the
Test solution, and the
Blank solution as follows. Place the flask in an ice bath for 5 minutes. Add 1.5 mL of 6 N hydrochloric acid, mix by swirling, add 1.0 mL of sodium nitrite solution (2 in 100), mix, and allow to stand in the ice bath for 4 minutes. Remove the flask from the ice bath, add 1.0 mL of 4% sulfamic acid solution, and swirl gently until gas evolution ceases. Add 1.0 mL of
N-(1-Naphthyl)ethylenediamine dihydrochloride solution, mix, dilute with water to volume, mix, and allow to stand for 5 minutes. Transfer the solution obtained from the
Test solution and the solution obtained from the
Standard solution to separate color-comparison tubes. The solution obtained from the
Test solution is lighter than the solution obtained from the
Standard solution: not more than 0.05% is found. If the solution obtained from the
Test solution is about the same color or darker than the solution obtained from the
Standard solution, proceed as follows. Concomitantly determine the absorbances of the solution obtained from the
Test solution, the solution obtained from the
Standard solution, and the solution obtained from the
Blank solution in 5-cm cells, at the wavelength of maximum absorbance at about 495 nm, using the solution obtained from the
Blank solution to zero the spectrophotometer. Calculate the percentage of free aromatic amine in the portion of Iodixanol taken by the formula:
(C/W)[(AU – AB)/(AS – AB)]
in which
C is the concentration, in µg per mL, of
USP Iohexol Related Compound B RS in the
Standard solution; W is the weight, in mg, of Iodixanol taken to prepare the
Test solution; and
AU,
AB, and
AS are the absorbances of the final solutions obtained from the
Test solution, the
Blank solution, and the
Standard solution, respectively: not more than 0.05% is found.
Change to read:
Limit of calcium—
Internal standard solution—
Accurately weigh about 3.067 g of scandium oxide, and dissolve in 1 L of water (each mL contains
2.0 mg
USP29 of scandium). Transfer 10.0 mL of this solution to a 100-mL volumetric flask, dilute with water to volume, and mix.
Blank solution—
Transfer 5.0 mL of Internal standard solution to a 50-mL volumetric flask, dilute with water to volume, and mix.
Standard solutions—
Prepare a solution having a concentration of 10 µg of calcium per mL. Add 0.5, 2.5, 5.0, and 10.0 mL of this solution to separate 50-mL volumetric flasks. To each flask, add 5.0 mL of Internal standard solution, dilute with water to volume, and mix.
Test solution—
Transfer about 2 g of Iodixanol, accurately weighed, to a 20-mL volumetric flask, add about 10 mL of water, and mix. Add 2.0 mL of Internal standard solution, dilute with water to volume, and mix.
Procedure—
Concomitantly determine the absorbances of each
Standard solution and the
Test solution at 393.366 nm, the calcium emission line, and at 361.38 nm, the scandium emission line, with an atomic absorption spectrophotometer, using the
Blank solution as the blank. Plot a standard curve of the ratio of the calcium absorption to the scandium absorption versus the respective calcium concentrations. From the graph so obtained, determine the calcium concentration,
C, in µg per mL, in the
Test solution. Calculate the content of calcium, in µg per g, in the portion of Iodixanol taken by the formula:
20(C/W)
in which
C is as obtained above; and
W is the weight, in g, of Iodixanol taken to prepare the
Test solution: not more than 5 µg per g is found.
Limit of ionic compounds—
[NOTE—Rinse all glassware with water.] Prepare a solution of Iodixanol in water (2 in 100). The specific conductance of this solution is not greater than that of a sodium chloride solution having a concentration of 4 µg per mL (equivalent to not more than 0.02% of ionic compounds, as NaCl).
Limit of methanol, isopropyl alcohol, and methoxyethanol—
Internal standard stock solution—
Transfer about 500 mg of secondary butyl alcohol to a 500-mL volumetric flask, dilute with water to volume, and mix.
Internal standard solution—
Transfer 1.0 mL of Internal standard stock solution to a 100-mL volumetric flask, dilute with water to volume, and mix.
Standard solution—
Transfer about 500 mg of methanol, accurately weighed, and about 1000 mg each of isopropyl alcohol and methoxyethanol, both accurately weighed, to a 500-mL volumetric flask, dilute with water to volume, and mix. Transfer 5.0 mL of this solution to a 100-mL volumetric flask, dilute with water to volume, and mix. Transfer 10.0 mL of this solution and 1.0 mL of Internal standard stock solution to a second 100-mL volumetric flask, dilute with water to volume, and mix. Transfer 1.0 mL to a headspace vial, and seal the vial with a septum and a crimp cap. This solution contains about 0.005 mg of methanol and about 0.01 mg each of isopropyl alcohol and methoxyethanol per mL.
Test solution—
Transfer about 250 mg of Iodixanol, accurately weighed, to a headspace vial. Add 1.0 mL of Internal standard solution, seal the vial with a septum and a crimp cap, and mix until dissolved.
Chromatographic system (see Chromatography 621)—
The gas chromatograph is equipped with a headspace injector and a 0.54-mm × 30-m capillary column coated with a 1-µm layer of phase G16. The carrier gas is helium, flowing at a rate of about 11 mL per minute. The column temperature is maintained at 40
for 3.0 minutes, then it is increased linearly at a rate of 8
per minute to 100
, and is maintained at 100
for 1 minute. The temperatures of the headspace injector and the detector are maintained at 150
and 200
, respectively. The
Standard solution and the
Test solution are maintained at about 105
, and the needle temperature and transfer temperature are maintained at about 130
and 140
, respectively. Chromatograph the
Standard solution, and record the peak responses as directed for
Procedure: the elution order is methanol, isopropyl alcohol, secondary butyl alcohol, and methoxyethanol; the resolution,
R, between methanol and isopropyl alcohol is not less than 1.0; and the relative standard deviation, determined from peak area ratios, for replicate injections is not more than 5% for methanol and isopropyl alcohol and not more than 10% for methoxyethanol.
Procedure—
Using a heated, gas-tight syringe, separately inject equal volumes (about 1 mL) of the headspace of the
Standard solution and the
Test solution into the chromatograph, record the chromatograms, and measure the peak areas. Calculate the percentage of methanol, isopropyl alcohol, and methoxyethanol in the portion of Iodixanol taken by the formula:
100(C/W)(RI / RS)
in which
C is the concentration, in mg per mL, of the relevant analyte in the
Standard solution; W is the weight, in mg, of Iodixanol taken to prepare the
Test solution; and
RI and
RS are the peak area ratios of the corresponding analyte to the internal standard obtained from the
Test solution and the
Standard solution, respectively: not more than 50 µg per g each of methanol, isopropyl alcohol, or methoxyethanol is found.
Related compounds—
TEST 1—
Solution A—
Prepare a solution containing 500 mL of acetonitrile and 500 mL of water, and degas.
Solution B—
Use 1000 mL of degassed water.
Mobile phase—
Use variable mixtures of
Solution A and
Solution B as directed for
Chromatographic system. Make adjustments if necessary (see
System Suitability under
Chromatography 621).
Blank solution—
Use water.
Standard stock solution 1—
Quantitatively dissolve an accurately weighed quantity of
USP Iodixanol RS in water to obtain a solution having a known concentration of about 12.5 mg of anhydrous iodixanol per mL.
Standard stock solution 2—
Quantitatively dissolve an accurately weighed quantity of
USP Iodixanol Related Compound C RS in water to obtain a solution having a known concentration of about 0.25 mg of anhydrous iodixanol related compound C per mL.
Standard stock solution 3—
Quantitatively dissolve an accurately weighed quantity of
USP Iodixanol Related Compound D RS in water to obtain a solution having a known concentration of about 0.025 mg of anhydrous iodixanol related compound D per mL.
Standard solution 1—
Dilute 2.0 mL of Standard stock solution 1 in water to 10.0 mL, and mix.
Standard solution 2—
Transfer 5.0 mL of Standard stock solution 1, 2.5 mL of Standard stock solution 2, and 2.5 mL of Standard stock solution 3 to a 25-mL volumetric flask, dilute with water to volume, and mix.
Test solution 1—
Dissolve an amount of Iodixanol, equivalent to about 500 mg of anhydrous iodixanol, in water, dilute with water to 20.0 mL, and mix.
Test solution 2—
Dilute 5.0 mL of Test solution 1 with water to 50.0 mL.
Control solution—
Dilute 5.0 mL of Test solution 1 and 2.5 mL of Standard stock solution 2 with water to 50 mL.
Chromatographic system (see Chromatography 621)—
The liquid chromatograph is equipped with a 254-nm detector and a 4.6-mm × 25-cm column that contains 5-µm 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 |
6 |
94 |
equilibration
(for about 20 minutes) |
| 0–30 |
6®20 |
94®80 |
linear gradient |
| 30–70 |
20®100 |
80®0 |
linear gradient |
| 70–80 |
100 |
0 |
isocratic |
| 80–81 |
100®6 |
0®94 |
linear gradient |
| 81–90 |
6 |
94 |
isocratic |
Chromatograph the solutions as directed for
Procedure, in the following injection sequence:
Blank solution, Standard solution 1, Control solution, and at least three replicates of
Standard solution 2.
The chromatogram obtained from Standard solution 1 exhibits two or three principal unresolved peaks. If the chromatogram exhibits two principal peaks, their relative areas are about 60% and 40%. If the chromatogram exhibits three principal peaks, their relative areas are about 60%, 38%, and 2%. The chromatogram obtained from Standard solution 2 exhibits two resolved peaks due to iodixanol related compound D that elute before the iodixanol peaks and one iodixanol related compound C peak between the two principal iodixanol peaks. The area of the two iodixanol related compound D peaks is between 0.075% and 0.125% of the total area. [NOTE—Disregard any peak due to the solvent front and any peak corresponding to those obtained from the Blank solution.] Add the areas of the two isomer peaks for iodixanol related compound D from each of the three injections of Standard solution 2, and calculate the relative standard deviation for the three summed areas: the relative standard deviation is not more than 5%. Measure the height of the iodixanol related compound C peak, and, if necessary, adjust the sensitivity of the amplifier to obtain a peak height between 80% and 100% of the full scale. Measure the height, A, above the baseline of the iodixanol related compound C peak and the height, B, above the baseline of the lowest part of the curve separating this peak from the first principal iodixanol peak: A is not less than 1.3B. In the chromatogram obtained from the Control solution, iodixanol related compound C exhibits a measurable peak.
Procedure—
Separately inject equal volumes (about 10 µL) of the
Blank solution, Test solution 1, and
Test solution 2.
HIGH-LOW CHROMATOGRAPHY—
Where it is specified to proceed as directed for
High-low chromatography, for the chromatogram obtained from
Test solution 1, calculate the percentage of each specified related compound in the portion of Iodixanol taken by the formula:
(10X)/(0.1Y + Z)
in which
X is the peak area for each of the specified related compounds obtained from
Test solution 1; Y is the total area of all the peaks eluted before and after iodixanol obtained from
Test solution 1, disregarding any peaks due to injection noise or solvent; and
Z is the sum of peak areas of iodixanol and any related compounds which are eluted together with, and between, the principal iodixanol peaks obtained from
Test solution 2.
IOHEXOL—
If iohexol is present, it exhibits two peaks, with retention times of about 0.37 and 0.39 relative to the main iodixanol peak, in the chromatogram obtained from Test solution 1. Draw a baseline at the height of the baseline obtained from the Blank solution. Calculate the total area of the two peaks and the percentage of iohexol in the portion of Iodixanol taken as directed for High-low chromatography.
IODIXANOL RELATED COMPOUND B1 —
If iodixanol related compound B is present, it elutes as a single peak with a retention time of about 0.34 relative to the main iodixanol peak, in the chromatogram obtained from
Test solution 1. Draw a baseline at the height of the baseline obtained from the
Blank solution. Calculate the area of the peak and the percentage of iodixanol related compound B in the portion of Iodixanol taken as directed for
High-low chromatography.
IODIXANOL RELATED COMPOUND C—
If iodixanol related compound C is present, only the first and larger peak, with a retention time of about 1.07 relative to the main iodixanol peak, is seen between the two principal iodixanol peaks in the chromatogram obtained from
Test solution 1; the second iodixanol related compound C peak co-elutes with iodixanol. The area of the first and larger peak corresponds to about 80% of the total area of iodixanol related compound C. Draw a vertical line through the minimum before the first and larger peak. Draw a horizontal baseline at the minimum after the first and larger peak. This encompasses the iodixanol related compound C peak area,
X2. Calculate the percentage of iodixanol related compound C in the portion of Iodixanol taken by the formula:
12.5X2 /(0.1Y + Z)
in which
Y and
Z are as defined for
High-low chromatography.
IODIXANOL RELATED COMPOUND F2 —
If iodixanol related compound F is present, only the first and smaller peak with a retention time of about 0.8 relative to the main iodixanol peak, can be seen in the chromatogram obtained from
Test solution 1; the second peak co-elutes with iodixanol. The area of the first and smaller peak corresponds to about 25% of the total area of iodixanol related compound F. Draw the baseline at the height of the baseline obtained from the
Blank solution. Calculate the percentage of iodixanol related compound F in the portion of Iodixanol taken by the formula:
40X1 /(0.1Y + Z)
in which
X1 is the actual observed area of the peak of iodixanol related compound F obtained from
Test solution 1; and
Y and
Z are as defined for
High-low chromatography.
IODIXANOL RELATED COMPOUND G3 —
If iodixanol related compound G is present, the second and larger peak, with a retention time of about 1.18 relative to the last iodixanol peak, is seen in the chromatogram obtained from
Test solution 1; the first peak co-elutes with iodixanol. The area of the second peak corresponds to about 85% of the total area of iodixanol related compound G. Draw the baseline at the height of the baseline obtained from the
Blank solution. Calculate the percentage of iodixanol related compound G in the portion of Iodixanol taken by the formula:
10X3 /[0.85(0.1Y + Z)]
in which
X3 is the peak area of iodixanol related compound G; and
Y and
Z are as defined for
High-low chromatography.
OVERALKYLATED RELATED COMPOUNDS—
These compounds elute after iodixanol related compound G, with a retention time greater than 1.18 relative to the last iodixanol peak. Draw the baseline at the height of the baseline obtained from the Blank solution, and determine the peak areas. Calculate the percentage of overalkylated related compounds as directed for High-low chromatography.
UNSPECIFIED RELATED COMPOUNDS—
Examine the chromatograms obtained from Test solution 1 and the area of each peak eluting before or after iodixanol, other than those of iodixanol, specified related compounds, specified impurities, and overalkylated related compounds. Draw the baseline at the height of the baseline obtained from the Blank solution. Calculate the percentage of the largest of these peaks as directed for High-low chromatography.
OTHER UNSPECIFIED RELATED COMPOUNDS—
Determine the area of any unspecified peak eluting between those of iodixanol. Draw the baseline between minima, and calculate the percentage as directed for
High-low chromatography.
Not more than 0.2% of iodixanol related compound B is found; not more than 0.4% of iodixanol related compound C is found; not more than 0.2% of iodixanol related compound F is found; not more than 0.2% of iodixanol related compound G is found; not more than 0.6% of iohexol is found; not more than 1.0% of overalkylated related compounds is found; not more than 0.2% of any individual unspecified related compound is found; not more than 0.5% of total unspecified related compounds is found; and not more than 1.5% of total related compounds is found.
TOTAL RELATED COMPOUNDS—
From each of the chromatograms obtained from
Test solution 1, calculate the percentage of all related compounds as the sum of the results for the peaks appearing between the two principal iodixanol peaks, and the area percent obtained by the formula:
[100(Y – X1 – X3 + X1/0.25 + X2/0.8 + X3/0.85)]/10(0.1Y + Z)
in which the variables are as defined above.
TEST 2—
Solution A—
Use 1000 mL of filtered and degassed acetonitrile.
Solution B and Mobile phase—
Proceed as directed for Test 1.
Blank solution—
Use water.
Standard stock solution 1—
Quantitatively dissolve an accurately weighed quantity of
USP Iodixanol RS in water to obtain a solution having a known concentration of about 12.5 mg of anhydrous iodixanol per mL.
Standard stock solution 2—
Quantitatively dissolve an accurately weighed quantity of
USP Iodixanol Related Compound D RS, equivalent to about 0.125 g of anhydrous iodixanol related compound D, in water, and dilute with water to 100.0 mL. Dilute 2.0 mL of this solution with water to 100.0 mL.
Standard stock solution 3—
Quantitatively dissolve an accurately weighed quantity of
USP Iodixanol Related Compound E RS in water to obtain a solution having a known concentration of about 2.5 mg per mL.
Standard solution 1—
Dilute 2.0 mL of Standard stock solution 1 with water to 10.0 mL.
Standard solution 2—
Transfer 5.0 mL of Standard stock solution 1 and 2.5 mL of Standard stock solution 2 to a 25-mL volumetric flask, dilute with water to volume, and mix.
Standard solution 3—
Transfer 1.0 mL of Standard solution 1 and 1.0 mL of Standard stock solution 3 to a 10-mL volumetric flask, dilute with water to volume, and mix.
Test solution—
Dissolve an accurately weighed quantity of Iodixanol, equivalent to about 125 mg of anhydrous iodixanol, in water, dilute with water to 50.0 mL, and mix.
Chromatographic system (see Chromatography 621)—
The liquid chromatograph is equipped with a 254-nm detector and a 4.6-mm × 25-cm column that contains 5-µm packing L8. The flow rate is about 2.5 mL per minute. The chromatograph is programmed as follows.
Time
(minutes) |
Solution A
(%) |
Solution B
(%) |
Elution |
| 0 |
85 |
15 |
equilibration |
| 0–25 |
85®66 |
15®34 |
linear gradient |
Chromatograph the solutions as directed for
Procedure. The chromatogram obtained from
Standard solution 1 exhibits three principal unresolved peaks: the relative areas are about 62%, 35%, and 3%; and the retention time of the last iodixanol peak is not more than 14 minutes.
The chromatogram obtained from Standard solution 2 exhibits two partially unresolved peaks due to iodixanol related compound D, with relative retention times of about 0.33 and 0.39, that elute before the iodixanol peaks: the peak area of iodixanol related compound D is between 0.075% and 0.125% of the total area. Disregard any peak due to the solvent.
Determine the sum of the peak areas of the two isomers of iodixanol related compound D for each of the three chromatograms obtained from Standard solution 2: the relative standard deviation for replicate injections is not more than 5%.
The chromatogram obtained from Standard solution 3 exhibits two unresolved peaks due to iodixanol related compound E, with relative retention times of about 0.67 and 0.72, that elute before the iodixanol peaks. Adjust the sensitivity of the amplifier so that the peak heights are between 90% and 100% of full scale of the highest peak: the resolution, R, between the first and largest iodixanol related compound E peak and the first principal iodixanol peak is not less than 5.0.
Procedure—
Separately inject equal volumes (about 10 µL) of
Standard solution 1, three times
Standard solution 2, Standard solution 3, and the
Test solution. For the first chromatogram obtained from
Standard solution 2, adjust the sensitivity of the amplifier to obtain a peak height of about 15% of the first and larger peak that corresponds to iodixanol related compound D. Use this sensitivity setting for the subsequent injections.
Compare the retention times of the peaks obtained from Standard solution 3 to those obtained from the Test solution. Iodixanol related compound E exhibits two peaks, the second of which may partly overlap with another peak; use only the area of the first and larger peak, which corresponds to about 60% of the total area of iodixanol related compound E. Draw a baseline at the height of the baseline obtained from the Blank solution. Calculate the percentage of iodixanol related compound E by dividing the area obtained from the Test solution by 0.6 and using area percent.
Iodixanol related compound H
4 appears as a single peak, with a shoulder, on the tail of the iodixanol peak. Calculate the percentage of iodixanol related compound H by using area percent. Not more than 0.3% of iodixanol related compound E is found; and not more than 0.6% of iodixanol related compound H is found.
Assay—
Transfer about 500 mg of Iodixanol, accurately weighed, to a glass-stoppered, 125-mL conical flask, add 25 mL of a sodium hydroxide solution (5 in 100) and 500 mg of powdered zinc, connect the flask to a reflux condenser, and reflux for 1 hour. Cool the flask to room temperature, and rinse the condenser with 20 mL of water, adding the rinsing to the refluxed solution. Filter the mixture, rinsing the flask and the filter with several small portions of water, and adding the filtered rinsings to the filtrate. Add 5 mL of glacial acetic acid, titrate with 0.1 N silver nitrate VS, and determine the endpoint potentiometrically. Each mL of 0.1 N silver nitrate is equivalent to 25.84 mg of C35H44I6N6O15.
1
5-(acetylamino)-N,N¢-bis(2,3-dihydroxypropyl)-2,4,6-triiodo-1,3-benzenedicarboxamide
2
2-[[acetyl[3,5-bis[[(2,3-dihydroxypropyl)amino]carbonyl]-2,4,6-triiodophenyl]amino]methyl]-N,N¢-bis(2,3-dihydroxypropyl)-2,3-dihydro-5,7-diiodo-4H-1,4-benzoxazine-6,8-dicarboxamide
3
4-acetyl-2-[[acetyl[3,5-bis[[(2,3-dihydroxypropyl)amino]carbonyl]-2,4,6-triiodophenyl]amino]methyl]-N,N¢-bis(2,3-dihydroxypropyl)-2,3-dihydro-5,7-diiodo-4H-1,4-benzoxazine-6,8-dicarboxamide
4
5-[[3-[[3-[[[3-[[3-[[3,5-bis-[[[2,3-dihydroxypropyl]amino]carbonyl]-2,4,6-triiodophenyl](acetylimino)]-2-hydroxypropyl](acetylimino)]-5-[[[2,3-dihydroxypropyl]amino]carbonyl]-2,4,6-triiodophenyl]carbonyl]amino]-2-hydroxypropyl]oxy]-2-hydroxypropyl](acetylimino)]-N,N¢-bis(2,3-dihydroxypropyl)-2,4,6-triiodo-1,3-benzendicarboxamide
;)
