Iohexol EUROPEAN PHARMACOPOEIA 10.

F. 2-[[acetyl[3,5-bis[(2,3-dihydroxypropyl)carbamoyl]- I. overalkylated impurities (an example) : 5-[acetyl[3-

2,4,6-triiodophenyl]amino]methyl]-N,N′-bis(2,3- [acetyl[3,5-bis[(2,3-dihydroxypropyl)carbamoyl]-2,4,6-tri-
dihydroxypropyl)-5,7-diiodo-3,4-dihydro-2H-1,4- iodophenyl]amino]-2-hydroxypropyl]amino]-N-
benzoxazine-6,8-dicarboxamide, [3-(2,3-dihydroxypropoxy)-2-hydroxypropyl]-N′-
(2,3-dihydroxypropyl)-2,4,6-triiodobenzene-1,3-
dicarboxamide.

01/2017:1114
corrected 10.0

IOHEXOL
Iohexolum

G. 4-acetyl-2-[[acetyl[3,5-bis[(2,3-dihydroxypropyl)-
carbamoyl]-2,4,6-triiodophenyl]amino]methyl]-N,N′-
bis(2,3-dihydroxypropyl)-5,7-diiodo-3,4-dihydro-2H-1,4-
benzoxazine-6,8-dicarboxamide, C19H26I3N3O9 Mr 821
[66108-95-0]
DEFINITION
5-[Acetyl(2,3-dihydroxypropyl)amino]-N,N′-bis(2,3-
dihydroxypropyl)-2,4,6-triiodobenzene-1,3-dicarboxamide.
The substance is a mixture of diastereoisomers and
atropisomers.
Content : 98.0 per cent to 101.0 per cent (anhydrous substance).
CHARACTERS
Appearance : white or greyish-white, hygroscopic powder.
Solubility : very soluble in water, freely soluble in methanol,
practically insoluble in methylene chloride.
IDENTIFICATION
A. Infrared absorption spectrophotometry (2.2.24).
Comparison : iohexol CRS.
B. Examine the chromatograms obtained in test A for related
substances.
Results : the principal peaks in the chromatogram obtained
with reference solution (b) are similar in retention time
and size to the peaks due to iohexol in the chromatogram
obtained with reference solution (a).
TESTS
H. 5-[acetyl[3-[acetyl[3-[[3-[3-[acetyl[3,5-bis[(2,3-
dihydroxypropyl)carbamoyl]-2,4,6-triiodophenyl]amino]- Solution S. Dissolve 5.0 g in water R and dilute to 50.0 mL
2-hydroxypropoxy]-2-hydroxypropyl]carbamoyl]-5-[(2,3- with the same solvent.
dihydroxypropyl)carbamoyl]-2,4,6-triiodophenyl]amino]- Appearance of solution. Solution S is clear (2.2.1) and not
2-hydroxypropyl]amino]-N,N′-bis(2,3-dihydroxypropyl)- more intensely coloured than reference solution Y7 (2.2.2,
2,4,6-triiodobenzene-1,3-dicarboxamide. Method II).

2974 See the information section on general monographs (cover pages)

EUROPEAN PHARMACOPOEIA 10.0 Iohexol

Related substances B. Thin-layer chromatography (2.2.27).

A. Liquid chromatography (2.2.29). Test solution. Dissolve 1.0 g of the substance to be examined
NOTE : iohexol gives rise to 2 non-resolved peaks in the in water R and dilute to 10.0 mL with the same solvent.
chromatogram due to endo-exo isomerism. In addition, Reference solution (a). Dissolve 50 mg of iohexol
a small peak (also due to iohexol) usually appears at the impurity J CRS and 50 mg of iohexol CRS in water R and
leading edge of the 1st principal peak. This small peak has a dilute to 10.0 mL with the same solvent.
retention time about 1.2 min less than the 1st principal peak. Reference solution (b). Dilute 1.0 mL of the test solution
Test solution. Dissolve 0.150 g of the substance to be to 10.0 mL with water R. Dilute 1.0 mL of this solution to
examined in water R and dilute to 100.0 mL with the same 50.0 mL with water R.
solvent. Plate : TLC silica gel F254 plate R.
Reference solution (a). Dissolve 15.0 mg of iohexol CRS Pretreatment : wash the plate with the mobile phase, dry at
and 15.0 mg of iohexol impurity A CRS in a mixture of room temperature for 30 min, then at 90 °C for 1 h.
0.05-0.1 mL of dilute sodium hydroxide solution R and
Mobile phase : concentrated ammonia R, methanol R,
10 mL of water R and dilute to 100.0 mL with water R.
2-propanol R, acetone R (16:16:28:40 V/V/V/V).
Dilute 1.0 mL of this solution to 10.0 mL with water R.
Application : 10 μL.
Reference solution (b). Dilute 1.0 mL of the test solution
to 100.0 mL with water R. Development : over 1/2 of the plate.
Reference solution (c). Dissolve 5.0 mg of iohexol for peak Drying : in air.
identification CRS (containing impurities B, C, D and E) in Detection : examine in ultraviolet light at 254 nm.
water R and dilute to 5.0 mL with the same solvent. System suitability : reference solution (a) :
Blank solution : water R. – the chromatogram shows 2 clearly separated spots.
Column : Limits :
– size : l = 0.25 m, Ø = 4.6 mm ; – any impurity : any spot in the chromatogram obtained
– stationary phase : octadecylsilyl silica gel for with the test solution, apart from the principal spot, is
chromatography R (5 μm). not more intense than the spot in the chromatogram
Mobile phase : obtained with reference solution (b) (0.2 per cent).
– mobile phase A : water R ; The thresholds indicated under Related substances
(Table 2034.-1) in the general monograph Substances for
– mobile phase B : acetonitrile R ; pharmaceutical use (2034) do not apply.
Time Mobile phase A Mobile phase B 3-Chloropropane-1,2-diol. Gas chromatography (2.2.28).
(min) (per cent V/V) (per cent V/V)
Test solution. Dissolve 1.0 g of the substance to be examined
0 - 60 99 → 87 1 → 13 in 1.0 mL of water R. Shake with 4 quantities, each of 2 mL,
of methyl acetate R. Dry the combined upper layers over
Flow rate : 1 mL/min. anhydrous sodium sulfate R. Filter and concentrate to about
Detection : spectrophotometer at 254 nm. 0.7 mL using a warm water-bath at 60 °C and a stream of
Injection : 10 μL. nitrogen and dilute to 1.0 mL with methyl acetate R.
Retention time : impurities A and H = about Reference solution. Dissolve 0.25 g of 3-chloropropane-1,2-
17 min ; iohexol (peaks corresponding to endo-exo diol R in 100.0 mL of methyl acetate R. Dilute 1.0 mL of this
isomerism) = about 20 min. solution to 100.0 mL with methyl acetate R.
System suitability : reference solution (a) : Column :
– resolution : minimum 5.0 between the peak due to – material : fused silica ;
impurity A and the 2nd and greater peak due to iohexol. – size : l = 25 m, Ø = 0.33 mm ;
Limits : – stationary phase : phenyl(50)methyl(50)polysiloxane R (film
– sum of impurities B, C, D and E (relative retention with thickness 1 μm).
reference to the 2nd and greater peak due to iohexol Carrier gas : helium for chromatography R.
between 1.1 and 1.4) : not more than 0.6 times the Flow rate : 1 mL/min.
total area of the principal peaks in the chromatogram
obtained with reference solution (b) (0.6 per cent) ; use Temperature :
the chromatogram obtained with reference solution (c) Time Temperature
to identify the corresponding peaks ; (min) (°C)
– sum of impurities A and H : not more than 0.5 times the Column 0-2 80
total area of the principal peaks in the chromatogram 2-8 80 → 170
obtained with reference solution (b) (0.5 per cent) ;
8 - 10 170
– unspecified impurities : for each impurity, not more than
0.1 times the total area of the principal peaks in the Injection port 230
chromatogram obtained with reference solution (b)
Detector 250
(0.10 per cent) ;
– total : not more than 1.5 times the total area of the Detection : flame ionisation.
principal peaks in the chromatogram obtained with Injection : 2 μL (splitless for 30 s).
reference solution (b) (1.5 per cent) ;
System suitability : reference solution :
– disregard limit : 0.03 times the total area of the principal
peaks in the chromatogram obtained with reference – retention time : 3-chloropropane-1,2-diol = about 8 min.
solution (b) (0.03 per cent). Limit :
The thresholds indicated under Related substances – 3-chloropropane-1,2-diol : not more than the area of the
(Table 2034.-1) in the general monograph Substances for principal peak in the chromatogram obtained with the
pharmaceutical use (2034) do not apply. reference solution (25 ppm).

General Notices (1) apply to all monographs and other texts 2975
Iohexol EUROPEAN PHARMACOPOEIA 10.0

Free aromatic amine : maximum 500 ppm. 1 mL of 0.1 M silver nitrate is equivalent to 27.37 mg
of C19H26I3N3O9.
Test solution. Transfer 0.200 g of the substance to be examined
to a 25 mL volumetric flask and dissolve in 15.0 mL of water R.
STORAGE
Reference solution. Dissolve 5.0 mg of iohexol impurity J CRS In an airtight container, protected from light and moisture.
in water R and dilute to 5.0 mL with water R. Dilute 1.0 mL
of the solution to 100.0 mL with water R. Mix 10.0 mL of this
solution with 5.0 mL of water R in a 25 mL volumetric flask. IMPURITIES
Specified impurities : A, B, C, D, E, H.
Blank solution. Transfer 15.0 mL of water R to a 25 mL
volumetric flask. Other detectable impurities (the following substances would,
if present at a sufficient level, be detected by one or other of
In conducting the following steps, keep the flasks in iced water the tests in the monograph. They are limited by the general
and protected as much as possible from light until all of the acceptance criterion for other/unspecified impurities. It
reagents have been added. is therefore not necessary to identify these impurities for
Place the 3 flasks containing respectively the test solution, demonstration of compliance. See also 5.10. Control of
the reference solution and the blank solution in iced water, impurities in substances for pharmaceutical use) : F, G, I, J, K,
protected from light, for 5 min. Add 1.5 mL of hydrochloric L, M, N, O, P, Q.
acid R1 and mix by swirling. Add 1.0 mL of a 20 g/L solution
of sodium nitrite R, mix and allow to stand for 4 min. Add
1.0 mL of a 40 g/L solution of sulfamic acid R, swirl gently
until gas liberation has ceased and allow to stand for 1 min.
(CAUTION : considerable pressure is produced). Add 1.0 mL of
a freshly prepared 3 g/L solution of naphthylethylenediamine
dihydrochloride R in a mixture of 30 volumes of water R and
70 volumes of propylene glycol R and mix. Remove the flasks
from the iced water, dilute to 25.0 mL with water R, mix
and allow to stand for 5 min. Simultaneously determine the
absorbance (2.2.25) at 495 nm of the solutions obtained from A. 5-(acetylamino)-N,N′-bis(2,3-dihydroxypropyl)-2,4,6-
the test solution and the reference solution in 5 cm cells, using triiodobenzene-1,3-dicarboxamide,
the blank as the compensation liquid. The absorbance of the
test solution is not greater than that of the reference solution.
Iodide : maximum 10 ppm.
Dissolve 6.000 g in water R and dilute to 20 mL with the
same solvent. Add 2.0 mL of 0.001 M potassium iodide.
Titrate with 0.001 M silver nitrate. Determine the end-point
potentiometrically (2.2.20), using a silver indicator electrode
and an appropriate reference electrode. Subtract the volume
of titrant corresponding to the 2.0 mL of 0.001 M potassium
iodide, determined by titrating a blank to which is added
2.0 mL of 0.001 M potassium iodide and use the residual value
to calculate the iodide content.
B. 5-[acetyl[3-(2,3-dihydroxypropoxy)-2-hydroxypropyl]-
1 mL of 0.001 M silver nitrate is equivalent to 126.9 μg of I−.
amino]-N,N′-bis(2,3-dihydroxypropyl)-2,4,6-
Ionic compounds (2.2.38) : maximum 0.01 per cent m/m triiodobenzene-1,3-dicarboxamide,
calculated as sodium chloride.
Rinse all glassware with distilled water R 5 times before use.
Test solution. Dissolve 1.0 g of the substance to be examined
in water R and dilute to 50.0 mL with the same solvent.
Reference solution. Dissolve 20.0 mg of sodium chloride R in
water R and dilute to 100.0 mL with the same solvent. Dilute
1.0 mL of this solution to 100.0 mL with water R.
Measure the conductivity of the test solution and the reference
solution using a suitable conductivity meter. The conductivity C. 5-[acetyl[2-(2,3-dihydroxypropoxy)-3-hydroxypropyl]-
of the test solution is not greater than that of the reference amino]-N,N′-bis(2,3-dihydroxypropyl)-2,4,6-
solution. triiodobenzene-1,3-dicarboxamide,
Water (2.5.12) : maximum 4.0 per cent, determined on 1.00 g.

ASSAY
To 0.500 g in a 125 mL round-bottomed flask add 25 mL of a
50 g/L solution of sodium hydroxide R, 0.5 g of zinc powder R
and a few glass beads. Boil under a reflux condenser for
30 min. Allow to cool and rinse the condenser with 20 mL
of water R, adding the rinsings to the flask. Filter through a
sintered-glass filter (2.1.2) and wash the filter with several
quantities of water R. Collect the filtrate and washings. Add D. 5-[acetyl(2,3-dihydroxypropyl)amino]-N-[3-
5 mL of glacial acetic acid R and titrate immediately with 0.1 M (2,3-dihydroxypropoxy)-2-hydroxypropyl]-N′-
silver nitrate. Determine the end-point potentiometrically (2,3-dihydroxypropyl)-2,4,6-triiodobenzene-1,3-
(2.2.20). dicarboxamide,

2976 See the information section on general monographs (cover pages)

EUROPEAN PHARMACOPOEIA 10.0 Iohexol

K. 5-amino-2,4,6-triiodobenzene-1,3-dicarboxylic acid,

E. 5-[acetyl(2,3-dihydroxypropyl)amino]-N-[2-
(2,3-dihydroxypropoxy)-3-hydroxypropyl]-N′-
(2,3-dihydroxypropyl)-2,4,6-triiodobenzene-1,3-
dicarboxamide,

L. 3,5-bis(chlorocarbonyl)-2,4,6-triiodobenzenamine,

F. 5-amino-N,N′-bis(2,3-dihydroxypropyl)diiodobenzene-
1,3-dicarboxamide,

M. N,N′-bis(2,3-dihydroxypropyl)-5-[(2,3-dihydroxy-
propyl)amino]diiodobenzene-1,3-dicarboxamide,

G. 5-(acetylamino)-N,N′-bis(2,3-dihydroxypropyl)diiodo-
benzene-1,3-dicarboxamide,

N. 5-[acetyl(2,3-dihydroxypropyl)amino]-N-[2-(acetyloxy)-
3-hydroxypropyl]-N′-(2,3-dihydroxypropyl)-2,4,6-
triiodobenzene-1,3-dicarboxamide,

H. 5-[acetyl(2,3-dihydroxypropyl)amino]-N,N′-bis(2,3-
dihydroxypropyl)diiodobenzene-1,3-dicarboxamide,

O. 5-[acetyl(2,3-dihydroxypropyl)amino]-N-[3-(acetyloxy)-
2-hydroxypropyl]-N′-(2,3-dihydroxypropyl)-2,4,6-
I. N,N′-bis(2,3-dihydroxypropyl)-2-(hydroxymethyl)- triiodobenzene-1,3-dicarboxamide,
5,7-diiodo-3,4-dihydro-2H-1,4-benzoxazine-6,8-
dicarboxamide,

P. 5-(diacetylamino)-N-[3-(2,3-dihydroxypropoxy)-
J. 5-amino-N,N′-bis(2,3-dihydroxypropyl)-2,4,6- 2-hydroxypropyl]-N′-(2,3-dihydroxypropyl)-2,4,6-
triiodobenzene-1,3-dicarboxamide, triiodobenzene-1,3-dicarboxamide,

General Notices (1) apply to all monographs and other texts 2977
Iopamidol EUROPEAN PHARMACOPOEIA 10.0

Reference solution (a). Dissolve 5.0 mg of iopamidol

impurity H CRS in water R and dilute to 100.0 mL with the
same solvent.
Reference solution (b). Dilute 2.0 mL of the test solution
to 20.0 mL with water R. Dilute 1.0 mL of this solution to
50.0 mL with water R.
Reference solution (c). Add 0.1 mL of the test solution to 20 mL
of reference solution (a) and dilute to 50 mL with water R.
Column : 2 columns coupled in series,
Q. 5-(diacetylamino)-N-[2-(2,3-dihydroxypropoxy)- – size : l = 0.25 m, Ø = 4.6 mm,
3-hydroxypropyl]-N′-(2,3-dihydroxypropyl)-2,4,6- – stationary phase : phenylsilyl silica gel for chromatography R
triiodobenzene-1,3-dicarboxamide. (5 μm),
– temperature : 60 °C.
01/2017:1115 Mobile phase :
corrected 10.0 – mobile phase A : water R,
– mobile phase B : acetonitrile R, water R (50:50 V/V),
Time Mobile phase A Mobile phase B
(min) (per cent V/V) (per cent V/V)
IOPAMIDOL 0 - 18 100 0

18 - 40 100 - 62 0 - 38
Iopamidolum
40 - 45 62 - 50 38 - 50

45 - 50 50 - 100 50 - 0

50 - 60 100 0

Flow rate : 2.0 mL/min.

Detection : spectrophotometer at 240 nm.
Injection : 20 μL.
Relative retention with reference to iopamidol (retention
C17H22I3N3O8 Mr 777 time = about 14.6 min) : impurity D = about 0.1 ;
[60166-93-0] impurity B = about 0.6 ; impurities I and H = about 0.9 ;
impurity G = about 1.1 ; impurity K = about 1.2 ;
DEFINITION impurity C = about 1.3 ; impurity J = about 1.5 ;
N,N′-Bis[2-hydroxy-1-(hydroxymethyl)ethyl]-5-[[(2S)- impurity A = about 1.8 ; impurity E = about 2.2 ;
2-hydroxypropanoyl]amino]-2,4,6-triiodobenzene-1,3- impurity F = about 2.3.
dicarboxamide. System suitability : reference solution (c) :
Content : 98.5 per cent to 101.0 per cent (dried substance). – resolution : minimum 2.0 between the peaks due to
CHARACTERS impurity H and iopamidol.
Appearance : white or almost white powder. Limits :
Solubility : freely soluble in water, very slightly soluble in – sum of impurities H and I : not more than the area of
methanol, practically insoluble in ethanol (96 per cent) and the principal peak in the chromatogram obtained with
in methylene chloride. reference solution (a) (0.5 per cent),
– impurities A, B, C, D, E, F, G, J, K : for each impurity, not
IDENTIFICATION more than 0.5 times the area of the principal peak in the
A. Infrared absorption spectrophotometry (2.2.24). chromatogram obtained with reference solution (b) (0.1 per
Comparison : iopamidol CRS. cent),
B. Loss on drying (see Tests). – any other impurity : for each impurity, not more than
C. Specific optical rotation (see Tests). 0.5 times the area of the principal peak in the chromatogram
obtained with reference solution (b) (0.1 per cent),
TESTS – sum of impurities other than H and I : not more than the
Appearance of solution. The solution is clear (2.2.1) and area of the principal peak in the chromatogram obtained
colourless (2.2.2, Method II). with reference solution (b) (0.2 per cent),
Dissolve 1 g in water R and dilute to 50 mL with the same – disregard limit : 0.05 times the area of the principal peak
solvent. in the chromatogram obtained with reference solution (b)
Acidity or alkalinity. Dissolve 10.0 g in carbon dioxide-free (0.01 per cent).
water R and dilute to 100 mL with the same solvent. Not more Free aromatic amines : maximum 200 ppm.
than 0.75 mL of 0.01 M hydrochloric acid or 1.4 mL of 0.01 M Keep the solutions and reagents in iced water, protected from
sodium hydroxide is required to adjust to pH 7.0 (2.2.3). bright light.
Specific optical rotation (2.2.7) : − 4.6 to − 5.2 (dried Test solution. In a 25 mL volumetric flask, dissolve 0.500 g of
substance), determined at 436 nm. the substance to be examined in 20.0 mL of water R.
Dissolve 10.0 g, with heating if necessary, in water R and dilute Reference solution. In a 25 mL volumetric flask, mix 4.0 mL
to 25.0 mL with the same solvent. of a 25.0 mg/L solution of iopamidol impurity A CRS with
Related substances. Liquid chromatography (2.2.29). 16.0 mL of water R.
Test solution. Dissolve 0.50 g of the substance to be examined Blank solution. Place 20.0 mL of water R in a 25 mL volumetric
in water R and dilute to 50.0 mL with the same solvent. flask.

2978 See the information section on general monographs (cover pages)