Hydrous Benzoyl Peroxide

(Ph. Eur. monograph 0704)

C14H10O4 242.2 94-36-0

Action and use

Used topically in the treatment of acne.

Preparations
Benzoyl Peroxide Cream
Benzoyl Peroxide Gel
Benzoyl Peroxide Lotion
Potassium Hydroxyquinoline Sulfate and Benzoyl Peroxide Cream

Ph Eur

DEFINITION
Content:
— dibenzoyl peroxide : 70.0 per cent to 77.0 per cent;

— water: minimum 20.0 per cent.

CHARACTERS

Appearance
White or almost white, amorphous or granular powder.

Solubility
Practically insoluble in water, soluble in acetone, soluble in methylene chloride with the separation of water, slightly
soluble in ethanol (96 per cent).
It loses water rapidly on exposure to air with a risk of explosion.
Mix the entire sample thoroughly before carrying out the following tests.

IDENTIFICATION
First identification B.
Second identification A, C, D.
A. Ultraviolet and visible absorption spectrophotometry (2.2.25).
Solution A Dissolve 80.0 mg in ethanol (96 per cent) R and dilute to 100.0 mL with the same solvent. Dilute 10.0 mL of
the solution to 100.0 mL with ethanol (96 per cent) R.
Solution B Dilute 10.0 mL of solution A to 100.0 mL with ethanol (96 per cent) R.
Spectral ranges 250-300 nm for solution A; 220-250 nm for solution B.
Absorption maxima At 274 nm for solution A; at 235 nm for solution B.
Shoulder At about 282 nm for solution A.
Absorbance ratio A235/A274 = 1.17 to 1.21.
B. Infrared absorption spectrophotometry (2.2.24).
Comparison Ph. Eur. reference spectrum of hydrous benzoyl peroxide.
C. Dissolve about 25 mg in 2 mL of acetone R. Add 1 mL of a 10 g/L solution of diethylphenylenediamine sulfate R and
mix. A red colour develops which quickly darkens and becomes dark violet within 5 min.
D. To 1 g add 5 mL of ethanol (96 per cent) R, 5 mL of dilute sodium hydroxide solution R and 10 mL of water R. Boil
the mixture under reflux for 20 min. Cool. The solution gives reaction (c) of benzoates (2.3.1).

TESTS

Acidity
Dissolve a quantity of the substance to be examined containing the equivalent of 1.0 g of dibenzoyl peroxide in 25 mL of
acetone R, add 75 mL of water R and filter. Wash the residue with two quantities, each of 10 mL, of water R. Combine
the filtrate and the washings and add 0.25 mL of phenolphthalein solution R1. Not more than 1.25 mL of 0.1 M sodium
hydroxide is required to change the colour of the indicator. Carry out a blank test.

Related substances
Liquid chromatography (2.2.29). Prepare the solutions immediately before use.
Test solution Dissolve a quantity of the substance to be examined containing the equivalent of 0.10 g of dibenzoyl
peroxide in acetonitrile R and dilute to 50 mL with the same solvent.
Reference solution (a) Dilute 1.0 mL of the test solution to 100.0 mL with acetonitrile R. Dilute 1.0 mL of this solution to
10.0 mL with acetonitrile R.
Reference solution (b) Dissolve 30.0 mg of benzoic acid R in the mobile phase and dilute to 100.0 mL with the mobile
phase. Dilute 1.0 mL of the solution to 10.0 mL with the mobile phase.
Reference solution (c) Dissolve 50.0 mg of ethyl benzoate R in the mobile phase and dilute to 100.0 mL with the mobile
phase. Dilute 1.0 mL of the solution to 100.0 mL with the mobile phase.
Reference solution (d) Dissolve 50.0 mg of benzaldehyde R in the mobile phase and dilute to 100.0 mL with the mobile
phase. Dilute 1.0 mL of the solution to 100.0 mL with the mobile phase.
Reference solution (e) Dissolve 30.0 mg of benzoic acid R and 30.0 mg of benzaldehyde R in the mobile phase and
dilute to 100.0 mL with the mobile phase. Dilute 1.0 mL of the solution to 10.0 mL with the mobile phase.
Column:
— size: l = 0.25 m, Ø = 4.6 mm;

— stationary phase: octadecylsilyl silica gel for chromatography R (10 µm).

Mobile phase glacial acetic acid R, acetonitrile R, water R (1:500:500 V/V/V).
Flow rate 1 mL/min.
Detection Spectrophotometer at 235 nm.
Injection 20 µL loop injector.
Run time 2 times the retention time of dibenzoyl peroxide.
Relative retention With reference to dibenzoyl peroxide (retention time = about 28.4 min): impurity B = about 0.15;
impurity A = about 0.2; impurity C = about 0.4.
System suitability Reference solution (e):

— resolution: minimum 6 between the peaks corresponding to benzoic acid and benzaldehyde.
Limits:
— impurity A: not more than the area of the principal peak in the chromatogram obtained with reference solution (d)
(0.25 per cent);

— impurity B: not more than the area of the principal peak in the chromatogram obtained with reference solution (b)
(1.5 per cent);

— impurity C: not more than the area of the principal peak in the chromatogram obtained with reference solution (c)
(0.25 per cent);
 — unspecified impurities: for each impurity, not more than the area of the principal peak in the chromatogram obtained
with reference solution (a) (0.10 per cent);

— disregard limit: 0.2 times the area of the principal peak in the chromatogram obtained with reference solution (a)
(0.02 per cent).

Chlorides (2.4.4)
Maximum 0.4 per cent.
Dissolve a quantity of the substance to be examined containing the equivalent of 0.5 g of dibenzoyl peroxide in 15 mL of
acetone R. Add, while stirring, 50 mL of 0.05 M nitric acid . Allow to stand for 10 min and filter. Wash the residue with 2
quantities, each of 10 mL, of 0.05 M nitric acid . Combine the filtrate and the washings and dilute to 100 mL with 0.05 M
nitric acid . Dilute 2.5 mL of the solution to 15.0 mL with water R.

ASSAY
Solution (a) Dissolve 2.500 g immediately before use in 75 mL of dimethylformamide R and dilute to 100.0 mL with the
same solvent.

Dibenzoyl peroxide
To 5.0 mL of solution (a) add 20 mL of acetone R and 3 mL of a 500 g/L solution of potassium iodide R and mix. Allow to
stand for 1 min. Titrate with 0.1 M sodium thiosulfate using 1 mL of starch solution R, added towards the end of the
titration, as indicator. Carry out a blank titration.
1 mL of 0.1 M sodium thiosulfate is equivalent to 12.11 mg of C14H10O4.

Water (2.5.12)
Carry out the semi-micro determination of water, using 5.0 mL of solution (a). Use as the solvent a mixture of 20.0 mL of
anhydrous methanol R and 3.0 mL of a 100 g/L solution of potassium iodide R in dimethylformamide R. After adding
solution (a), stir for 5 min before starting the titration. Carry out a blank determination.
Calculate the percentage content of water using the following expression:

n1 = number of millilitres of iodosulfurous reagent R used in the sample determination,

n2 =number of millilitres of iodosulfurous reagent R used in the blank determination,
w = water equivalent of iodosulfurous reagent R in milligrams of water per millilitre of reagent,
m = mass of the substance to be examined used for the preparation of solution (a) in grams,
p = percentage content of dibenzoyl peroxide.

STORAGE
In a container that has been treated to reduce static discharge and that has a device for release of excess pressure, at a
temperature of 2 °C to 8 °C, protected from light.

IMPURITIES

A. R = H: benzaldehyde,
B. R = OH: benzoic acid,
C. R = O-CH2-CH3: ethyl benzoate.
Ph Eur
 Benzoyl Peroxide Cream

DEFINITION
Benzoyl Peroxide Cream contains Hydrous Benzoyl Peroxide in a suitable basis.
The cream complies with the requirements stated under Topical Semi-solid Preparations and with the following
requirements.

Content of anhydrous benzoyl peroxide, C14H10O4

90.0 to 110.0% of the stated amount.

IDENTIFICATION
Carry out the method for thin-layer chromatography, Appendix III A, using the following solutions.
(1) Shake a quantity of the preparation being examined containing the equivalent of 50 mg of anhydrous benzoyl
peroxide with 10 mL of chloroform and filter.
(2) 0.5% w/v of benzoyl peroxide in chloroform.

CHROMATOGRAPHIC CONDITIONS

(a) Use as the coating silica gel F254.

(b) Use the mobile phase as described below.
(c) Apply 5 µL of each solution.
(d) Develop the plate to 15 cm.
(e) After removal of the plate, allow it to dry in air and examine under ultraviolet light (254 nm).

MOBILE PHASE

1 volume of glacial acetic acid , 2 volumes of dichloromethane and 50 volumes of toluene.

CONFIRMATION

The principal spot in the chromatogram obtained with solution (1) corresponds to that in the chromatogram obtained with
solution (2).

TESTS

Related substances
Carry out the method for liquid chromatography, Appendix III D, using the following solutions.
(1) Disperse a quantity of the preparation being examined containing the equivalent of 0.10 g of anhydrous benzoyl
peroxide in 25 mL of acetonitrile, add sufficient water to produce 50 mL, mix and filter.
(2) Dilute 1 volume of solution (1) to 100 volumes with the mobile phase.
(3) 0.020% w/v of benzoic acid in the mobile phase.
(4) 0.0020% w/v of ethyl benzoate in the mobile phase.
(5) 0.0020% w/v of benzaldehyde in the mobile phase.

CHROMATOGRAPHIC CONDITIONS

(a) Use a stainless steel column (25 cm × 4.6 mm) packed with octadecylsilyl silica gel for chromatography (10 µm)
(Spherisorb ODS 1 is suitable).
(b) Use isocratic elution and the mobile phase described below.
(c) Use a flow rate of 1 mL per minute.
(d) Use an ambient column temperature.
(e) Use a detection wavelength of 235 nm.
(f) Inject 20 µL of each solution.

MOBILE PHASE
1 volume of glacial acetic acid , 500 volumes of acetonitrile and 500 volumes of water.

LIMITS

In the chromatogram obtained with solution (1):

the areas of any peaks corresponding to benzoic acid, ethyl benzoate and benzaldehyde are not greater than the areas
of the principal peaks in the chromatograms obtained with solutions (3) (10%), (4) (1%) and (5) (1%) respectively;
the area of any other secondary peak is not greater than the area of the principal peak in the chromatogram obtained
with solution (2) (1%).

ASSAY
Mix a quantity of the preparation being examined containing the equivalent of 0.25 g of anhydrous benzoyl peroxide with
50 mL of acetone and add sufficient acetone to produce 100 mL. To 10 ml add 25 mL of a 20% w/v solution of potassium
iodide, mix, stopper the flask and allow to stand for 15 minutes protected from light. Add 25 ml of acetone and titrate with
0.01M sodium thiosulfate VS using starch mucilage, added towards the end of the titration, as indicator. Repeat the
operation without the substance being examined. The difference between the titrations represents the amount of sodium
thiosulfate required. Each mL of 0.01M sodium thiosulfate VS is equivalent to 1.211 mg of C14H10O4.

LABELLING
The quantity of active ingredient is stated in terms of the equivalent amount of anhydrous benzoyl peroxide.

Benzoyl Peroxide Gel

DEFINITION
Benzoyl Peroxide Gel is a solution of Hydrous Benzoyl Peroxide in a suitable water- soluble basis.
The gel complies with the requirements stated under Topical Semi-solid Preparations and with the following
requirements.

Content of anhydrous benzoyl peroxide, C14H10O4

90.0 to 110.0% of the stated amount.

IDENTIFICATION
Carry out the method for thin-layer chromatography, Appendix III A, using the following solutions.
(1) Shake a quantity of the preparation being examined containing the equivalent of 50 mg of anhydrous benzoyl
peroxide with 10 mL of chloroform and filter.
(2) 0.5% w/v of benzoyl peroxide in chloroform.

CHROMATOGRAPHIC CONDITIONS

(a) Use as the coating silica gel F254.

(b) Use the mobile phase as described below.
(c) Apply 5 µL of each solution.
(d) Develop the plate to 15 cm.
(e) After removal of the plate, allow it to dry in air and examine under ultraviolet light (254 nm).

MOBILE PHASE

1 volume of glacial acetic acid , 2 volumes of dichloromethane and 50 volumes of toluene.

CONFIRMATION

The principal spot in the chromatogram obtained with solution (1) corresponds to that in the chromatogram obtained with
solution (2).

TESTS
Related substances
Carry out the method for liquid chromatography, Appendix III D, using the following solutions.
(1) Disperse a quantity of the preparation being examined containing the equivalent of 0.10 g of anhydrous benzoyl
peroxide in 25 mL of acetonitrile, add sufficient water to produce 50 mL, mix and filter.
(2) Dilute 1 volumes of solution (1) to 100 volumes with the mobile phase.
(3) 0.020% w/v of benzoic acid in the mobile phase.
(4) 0.0020% w/v of ethyl benzoate in the mobile phase.
(5) 0.0020% w/v of benzaldehyde in the mobile phase.

CHROMATOGRAPHIC CONDITIONS

(a) Use a stainless steel column (25 cm × 4.6 mm) packed with octadecylsilyl silica gel for chromatography (10 µm)
(Spherisorb ODS 1 is suitable).
(b) Use isocratic elution and the mobile phase described below.
(c) Use a flow rate of 1 mL per minute.
(d) Use an ambient column temperature.
(e) Use a detection wavelength of 235 nm.
(f) Inject 20 µL of each solution.
MOBILE PHASE

1 volume of glacial acetic acid , 500 volumes of acetonitrile and 500 volumes of water.

LIMITS

In the chromatogram obtained with solution (1):

the areas of any peaks corresponding to benzoic acid, ethyl benzoate and benzaldehyde are not greater than the areas
of the principal peaks in the chromatograms obtained with solutions (3) (10%), (4) (1%) and (5) (1%) respectively;
the area of any other secondary peak is not greater than the area of the principal peak in the chromatogram obtained
with solution (2) (1%).

ASSAY
Mix a quantity of the preparation being examined containing the equivalent of 0.25 g of anhydrous benzoyl peroxide with
50 mL of acetone and add sufficient acetone to produce 100 mL. To 10 mL add 25 mL of a 20% w/v solution of
potassium iodide, mix, stopper the flask and allow to stand for 15 minutes protected from light. Add 25 mL of acetone
and titrate with 0.01M sodium thiosulfate VS using starch mucilage, added towards the end of the titration, as indicator.
Repeat the operation without the substance being examined. The difference between the titrations represents the
amount of sodium thiosulfate required. Each ml of 0.01 M sodium thiosulfate VS is equivalent to 1.211 mg of C14H10O4.

LABELLING
The quantity of active ingredient is stated in terms of the equivalent amount of anhydrous benzoyl peroxide.

Benzoyl Peroxide Lotion

Benzoyl Peroxide Cutaneous Suspension

DEFINITION
Benzoyl Peroxide Lotion is a cutaneous suspension. It contains Hydrous Benzoyl Peroxide in a suitable non-greasy
vehicle.
The lotion complies with the requirements stated under Liquids for Cutaneous Application and with the following
requirements.

Content of anhydrous benzoyl peroxide, C14H10O4

90.0 to 110.0% of the stated amount.
IDENTIFICATION
Carry out the method for thin-layer chromatography, Appendix III A, using the following solutions.
(1) Shake a quantity of the preparation being examined containing the equivalent of 50 mg of anhydrous benzoyl
peroxide with 10 mL of chloroform and filter.
(2) 0.5% w/v of benzoyl peroxide in chloroform.

CHROMATOGRAPHIC CONDITIONS

(a) Use as the coating silica gel F254.

(b) Use the mobile phase as described below.
(c) Apply 5 µL of each solution.
(d) Develop the plate to 15 cm.
(e) After removal of the plate, allow it to dry in air and examine under ultraviolet light (254 nm).

MOBILE PHASE

1 volume of glacial acetic acid , 2 volumes of dichloromethane and 50 volumes of toluene.

CONFIRMATION

The principal spot in the chromatogram obtained with solution (1) corresponds to that in the chromatogram obtained with
solution (2).

RELATED SUBSTANCES
Carry out the method for liquid chromatography, Appendix III D, using the following solutions.
(1) Disperse a quantity of the preparation being examined containing the equivalent of 0.10 g of anhydrous benzoyl
peroxide in 25 mL of acetonitrile, add sufficient water to produce 50 mL, mix and filter.
(2) Dilute 1 volume of solution (1) to 100 volumes with the mobile phase.
(3) 0.020% w/v of benzoic acid in the mobile phase.
(4) 0.0020% w/v of ethyl benzoate in the mobile phase.
(5) 0.0020% w/v of benzaldehyde in the mobile phase.

CHROMATOGRAPHIC CONDITIONS

(a) Use a stainless steel column (25 cm × 4.6 mm) packed with octadecylsilyl silica gel for chromatography (10 µm)
(Spherisorb ODS 1 is suitable).
(b) Use isocratic elution and the mobile phase described below.
(c) Use a flow rate of 1 mL per minute.
(d) Use an ambient column temperature.
(e) Use a detection wavelength of 235 nm.
(f) Inject 20 µL of each solution.

MOBILE PHASE

1 volume of glacial acetic acid , 500 volumes of acetonitrile and 500 volumes of water.

LIMITS

In the chromatogram obtained with solution (1):

the areas of any peaks corresponding to benzoic acid, ethyl benzoate and benzaldehyde are not greater than the areas
of the principal peaks in the chromatograms obtained with solutions (3) (10%), (4) (1%) and (5) (1%) respectively;
the area of any other secondary peak is not greater than the area of the principal peak in the chromatogram obtained
with solution (2) (1%).

ASSAY
Mix a quantity of the preparation being examined containing the equivalent of 0.25 g of anhydrous benzoyl peroxide with
50 mL of acetone and add sufficient acetone to produce 100 mL. To 10 mL add 25 mL of a 20% w/v solution of
potassium iodide, mix, stopper the flask and allow to stand for 15 minutes protected from light. Add 25 mL of acetone
and titrate with 0.01M sodium thiosulfate VS using starch mucilage, added towards the end of the titration, as indicator.
Repeat the operation without the substance being examined. The difference between the titrations represents the
amount of sodium thiosulfate required. Each mL of 0.01M sodium thiosulfate VS is equivalent to 1.211 mg of C14H10O4.

LABELLING
The quantity of active ingredient is stated in terms of the equivalent amount of anhydrous benzoyl peroxide.