Appendix XX C. Polyethylene

1. Polyethylene Without Additives for Containers for Parenteral Preparations and for Ophthalmic Preparations

(Ph. Eur. method 3.1.4)

DEFINITION

Polyethylene without additives is obtained by polymerisation of ethylene under high pressure in the presence of oxygen or free-radical-forming initiators as catalyst.

CHARACTERS

Appearance

Beads, granules, powder or, after transformation, translucent sheets of varying thickness or containers.

Solubility

Practically insoluble in water, soluble in hot aromatic hydrocarbons, practically insoluble in anhydrous ethanol, in hexane and in methanol.

It softens at temperatures beginning at 65 °C.

Relative density

0.910 to 0.937.

IDENTIFICATION

If necessary, cut the samples of the material to be examined into pieces of maximum dimension on a side of not greater than 1 cm.

A. Infrared absorption spectrophotometry (2.2.24).

Preparation To 0.25 g add 10 mL of toluene R and boil under a reflux condenser for about 15 min. Place a few drops of the solution on a sodium chloride slide or on a disc of potassium bromide R and evaporate the solvent in an oven at 80 °C.

Alternatively, the spectrum may be recorded directly on a cut piece of suitable size (sheets), granules or hot pressed films by attenuated total reflection (ATR).

Absorption maximaat some of the following wavenumbers (tolerance: ± 5 cm^-¹): at 2915 cm^-¹, 2848 cm^-¹, 1471 cm^-¹, 1465 cm^-¹, 729 cm^-¹ and 719 cm^-¹.

The spectrum obtained is identical to that obtained with the material selected for the type sample (see chapter 3.2.2. Plastic containers and closures for pharmaceutical use for the definition of ‘type sample’).

B. Additives (see Tests).

TESTS

If necessary, cut the samples of the material to be examined into pieces of maximum dimension on a side of not greater than 1 cm.

Solution S1

Place 25 g in a borosilicate-glass flask with a ground-glass neck. Add 500 mL of water R and heat under a reflux condenser for 5 h. Allow to cool and decant. Keep part of the solution for the test for appearance of solution. Filter the rest through a sintered glass filter (16) (2.1.2). Use within 4 h of preparation.

Solution S2

Place 2.0 g in a conical borosilicate-glass flask with a ground-glass neck. Add 80 mL of toluene R and boil under a reflux condenser with constant stirring for 90 min. Allow to cool to 60 °C and add, with contant stirring, 120 mL of methanol R. Filter the solution through a sintered-glass filter (16) (2.1.2). Rinse the flask and the filter with 25 mL of a mixture of 40 mL of toluene R and 60 mL of methanol R, add the rinsings to the filtrate and dilute to 250 mL with the same mixture of solvents. Prepare a blank solution.

Solution S3

Place 100 g in a conical borosilicate-glass flask with a ground-glass neck. Add 250 mL of 0.1 M hydrochloric acid and boil under a reflux condenser with constant stirring for 1 h. Allow to cool and decant the solution.

Appearance of solution

Solution S1 is clear (2.2.1) and colourless (2.2.2, Method II).

Acidity or alkalinity

To 100 mL of solution S1 add 0.15 mL of BRP indicator solution R. Not more than 1.5 mL of 0.01 M sodium hydroxide is required to change the colour of the indicator to blue. To 100 mL of solution S1 add 0.2 mL of methyl orange solution R. Not more than 1.0 mL of 0.01 M hydrochloric acid is required to reach the beginning of the colour change of the indicator from yellow to orange.

Absorbance (2.2.25)

Maximum 0.2, determined between wavelengths of 220 nm and 340 nm on solution S1.

Reducing substances

To 20 mL of solution S1 add 1 mL of dilute sulfuric acid R and 20 mL of 0.002 M potassium permanganate. Boil under a reflux condenser for 3 min and cool immediately. Add l g of potassium iodide R and titrate immediately with 0.01 M sodium thiosulfate, using 0.25 mL of starch solution R as indicator. Carry out a blank titration. The difference between the titration volumes is not more than 0.5 mL.

Additives

Thin-layer chromatography (2.2.27).

Test solution Evaporate 50 mL of solution S2 to dryness in vacuo at 45 °C. Dissolve the residue in 5 mL of methylene chloride R. Prepare a blank solution from the blank solution corresponding to solution S2.

Reference solution Dissolve 20 mg of plastic additive 15 CRS and 20 mg of plastic additive 08 CRS in methylene chloride R and dilute to 10 mL with the same solvent.

PlateTLC silica gel G plate R.

Mobile phase Ahexane R.

Mobile phase Bmethanol R,methylene chloride R (5:95 V/V).

Application 10 µL.

Development A Over a path of 13 cm with mobile phase A.

Drying A In air.

Development B Over a path of 10 cm with mobile phase B.

Drying B In air.

Detection Spray with a 40 g/L solution of phosphomolybdic acid R in ethanol (96 per cent) R and heat at 120 °C until the spots appear in the chromatogram obtained with the reference solution.

System suitability: reference solution:

— the chromatogram shows 2 separated spots.

Limit No spot appears in the chromatogram obtained with the test solution, except for a spot which may be at the solvent front from the first development and which corresponds to oligomers. Disregard any spots corresponding to those obtained in the chromatogram with the blank solution.

Extractable heavy metals (2.4.8)

Maximum 2.5 ppm.

Evaporate 50 mL of solution S3 to about 5 mL on a water-bath and dilute to 20 mL with water R. 12 mL of solution complies with test A. Prepare the reference solution using 2.5 mL of lead standard solution (10 ppm Pb) R.

Sulfated ash (2.4.14)

Maximum 0.02 per cent, determined on 5.0 g.

2. Polyethylene With Additives for Containers for Parenteral Preparations and for Opthalmic Preparations

(Ph. Eur. method 3.1.5)

DEFINITION

Polyethylene with additives is obtained by polymerisation of ethylene under pressure in the presence of a catalyst or by copolymerisation of ethylene with not more than 25 per cent of higher alkene homologues (C₃ to C₁₀).

PRODUCTION

Depending on the intended use of the polymers, they may contain additives to optimise their processing or their chemical, physical and mechanical properties. Unless otherwise justified and authorised, these additives are chosen from the following list, which specifies for each substance the maximum permitted content.

Polyethylene may contain at most 3 antioxidants, 1 or several lubricants or antiblocking agents as well as titanium dioxide as an opacifying agent when the material must provide protection from light.

— butylhydroxytoluene (plastic additive 07): maximum 0.125 per cent;

— pentaerythrityl tetrakis[3-(3,5-di-tert-butyl-4-hydroxyphenyl)propionate] (plastic additive 09): maximum 0.3 per cent;

— 1,3,5-tris(3,5-di-tert-butyl-4-hydroxybenzyl)-S-triazine2,4,6(1H,3H,5H)-trione (plastic additive 13): maximum 0.3 per cent;

— octadecyl 3-(3,5-di-tert-butyl-4-hydroxyphenyl)propionate (plastic additive 11): maximum 0.3 per cent;

— ethylene bis[3,3-bis[3-(1,1-dimethylethyl)-4-hydroxyphenyl]butanoate] (plastic additive 08): maximum 0.3 per cent;

— dioctadecyl disulfide (plastic additive 15): maximum 0.3 per cent;

— 4,4′,4″-(2,4,6-trimethylbenzene-1,3,5-triyltrismethylene)tris[2,6-bis(1,1-dimethylethyl)phenol] (plastic additive 10): maximum 0.3 per cent;

— 2,2′-bis(octadecyloxy)-5,5′-spirobi[1,3,2-dioxaphosphinane] (plastic additive 14): maximum 0.3 per cent;

— didodecyl 3,3′-thiodipropionate (plastic additive 16): maximum 0.3 per cent;

— dioctadecyl 3,3′-thiodipropionate (plastic additive 17): maximum 0.3 per cent;

— tris [2,4-bis(1,1-dimethylethyl)phenyl] phosphite (plastic additive 12): maximum 0.3 per cent.

The total of antioxidant additives listed above does not exceed 0.3 per cent.

— hydrotalcite: maximum 0.5 per cent;

— alkanamides: maximum 0.5 per cent;

— alkenamides: maximum 0.5 per cent;

— sodium silico-aluminate: maximum 0.5 per cent;

— silica (natural or synthetic, coated or uncoated): maximum 0.5 per cent;

— sodium benzoate: maximum 0.5 per cent;

— fatty acid esters or salts: maximum 0.5 per cent;

— trisodium phosphate: maximum 0.5 per cent;

— liquid paraffin: maximum 0.5 per cent;

— zinc oxide: maximum 0.5 per cent;

— magnesium oxide: maximum 0.2 per cent;

— calcium stearate or zinc stearate or a mixture of both: maximum 0.5 per cent;

— titanium dioxide only for materials for containers for ophthalmic use: maximum 4 per cent.

The supplier of the material must be able to demonstrate that the qualitative and quantitative composition of the type sample (see chapter 3.2.2. Plastic containers and closures for pharmaceutical use for the definition of ‘type sample’) is satisfactory for each production batch.

CHARACTERS

Appearance

Powder, beads, granules or, after transformation, translucent sheets of varying thicknesses or containers.

Solubility

Practically insoluble in water, soluble in hot aromatic hydrocarbons, practically insoluble in anhydrous ethanol, in hexane and in methanol.

It softens at temperatures between 70 °C and 140 °C.

Relative density: 0.890 to 0.965.

IDENTIFICATION

If necessary, cut the samples of the material to be examined into pieces of maximum dimension on a side of not greater than 1 cm.

A. Infrared absorption spectrophotometry (2.2.24).

Alternatively, the spectrum may be recorded directly on a cut piece of suitable size (sheets), granules or hot pressed films by attenuated total reflection (ATR).

Absorption maxima at some of the following wavenumbers (tolerance: ± 5 cm^-¹): at 2915 cm^-¹, 2848 cm^-¹, 1471 cm^-¹, 1465 cm^-¹, 729 cm^-¹ and 719 cm^-¹.

The spectrum obtained is identical to that obtained with the material selected for the type sample.

B. It complies with the supplementary tests corresponding to the additives present (see Tests).

C. (To be performed only on opacified material.) In a platinum crucible, mix about 20 mg with 1 g of potassium hydrogen sulfate R and heat until completely melted. Allow to cool and add 20 mL of dilute sulfuric acid R. Heat gently. Filter the resulting solution. To the filtrate add 1 mL of phosphoric acid R and 1 mL of strong hydrogen peroxide solution R. If the substance is opacified with titanium dioxide, an orange-yellow colour develops.

TESTS

If necessary, cut the samples of the material to be examined into pieces of maximum dimension on a side of not greater than 1 cm.

Solution S1

Place 25 g in a borosilicate-glass flask with a ground-glass neck. Add 500 mL of water R and boil under a reflux condenser for 5 h. Allow to cool and decant. Reserve a portion of the solution for the test for appearance of solution and filter the rest through a sintered-glass filter (16) (2.1.2). Use within 4 h of preparation.

Solution S2

Place 2.0 g in a conical borosilicate-glass flask with a ground-glass neck. Add 80 mL of toluene R and boil under a reflux condenser for 90 min with constant stirring. Allow to cool to 60 °C and add with continued stirring 120 mL of methanol R. Filter the solution through a sintered-glass filter (16) (2.1.2). Rinse the flask and the filter with 25 mL of a mixture of 40 mL of toluene R and 60 mL of methanol R, add the rinsings to the filtrate and dilute to 250 mL with the same mixture of solvents. Prepare a blank solution.

Solution S3

Place 100 g in a conical borosilicate-glass flask with a ground-glass neck. Add 250 mL of 0.1 M hydrochloric acid and boil under a reflux condenser for 1 h with constant stirring. Allow to cool and decant the solution.

Appearance of solution

Solution S1 is clear (2.2.1) and colourless (2.2.2, Method II).

Acidity or alkalinity

Absorbance (2.2.25)

Maximum 0.2, determined between wavelengths of 220 nm and 340 nm on solution S1.

Reducing substances

To 20 mL of solution S1 add 1 mL of dilute sulfuric acid R and 20 mL of 0.002 M potassium permanganate. Boil under a reflux condenser for 3 min and cool immediately. Add 1 g of potassium iodide R and titrate immediately with 0.01 M sodium thiosulfate, using 0.25 mL of starch solution R as indicator. Carry out a blank titration. The difference between the titration volumes is not more than 0.5 mL.

Extractable aluminium

Maximum 1 ppm.

Inductively coupled plasma-atomic emission spectrometry (2.2.57).

Test solution Use solution S3.

Reference solutions Prepare the reference solutions using aluminium standard solution (200 ppm Al) R, diluting with 0.1 M hydrochloric acid.

Wavelength Use the emission of aluminium at 396.15 nm, the spectral background being taken as 396.25 nm.

Verify the absence of aluminium in the hydrochloric acid used.

Extractable chromium

Maximum 0.05 ppm.

Inductively coupled plasma-atomic emission spectrometry (2.2.57).

Test solution Use solution S3.

Reference solutions Prepare the reference solutions using chromium standard solution (100 ppm Cr) R, diluting with a mixture of 2 volumes of hydrochloric acid R and 8 volumes of water R.

Wavelength Use the emission of chromium at 205.55 nm, the spectral background being taken as 205.50 nm.

Verify the absence of chromium in the hydrochloric acid used.

Extractable titanium

(not for materials opacified with titanium dioxide): maximum 1 ppm.

Inductively coupled plasma-atomic emission spectrometry (2.2.57).

Test solution Use solution S3.

Reference solutions Prepare the reference solutions using titanium standard solution (100 ppm Ti) R, diluting with 0.1 M hydrochloric acid.

Wavelength Use the emission of titanium at 336.12 nm, the spectral background being taken as 336.16 nm.

Verify the absence of titanium in the hydrochloric acid used.

Extractable vanadium

Maximum 0.1 ppm.

Inductively coupled plasma-atomic emission spectrometry (2.2.57).

Test solution Use solution S3.

Reference solutions Prepare the reference solutions using vanadium standard solution (1 g/L V) R, diluting with a mixture of 2 volumes of hydrochloric acid R and 8 volumes of water R.

Wavelength Use the emission of vanadium at 292.40 nm, the spectral background being taken as 292.35 nm.

Verify the absence of vanadium in the hydrochloric acid used.

Extractable zinc

Maximum 1 ppm.

Atomic absorption spectrometry (2.2.23, Method I).

Test solution Use solution S3.

Reference solutions Prepare the reference solutions using zinc standard solution (10 ppm Zn) R, diluting with 0.1 M hydrochloric acid.

Source Zinc hollow-cathode lamp.

Wavelength 213.9 nm.

Atomisation device Air-acetylene flame.

Verify the absence of zinc in the hydrochloric acid used.

Extractable zirconium

Maximum 0.1 ppm.

Inductively coupled plasma-atomic emission spectrometry (2.2.57).

Test solution Use solution S3.

Reference solutions Prepare the reference solutions using zirconium standard solution (1 g/L Zr) R, diluting with a mixture of 2 volumes of hydrochloric acid R and 8 volumes of water R.

Wavelength Use the emission of zirconium at 343.82 nm, the spectral background being taken as 343.92 nm.

Verify the absence of zirconium in the hydrochloric acid used.

Extractable heavy metals (2.4.8)

Maximum 2.5 ppm.

Evaporate 50 mL of solution S3 to about 5 mL on a water-bath and dilute to 20.0 mL with water R. 12 mL of the solution complies with test A. Prepare the reference solution using 2.5 mL of lead standard solution (10 ppm Pb) R.

Sulfated ash (2.4.14)

Maximum 1.0 per cent, determined on 5.0 g. This limit does not apply to material opacified with titanium dioxide.

SUPPLEMENTARY TESTS

These tests are to be carried out, in whole or in part, only if required by the stated composition of the material.

Phenolic antioxidants

Liquid chromatography (2.2.29).

Solvent mixtureacetonitrile R,tetrahydrofuran R (50:50 V/V).

Test solution S21 Evaporate 50 mL of solution S2 to dryness in vacuo at 45 °C. Dissolve the residue in 5.0 mL of the solvent mixture. Prepare a blank solution from the blank solution corresponding to solution S2.

Of the following reference solutions, only prepare those that are necessary for the analysis of the phenolic antioxidants stated in the composition of the substance to be examined.

Reference solution (a) Dissolve 25.0 mg of butylhydroxytoluene CRS (plastic additive 07) and 60.0 mg of plastic additive 08 CRS in 10.0 mL of the solvent mixture. Dilute 2.0 mL of this solution to 50.0 mL with the solvent mixture.

Reference solution (b) Dissolve 60.0 mg of plastic additive 09 CRS and 60.0 mg of plastic additive 10 CRS in 10.0 mL of the solvent mixture. Dilute 2.0 mL of this solution to 50.0 mL with the solvent mixture.

Reference solution (c) Dissolve 60.0 mg of plastic additive 11 CRS and 60.0 mg of plastic additive 12 CRS in 10.0 mL of the solvent mixture. Dilute 2.0 mL of this solution to 50.0 mL with the solvent mixture.

Reference solution (d) Dissolve 25.0 mg of butylhydroxytoluene CRS (plastic additive 07) in 10.0 mL of the solvent mixture. Dilute 2.0 mL of this solution to 50.0 mL with the solvent mixture.

Reference solution (e) Dissolve 60.0 mg of plastic additive 08 CRS in 10.0 mL of the solvent mixture. Dilute 2.0 mL of this solution to 50.0 mL with the solvent mixture.

Reference solution (f) Dissolve 60.0 mg of plastic additive 13 CRS in 10.0 mL of the solvent mixture. Dilute 2.0 mL of this solution to 50.0 mL with the solvent mixture.

Reference solution (g) Dissolve 60.0 mg of plastic additive 09 CRS in 10.0 mL of the solvent mixture. Dilute 2.0 mL of this solution to 50.0 mL with the solvent mixture.

Reference solution (h) Dissolve 60.0 mg of plastic additive 10 CRS in 10.0 mL of the solvent mixture. Dilute 2.0 mL of this solution to 50.0 mL with the solvent mixture.

Reference solution (i) Dissolve 60.0 mg of plastic additive 11 CRS in 10.0 mL of the solvent mixture. Dilute 2.0 mL of this solution to 50.0 mL with the solvent mixture.

Reference solution (j) Dissolve 60.0 mg of plastic additive 12 CRS in 10.0 mL of the solvent mixture. Dilute 2.0 mL of this solution to 50.0 mL with the solvent mixture.

A. If the substance to be examined contains plastic additive 07 and/or plastic additive 08, proceed as follows.

Column:

— size: l = 0.25 m, Ø = 4.6 mm;

— stationary phase: octadecylsilyl silica gel for chromatography R (5 µm).

Mobile phasewater R,acetonitrile R (30:70 V/V).

Flow rate 2 mL/min.

Detection Spectrophotometer at 280 nm.

Injection 20 µL of test solution S21, the corresponding blank solution, reference solution (a), and either reference solution (d) or (e), or reference solutions (d) and (e).

Run time 30 min.

System suitability:

— resolution: minimum 5.0 between the peaks due to plastic additive 07 and plastic additive 08 in the chromatogram obtained with reference solution (a).

The chromatogram obtained with test solution S21 only shows peaks due to antioxidants stated in the composition and minor peaks that also appear in the chromatogram obtained with the blank solution.

Calculation of percentage contents:

For plastic additive 07 and/or 08, use the concentration of the corresponding reference substance in reference solutions (d) and/or (e).

Limits:

— plastic additive 07: maximum 0.125 per cent;

— plastic additive 08: maximum 0.3 per cent.

B. If the substance to be examined contains one or more of the following antioxidants:

— plastic additive 09;

— plastic additive 10;

— plastic additive 11;

— plastic additive 12;

— plastic additive 13;

proceed as described above with the following modifications.

Mobile phasewater R,tetrahydrofuran R,acetonitrile R (10:30:60 V/V/V).

Flow rate 1.5 mL/min.

Injection 20 µL of test solution S21, the corresponding blank solution, reference solution (b), reference solution (c) and the reference solutions of the antioxidants in the list above that are stated in the composition.

System suitability:

— resolution: minimum 2.0 between the peaks due to plastic additive 09 and plastic additive 10 in the chromatogram obtained with reference solution (b); minimum 2.0 between the peaks due to plastic additive 11 and plastic additive 12 in the chromatogram obtained with reference solution (c).

The chromatogram obtained with test solution S21 only shows peaks due to antioxidants stated in the composition and minor peaks that also appear in the chromatogram obtained with the blank solution.

Calculation of percentage contents:

For plastic additive 09, 10, 11, 12 and/or 13, use the concentration of the corresponding reference substance in reference solutions (g), (h), (i), (j) and/or (f).

Limits:

— plastic additive 09: maximum 0.3 per cent;

— plastic additive 10: maximum 0.3 per cent;

— plastic additive 11: maximum 0.3 per cent;

— plastic additive 12: maximum 0.3 per cent;

— plastic additive 13: maximum 0.3 per cent.

Non-phenolic antioxidants

Thin-layer chromatography (2.2.27).

Test solution S22 Evaporate 100 mL of solution S2 to dryness in vacuo at 45 °C. Dissolve the residue in 2 mL of acidified methylene chloride R.

Reference solution (k) Dissolve 60 mg of plastic additive 14 CRS in methylene chloride R and dilute to 10 mL with the same solvent. Dilute 2 mL of this solution to 10 mL with acidified methylene chloride R.

Reference solution (l) Dissolve 60 mg of plastic additive 15 CRS in methylene chloride R and dilute to 10 mL with the same solvent. Dilute 2 mL of this solution to 10 mL with acidified methylene chloride R.

Reference solution (m) Dissolve 60 mg of plastic additive 16 CRS in methylene chloride R and dilute to 10 mL with the same solvent. Dilute 2 mL of this solution to 10 mL with acidified methylene chloride R.

Reference solution (n) Dissolve 60 mg of plastic additive 17 CRS in methylene chloride R and dilute to 10 mL with the same solvent. Dilute 2 mL of this solution to 10 mL with acidified methylene chloride R.

Reference solution (o) Dissolve 60 mg of plastic additive 16 CRS and 60 mg of plastic additive 17 CRS in methylene chloride R and dilute to 10 mL with the same solvent. Dilute 2 mL of this solution to 10 mL with acidified methylene chloride R.

PlateTLC silica gel F₂₅₄ plate R.

Mobile phase Ahexane R.

Mobile phase Bmethylene chloride R.

Application 20 µL of test solution S22, reference solution (o) and reference solutions corresponding to all the phenolic and non-phenolic antioxidants mentioned in the type composition of the material to be examined.

Development A Over a path of 18 cm with mobile phase A.

Drying A In air.

Development B Over a path of 17 cm with mobile phase B.

Drying B In air.

Detection Examine in ultraviolet light at 254 nm, spray with alcoholic iodine solution R and examine in ultraviolet light at 254 nm after 10-15 min.

System suitability Reference solution (o):

— the chromatogram shows 2 clearly separated spots.

Limits Any spots in the chromatogram obtained with test solution S22 are not more intense than the spots in the same positions in the chromatograms obtained with the reference solutions.

Amides and stearates

Thin-layer chromatography (2.2.27).

Test solution Use test solution S22 described in the test for non-phenolic antioxidants.

Reference solution (p) Dissolve 20 mg of stearic acid CRS (plastic additive 19) in methylene chloride R and dilute to 10 mL with the same solvent.

Reference solution (q) Dissolve 40 mg of plastic additive 20 CRS in methylene chloride R and dilute to 20 mL with the same solvent.

Reference solution (r) Dissolve 40 mg of plastic additive 21 CRS in methylene chloride R and dilute to 20 mL with the same solvent.

PlatesTLC silica gel F₂₅₄ plate R (2 plates).

A. Mobile phase: anhydrous ethanol R,trimethylpentane R (25:75 V/V).

Application 10 µL of test solution S22 and reference solution (p).

Development Over a path of 10 cm.

Drying In air.

Detection Spray with a 2 g/L solution of dichlorophenolindophenol, sodium salt R in anhydrous ethanol R and heat in an oven at 120 °C for a few minutes to intensify the spots.

Limit Any spot corresponding to plastic additive 19 in the chromatogram obtained with test solution S22 is similar in position (R_F = about 0.5) but not more intense than the corresponding spot in the chromatogram obtained with reference solution (p).

B. Mobile phase A: hexane R.

Mobile phase Bmethanol R,methylene chloride R (5:95 V/V).

Application 10 µL of test solution S22 and reference solutions (q) and (r).

Development A Over a path of 13 cm with mobile phase A.

Drying A In air.

Development B Over a path of 10 cm with mobile phase B.

Drying B In air.

Detection Spray with a 40 g/L solution of phosphomolybdic acid R in anhydrous ethanol R and heat in an oven at 120 °C until spots appear.

Limits Any spots corresponding to plastic additive 20 or plastic additive 21 in the chromatogram obtained with the test solution S22 are similar in position (R_F = about 0.2) but not more intense than the corresponding spots in the chromatograms obtained with reference solutions (q) and (r).