Changes from plenary report to adopted text

Added2.2.2. Neutral methylene blue solution

AddedDissolve 0,35 g methylene blue AR in deionised water and dilute to 1000 ml. Prepare the solution at least twenty-four hours before use. The absorbency of the blank chloroform phase, measured against chloroform must not exceed 0,015 per 1 cm of layer thickness at 650 nm.

Added2.2.3. Acidic methylene blue solution

AddedDissolve 0,35 g methylene blue AR in 500 ml deionised water and mix with 6,5 ml H2SO4 (d = 1,84 g/ml). Dilute to 1000 ml with deionised water. Prepare the solution at least twenty-four hours before use. The absorbency of the blank chloroform phase, measured against chloroform must not exceed 0,015 per 1 cm of layer thickness at 650 nm.

Added2.2.4. Chloroform (trichloromethane) AR freshly distilled

Added2.2.5. Dodecyl benzene sulphonic acid methyl ester

Added2.2.6. Ethanolic potassium hydroxide solution, KOH 0,1 M

Added2.2.7. Ethanol pure, C2H5OH

Added2.2.8. sulphuric acid, H2SO4 0,5 M

Added2.2.9. Phenolphthalein solution

AddedDissolve 1 g phenolphthalein in 50 ml ethanol and add 50 ml deionised water while stirring continuously. Filter off any precipitate obtained.

Added2.2.10. Methanolic hydrochloric acid: 250 ml hydrochloric acid AR and 750 ml methanol

Added2.2.11. Separating funnel, 250 ml

Added2.2.12. Graduated flask, 50 ml

Added2.2.13. Graduated flask, 500 ml

Added2.2.14. Graduated flask, 1000 ml

Added2.2.15. Round-bottomed flask with ground glass stopper and reflux condenser, 250 ml; boiling granules

Added2.2.16. pH meter

Added2.2.17. Photometer for measurements at 650 nm, with 1 to 5 cm cells

Added2.2.18. Qualitative grade filter paper

Added2.3. Procedure

AddedThe samples for analysis must not be taken through a layer of foam.

AddedAfter thorough cleaning with water, the equipment used for the analysis must be thoroughly rinsed with methanolic hydrochloric acid (point 2.2.10) and then with deionised water before using.

AddedFilter the activated sludge plant influent and effluent to be examined immediately on sampling. Discard the first 100 ml of the filtrates.

AddedPlace a measured volume of the sample, neutralised if necessary, into a 250 ml separating funnel (point 2.2.11). The volume of sample should contain between 20 and 150 g of MBAS. At the lower MBAS content, up to 100 ml of sample may be used. When using less than 100 ml, dilute to 100 ml with deionised water. Add to the sample 10 ml of buffer solution (point 2.2.1), 5 ml of neutral methylene blue solution (point 2.2.2) and 15 ml of chloroform (point 2.2.4). Shake the mixture uniformly and not too vigorously for one minute. After phase separation, run the chloroform layer into a second separating funnel, containing 110 ml of deionised water and 5 ml of acidic methylene blue solution (point 2.2.3). Shake the mixture for one minute. Pass the chloroform layer through a cotton-wool filter previously cleaned and wetted with chloroform into a graduated flask (point 2.2.12).

AddedExtract the alkaline and acid solutions three times, using 10 ml of chloroform for the second and third extractions. Filter the combined chloroform extracts through the same cotton wool filter and dilute to the mark in the 50 ml flask (point 2.2.12) with chloroform used for rewashing the cotton wool. Measure the absorbency of the chloroform solution with a photometer at 650 nm in 1 to 5 cm cells against chloroform. Run a blank determination through the whole procedure.

Added2.4. Calibration curve

AddedPrepare a calibration solution from the standard substance dodecylbenzene sulphonic acid methyl ester (tetrapropylene type mol. wt. 340) after saponification into the potassium salt. The MBAS is calculated as sodium dodecyl benzene sulphonate (mol. wt. 348).

AddedFrom a weighing pipette, weigh 400 to 450 mg of dodecyl-benzene-sulphonic-acid-methyl-ester (point 2.2.5) to the nearest 0,1 mg in a round-bottomed flask and add 50 ml of ethanolic potassium hydroxide solution (point 2.2.6) and some boiling granules. After mounting the reflux condenser, boil for one hour. After cooling, wash the condenser and ground glass joint with about 30 ml of ethanol, and add these washings to the contents of the flask. Titrate the solution with sulphuric acid against phenolphthalein until it becomes colourless. Transfer this solution to a 1000 ml graduated flask (point 2.2.14), dilute to the mark with deionised water and mix.

AddedPart of this surfactant stock solution is then further diluted. Withdraw 25 ml, transfer to a 500 ml graduated flask (point 2.2.13), dilute to the mark with deionised water and mix.

AddedThis standard solution contains:

Addedwhere E is the sample weight in mg.

AddedTo establish the calibration curve, withdraw 1, 2, 4, 6, 8 ml portions of the standard solution and dilute each to 100 ml with deionised water. Then proceed as stated under point 2.3 including a blank determination.

Added2.5. Calculation of results

AddedThe amount of anionic surfactant (MBAS) in the sample is read from the calibration curve (point 2.4). The MBAS content of the sample is given by:

Addedwhere: V = ml volume of the sample used.

AddedExpress the results as sodium dodecylbenzene sulphonate (MW 348).

Added2.6. Expression of results

AddedExpress the results as MBAS mg/l to the nearest 0,1.

Added3. Determination of non-ionic surfactants in biodegradation test liquors

Added3.1. Principle

AddedSurface active agents are concentrated and isolated by gas stripping. In the sample used, the quantity of non-ionic surfactant should be in the range 250-800 g.

AddedThe stripped surfactant is dissolved in ethyl acetate.

AddedAfter phase separation and evaporation of the solvent, the non-ionic surfactant is precipitated in aqueous solution with modified Dragendorff reagent (KBiI4 + BaCl2 + glacial acetic acid).

AddedThe precipitate is filtered, washed with glacial acetic acid and dissolved in ammonium tartrate solution. The bismuth in the solution is titrated potentiometrically with pyrrolidinedithiocarbamate solution at pH 4-5 using a bright platinum indicator electrode and a calomel or silver/silver chloride reference electrode. The method is applicable to non-ionic surfactants containing 6-30 alkylene oxide groups.

AddedThe titration result is multiplied by the empirical factor of 54 for conversion to the reference substance nonylphenol condensed with 10 mols ethylene oxide (NP 10).

Added3.2. Reagents and Equipment

AddedReagents are to be made up in deionised water.

Added3.2.1. Pure ethyl acetate, freshly distilled.

Added3.2.2. Sodium bicarbonate, NaHCO3 AR.

Added3.2.3. Dilute hydrochloric acid [20 ml concentrated acid (HCl) diluted to 1000 ml with water]

Added3.2.4. Methanol AR, freshly distilled, stored in a glass bottle.

Added3.2.5. Bromocresol purple, 0,1 g in 100 ml methanol.

Added3.2.6. Precipitating agent: the precipitating agent is a mixture of two volumes of solution A and one volume of solution B. The mixture is stored in a brown bottle and can be used for up to one week after mixing.

Added3.2.6.1. Solution A

AddedDissolve 1,7 g bismuth nitrate, BiONO3.H2O AR, in 20 ml glacial acetic acid, and make up to 100 ml with water. Then dissolve 65 g potassium iodide AR in 200 ml water. Mix these two solutions in a 1000 ml measuring flask, add 200 ml glacial acetic acid (point 3.2.7) and make up to 1000 ml with water.

Added3.2.6.2. Solution B

AddedDissolve 290 g barium chloride, BaCl2.2H2O AR, in 1000 ml of water.

Added3.2.7. Glacial acetic acid 99-100 % (lower concentrations are unsuitable).

Added3.2.8. Ammonium tartrate solution: mix 12,4 g tartaric acid AR and 12,4 ml of ammonia solution AR (d = 0,910 g/ml) and make up to 1000 ml with water (or use the equivalent amount of ammonium tartrate AR).