⚗️ Anionic surfactants in water and wastewater — the methylene blue index (MBAS): extraction with chloroform and measurement of the absorbance at 650 nm

Physicochemistry PN-EN 903

Overview

PN-EN 903:2002 (EN 903:1993, based on a modified ISO 7875-1:1984) specifies a spectrometric method for determining anionic surfactants by measuring the methylene blue index (MBAS) in the aquatic environment. Natural and synthetic anionic surface-active substances are determined as substances active towards methylene blue — a sum parameter called the MBAS index. The method (its scope described after the newer edition, ISO 7875-1:1996) applies to drinking water, surface water and wastewater, for example for assessing the primary biodegradation of surfactants in test systems with natural or synthetic wastewater, on a laboratory and on a technical scale; in the effluents of municipal treatment plants the MBAS index covers not only synthetic but, to a considerable extent, also natural anionic surface-active substances. The concentration range is 0.1–5.0 mg/l and the limit of detection is about 0.05 mg/l for standard surfactants in distilled water. The 2019 regulation (Dz.U. 2019 poz. 1311 [Journal of Laws 2019, item 1311]) names PN-EN 903 (molecular absorption spectrophotometry) as the reference method for anionic surfactants in wastewater.

The determination consists in the formation of ion pairs between the methylene blue cation and the surfactant anion, which can be extracted quantitatively with chloroform; what is determined above all are organic sulfates and sulfonates, but positive interferences may occur (organic sulfates, sulfonates, carboxylates, phenols, inorganic anions such as cyanate, nitrate, thiocyanate and sulfide) as well as negative ones (cationic substances — quaternary ammonium compounds, proteins — that form complexes with the anionic surfactants); it is therefore more correct to describe the result as the MBAS index than as "anionic surfactants". Samples are not taken through a layer of foam; the glass bottles are cleaned with methanol; short storage at 4 °C, longer storage with a preservative (1 % of 40 % formaldehyde for up to 4 days, saturation with chloroform for up to 8 days); in surface waters of unknown composition the surfactants can be separated from other active substances by gas stripping (sublation) in an apparatus with ethyl acetate.

Method principle

The formation of the methylene blue salts of the anionic surfactants in an alkaline medium, the extraction of these salts with chloroform and the acid treatment of the chloroform solution; the removal of interferences by extracting the surfactant–methylene blue complex from the alkaline solution and shaking it with an acidic solution of the blue. The absorbance of the separated organic phase is measured at the absorption maximum of 650 nm and evaluated against a calibration curve. The standard is the methyl ester of dodecylbenzenesulfonic acid (the tetrapropylene type, relative molecular mass 340; 400–450 mg is saponified for 1 h under a reflux condenser in 50 ml of ethanolic NaOH, neutralised with H₂SO₄ 0.5 mol/l against phenolphthalein and diluted to 1000 ml; stable for at least 6 months; the sodium salts of dodecane-1-sulfonic acid, dodecane-1-sulfuric acid or dioctylsulfosuccinic acid are also allowed), and the MBAS index is calculated as sodium dodecylbenzenesulfonate. Reagents: neutral methylene blue solution (0.350 g/l, prepared at least 24 h before use, stable for at least 2 weeks; the absorbance of the blank chloroform phase must be ≤ 0.2 per 10 mm, otherwise it is purified by extraction: 200 ml of buffer and 200 ml of chloroform per 100 ml of the solution, shaking for 30 s, running off the chloroform layer and rinsing the aqueous layer with 60 ml of chloroform, twice), acidic methylene blue solution (0.350 g in 500 ml of water + 6.50 ml of H₂SO₄ of 1.84 g/ml, made up to 1000 ml; blank ≤ 0.02 per 10 mm), pH 10 buffer (24 g of NaHCO₃ + 27 g of anhydrous Na₂CO₃ per 1000 ml; for very hard waters, alternatively an alkaline borate solution of equal volumes of 0.05 mol/l Na₂B₄O₇·10H₂O and 0.1 mol/l NaOH), chloroform, ethanol 95 %, methanol, H₂SO₄ 0.5 mol/l, ethanolic NaOH and phenolphthalein. Apparatus: a spectrometer able to measure at 650 nm with 10 mm cells, separating funnels (e.g. 250 ml), volumetric flasks of 50 ml, glass-fibre or soft cellulose filters and — optionally — a gas stripping (sublation) apparatus.

Applications

• Municipal and industrial wastewater (laundries, cosmetics, detergents) — the reference method for anionic surfactants under Dz.U. 2019 poz. 1311
• Effluents from treatment plants — assessment of the primary biodegradation of surfactants on a laboratory and a technical scale
• Surface waters (rivers, lakes) — monitoring of pollution by detergents; with sublation where the matrix is unknown
• Drinking water — confirmation that there are no residues of anionic surfactants (down to 0.05 mg/l)
• Waterworks and environmental laboratories — the classical extraction with spectrophotometric measurement

Key parameters

Standard

Standard number

PN-EN 903:2002 (EN 903:1993; ISO 7875-1:1984 MOD)

Title (PL)

Jakość wody — Oznaczanie surfaktantów anionowych przez pomiar indeksu błękitu metylenowego MBAS

Title (EN)

Water quality — Determination of anionic surfactants by measurement of the methylene blue index MBAS

Step-by-step procedure

1. Sampling and storage

   Take the sample, not through the layer of foam, into a clean glass bottle cleaned with methanol; cool it to 4 °C; for storage longer than 24 h add 1 % (V/V) of 40 % formaldehyde (up to 4 days) or saturate it with chloroform (up to 8 days). Suspended matter can be centrifuged off (surfactant adsorbed on the suspended matter will not be determined).

2. Extraction

   Transfer into a separating funnel a measured volume of the sample containing 20–200 µg of MBAS (make up less than 100 ml with water to 100 ml), add 5.0 ml of the neutral methylene blue solution, 10 ml of the pH 10 buffer and 15 ml of chloroform; shake evenly (preferably in the vertical plane) for at least 1 min and leave the phases to separate.

3. Washing the extract

   Transfer the chloroform layer as completely as possible into a second funnel containing 110 ml of water and 5 ml of the acidic methylene blue solution; shake for at least 1 min and leave it to separate.

4. Filtration and repetition

   Filter the chloroform layer into a 50 ml volumetric flask through a glass-fibre filter moistened with chloroform; repeat the extraction from the first and the second funnel (e.g. twice with 10 ml of chloroform), filtering into the same flask; make up to the mark with chloroform and mix.

5. Measurement

   Measure the absorbance of the extract at 650 nm in a 10 mm cell against pure chloroform; for each batch of samples run a blank with 100 ml of distilled water taken through the whole procedure.

6. Calibration

   Add 0; 1; 2; 4; 6; 8 ml of the diluted standard solution (e.g. 20 µg/ml of sodium dodecylbenzenesulfonate) to the funnels, make up with water to 100 ml and proceed as with a sample; plot the absorbance-against-concentration curve (including the zero solution).

7. Calculation

   Read the MBAS index (as sodium dodecylbenzenesulfonate) from the curve or calculate it: c = (A₁ − A₀)·f₁/V, where A₁ is the absorbance of the sample, A₀ that of the blank, f₁ the calibration factor (the mass of MBAS giving an absorbance of 1.000) and V the volume of the sample; report it in mg/l (µg/l).

Required equipment and apparatus

Reagents, media and consumables

Health and safety (OHS)

• Chloroform is toxic and suspected of being carcinogenic — carry out all the extractions in a fume hood, wear gloves resistant to solvents, dispose of the chloroform waste
• Ethyl acetate (sublation) — flammable; keep the apparatus in a well-ventilated fume hood
• Formaldehyde (preservation) — work in a fume hood, gloves
• Sulfuric acid and ethanolic NaOH — goggles and gloves

Frequently asked questions