🧪 Phenolic index of water
In short
Determination of the phenol index in drinking water, surface water and waste water after steam distillation — reaction of phenolic compounds with 4-aminoantipyrine in the presence of potassium hexacyanoferrate(III); method A is direct, method B uses extraction with chloroform. The test is performed according to PN-ISO 6439:1994; Method A (direct): more than 0.10 mg/l in the aqueous phase; 510 nm (clause 1, 4.1). The procedure comprises 7 steps; it is used for: Waste water — reference method for the phenol index (Journal of Laws 2019, item 1311, Annex 12 item 25), Monitoring of surface and ground waters — volatile phenols, phenol index (Journal of Laws 2021, item 1576, Annex 8), Drinking and surface waters — method B for an index of about 0.002–0.10 mg/l (clause 1).
At a glance
- Standard: PN-ISO 6439:1994
- Category: Physicochemistry
- Procedure steps: 7
- Method A (direct): more than 0.10 mg/l in the aqueous phase; 510 nm (clause 1, 4.1)
- Method B (chloroform extraction): about 0.002–0.10 mg/l without dilution; 460 nm (clause 1; wavelength according to the 1984 edition)
- Reaction conditions: pH 10.0 ± 0.2, 4-aminoantipyrine and potassium hexacyanoferrate(III); measurement after 15 min (4.1, 4.6.3.3)
Overview
STATUS (as of 29.10.2026). PN-ISO 6439:1994 — Polish version, published 23.12.1994, implements ISO 6439:1990 [IDT]; it replaced PN-C-04602-02:1972 and PN-C-04602-03:1972. The standard is current — no withdrawal note. ISO 6439:1990 (second edition, 15.05.1990) is published in the ISO catalogue, with no newer edition; it replaced the first edition of 1984, of which it is a minor revision.
SCOPE (clause 1). Methods for determining the phenol index in drinking waters, surface waters and waste waters. After a preliminary distillation: method A (direct colorimetric) — samples containing more than 0.10 mg/l in the aqueous phase, without chloroform extraction; method B (chloroform extraction) — without dilution from about 0.002 to about 0.10 mg/l, when the coloured product is concentrated in chloroform. Phenol is the standard for both methods. According to a German interlaboratory trial with a method almost identical to method B, the lower limit of detection is 0.01 mg/l. The standard also notes that the limits of detection of both methods are insufficient for checking compliance with the drinking water limits of Directive 80/778/EEC.
WHAT THE PHENOL INDEX MEASURES (introduction, clause 3). The phenol index is a number, expressed in milligrams of phenol per litre, corresponding to the intensity of colour produced with 4-aminoantipyrine by various phenolic compounds (hydroxy derivatives of benzene and its analogues). The composition of phenolic compounds in a sample is unpredictable, so the colour of all of them is measured as phenol (C₆H₅OH); the methods do not differentiate between kinds of phenols. Some phenols with an alkyl, aryl or nitro substituent in the para position do not produce colour; those with a carboxyl, halogen, hydroxyl, methoxyl or sulfonic acid substituent in the para position do.
INTERFERENCES (according to the 1984 edition, 4.8). Phenol-decomposing bacteria, oxidizing and reducing substances and strongly alkaline conditions of the sample; biological degradation is inhibited by copper(II) sulfate, and acidification with phosphoric acid assures the presence of copper(II) ions. Oxidizing agents (smell of chlorine, liberation of iodine from potassium iodide on acidification) are removed immediately after sampling with iron(II) sulfate or ascorbic acid. Oils and tar are removed by an alkaline extraction (pH 12–12.5, without copper(II) sulfate), compounds liberating hydrogen sulfide — by copper(II) sulfate, reducing agents — by an excess of potassium hexacyanoferrate(III), and the high bias from amines is minimized by distillation below pH 0.5. Removing interferences may cause losses of some phenolic compounds.
LEGAL REQUIREMENTS. Waste water: PN-ISO 6439 is a reference method for phenol (phenol index), alongside PN-EN ISO 14402 (flow analysis) (Journal of Laws 2019, item 1311, Annex 12 item 25). Monitoring of surface and ground waters: method for the indicator "volatile phenols — phenol index" (Journal of Laws 2021, item 1576, Annex 8, indicator 3.6.9). The regulation on water intended for human consumption (Journal of Laws 2026, item 748) does not include the phenol index.
WHAT WE DO NOT GIVE HERE AND WHERE THE PROCEDURE DETAILS COME FROM. The available text of the 1990 edition ends at 4.6.3.3 (measurement of the calibration solutions of method A). The determination (4.6.4), expression of results, interferences and method B are given according to the full text of the 1984 edition. In the part common to both editions they differ, among other things, in the scope (the 1984 edition also lists brines and domestic waters), the recipe for the pH 10 buffer and the stability of the phenol stock solution — so figures marked "according to the 1984 edition" must be checked in the current text of the standard. We do not describe the titrimetric check of the phenol solution concentration (Annex A).
Method principle
Phenolic compounds are separated from impurities and preservatives by distillation; phenols volatilize gradually, so the volume of the distillate must equal the volume of the sample distilled (4.1). The steam-distillable phenolic compounds react with 4-aminoantipyrine at pH 10.0 ± 0.2 in the presence of potassium hexacyanoferrate(III) to form an antipyrine dye.
Method A: the absorbance of the dye is measured at 510 nm; the minimum detectable quantity corresponds to 0.01 mg of phenol with a 50 mm cell and 100 ml of distillate. Method B (according to the 1984 edition): the dye is extracted with chloroform and measured at 460 nm; the minimum detectable quantity is 0.005 mg of phenol (25 ml of chloroform and a 50 mm cell or 50 ml of chloroform and a 100 mm cell), i.e. an index of 0.002 mg/l in 500 ml of distillate. The result is given in milligrams of phenol per litre.
Applications
- Waste water — reference method for the phenol index (Journal of Laws 2019, item 1311, Annex 12 item 25)
- Monitoring of surface and ground waters — volatile phenols, phenol index (Journal of Laws 2021, item 1576, Annex 8)
- Drinking and surface waters — method B for an index of about 0.002–0.10 mg/l (clause 1)
Key parameters
| Parameter | Value |
|---|---|
| Method A (direct) | more than 0.10 mg/l in the aqueous phase; 510 nm (clause 1, 4.1) |
| Method B (chloroform extraction) | about 0.002–0.10 mg/l without dilution; 460 nm (clause 1; wavelength according to the 1984 edition) |
| Reaction conditions | pH 10.0 ± 0.2, 4-aminoantipyrine and potassium hexacyanoferrate(III); measurement after 15 min (4.1, 4.6.3.3) |
| Distillation | 500 ml of sample at pH 1–2 (phosphoric acid), with copper(II) sulfate; distillate 500 ml (4.5.1) |
| Sample preservation | if analysed later than 4 h after collection: pH about 4.0, 1.0 g of CuSO₄·5H₂O per litre, 5–10 °C, analysis within 24 h (4.4) |
| Result | mg of phenol (C₆H₅OH) per litre; the methods do not differentiate between kinds of phenols |
Standard
- Standard number
- PN-ISO 6439:1994
- Title (PL)
- Jakość wody — Oznaczanie indeksu fenolowego — Metody spektrometryczne z 4-aminoantypiryną po destylacji
- Title (EN)
- Water quality — Determination of phenol index — 4-Aminoantipyrine spectrometric methods after distillation
Step-by-step procedure
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Sampling and preservation
According to ISO 5667-1, -2 and -3, into glass bottles. If the analysis is not carried out within 4 h of collection: acidify with 1 + 9 phosphoric acid to pH about 4.0 (methyl orange or pH meter), add 1.0 g of copper(II) sulfate pentahydrate per litre, store at 5–10 °C and analyse within 24 h of collection (4.4).
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Distillation
Measure 500 ml of sample; if it was not preserved with copper(II) sulfate, add 5 ml of copper(II) sulfate solution; adjust the pH to 1–2 with phosphoric acid. Distil 400 ml into a 500 ml graduated cylinder, stop, add 100 ml of water to the flask when boiling ceases and continue until a total of 500 ml has been collected (4.5.1).
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Turbid distillate
Acidify with phosphoric acid, add 5 ml of copper(II) sulfate solution and distil again; if this distillate is also turbid — extract a new sample: acidify 500 ml with sulfuric acid (methyl orange), add 150 g of sodium chloride, shake with five portions of chloroform (40 ml, then 4 × 25 ml), back-extract the phenols from the combined extracts with three portions of sodium hydroxide (4.0 ml and 2 × 3.0 ml), remove the chloroform on a water bath, dilute to 500 ml and distil as above (4.5.2–4.5.3).
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Test portion, blank, calibration — method A
100 ml of distillate or a portion containing not more than 0.5 mg of phenol diluted to 100 ml (smallest practical portion 10 ml), at room temperature; blank — 100 ml of water. Calibration solutions: in seven 500 ml flasks 0, 25, 50, 100, 150, 200 and 250 ml of 0.01 g/l phenol standard solution, made up with water and distilled like the sample (4.6.1–4.6.3.1).
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Colour reaction and measurement — method A
According to the 1984 edition (4.6.4): add 5 ml of buffer to the portion (or 5 ml of ammonium chloride solution and adjust to pH 10 ± 0.2 with ammonium hydroxide), then 2.0 ml of 4-aminoantipyrine solution and mix immediately, then 2.0 ml of hexacyanoferrate(III) solution and mix immediately again. After 15 min measure the absorbance at about 510 nm against water (4.6.3.3).
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Method B — chloroform extraction
According to the 1984 edition (clause 5): 500 ml of distillate or a portion with not more than 0.05 mg of phenol (at least 50 ml) diluted to 500 ml; blank — 500 ml of water; calibration solutions with 0, 1, 2, 5, 10, 20, 30, 40 and 50 ml of 0.001 g/l standard solution. Add 20 ml of buffer (pH 10 ± 0.2), 3.0 ml of 4-aminoantipyrine and 3.0 ml of hexacyanoferrate(III), after 15 min shake for 1 min with exactly 25 ml of chloroform (50 ml for a 100 mm cell), filter the extract through 5 g of sodium sulfate into a volumetric flask and make up with chloroform; measure within 1 h at 460 nm against chloroform.
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Calculation
The mass of phenol in the portion is read from the calibration graph (absorbance against mass of phenol in mg) after subtracting the blank; phenol index, mg/l = m × 1000/V₀, where V₀ is the volume of the portion in ml (according to the 1984 edition, 4.7 and 5.6).
Required equipment and apparatus
| Equipment | Example | Indicative price |
|---|---|---|
| Distillation apparatus | all glass, borosilicate, 1 l, with a Graham condenser or equivalent (4.3.1) | — |
| pH meter | with suitable electrodes (4.3.2) | — |
| Spectrometer | for measurements at 510 nm (method B — at 460 nm), with cells of 10–100 mm path length; if the absorbance exceeds 1.0, a cell with a shorter path is used (4.3.3, 5.3.1) | — |
| Sample bottles | glass (4.4) | — |
| For method B | 1 l separating funnels, Büchner funnel with a fritted disc or a phase separator filter, 25 ml or 50 ml volumetric flasks (according to the 1984 edition, 5.3, 5.5.4) | — |
Reagents, media and consumables
| Reagent | CAS | Details |
|---|---|---|
| 4-Aminoantipyrine, 20 g/l | 83-07-8 | dissolve 2.0 g in water and dilute to 100 ml; prepare just before use; if red particles remain, the solution cannot be used (4.2.1) |
| Potassium hexacyanoferrate(III), 80 g/l | 13746-66-2 | dissolve 8.0 g in water and dilute to 100 ml, filter if necessary; use within one week of preparation (4.2.14) |
| Tartrate buffer pH 10 | — | dissolve 34 g of ammonium chloride and 200 g of potassium sodium tartrate in 700 ml of water, add 150 ml of ammonium hydroxide (ρ = 0.90 g/ml) and dilute to 1000 ml (4.2.4); alternatively 20 g/l ammonium chloride solution and ammonium hydroxide to pH 10 ± 0.2 (4.2.2–4.2.3; according to the 1984 edition, 4.6.4) |
| Phosphoric acid, ρ = 1.70 g/ml, and solution 1 + 9 | 7664-38-2 | 1 volume of acid to 9 volumes of water; for preservation and distillation (4.2.12–4.2.13) |
| Copper(II) sulfate pentahydrate and 100 g/l solution | 7758-99-8 | solid — for preservation (1.0 g/l of sample); solution — 190 g of CuSO₄·5H₂O dissolved and diluted to 1000 ml (4.2.5–4.2.6) |
| Phenol — solutions | 108-95-2 | stock 1.00 g/l in freshly boiled and cooled water (stable for about one week; the phenol must not be liquid or discoloured, its concentration may be checked by titration according to Annex A); standard 0.01 g/l — on the day of use; standard 0.001 g/l — within 2 h of use (4.2.9–4.2.11) |
| Methyl orange | — | 0.5 g in 1000 ml of water — indicator for acidification (4.2.8) |
| For turbid distillates and method B | — | 0.5 mol/l sulfuric acid, sodium chloride, 2.5 mol/l sodium hydroxide (10 g in 100 ml of water), chloroform, anhydrous granular sodium sulfate (4.2.15–4.2.16) |
Health and safety (OHS)
- Phenol must not come into contact with the skin (caution at 4.2.9)
- Chloroform is toxic and a suspected carcinogen — do not breathe the vapour, avoid contact with skin and eyes (warning at 4.2.16.4)
- Handle phosphoric acid, ammonium hydroxide and potassium hexacyanoferrate(III) according to the safety data sheets
Frequently asked questions
Which standard describes this test?
The test is performed according to PN-ISO 6439:1994 — “Water quality — Determination of phenol index — 4-Aminoantipyrine spectrometric methods after distillation”.
How does this method work?
Phenolic compounds are separated from impurities and preservatives by distillation; phenols volatilize gradually, so the volume of the distillate must equal the volume of the sample distilled (4.1).
What is the measuring range and accuracy?
Method A (direct): more than 0.10 mg/l in the aqueous phase; 510 nm (clause 1, 4.1); Method B (chloroform extraction): about 0.002–0.10 mg/l without dilution; 460 nm (clause 1; wavelength according to the 1984 edition); Reaction conditions: pH 10.0 ± 0.2, 4-aminoantipyrine and potassium hexacyanoferrate(III); measurement after 15 min (4.1, 4.6.3.3); Distillation: 500 ml of sample at pH 1–2 (phosphoric acid), with copper(II) sulfate; distillate 500 ml (4.5.1).
How long does the test take?
The procedure comprises 7 steps. The standard does not give a duration for every stage — the laboratory's own procedure decides.
What equipment is required?
Distillation apparatus, pH meter, Spectrometer, Sample bottles, For method B.
Where is this test used?
Waste water — reference method for the phenol index (Journal of Laws 2019, item 1311, Annex 12 item 25); Monitoring of surface and ground waters — volatile phenols, phenol index (Journal of Laws 2021, item 1576, Annex 8); Drinking and surface waters — method B for an index of about 0.002–0.10 mg/l (clause 1).
What safety precautions apply?
Phenol must not come into contact with the skin (caution at 4.2.9); Chloroform is toxic and a suspected carcinogen — do not breathe the vapour, avoid contact with skin and eyes (warning at 4.2.16.4); Handle phosphoric acid, ammonium hydroxide and potassium hexacyanoferrate(III) according to the safety data sheets.
Which laboratory can perform this test?
The test is performed by laboratories accredited to ISO/IEC 17025. On LabCoda you can find them by the standard number PN-ISO 6439.