🔬 Phosphates / Total Phosphorus in Water
Spectrophotometric determination of orthophosphates and total phosphorus in water by ammonium molybdate and ascorbic acid method (molybdenum blue). A key eutrophication parameter.
Overview
Phosphorus is a key biogenic element responsible for eutrophication of surface waters — excessive algal growth, cyanobacterial blooms, oxygen deficiency, and aquatic ecosystem degradation. In natural waters phosphorus occurs as: orthophosphates (PO₄³⁻, reactive, directly assimilable by organisms), polyphosphates (hydrolyze to orthophosphates), and organic phosphorus (bound to organic matter).
Total phosphorus = orthophosphates + polyphosphates + organic phosphorus (after mineralization).
Standards: drinking water ≤0.4 mg PO₄³⁻/L, treated wastewater ≤2 mg P/L (>10 000 PE), surface waters class I ≤0.2 mg P/L. Urban Wastewater Directive requires phosphorus reduction in treatment plants above 10 000 PE.
ISO 6878 method (molybdenum blue with ascorbic acid) is reference method — highly sensitive, range 0.005–0.8 mg P/L.
Method principle
Orthophosphate ions react with ammonium molybdate and antimony potassium tartrate in acidic environment (H₂SO₄), forming heteropolyacid phosphomolybdic complex (antimony-phosphomolybdate). Complex is reduced by ascorbic acid to intensely blue complex (molybdenum blue).
PO₄³⁻ + Mo(VI) + Sb³⁺ → (H⁺) → phosphomolybdate complex Complex + ascorbic acid → molybdenum blue (intensely blue)
Absorbance is measured at 880 nm (maximum sensitivity) or 700 nm (with lower sensitivity). Color develops in 10–30 minutes and is stable for 24 hours.
For total phosphorus: before analysis sample is mineralized (K₂S₂O₈ + H₂SO₄, 120°C/30 min) to convert all phosphorus forms to orthophosphates.
Applications
- Surface water eutrophication monitoring
- Treated wastewater control (limit ≤1–2 mg P/L)
- Drinking water quality control
- Chemical phosphorus removal process monitoring (coagulation)
- Enhanced biological phosphorus removal (EBPR) process control
- Groundwater testing (agricultural impact)
Key parameters
| Parameter | Value |
|---|---|
| Measurement range | 0.005–0.8 mg P/L (without dilution) |
| Detection limit | 0.005 mg P/L (direct), 0.0005 mg P/L (after extraction) |
| Wavelength | 880 nm (optimum) or 700 nm |
| Color development time | 10–30 minutes (stable 24 h) |
| Precision (CV) | 2–5% |
| Mineralization (total phosphorus) | K₂S₂O₈ + H₂SO₄, 120°C, 30 min |
Standard
- Standard number
- PN-EN ISO 6878:2006+Ap1:2010+Ap2:2010
- Title (PL)
- Jakość wody — Oznaczanie fosforu — Metoda spektrometryczna z molibdenianem amonu
- Title (EN)
- Water quality — Determination of phosphorus — Ammonium molybdate spectrometric method
Step-by-step procedure
1. Mixed reagent preparation
Mix in sequence: 50 mL H₂SO₄ 4.5M + 5 mL antimony potassium tartrate solution + 15 mL ammonium molybdate solution + 30 mL ascorbic acid solution. Prepare on day of use!
2. Mineralization (total phosphorus only)
To 40 mL sample add 5 mL K₂S₂O₈ solution and 1 mL H₂SO₄ 4.5M. Mineralize in autoclave 120°C/30 min or thermoreactor. Cool to room temperature.
3. Sample preparation (orthophosphates)
Measure 40 mL sample into 50 mL volumetric flask. For turbid samples — filter through 0.45 μm filter. Sample must be at room temperature.
4. Reagent addition
Add 1 mL ascorbic acid, then 2 mL acidic molybdate solution (mixed reagent). Fill to 50 mL with water. Mix thoroughly.
5. Color development
Wait 10–30 minutes for full blue color development. Color stable up to 24 hours.
6. Absorbance measurement
Measure absorbance at 880 nm (or 700 nm) against blank. Read P concentration from calibration curve. Cuvette 10–40 mm (select for concentration).
7. Calculations
P concentration [mg/L] = curve reading × dilution factor. Conversion: PO₄³⁻ [mg/L] = P × 3.066.
8. Quality control
Control standard (e.g., 0.5 mg P/L) — deviation max ±10%. Blank — absorbance <0.01. Duplicates — RPD <10%.
Required equipment and apparatus
| Equipment | Example | Indicative price |
|---|---|---|
| UV-VIS spectrophotometer | Hach DR6000, Shimadzu UV-1900, Analytik Jena Specord 250 Plus | 15 000–45 000 PLN |
| Optical cuvettes 10–50 mm | Glass or quartz cuvettes, 10, 20, 40 mm | 50–300 PLN/pc |
| Autoclave or thermoreactor | For total phosphorus mineralization (120°C/30 min) | 4 000–30 000 PLN |
| Automatic pipettes | Eppendorf Research Plus 1–10 mL, 100–1000 μL | 800–2 000 PLN |
| Volumetric flasks 50 mL | Class A, borosilicate glass | 30–80 PLN/pc |
Reagents, media and consumables
| Reagent | CAS | Details |
|---|---|---|
| Ammonium molybdate (NH₄)₆Mo₇O₂₄·4H₂O | 12054-85-2 | Acidic solution: 13 g molybdate in 100 mL H₂SO₄ 4.5 mol/L + 400 mL water. Solution I. |
| Ascorbic acid C₆H₈O₆ | 50-81-7 | Reducer: 17.6 g/L in water. Prepare fresh (unstable — oxidizes). Valid max 1 week refrigerated. |
| Antimony potassium tartrate K(SbO)C₄H₄O₆·½H₂O | 28300-74-5 | Reaction catalyst: 0.34 g/L in water. Solution II. Shelf life: several months. |
| Sulfuric acid H₂SO₄ | 7664-93-9 | Solution 4.5 mol/L (diluted 1+3). Acidic reaction medium. |
| Potassium persulfate K₂S₂O₈ | 7727-21-1 | For total phosphorus mineralization: 5 g/100 mL. Strong oxidizer decomposing organic P forms. |
| PO₄³⁻ standard solution 1000 mg P/L | 7778-77-0 | KH₂PO₄ in water, certified CRM. For calibration curve. |
Health and safety (OHS)
- Sulfuric acid — highly corrosive, causes severe burns. Use goggles and gloves.
- Antimony potassium tartrate — contains antimony (Sb), toxic. Avoid inhalation and ingestion.
- Potassium persulfate — strong oxidizer, irritating. Store away from combustibles.
- Safety goggles, nitrile gloves, and coat mandatory.
- Molybdenum and antimony-containing waste collect separately.