ISO 13395:1996

Overview

ISO 13395:1996 specifies automated methods for the determination of nitrite nitrogen (nitrite‑N), nitrate nitrogen (nitrate‑N) and the sum of both (nitrite/nitrate‑N) in waters using flow analysis (FIA and CFA) with spectrometric (photometric) detection. The standard describes reagents, calibration, apparatus and procedures for continuous flow analysis (CFA) and flow injection analysis (FIA), including the common step of cadmium reduction of nitrate to nitrite and subsequent diazotization/coupling to form a colored dye measured at ~520–560 nm.

Key topics and technical requirements

• Analytical principle: sample mixed into a carrier stream (FIA) or continuously blended (CFA); nitrate reduced to nitrite with metallic cadmium, then diazotized and coupled to form a detectable dye.
• Working ranges:
  • Nitrite (N): two ranges - 0.01–0.1 mg/L and 0.1–1.0 mg/L.
  • Nitrite/nitrate (N): 0.2–2 mg/L and 2–20 mg/L.
• Apparatus essentials: low‑pulsation pumps, injector (10–300 µL), cadmium reductor (minimum 90% reduction efficiency), flow photometer with flow cell (520–560 nm), reaction coils, tubing and optional autosampler.
• Reagents & solutions: phosphoric acid buffer, sulfanilamide, N‑(1‑naphthyl)ethylenediamine reagent, imidazole or ammonium buffer, copper sulfate, and prepared calibration standards. Many reagent stability and preparation notes are specified.
• Quality checks: calibration with at least five standards per range; a low‑range calibration solution should produce absorbance ≥0.04 per 10 mm path length. Degassing of carrier/reagent solutions and control of surfactant content are required.
• Interferences & sample handling: pH control (ensure pH ~6.5–7.5 through reducer), surfactant levels >10 mg/L can interfere (dialysis recommended), filtration for particulate matter, acidification and cold storage for some samples, and safe disposal of cadmium waste.

Applications and users

ISO 13395 is used for routine and high‑throughput analysis in:

• Environmental and regulatory laboratories monitoring surface water, groundwater, drinking water and wastewater for nitrogen species.
• Water utilities performing compliance testing and process monitoring.
• Contract testing and research labs needing automated, reproducible nitrite/nitrate analysis.
• Instrument manufacturers and method developers designing or validating FIA/CFA analysers.

Practical benefits include high sample throughput (up to ~100 samples/hour), automation of wet chemistry, and flexibility of working ranges by adjusting operating conditions.

Related standards

• ISO 3696: Water for analytical laboratory use - specifications
• ISO 5667 series: Sampling guidance (Parts 1–3 cited)
• ISO 6777: Alternative spectrometric nitrite method
  (ISO 13395 was prepared by ISO/TC 147/SC 2 and includes informative annexes on system configurations and reagents.)

Keywords: ISO 13395, water quality, nitrite, nitrate, flow analysis, FIA, CFA, spectrometric detection, cadmium reduction, flow photometer.