ISO 13641-1:2003 pdf download
ISO 13641-1:2003 pdf download.Water quality -Determination of
inhibition of gas production of anaerobic bacteria— Part 1: General test.
5.2 Insulated containers, for transport of sludge.
5.3 Three-way pressure-release valves, capable of being fitted to the caps of inoculum collection bottles.
5.4 Centrifuge, for determination of the mass concentration of inoculum solids.
5.5 Sieve, having a mesh size of 1 mm2.
5.6 Reservoir for digesting sludge, consisting of a glass or plastic bottle (capacity about 5 I) fitted with a stirrer and facilities for passing a stream of nitrogen gas through the headspace.
5.7 Membrane filters, with a pore size of 0,2 pm for sterilizing the substrate.
5.8 Microsyringes, for the gas-tight connection of the pressure meter to the headspace in the bottles; also for adding insoluble liquid test materials into the bottles.
5.9 Pressure-resistant gas-tight closed glass test bottles, with an appropriate nominal size (0,1 Ito 11).
Use, for example, nominal 125-mI serum bottles with an actual total volume of 160 ml, gas-tight sealed with septa and crimped aluminium rings. Use septa preferably made of silicone or polytetrafluoroethene-coated butyl rubber capable of withstanding a pressure of about 2 x iO Pa. Gas-tightness of the caps used, especially butyl rubber septa, should be tested in advance because several commercially available septa are not sufficiently gas-tight against methane, and some septa do not stay tight when they are pierced with a needle as required under the conditions of this test.
5.10 Precision pressure meter for measuring total biogas production (methane plus carbon dioxide).
A needle attachment is adapted to enable measurement and venting of the biogas produced. An example of a suitable instrument is a hand-held precision pressure meter connected to a suitable syringe needle; a 3-way gas-tight valve facilitates the release of excess pressure. Calibrate the meter (see Annex A) in order to allow, if necessary, the conversion of pressure measurements to gas volumes. The internal volume of the pressure meter tubing and valve should be kept as low as practically possible, so that errors introduced by neglecting the volume of the equipment are insignificant.
If a pressure meter of the described quality is used (for example capsulated with a steel membrane), no calibration is necessary in the laboratory. It should be calibrated by a licensed institute at the intervals recommended by the manufacturer. The accuracy of the calibration can be checked at the laboratory with a one-point measurement at 1 x iO Pa against a pressure meter with a mechanical display. When this point is measured correctly, the linearity will also be unaltered. If other measurement devices are used (without certified calibration by the manufacturer), calibration is recommended over the total range at regular intervals.
5.11 Glove box (optional), with a slight positive pressure of nitrogen.
5.12 Spark-free incubator, preferably equipped with a shaking device, and capable of maintaining the
temperature at 35 °C ± 1 °C.
6 Test environment and interferences
6.1 Test environment
Carry out the test by incubating the sealed test bottles at a constant temperature of 35 °C ± 1 °C in the absence of oxygen, initially in an atmosphere of nitrogen, in the dark or in diffused light. Use nitrogen of high purity (4.1.2). In special cases, the test may be conducted at other temperatures.
6.2.3 pH of the medium
Activity of anaerobic cultures is very sensitive to the pH value. Ensure that the pH of the reaction mixture is
adjusted to 7 ± 0,5 and remains within the range of 6,2 to 7,5 until the end of incubation (see 7.5).
6.2.4 Quality of the caps of the bottles
Different types of septa are commercially available. Many of them do not remain gas-tight after being pierced with a needle under the test conditions. Sometimes the pressure falls very slowly once the septa have been pierced with the syringe needle.
6.2.5 Remaining substrates in the sludge
The anaerobic biogas production and the sensitivity of the sludge are influenced by substrates, which are transferred with the inoculum into the test bottles. Digested sludge from domestic anaerobic digesters often contains materials such as sand, hair or plant residues of cellulose. Sieving the sludge will remove gross insoluble matter, thus making it easier to take representative samples.
6.2.6 Volatile chemicals
Volatile chemicals are released into the headspace of the serum bottles. This can result in the loss of some of the test material from the system during venting after pressure measurements, yielding falsely high EC50 values. For details see Reference [1] in the Bibliography.
7 Procedure
7.1 Test set-up and preliminary test.
