ISO 22404:2019 download free.Plastics — Determination of the aerobic biodegradation of nonfloating materials exposed to marine sediment — Method by analysis of evolved carbon dioxide.
Introduction
Products made with biodegradable plastics and other biodegradable materials are designed to be recoverable by means of organic recycling In composring plants or in anaerobic digesters. The uncontrolled dispersion of biodegradable plastics in natural environments Is not desirable. The biodegradability of products cannot be considered as an excuse to spread wastes that should be recovered and recycled, However, Lest methods to measure rate and level of biodegradation in natural environments (such as soil or the marine environment) are of interest In order to better characterize the behaviour of plastics in these very particular environments. As a matter of fact, some plastics are used in products that are applied in the sea (for example, fishing gear) and sometimes they can get lost or put willingly in marine environment, The characterization of biodegradable plastic materials can be enlarged by applying specilic test methods that enable the quantitative assessment of biodegradation of plastics exposed to marine sediment and seawater. In order to carry out a proper product design, it is important to know whether a plastic material is inherently biodegradable when exposed to marine inocub.
ISO 22404 provides a test method for calculating and reporting biodegradation level obtained under laboratory conditions using a marine inoculum. The marine Inoculum Is sediment sampled at the tidal zone. The plastic material Is exposed to this environmental matrix and biodegradation is followed by measuring the evolved CO2.
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theoretical amount of evolved carbon dioxide ThC02
maxtmum theoretical amount ol carbon dioxide evolved after completely oxldesrng a chemical compound, calculated from the molecular formula or from determrnatlon of total organic carbon (TOC) (3.3) and expressed as milligrams of carbon dioxide evolved per milligram or gram of test compound
3.3
total organic carbon
TOC
amount of carbon bound in an organic compound
Note Ito entry; Total organic carbon Is expressed as milligrams of carbon per 100 mIlligrams of the compound.
3.4
dissolved organic carbon
DOC
part of the organic carbon in water which cannot be removed by specified phase separation methods, for example by centrifugation at 40 000 ms2 for 15 mm or by membranes with pores of 0.2 urn to 0,45 trn diameter
3.5
pre-conditioning phase
pre-incubatlon clan inoculum under the conditions ofthe subsequent test In the absence of test material, with the aim to consume potential organic matter present in excess that could disturb biodegradation measurement and to improve the acclimatization of the microorganisms to the test conditions
4 Principle
This test method is based on the determination of the evolved carbon dioxide and derives from ISO 19679. The testing medium is based on a sandy marine sediment laid in the bottom of a closed flask; the sediment Is kept wet with natural sea water, The test material is prererably in the form of a powder.
The carbon dioxide evolved during the microbial degradation Is determined by a suitable analytical method. The level of biodegradation Is determined by comparing the amount of carbon dioxide evolved with the theoretical amount (ThCO2) and expressed in per cent. The test result is the maximum level of biodegradation, determined from the plateau phase of the biodegradation curve. The principle of a system for measuring evolved carbon dioxide is given in ISO 14852:2018. Annex A.
5 Test environment
Incubation shall take place in the dark in an enclosure which is free from vapours inhibitory to microorganisms and which is maintained at a constant temperature, preferably between 15°C to 25 °C, but not exceeding 28 C. to an accuracy of ±2 °C. Any change in temperature shall be justified and clearly Indicated in the test report.
NOTE Test results are obtained for temperature that can be different from real conditions In marine environment.
6 Reagents
6.1 Water, dLstdled or deionized water, free of toxic substances (copper in particular) and containing less than 2 mgjl of DOC
The form and shape of the test material may influence its biodegradation speed. Similar shapes and thicknesses should preferably be used if different kinds of plastic materials are to be compared. A particle size distribution with the maximum at 250 pm diameter Is recommended.
The test material may also be introduced as a film. When the test material in form of film is buried in the sediment it could limit the gas exchange between the water body and the sediment, promoting the formation of anaerobic zones under the test material. In order to reduce this elfea, it is possible to perforate the film sample homogeneously over the entire surface.
8.2 Reference material
Use microcrystalline cellulose or ashless cellulose filters as a reference material’). If possible, the TOC, form and size should be comparable to that of the test material. As a negative control, a non- biodegradable polymer (e.g. polyethylene) in the same form as the test material should be used.
8.3 Preparation of the sediment
Filter the sediment in a funnel with a coarse filter paper to eliminate excess seawater. Sediment is ready for testing when dripping of sea water is ended. Sediment after filtering is named “wet sediment” hereafter. Nitrogen (such as NH4CI or NaNO3) can be added to the sediment If this is considered as a factor limiting biodegradation, It is suggested to add 0,1 g of N per g of TOC of test item added to the test system. These additions shall be reported In the teSt report.
It is recommended that the ratio between organic carbon and nitrogen (C/N ratio) of the test mixture is optimized so as to ensure optimum biodegradation conditions. The C/N ratio for the test mixture should preferably be between 10 and 40.
8.4 Test setup
Provide a number of flasks, so that the test includes at least the following:
a) three flasks for the test material (symbol F.,,); b) three flasks for the blank (symbol Fn);
c) three flasks for reference material (symbol Fr). In addition, it is recommended to add:
d) three flasks for negative control (symbol FN).
Two flasks for test material, blank, reference material and negative control may be used instead of three for screening purposes
8.5 Pre-conditlonlng phase
in a typical case, use a test flask with a volume of 3 1. Laid down equal amount (between 200 g and 600g. typically 400g) of the wet sediment on the bottom of each flask to form a homogenous layer. Do not press or compact the sediment.
Add carbon dioxide absorber to the absorber compartments of the test flask in a typical case 30 ml of KOH 0,5 N or 100 ml of Ba(OH)2 0,025 N. Place the flasks In a constant-temperature environment and allow all vessels to reach the desired temperature. Take the necessary readings and monitor the CO2 evolution,
10 Validity of results The test is considered valid if:
a) the amount of evolved CO2 of the reference material (Fe) is>60 % of the ThCO2 after 1110 days
b) the evolved CO2 oldie blank FH at the end or the test does not exceed a 3,5 mg C02/g wet sediment (see i1) after six months;
c) theamountofcarbon dioxide evolved from the three blanks FH are within 20 %of the mean at the plateau phase or at the end of the test;
d) the difference between the percentage biodegradation of the reference material in the different vessels is less than 20 % of the mean at the end of the test.
II a negative control (flasks FM) has been performed no significant biodegradation shall be observed
(<10 %).
II these criteria are not fulfilled, repeat the test using another sediment.
11 Test report
The rest report shall contain at least the following information:
a) a reference to this document, ic. ISO 22404:201%
b) all information necessary to identify the test material, reference material, and negative control (if applicable), including their TOC, ThCO2, chemical composition and formula (if known);
c) the main test parameters, including test volume, test medium used, incubation temperature and final pH;
d) the source, amount, and characteristics of the marine sediment used, Indudlng p11, ash content, TOC. C:N ratio, date ol collection, storage conditions, handling, and potential acclimation to test material;
e) the analytical techniques used, including the principle of the respirometer and the TOC;
1) all the test results obtained for the test material, reference material, and negative control (if applicable), in tabular and graphical form, Including the evolved CO, the percentage biodegradation values;
g) the duration of the lag phase, biodegradation phase and maximum level of degradatian, as well as the total test duration; and, optionally, if rin or determined: the negative control FM:
h) any other relevant data (e.g. result of the visual final inspection and analysis 01 final samples, ii still retrievable; photos of the final samples);
I) details of the methods used during the test period In order to support microbial diversity or to avoid nutrient deflc,cncy.