EN ISO 14240-2:2011
(Main)Soil quality - Determination of soil microbial biomass - Part 2: Fumigation-extraction method (ISO 14240-2:1997)
Soil quality - Determination of soil microbial biomass - Part 2: Fumigation-extraction method (ISO 14240-2:1997)
Gives a method for the determination of microbial biomass of soils by measurement of total extractable organic biomass material mainly from freshly killed microorganisms. It is also applicable to the estimation of microbial nitrogen and ninhydrin-reactive nitrogen in soil.
Bodenbeschaffenheit - Bestimmung der mikrobiellen Biomasse von Böden - Teil 2: Fumigations-Extraktionsverfahren (ISO 14240-2:1997)
Dieser Teil von ISO 14240 legt ein Verfahren zur Bestimmung der mikrobiellen Biomasse von Böden durch Messung der gesamten extrahierbaren organischen Biomasse aus frisch abgetöteten Mikroorganismen fest. Das Verfahren kann auch für die Bestimmung des mikrobiellen Stickstoffs und mikrobiellen Ninhydrin-reaktiven Stickstoffs in Böden verwendet werden, jedoch ist im Rahmen dieses Teils von ISO 14240 nur die Messung von extrahierbarem organischem Kohlenstoff beschrieben. Das Fumigations-Extraktions-Verfahren ist auf aerobe und anaerobe (vernässte, Reisfeld-) Böden über den gesamten pH-Bereich anwendbar. Die Biomasse kann auch in Böden gemessen werden, die leicht zersetzbare Substrate enthalten und in Böden, die mit Kaliumsulfat-Lösung übersättigt sind.
ANMERKUNG Die Fumigation mit Chloroform beeinflusst ebenfalls die Bodenfauna. Der Kohlenstoffbeitrag dieser Organismen ist üblicherweise gering (< 5 %) und kann in der Regel vernachlässigt werden.
Qualité du sol - Détermination de la biomasse microbienne du sol - Partie 2 : Méthode par fumigation-extraction (ISO 14240-2:1997)
La présente partie de l'ISO 14240 prescrit une méthode pour l'estimation de la biomasse microbienne des sols par mesurage de la matière extractible totale de biomasse organique, essentiellement à partir de microorganismes venant d'être tués. La méthode est également applicable à l'estimation de l'azote microbien et de l'azote microbien réagissant à la ninhydrine dans les sols; toutefois, seul le mesurage du carbone organique extractible est décrit dans le cadre de la présente partie de l'ISO 14240. La méthode par fumigation-extraction (FE) est applicable aux sols aérobies et anaérobies (saturés, irrigués) dans toute la gamme des pH du sol. La biomasse peut également être mesurée dans des sols contenant des substrats en décomposition active et dans des sols sursaturés de solution de sulfate de potassium.
NOTE -- La fumigation au chloroforme affecte également la faune du sol. La contribution du carbone de ces organismes est généralement faible (< 5 %) et peut en règle générale être négligée.
Kakovost tal - Določevanje talne mikrobne biomase - 2. del: Ekstrakcijska metoda z zaplinjanjem (ISO 14240-2:1997)
Ta del IS0 14240 določa metodo za ocenjevanje mikrobne biomase v tleh z merjenjem skupne izločljive organske biomase, ki v glavnem prihaja iz sveže ubitih mikroorganizmov. Metoda se uporablja tudi za ocenjevanje mikrobnega dušika in mikrobnega ninhidronsko reaktivnega dušika v tleh, vendar ta del IS0 14240 opisuje le merjenje izločljivega organskega ogljika. Ekstrakcijska metoda z zaplinjanjem (FE) se uporablja za aerobna in anaerobna (prepojena, namočena z vodo) tla v celotnem razponu pH tal. Biomasa se lahko meri tudi v tleh, ki vsebujejo aktivno razpadajoče substrate, in tleh, prenasičenih z raztopino kalijevega sulfata.
General Information
Standards Content (Sample)
2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.Bodenbeschaffenheit - Bestimmung der mikrobiellen Biomasse von Böden - Teil 2: Fumigations-Extraktionsverfahren (ISO 14240-2:1997)Qualité du sol - Détermination de la biomasse microbienne du sol - Partie 2 : Méthode par fumigation-extraction (ISO 14240-2:1997)Soil quality - Determination of soil microbial biomass - Part 2: Fumigation-extraction method (ISO 14240-2:1997)13.080.30Biološke lastnosti talBiological properties of soilsICS:Ta slovenski standard je istoveten z:EN ISO 14240-2:2011SIST EN ISO 14240-2:2011en,fr,de01-november-2011SIST EN ISO 14240-2:2011SLOVENSKI
STANDARD
EUROPEAN STANDARD NORME EUROPÉENNE EUROPÄISCHE NORM
EN ISO 14240-2
June 2011 ICS 13.080.30 English Version
Soil quality - Determination of soil microbial biomass - Part 2: Fumigation-extraction method (ISO 14240-2:1997)
Qualité du sol - Détermination de la biomasse microbienne du sol - Partie 2 : Méthode par fumigation-extraction (ISO 14240-2:1997)
Bodenbeschaffenheit - Bestimmung der mikrobiellen Biomasse von Böden - Teil 2: Fumigations-Extraktionsverfahren (ISO 14240-2:1997) This European Standard was approved by CEN on 3 June 2011.
CEN members are bound to comply with the CEN/CENELEC Internal Regulations which stipulate the conditions for giving this European Standard the status of a national standard without any alteration. Up-to-date lists and bibliographical references concerning such national standards may be obtained on application to the CEN-CENELEC Management Centre or to any CEN member.
This European Standard exists in three official versions (English, French, German). A version in any other language made by translation under the responsibility of a CEN member into its own language and notified to the CEN-CENELEC Management Centre has the same status as the official versions.
CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia, Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway, Poland, Portugal, Romania, Slovakia, Slovenia, Spain, Sweden, Switzerland and United Kingdom.
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INTERNATIONAL STANDARD IS0 14240-2 First edition 1997-01-I 5 Soil quality - Determination of soil microbial biomass - Part 2: Fumigation-extraction method Qualit6 du sol - Dktermination de la biomasse microbienne du sol - Partie 2: Mkthode d/extraction par fumigation Reference number IS0 14240-2: 1997(E) SIST EN ISO 14240-2:2011
IS0 14240=2:1997(E) 0 IS0 Foreword IS0 (the llnternational Organization for Standardization) is a worldwide federation of national standards bodies (IS0 member bodies). The work of preparing International Standards is normally carried out through IS0 technical committees. Each member body interested in a subject for which a technical committee has been established has the right to be represented on that committee. International organizations, governmental and non-governmental, in liaison with ISO, also take part in the work. IS0 collaborates closely with the International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization. Draft International Standards adopted by the technical committees are circulated to the member bodies for voting. Publication as an International Standard requires approval by at least 75 % of the member bodies casting a vote. International Standard IS0 14240-2 was prepared by Technical Committee ISOmC 190, Soil quality, Subcommittee SC 4, Biological methods. IS0 14240 consists of the following parts, under the general title Soil quality - Determination of soil microbial” biomass: - Part 1: Substrate-induced respiration method Part 2: Fumigation-extraction method 0 IS0 1997 All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying and microfilm, without permission in writing from the publisher. International Organization for Standardization Case postale 56 l CH-1211 Geneve 20 l Switzerland Internet central @ iso.ch x.400 c=ch; a=400net; p=iso; o=isocs; s=central Printed in Switzerland ii SIST EN ISO 14240-2:2011
0 IS0 IS0 14240=2:1997(E) Introduction Soil consists of both living and nonliving components which exist in a complex and heterogeneous environment. Soil microflora is responsible for the degradation of organic matter, stability of aggregates and most nutrient cycling which occurs in soils. The purpose of determining the microbial biomass of soils is to allow assessment of the continued maintenance of soil fertility, the potential ability to degrade added organic materials, and the effects of added materials on the natural microbial population. SIST EN ISO 14240-2:2011
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INTERNATIONAL STANDARD o IS0 IS0 14240-2: 1997(E) Soil quality - Determination of soil microbial biomass - Part 2: Fumigation-extraction method 1 Scope This part of IS0 14240 specifies a method for the estimation of microbial biomass of soils by measurement of total extractable organic biomass material mainly from freshly killed microorganisms. The method is also applicable to the estimation of microbial nitrogen and microbial ninhydrin-reactive nitrogen in soil, but this part of IS0 14240 describes only the measurement of extractable organic carbon. The fumigation-extraction (FE) method is applicable to aerobic and anaerobic (water-logged, paddy) soils over the whole range of soil pH. Biomass can be also measured in soils containing actively decomposing substrates and soils supersaturated with potassium sulfate solution. NOTE - Chloroform fumigation also affects soil fauna. The contribution of carbon from these organisms is generally small (< 5 %) and can usually be neglected. 2 Normative references The following standards contain provisions which, through reference in this text, constitute provisions of this part of IS0 14240. At the time of publication, the editions indicated were valid. All standards are subject to revision, and parties to agreements based on this part of IS0 14240 are encouraged to investigate the possibility of applying fhe most recent editions of the standards indicated below. Members of IEC and IS0 maintain registers of currently valid International Standards. IS0 10381-6:1993, Soil quality - Sampling - Part 6: Guidance on the collection, handling and storage of soil for the assessment of aerobic microbial processes in the laboratorym IS0 10694:1995, Soil quality - analysis). Determination of organic and total carbon after dry combustion (elementary IS0 11465:1993, Soil quality - Determination of dry matter and water content on a mass basis - Gravimetric method. 3 Definition For the purposes of this part of IS0 14240, the following definition applies. 3.1 soil microbial biomass mass of intact microbial cells in a given soil NOTE - This parameter can be estimated from the measurement of the carbon or nitrogen content of these cells or by the measurement of their ability to mineralize an added carbon source. Dead cells and cell fragments may be detected when carbon or nitrogen analysis is used but only intact cells will be detected when respiration is measured. SIST EN ISO 14240-2:2011
IS0 14240-2: 1997(E) 0 IS0 4 Principle Through fumigation of the soil sample, intact microbial cells are lysed and the microbial organic matter released. Nonliving soil organic matter is not seriously affected by such fumigation. Soil samples are fumigated with chloroform for 24 h. The organic carbon extracted using 05 mol/l potassium sulfate is determined in fumigated and unfumigated samples, and the difference in extracted organic carbon is used to determine microbial biomass carbon. The method is usually termed fumigation-extraction (FE). 5 Materials and reagents 5.6 Soils Guidance for the collection, handling and storage of soil (IS0 10381-6) shall be followed, as far as applicable. Sieve the soil samples at approximately 40 % water-holding capacity (WHC) if homogeneous samples are required. Determine the soil WHC in accordance with annex A. The water content of samples shall be higher than 30 % WHC to ensure uniform chloroform distribution and effective fumigation. Take care to avoid smearing and compaction of wet soil in this method. Samples of waterlogged soils do not have to be dried prior to analysis. 5.2 Reagents Reagents of recognized analytical grade shall be used, including: 5.2.1 Silicone grease (medium viscosity). 5.2.2 Ethanol-free chloroform. WARNING - In the presence odourless and highly toxic. of light, ethanol-free chloroform degrades rapidly to form phosgene 5.2.3 Potassium sulfate solution, c(K,SO,) = 05 mol/l (p = 87,135 g/l)= 5.2.4 Soda lime. 6 Apparatus 6.1 Room, or incubator, capable of being maintained at (25 * 2) “C. 6.2 Implosion-protected desiccator. 6.3 Filter paper’). 6.4 Glass beakers. 6.5 Petri dishes. 6.6 Polyethylene bottles, of capacity 250 ml. is 1) Whatman No. 42, Schleicher & Schijll 595 l/2, Machery & Nagel 261 G l/4, are examples of suitable products available commercially. This information is given for the convenience of users of this part of IS0 14240 and does not constitute an endorsement by IS0 of these products. 2 SIST EN ISO 14240-2:2011
0 IS0 IS0 14240-2: 1997(E) 6.7 Vacuum line (water-pump or electric pump). 6.8 Horizontal or overhead shaker. 6.9 Freezer, operation at (-15 to -20) “C. 6.10 Antibumping granules. 7 Fumigation and extraction 7.1 Fumigation. Line the desiccator (6.2) with moist filter paper (6.3) for fumigation of the soil samples. Weigh at least three samples of moist soil (5.1), each containing a mass equivalent to 25 to 50 g of oven-dry soil, into glass beakers (6.4) or Petri dishes (6.5) and place them in the desiccator with a beaker containing 25 ml of ethanol-free chloroform (5.2.2) and a few antibumping granules (6.10), and a beaker with soda lime (5.2.4). Evacuate the desiccator until the chloroform has boiled vigorously for approximately 2 min. Close the vacuum tap on the desiccator and incubate in the dark (6.1) at (25 t 2) “C for 22 h to 24 h. If there is insufficient soil available, use smaller samples provided the ratio of soil mass to volume of extractant is kept the same (1:4). In soils containing more than 20 % organic material (as determined according to IS0 10694), increase the ratio soil:extract above I:4 (to a maximum of I:30 for soils containing 95 % organic matter, e.g. L-layer) in order to obtain sufficient extracted matter. Record the mass of soil used. After fumigation is completed, remove the beaker containing the chloroform and the filter paper from the desiccators. Remove the chloroform vapour from the soil by repeated evacuation (6 times for 2 min each) in the desiccator. The samples are ready for extraction. Place three unfumigated, moist control samples (50 g dry mass) into polyethylene bottles (6.6) and immediately extract with 200 ml of the potassium sulfate (5.2.3), as indicated in 7.2. 7.2 Extraction To extract organic carbon, transfer the soil quantitatively to polyethylene bottles (6.6), add 200 ml of potassium sulfate (5.2.3), shake the bottles on a horizontal shaker (6.8) at 200 r/min for 30 min or an overhead shaker at 60 r/min for 45 min and filter the extracts through a folded filter paper (6.3). Extract the unfumigated controls and filter the extracts in the same way. If not analyzed at once, store the extracts of fumigated and unfumigated soil samples in the freezer (6.9) at between -15 “C and -20 “C. Homogenize frozen extracts before use, after thawing at room temperature. NOTES 1 A white precipitate occurs during storage of the potassium sulfate soil extracts (especially if the samples are frozen) because they are usually supersaturated with calcium sulfate (CaSO,). It is unnecessary to dissolve this excess CaSO, because it does not interfere with any of the analytical procedures described in this method. 2 Cell membranes of young, living roots are also affected by chloroform fumigation. In soils containing large amounts of living roots, the pre-extraction procedure given in annex B should be used. 8 Determination of carbon in the extracts The carbon content of microorganisms in a soil sample is determined analytically and can be used to make comparisons between different soil samples. Where a figure for actual microbial biomass is required, then such analyses are multiplied by a conversion factor derived from experiments correlating a known cell mass to carbon analysis after fumigation-extraction. All conversion factors used are related to this initial factor. 3 SIST EN ISO 14240-2:2011
IS0 14240-2: 1997(E) 0 IS0 Determine the carbon content in the extracts using either dichromate oxidation (8.1) or instrumental analysis (8.2). 8.1
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