ISO/TS 29843-2:2021 土壌の質 — 土壌微生物の多様性の決定 — Part 2: 単純なPLFA抽出法を使用したリン脂質脂肪酸分析（PLFA）による方法 | ページ 2

Foreword

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This document was prepared by Technical Committee ISO/TC 190, Soil quality, Subcommittee SC 4, Biological characterization,in collaboration with the European Committee for Standardization (CEN) Technical Committee CEN/TC 444, Environmental characterization of solid matrices,in accordance with the Agreement on technical cooperation between ISO and CEN (Vienna Agreement).

This second edition cancels and replaces the first edition (ISO/TS 29843-2:2011), which has been technically revised.

The main changes compared to the previous edition are as follows:

• addition of specification for qualitative and quantitative analysis of PLFAs;
• use of BAME (qualitative) or FAME (quantitative) standards;
• use of GC-MS apparatus;
• precisions in 7.2 and 7.3;
• possibility to use commercial cartridges in addition to, or replacement of, home-made cartridges;
• update of bibliographic references.

A list of all the parts in the ISO/TS 29843 series can be found on the ISO website.

Introduction

Phospholipids are essential components of membranes of all living cells. Extracted from soil samples in fatty acid form (phospholipid fatty acids, PLFA) or ether-linked isoprenoid side chains (phospholipid ether lipid, PLEL), they provide quantitative and qualitative insights into the soil's viable/active microbial biomass. Cellular enzymes hydrolyse and release the phosphate group within minutes to hours following cell death^[1]. The determination of total PLFA and PLEL provides a quantitative measure of the viable biomass of soil, i.e. microorganisms of all three primary domains of the biosphere (bacteria, archaea and microeukaryota). PLFA and PLEL can also allow for rough taxonomic differentiation within complex microbial communities^[2],[3]. Each microbial species contains several fatty acids, with a total composition in PLFA subject to the environmental conditions^[4]. The approach is performed to evaluate biomass and shifts in microbial community composition^[5], in what regards dominance of main groups of organisms^[6]. Furthermore, combined with the use of isotope (¹³C or ¹⁴C) labelled substrates, the lipid methods can also be used to identify the metabolically active part of the microbial community. This approach is well established in soil ecology and serves as a phenotypic, and thus complementary, tool to genotyping approaches for determining microbial diversity. Apart from taxonomic descriptions, the PLFA technique enables the determination of physiological changes within microbial consortia. For instance, the monoenic PLFA 16:1ω7c and 18:1ω7c are increasingly converted to the cyclopropyl fatty acids cy17:0 and cy19:0 in Gram-negative bacteria in response to environmental stress^[7].

Different methodologies are available for the determination of soil fatty acids. These methodologies present different levels of complexity when applied and provide different levels of resolution in the description of soil microbial communities. ISO/TS 29843-1 deals with the generally called “extended PLFA extraction method” while this document deals with the generally called “simple PLFA extraction method”^[8],[9].

This document is accessible to most research and analytical laboratories involved in soil sciences. This methodology can be used for a wide range of soils. It provides a broad diversity measurement of a soil microbial community at the phenotypic level. It can be applied to biomass estimation and can be used to differentiate microbial communities among different soil samples (with the aid of an adapted statistical method). This method is especially adapted for detecting rapid changes in the soil microbial community structure. It can also be used to give a rough description of microbial groups present in soil samples (e.g. Gram-positive bacteria, actinomycetes, fungi^[6]). Table 1 (adapted from Table 1 in Reference [8]), presents a comparison of the sensitivity of the “extended PLFA” versus “simple PLFA” techniques.

1 Scope

This document specifies a simple method for the extraction of only phospholipid fatty acids (PLFA) from soils.

2 Normative references

The following documents are referred to in the text in such a way that some or all of their content constitutes requirements of this document. For dated references, only the edition cited applies. For undated references, the latest edition of the referenced document (including any amendments) applies.

3 Terms and definitions

No terms and definitions are listed in this document.

ISO and IEC maintain terminology databases for use in standardization at the following addresses:

Bibliography