7. Stable Isotope Probing (SIP)

7.1 Summary of SIP

The stable isotope probing (SIP)A synthesized form of the contaminant containing a stable isotope (e.g., ¹³C label) is added. If biodegradation is occurring the isotope will be detected in biomolecules (e.g., phospholipids, DNA). method includes a family of techniques that all use individual contaminants (probes) that are enriched with stable isotopesForms of an element that do not undergo radioactive decay at a measureable rate. (such as 13C-labeled benzene or 15N-labeled RDX) to characterize contaminant-specific biodegradationA process by which microorganisms transform or alter (through metabolic or enzymatic action) the structure of chemicals introduced into the environment (USEPA 2011). processes. The underlying principle of all SIP techniques is that biodegradation of isotopically-enriched contaminants results in the selective labeling of biomoleculesClasses of compounds produced by or inherent to living cells including phospholipids, nucleic acids (e.g., DNA, RNA), and proteins. such as DNA, RNA, or phospholipidA type of biomolecule that is a primary structural component of the membranes of almost all cells. fatty acids (resulting in 13C- or 15N-lableled DNA or 13C-labeled fatty acids) in organisms that are responsible for degradation. A flow diagram showing the basic protocol for SIP studies is provided in Figure 7-1.

Additional detailed information about the types of SIP analyses and their methodologies is available in the SIP Fact Sheet and in Section 7.4

7.2 Applications

Most current applications of SIP technique make use of contaminants that have enriched levels of 13C. 13C-based SIP techniques can potentially be applied to any type of environmental sample and can also be used to examine the biodegradation of any compound that is used by microorganisms as a sole (or predominant) source of carbon for growth. However, in current practice, SIP is used for two main purposes; (a) demonstrating that a specific contaminant undergoes biodegradation in a specific environment (PLFAPhospholipid fatty acids derived from the two hydrocarbon tails of phospholipids.-SIP) and (b) identifying organisms involved in specific biodegradation processes (DNA/RNA-SIP).

7.3 Data Interpretation

The application, analysis, and interpretation of microbiology-based EMD methods differ from typical soil and groundwater geochemical measurement in a number of ways. For example, microbial biomarkers cannot easily be preserved, and sample handling and processing requires special care. The analysis of microbial parameters requires specific data quality considerations for sampling plans, sample collection and handling, quality control and laboratory procedures, and these are discussed in Section 10. Included below is a brief introduction to how SIP data are typically reported and some specific examples of how the data would be interpreted in answering the questions presented in Table 2-3.

7.3.1 How are the data reported?

Included in Table 7-2 below is information that should be provided in laboratory reports of SIP data including common laboratory report information, recommended information about the SIP method, and desirable information about the SIP method and results.

Additional information regarding sample handling and collection can be found Section 10.4 and Section 10.5

7.3.2 How are the data interpreted?

For the most part, SIP techniques are not inherently quantitative and their primary use in contaminant biodegradation studies is to demonstrate whether the contaminant biodegrades under a given set of environmental conditions. More elaborate studies can refine this information and identify which organisms are responsible for any observed biodegradation activity. To illustrate interpretation of SIP results, each question relevant to SIP in Table 2.3 is discussed.

7.3.3 Practical considerations

7.4 Additional Information

7.4.1 Method Details

7.4.2 Permitting and Regulatory Issues Specific to SIP

7.4.3 Variations or newer methods that the user may encounter

7.4.4 Additional Information

Further reading specific to SIP is provided in Appendix F.

Publication Date: April 2013

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