• People
• Research
• Publications
• Dissertations
• Posters
• Facilities
• Analysis
• History
• Data
• Photos
• Links
• Contacts

• Definitions
• Reference Materials
• Internal

Reference Materials: We use nitrates USGS35, USGS34, and IAEA-NO-3 as our international standards for this method.

Sample submission: We prefer 20 nmoles of nitrate / nitrite (minimum 5 nmoles) to complete the δ¹⁵N and δ¹⁸O analysis. We prefer 200 nmoles of nitrate / nitrite (minimum 10 nmoles) to complete the Δ¹⁷O analysis. It is most efficient to have the nitrate / nitrite concentration prior to isotope analysis. Please have your samples filtered and frozen in a plastic container capable of freezing. Please include in your shipment container the following form (pdf or doc).

Exhaustive description of analysis: Pseudomonas aureofaciens denitrifying bacteria are grown from freezer stocks on agar petri plates and used to inoculate 500 mL of trypic soy broth growth media. After 6-8 days bacterial cells are harvested by centrifugation at 8000 rpm and 18 °C. Bacterial cell pellets are resuspended in nitrate free media and divided into 22 20 mL crimp-seal vials. Vials are crimped and purged with helium for 4-5 hours to purge the vial completely of oxygen and to allow the cells to process any remaining nitrate to nitrous oxide. A volume of sample and standard is injected to yield a total of 20 nmoles of nitrate for direct analysis of N₂O or 200 nmoles of nitrate for N₂O pyrolysis. Vials are inverted and placed on a reciprical shaker overnight. After at least 8 hours, 0.2 mL of 10 M NaOH is injected into each vial to lyse the cells and several drops of Antifoam are added. Vials are then loaded into a GC PAL autosampler. Each vial is purged with helium, forcing the effluent through a cold water trap (-60 °C), a carbon dioxide, water chemical trap (Ascarite, Magneiusm perchlorate), a volatile organic compound trap, and a liquid nitrogen trap, before a vent. All sample / standard N₂O remains in the liquid nitrogen trap and is moved into a secondary focusing liquid nitrogen trap. A dochotomy exists from here.

Intact N₂O for δ¹⁵N and δ¹⁸O - If samples are to be analyzed for δ¹⁵N and δ¹⁸O only, the cryofocused N₂O moves through a porapaq Q gas chromatography column for separation with any remaining CO₂ and then into a Thermo Finnigan Delta Plus for mass / charge 44, 45, and 46 measurement.

Pyrolyzed N₂O for δ¹⁵N, δ¹⁷O and δ¹⁸O - If samples are to be analyzed for δ¹⁵N, δ¹⁷O and δ¹⁸O, the cryofocused N₂O moves through a gold tube held at 800 °C which pyrolyzes the N₂O into O₂ and N₂. These two species are then separated in a 5A molecular sieve gas chromatography column and then sent into the isotope ratio mass spectrometer for mass / charge 32, 33, 34 and then 28, 29 measurement.

Calculations: Intact N₂O for δ¹⁵N and δ¹⁸O - Raw δ¹⁵N values are corrected to the Air-N2 scale by generating a linear curve from measured versus accepted USGS34 and IAEANO3 international reference materials with values of -1.8 ‰ and +4.76 permil, respectively. The IsoDat software assumes all oxygen measurements are mass dependent. Because USGS35 exhibits mass independent fractionation between O¹⁷ and O¹⁸, δ values must be recalculated with an appropriate alpha value. USGS35 δ¹⁵N values are then corrected to the USGS34 and IAEANO3 curve and used as a quality control reference. This method assumes all samples have a Δ¹⁷O equal to 0.

Pyrolyzed N₂O for δ¹⁵N, δ¹⁷O and δ¹⁸O - Raw δ¹⁵N values are corrected to the Air-N2 scale as in the intact N₂O method above without any corrections concerning mass dependence because all of the oxygen istopes are measured directly when N₂O is pyrolyzed. δ¹⁷O and δ¹⁸O raw values are corrected to VSMOW by generating a linear curve from measured versus accepted USGS35 and USGS34 international reference materials with values 51.1 ‰ and -14.6 ‰ for δ¹⁷O and +56.81 ‰ and -27.78 ‰ for δ¹⁸O, respectively (using CIAAW δ¹⁸O).

Suggested reading: