Hinrichs Lab - Isotope Ratio Mass Spectrometry (IRMS)
Introduction
Two dedicated on-line IRMS systems are available in our group: (1) a newly acquired IRMS system (Delta V Plus) equipped with the GC Isolink interface (GC-IRMS) and an elemental analyzer (EA-IRMS) and (2) an older system (Delta Plus XP) used either in connection with a GC combustion interface III or the liquid chromatography (LC) Isolink interface (LC-IRMS). The former system enables the determination of both C and H isotopes which are transformed after GC separation either via oxidation or high temperature pyrolysis to CO2 or H2, respectively. This new system can be also switched over to an EA thus giving the possibility to determine C and N isotope compositions of bulk organic matter and enriched or pure organic compounds. The latter IRMS system is dominantly used for the determination of C and H isotopes of natural gaseous compounds in the C1 to C6 range, e.g. methane, ethane, dimethyl sulfide, methyl amine etc., but is routinely switched over to the LC Isolink interface for carbon isotopic measurements of VFAs and other water-soluble organic compounds.
Gas chromatography coupled to isotope ratio mass spectrometry (GC-IRMS)
GC-IRMS is a highly specialized instrumental technique and is used in our lab to determine the relative ratio of light stable isotopes of hydrogen (2H/1H) and carbon (13C/12C) in individual compounds of complex sample fractions or mixtures. The stable isotopic composition of organic molecules in natural materials (sediment, soil, cell biomass etc.) varies slightly as a result of isotopic fractionation during physical, chemical and biological processes. These processes are controlled by kinetic or equillibrium isotope effects (KIE or EIE, respectively) with KIEs dominanting in biogeochemical research. In some cases, the relative isotopic ratio of specific compounds is highly diagnostic of key environmental processes (e.g. photosynthesis or methanogenesis). By the use of growth substrates artificially enriched in the heavier isotope and its uptake into living biomass, IRMS determinations aid in the deconvolution highly complex and often enigmatic biogeochemical pathways.
The primary prerequisite for GC-IRMS is that the compounds constituting the sample mixture are amenable to GC, i.e. they are suitably volatile and thermally stable. Polar compounds may require further chemical modification (derivatization) and in such cases the relative stable isotope ratio of the derivatization agent must also be determined. Just as for other GC techniques the sample solution is injected via the GC injector and separated on the capillary column. Single organic compounds eluting from the column then pass through an oxidizing combustion reactor fitted with an alumina tube maintained at 960ºC and containing Cu, Ni and Pt wires, resulting in the formation of CO2 and H2O. Water is then removed by passing the gas stream through a tube fitted with a water permeable Nafion membrane. For hydrogen isotope measurements, instead of combustion a high temperature thermal conversion reactor tube (open alumina) is operated at 1440°C, resulting in the formation of CO, elemental C and H2. The final analyte gas stream (CO2 or H2) is introduced into the ion source of the IRMS and ionization is achieved using electron ionisation (EI) at 150 eV. Ionized gas isotopes are separated in a single magnetic sector analyzer and detected by Faraday cups. Stable isotope ratios are calculated relative to standards of known isotopic composition and expressed using the dimensionless "per mil" notation against the Vienna Pee Dee Belemnite (VPDB) standard for carbon or Vienna Standard Mean Ocean Water (VSMOW) for hydrogen.
Elemental Analyzer coupled to isotope ratio mass spectrometry (EA-IRMS)
The homogenized samples are weighed into tin capsules, tightly closed and introduced into the EA via an autosampler. The samples are combusted at 999°C in the oxidation oven containing chromium and other metal oxides. This leads to a conversion of sample organic matter into CO2, H2O and N2/NOx. NOx components are subsequently reduced to N2 with elemental copper in the reduction oven at 680°C. Water is eliminated using a chemical trap filled with magnesium perchlorate and target gases CO2 and N2 are separated from each other on a gas chromatographic column. Before entering the IRMS ion source, gas peak areas are automatically adjusted in the Conflo IV interface by helium dilution. Sample's preliminary isotope ratios are measured relative to reference gas pulses (CO2, N2) analyzed during each run with precision routinely checked by standard reference materials (i.e., IAEA-CH-6 and IAEA-N-2). In order to achive C and N concentrations, a daily calibration with a large-volume sediment laboratory standard is performed.
Liquid chromatography coupled to isotope ratio mass spectrometry (LC-IRMS)
The LC-IRMS technique opens a new field of carbon isotopic studies in biogeochemistry with target compounds of interest including for example volatile fatty acids (VFAs), amino acids, amino sugars, carbohydrates, and nucleotides. By appling this technique, we developed and validated a reversed-phase LC method that is suitable for highly-precise carbon isotope analysis of VFAs from sedimentary pore waters and other aqueous solutions (Heuer et al., 2006).
Thermo Scientific Delta V Plus with GC Isolink and EA
Specifications:
Sensitivity (continuous flow mode): 1100 molecules CO2 per mass 44 ion
Isotope Ratio Linearity: 0.02 %/nA ion current (m/z 44)
Mass Range: 1-96 Daltons at 3 kV
Mass Resolution: m/Δm = 110 (10% valley)
System Stability: <10 ppm
H3+ Factor: <10 ppm/nA, stability <0.03 ppm/nA/h
Thermo Finnigan Delta Plus XP with Trace GC and LC Isolink
Specifications:
Sensitivity (continuous flow mode): 1500 molecules CO2 per mass 44 ion
Isotope Ratio Linearity: 0.02 %/nA ion current (m/z 44)
Mass Range: 1-70 Daltons at 3 kV
Mass Resolution: CNOS: m/Δm = 95 (10% valley), H/D: m/Δm = 10 (10% valley)
System Stability: <10 ppm
H3+ Factor: <10 ppm/nA, stability <0.03 ppm/nA/h