1. Delta Units (δ): are expressed in molecules per thousand (‰), or “per mil”. For example, δ 15 N Air = 12 per mil means that the sample was analyzed against a reference material and found to be 12 molecules per thousand more abundant than in air – the accepted zero point for expression of nitrogen-15 in per mil notation. For the calculation: δ = [ (R s / R r ) - 1 ] * 1000 where R s is the ratio of the heavy isotope to the light isotope of the sample and R r is the ratio of the heavy isotope to the light isotope of the reference. Expression of Stable Isotope Values – Delta Notation:

2. Isotope Ratio Mass Spectrometer – “IRMS” A mass spectrometer is an instrument which separates charged molecules by mass. An isotope ratio mass spectrometer (IRMS) works on this principle, but unlike other conventional mass spectrometers it has been specifically designed to measure the proportions of particular isotopes. An IRMS will be much more precise, but much less sensitive than other mass spectrometers. Ionisation bending magnet

3. Principle of a Quadrupole Mass Spectrometer A quadrupole has four parallel rods that have fixed DC and alternating RF potentials applied to them. Ions produced in the source of the instrument are then focused and passed along the middle of the quadrupoles. For mass separation, quadrupole systems use a high-frequency alternating electrical field. A Faraday cup and a secondary electron multiplier serve as detectors. Commonly used in conjunction with either gas-chromatography or liquid-chromatography, and more recently with ICP, as a simple high throughput screening system.

4. Element Isotopes Hydrogen 1 H, 2 H Boron 10 B, 11 B Carbon 12 C, 13 C Nitrogen 14 N, 15 N Oxygen 16 O, 17 O, 18 O Sulfur 32 S, 33 S, 34 S, 36 S The most widely studied stable isotopes are: α is typically calculated in K (degrees Kelvin) which is equal to o C + 273. α is the isotope fractionation factor, (e.g. α for water liquid and vapor is 10‰ i.e., the δ 18 O of the liquid is 10‰ heavier). Rayleigh Effects two species (or phases as in water) that are in equilibrium with one another, or where the reactants and products become separated from one another Kinetic Effects kinetic effects are most commonly seen in processes that are influenced by biologic processes, during unidirectional processes Diffusion the light isotope of an element will diffuse more rapidly than the heavy isotope Stable Isotopes – Mass Dependant Fractionation

5. Natural log notation, 1000 ln (α): The epsilon notation, ε : The epsilon notation has the advantage over the 1000ln(α) notation in that it is an exact expression of the per mil fractionation. α is the isotope fractionation factor, (e.g. α for water liquid and vapor is 10 ‰, i.e., the δ 18 O of the liquid is 10‰ heavier). 1000ln(α) – 1000 ln (1- ε ) = ε The capital delta notation, Δ : The size of this isotopic fractionation can be expressed in several ways: Δ A-B = δ A - δ B ε A-B = ( α A-B – 1) * 1000

6. Some General “Rules” Isotope fractionation factors are greater at lower temperatures. Light isotopes are enriched in biogenic compounds. Light isotopes are enriched in reduced species and heavy isotopes are enriched in oxidized species (e.g., the δ 13 C of CO 2 is higher than that of CH 4 ). It follows from this that reactions which involve a change in oxidation state result in a greater degree of stable isotope fractionation than those that do not. Where two minerals of the same oxidation state are in equilibrium with one another, the mineral with the heaviest cation will have the lightest stable isotope composition (e.g., the δ 34 S of ZnS (sphalerite) is higher than that of PbS (galena)). The extent of stable isotope fractionation is inversely proportion to the square of the relative mass difference between two isotopes. This means that the extent of stable isotope fractionation between 100 Ru and 101 Ru is less than 1% of that between 10 B and 11 B. In practice this has meant that stable isotope fractionation is effectively below detection limits for elements with masses greater than 40 (i.e., for elements with masses greater than that of Ca). Recent advances in mass spectrometry are increasing the range of elements for which stable isotope variations can be detected.

7. Oxygen isotopes of meteoric water are generally lighter than those of seawater (Hoefs, 1980, fig. 10). δ 18 O (‰ VSMOW) 40-70-60-50-40-30-20-100102030 extraterrestrial materials (meteorites and lunar rocks) basaltic rocks granitic rocks metamorphic rocks sedimentary rocks ocean water – VSMOW = 0 ‰ meteoric waters Ranges of O Isotopic Values in Geologic Systems

8. Rainout effect on δ 2 H and δ 18 O values; ( based on Hoefs 1997 and Coplen et al. 2000 ). Change in the 18 O content of rainfall according to a Rayleigh distillation, starting with δ 18 O vapor = -11‰, temp. = 25°C, and final temp. of -30°C. Note that at 0°C, fractionation between snow and water vapor replaces rain- vapor fractionation. The fraction remaining has been calculated from the decrease in moisture carrying capacity of air at lower temperatures, starting at 25°C. Dashed lines link δ 18 O of precipitation with temperature of condensation. (Reproduced from Clark and Fritz 1997, p.48)

9. http://www.sahra.arizona.edu/programs/isotopes/images/diagram7.gif (Clark and Fritz 1997, p. 37, as compiled in Rozanski et al. 1993, modified by permission of American Geophysical Union). W Meteoric Water Line - MWL Ocean Water

10. The isotopic composition of VSMOW water is specified as ratios of the molar abundance of the rare isotope in question divided by that of its most common isotope and is expressed as parts per million (ppm). For instance 16 O (the most common isotope of oxygen with eight protons and eight neutrons) is roughly 2,632 times more prevalent in sea water than is 17 O (with an additional neutron). The isotopic ratios of VSMOW water are defined as follows: 2 H/ 1 H = 155.76 ±0.1 ppm (a ratio of 1 part per approximately 6420 parts) 3 H/ 1 H = 1.85 ±0.36 × 10 −11 ppm (a ratio of 1 part per approximately 5.41 × 10 16 parts, ignored for physical properties-related work) 18 O/ 16 O = 2,005.20 ±0.43 ppm (a ratio of 1 part per approximately 498.7 parts) 17 O/ 16 O = 379.9 ±1.6 ppm (a ratio of 1 part per approximately 2,632 parts) Vienna Standard Mean Ocean Water (VSMOW) is a water standard defining the isotopic composition of water. It was promulgated by the International Atomic Energy Agency in 1968. Despite the misleading designation ocean water, VSMOW does not include any salt or other substances usually found in seawater and refers to pure water with a particular composition of isotopes. VSMOW serves as a reference standard for comparing hydrogen and oxygen isotope ratios, mostly in water samples. Very pure, distilled VSMOW water is also used for making high accuracy measurement of water's physical properties and for defining laboratory standards since it is considered to be representative of average ocean water, in effect representing all water on Earth. http://www.sahra.arizona.edu/programs/isotopes/images/diagram7.gif 0‰

11. Variation in the closely spaced rings of a bristlecone pine correspond to annual changes in rainfall and temperature. ( Photograph copyright Henri D. Grissino-Mayer ) Annual Climate Records: An ice core with a layer of algae. A Greenland ice core with a layer of algae. global-warming.accuweather.com/ice_core_algae Water vapor gradually loses 18 O as it travels from the equator to the poles. Because water molecules with heavy 18 O isotopes in them condense more easily than normal water molecules, air becomes progressively depleted in 18 O as it travels to high latitudes and becomes colder and drier. In turn, the snow that forms most glacial ice is also depleted in 18 O. As glacial ice melts, it returns 16 O-rich fresh water to the ocean. Therefore, oxygen isotopes preserved in ocean sediments provide evidence for past ice ages and records of salinity. ( Illustration by Robert Simmon, NASA GSFC based on data provided by Cole et. al. 2000, archived at the World Data Center for Paleoclimatology ). http://earthobservatory.nasa.gov/Features/Paleoclimatology_Evidence/ sclectarian corals

12. From Gercke, in preparation

13. Perissocytheridea sp. Cyprideis sp. δ 18 O Age y.b.p. From Gercke, in preparation

14. http://esp.cr.usgs.gov/info/eolian/task2.html Diagram showing the nature of the loess stratigraphic record. In most regions, including much of North America, Europe, and China, loess was deposited during glacial periods and soils were formed during interglacial periods. Soils that become buried by younger loess are called "paleosols." Large scale Climate Records:

15. http://earthobservatory.nasa.gov/Features/Paleoclimatology_Evidence/ Global Records:

16. Shriner, C.M., Elswick, E.R., Ripley, E.M., Shimmelmann, A., and Murray, H.H., (in press) Natural Environment as a Determinative Factor in Greek Early Helladic Cultural Change on the Argive Plain: in Katsonopoulou, D. Ed., The Early Helladic Peloponnesos, Helike IV; The Heike Society; Athens, Greece.

18. Carbon isotopes in geologic systems. Carbonate carbon derived from seawater is much heavier than organic carbon, and hence than carbonate formed by oxidation of organic matter (Hoefs, 1980, fig. 9; Trumbore and Druffel, 1995). δ1 3 C (‰ VPDB) 4050-50-40-30-20-10010203060 marine organic C extraterrestrial materials (meteorites and lunar rocks) atmospheric CO 2 marine carbonates freshwater carbonates carbonatites, diamonds marine and non-marine organisms sedimentary organic matter, petroleum, coal soil CO 2 biogenic methane volcanic CO 2 soil organic C land plants freshwater Σ CO 2 shallow ocean Σ CO 2 deep ocean Σ CO 2 Ranges of C Isotopic Values in Geologic Systems

19. Diagrammatic Representation of the Exogenic Cycles of Carbon and Sulfur S - CycleC - Cycle Car Carbonate Carbon 6460 x 10 18 moles δ 13 C = -0.4 Organic Carbon 1180 x 10 18 moles δ 13 C = -27 Gypsum 166 x 10 18 moles δ 34 S= +8.4 Pyrite 180 x 10 18 moles δ 34 S= -7.9 Carbonate, DIC 3.3 x 10 18 moles δ 13 C = +0.46 SW sulfate 40 x 10 18 moles δ 34 S= +20 Ocean Gregor et al.. 1988

20. http://homepage.mac.com/uriarte/carbon13.html Most plants (85%) (e.g. trees and crops) follow the C 3 photosynthesis pathway and have lower values of δ 13 C, between -22‰ and -30‰. The remaining 15% of the plants are of type C 4. The majority are tropical herbs and have high values of δ 13 C, between –10 ‰ and –14 ‰. The denominations are because in the plants of group C 3, the first photosynthesized organic compound has 3 atoms of carbon while in group C 4, there are 4. (There is also a third, very minor, group called CAM, a combination of C 3 and C 4 where some cactus and succulents belong to.)

21. Carbon-13 ---------------------C 3 and C 4 plants The isotopes of a chemical element are the various configurations of its atoms. There are three carbon isotopes in nature: 12 C, 13 C and 14 C. These are three varieties of the same chemical element, carbon, whose nuclei contain the same number of protons (six), but a different number of neutrons (six, seven and eight, respectively). Thus, besides having the same chemical properties, the isotopes have different atomic masses: twelve, thirteen and fourteen, respectively. Almost 99% of atmospheric CO 2, contains the less heavy carbon, 12 C. A small part, 1.1% of CO 2, is somewhat heavier, since it contains 13 C. Terrestrial vegetation and marine phytoplankton, in the process of photosynthetic absorption of CO 2, discriminate against heavy molecules preferring 12 C to 13 C. In this way, the carbon trapped in continental flora contains a smaller proportion of 13 C than the carbon in atmospheric CO 2. -30 -20 -10 C3C3 C4C4 CAM δ 13 C (‰ VPDB) frequency

22. An illustration of the stomatal CO 2 proxy. (Left) Photomicrograph of fossil leaf cuticle of the fern aff. Stenochlaena from just after the Cretaceous/Tertiary (K/T) boundary. (Right) The fern's nearest living relative, Stenochlaena palustris. The stomatal index of the fossil cuticle is considerably lower than the extant cuticle, indicating that CO 2 was higher directly after the K/T boundary than today (21). Photos courtesy of Barry Lomax (University of Sheffield, Sheffield, U.K.). (Scale bars, 10 μm.) www.pnas.org/content/105/2/407/F2.large.jpg www.pnas.org/content/105/2/407/F2.large.jpg C 3 – plants The isotopic signature of C 3 plants shows higher degree of 13 C depletion than the C 4 plants. Examples of C 3 plants include wheat, rice, soybeans, maples.

23. The formula for calculating δ 13 C (in ‰) is as follows: ( 13 C/ 12 C)sampled – ( 13 C/ 12 C)standard ——————————––––––––––––––– x 1.000 ( 13 C/ 12 C)standard C 4 – plants Over 8000 species of angiosperms have developed adaptations which minimize the losses to photorespiration developed in the Oligocene (25-32 mya) and became ecologically important in the Miocene (5-12 mya). These C 4 plants are well adapted to (and likely to be found in) habitats with high daytime temperatures and intense sunlight, as well as potential nitrogen or CO 2 limitations. Some examples: crabgrass, corn (maize), sugarcane, sorghum Although only ~3% of the angiosperms, C 4 plants are responsible for ~25% of all the photosynthesis on land.

24. Increase in δ 13 C from palaeosols and tooth enamel showing apparent synchronicity in the transition to C 4 - dominated terrestrial ecosystems across continents. Data for (a) from Cerling et al. (1997), (b) from Quade & Cerling (1995) and (c) from Passey et al. (2002). wetter  dryer

25. www.illinoistimes.com www.enviroone.com/images/crops/corn.jpg Present-day C 4 plants are concentrated in the tropics (below latitudes of 45°) where the high air temperature contributes to higher possible levels of oxygenase activity by rubisco. Monocot examples: Forty-six percent of grasses are C 4 and together account for 61% of C 4 species. Dicot examples: Members of the sedge family Cyperaceae, and numerous families of Eudicots, including the daisies Asteraceae, cabbages Brassicaceae, and spurges Euphorbiaceae also use C 4.

26. The common reference for delta 13 C Marine Carbonate Standard was obtained from a Cretaceous marine fossil, Belemnitella americana, from the PeeDee Formation in South Carolina. This material has a higher 13 C/ 12 C ratio than nearly all other natural carbon-based substances. For convenience it is assigned a delta 13 C value of zero, giving almost all other naturally-occurring samples a positive delta value. The original sample was used up long ago, but the IAEA calibrated a new reference sample to the original fossil, giving rise to the widespread use of the term Vienna- PeeDee Belemnite standard, abbreviated to V-PDB. Vienna- PeeDee Belemnite standard -- V-PDB

27. Bullet-shaped fossils called belemnites occur commonly in rocks of Jurassic and Cretaceous age (65-205 million years old). They can be found weathered out of clays and chalks in great abundance at some localities. These examples are from the Pee Dee Formation, Florence County, South Carolina. ucmp.berkeley.edu www.ucmp.berkeley.edu/.../belemnite_anatomy.gif www.nhm.ac.uk/.../fossil_types/belemnites.htm www.blackriverfossils.org Pee Dee Formation, Florence County, SC

28. δ 34 S (‰ VCDT) 4050-50-40-30-20-10010203060 extraterrestrial materials (meteorites and lunar rocks) granitic rocks metamorphic rocks sedimentary rocks basaltic rocks ocean water evaporate sulfate Ranges of S Isotopic Values in Geologic Systems Range of sulfur isotopes in geologic systems. Note the difference between mantle-derived S and Sedimentary sulfides (Hoefs, 1980, fig. 12).

29. Diagrammatic Representation of the Exogenic Cycles of Carbon and Sulfur S - CycleC - Cycle Car Carbonate Carbon 6460 x 10 18 moles δ 13 C = -0.4 Organic Carbon 1180 x 10 18 moles δ 13 C = -27 Gypsum 166 x 10 18 moles δ 34 S= +8.4 Pyrite 180 x 10 18 moles δ 34 S= -7.9 Carbonate, DIC 3.3 x 10 18 moles δ 13 C = +0.46 SW sulfate 40 x 10 18 moles δ 34 S= +20 Ocean Gregor et al.. 1988

30. http://www.libraryindex.com/article_images/www.libraryindex.com/sulfur.01.jpg&imgrefurl=http://www.libraryindex.com/pages/3406/Sulfur- weathering 72 volatile biological emissions 22 deposition over continents 32 volcanic emissions 10 volatile biological emissions 43 MOR volcanic emissions 10 deposition over oceans 207 some as OCS to stratosphere sea-salt sulfur 144 some from explosive volcanism to stratosphere eolian erosion 10 precipitation of evaporites BSR to pyrite Natural Sulfur Cycle 104

31. GENERALIZED STRATIGRAPHY (YAX-1) AND SAMPLE DISTRIBUTION (n=46) Depth (m) rst.gsfc.nasa.gov/Sect18/Chicxulub

32. (Keller et al., 2004)

33. Biomineral formation by fungi and sulphate-reducing bacteria. (A) a cord-forming fungus growing on copper phosphate (B) light micrograph of moolooite crystals (copper oxalate, CuC 2 O 4.xH 2 O) around the hyphal cords (C) scanning electron micrograph of moolooite crystals associated with hyphal cord and mucilaginous sheath (Fomina, M. et al. (2005) Applied and Environmental Microbiology 71, 371-381) (D) a crust of calcium oxalate (weddelite and whewellite) crystals and tubular crystalline sheath around fungal hyphae (ESEM dry mode) (Gadd, G.M. et al. (2006) In: Fungi in the Environment, Cambridge University Press. (E) hydrated sulphate-reducing bacterial biofilm (Desulphomicrobium sp.) transforming selenite to abundant Se/S granules. Inset shows granules associated with the surface of an individual bacterium and precipitation in the extracellular matrix (Hockin, S.L. & Gadd, G.M. (2003) Applied and Environmental Microbiology 69, 7063-7072). Evolution of the sulfide and sulfate reservoirs Evolution of the sulfide and sulfate reservoirs. BSR – bacterial sulfate reduction www.lifesci.dundee.ac.uk

34. Vienna- Canyon Diablo Triolite -- V-CDT The Canyon Diablo meteorite comprises many fragments of the asteroid that impacted at Barringer Crater (Meteor Crater), Arizona, about 50,000 years ago. Meteorites have been found around the crater rim, and are named for nearby Canyon Diablo, which lies about three to four miles west of the crater. Pyrrhotite var. Triolite: An unusual i ron sulfide mineral in which the ratio of iron to sulfur atoms is somewhat variable Fe (1-x) S (x = 0 to 0.2) but is always slightly less than 1. It commonly is found in association with other sulfides. The variety troilite, with a composition near that of iron sulfide (FeS), is an important constituent of some iron-nickel meteorites. Canyon Diablo Troilite (CDT) is used as a zero standard of relative concentration of different isotopes of sulfur. A meteoritic standard was chosen because of the constancy of the sulfur isotopic ratio in meteorites, while the sulfur isotopic composition in Earth materials varies owing to bacterial activity. VCDT is the synthetic standard -0.3‰ more depleted than the original CDT.