STABLE CARBON ISOTOPES, C3 AND C4 PLANTS, AND CLIMATE CHANGE

Annotated Bibliography

by

Todd Martin

Cerling, T.E., Quade, J., Wang, Y., and Bowman, J.R., 1989, Carbon isotopes in soils

and palaeosols as ecology and palaeoecology indicators: Nature, v. 341, p. 138-139.

An analysis of 10 North American soils with simple pedogenic histories are used here to demonstrate the usefulness of stable carbon isotopes (12C, 13C) as palaeoecological indicators. Past studies have invoked complex processes to describe the systematics of carbon isotopes based on a lack of data for cogenic organic carbon and pedogenic carbonate. These previous studies made necessary assumptions due to extensive cultivation, grazing or long periods of pre-Holocene vegetation and soil development. The present study, having selected sites with no disturbances, and assuming constant vegetative cover allows for a determination of definite carbon isotopic signatures.

Assuming no major climate changes and no anthropogenic activity leads to a model based on isotopic equilibrium between carbon species attributable to organic matter (OM) and soil carbonate. Soils were examined in areas of > 35cm/yr rainfall to negate the diffusive mixing of atmosphere and soil that would result in an enriched sample. The results shows a definite difference in isotopic composition. Pedogenic carbonate values average 0.5 , while OM values averaged 15. This difference of ª 14, according to the authors "constitutes a simple test of the state of preservation of the original palaeoecological signal in soils". The difference is thus a measure of the fractionation of carbon due to C3 and C4 plant pathways.

This paper appears to have been published in part to lay the groundwork for a logical extension in using C3 and C4 plant fractionation of carbon as a proxy for climate change. The authors of this paper do cite their own contemporary study of 2 sites that seem to show this "climate change" but caution that based on sample size, the relationship between organic carbon, soil carbonate and climate is premature. Overall, this paper was quite readable and serves as a good introduction to soils and carbon isotopes.

Cerling, T.E., Wang, Y., and Quade, J., 1993, Exapnsion of C4 ecosystems as an

indicator of global ecological change in the late Miocene: Nature, v. 361, p. 344-345.

Fossil soils and fossil teeth are used as "important" indicators of the presence of C4 biomass in local ecosystems. Soil carbonate is enriched by 14 - 17 in 13C compared to local biomass while carbon isotope composition of tooth enamel is enriched by about 12 compared to diet. Both of these variables are reported as "robust" indicators of palaeoenvironments and paleaeodiets respectively.

While it is commonly known that average 13C values for C3 plants are about -26, and about -13 for C4, the carbon isotope composition for C3 ecosystems has been reported to be as low as -23 to -20 in arid environments due to moisture stress. Therefore, reported values for soil carbonate as high as -8, and tooth enamel values as high as

-10 can still indicate a primary biomass of C3. Palaeosols from Pakistan reflect a definite change in isotopic composition from -9 to -12 beginning about 7.4 Myr, to -2 to +2 by about 4 Myr. This evidence indicates a change from C3 vegetation to C4 vegetation. Tooth enamel from Pakistan and horses in North America, indicate C4 plants became an important part of diets starting about 7-6 Myr., which correlates to the time C4 biomass became important in Pakistan. These data suggest a rapid expansion of C4 ecosystems in both the New and Old World, and thus the authors suggest a change in global conditions as opposed to local development and a gradual expansion around the world. The authors further assume that since the C4 pathway evolved as a more effective metabolic pathway due to decreased atmospheric CO2, that the fossil evidence affirms a global change likely. This they say is confirmed by a reported change in ocean isotope composition during the same time. Additionally, the authors cite their 1989 paper as a reflection of this global change.

Overall, I liked this paper for its discussion of more enriched isotopic values with C3 biomass due to stressed conditions, and the additional link to tooth enamel (I passed up several other papers that used this evidence). I do not believe however that they adequately presented the other evidence that justified the leap from isotopic analysis useful at a local scale to the global scale.

Ehleringer, J.R., Sage, R.F., Flanagan, L.B., and Pearcy, R.W., 1991, Climate change

and the evolution of C4 photosynthesis: Trends in Evolutionary Ecology, v. 6, no. 3,

p. 95-99.

C3 plants dominate most of the terrestrial ecosystem, comprising 85% of all plant species. The CAM plants, those that use both C3 and C4 pathways account for 10%. The remaining 5% include the C4 plants, which dominate in warm to hot, open sites typically tropical and temperate grasslands. C4 plants are thought to have evolved from C3 plants as recently as 50-60 Mya. due to decreased atmospheric C02 concentrations. The authors discuss the efficiencies of the two different metabolic pathways, the benefits associated with C4 photosynthesis, the diversification of taxa, the conditions favoring C4 evolution and the "geologic" evidence for C4 evolution.

Carbon fixation in C3 plants is catalyzed by Rubisco (ribulose-1,5-biphosphate carboxylase/oxygenase). The result is a 3 carbon compound phosphoglycerate (PGA) which is then easily metabolized for photosynthesis. Rubisco can also catalyze PGA and phosphoglycolate which then releases CO2 (photorespiration), and can severely reduce the efficiency of metabolism. This efficiency is dependent on pCO2, pO2, and leaf temperature. C4 plants initially fix carbon by phosphoenol pyruvate carboxylase (PEP), producing a 4 carbon compound, and ultimately produces CO2 which is then metabolized in the same manner as the C3 pathway. Unlike C3 plants, C4 plants do not lose CO2 (photorespiration).

Since C4 plants are able to recycle CO2, the benefits of C4 metabolism allows plants at low CO2 levels and higher temperatures to more efficiently use CO2. Because photorespiration does not occur, the C4 pathways should be more efficient, also contributing to improvements in water-use efficiency and nitrogen-use efficiency.

C4 plants contain no enzymes or anatomical structures not also present in C3 plants, and C4 photosynthesis occurs in diverse and distantly related angiosperms with no common ancestor. Even within genera, photosynthesis likely evolved independently several times, as evidenced by the grass genus Panicum, which exhibits "enough biochemical variation in C4 photosynthesis" to indicate that C4 plants evolved from C3 plants, and that this evolution occurred independently many times.

Assuming these variations in C4 photosynthesis represent evolution from the C3 pathway, what could have caused this? The authors refer to various global C02 models which indicate "dramatic changes in atmospheric CO2 concentrations. Atmospheric CO2 levels are generally accepted to have been much greater that they are presently, having been relatively low for the last 50 My. A rapid drop in CO2 levels "during the period of major expansion of the angiosperms would have provided strong selection pressure for increased" efficiency of photosynthetic pathways.

Although geological evidence for the first appearance of C4 plants is "scanty", plant fossils with well developed anatomy (C4), and carbon isotopic composition of soil layers suggests that C4 plants could have appeared as early as the Cretaceous, but certainly no later than the late Miocene. Pollen evidence also indicates the worlds' major grasslands developed during the Miocene, further supporting the authors claim.

A very good article for some introductory plant biology. Several figures as well as their explanations provide a good framework for understanding the different metabolic pathways for C3 and C4 photosynthesis. The conclusions however are based on world climatic models and laboratory investigations of plant productivity, so caution is urged. Overall I think I like the idea, but would like to see more hard data on global CO2 levels that could support the evolutionary hypothesis.

Fredlund, G.G., and Tieszen, L.L., 1997, Phytolith and carbon isotope evidence for late

Quaternary vegetation and climate change in the southern Black Hills, South Dakota:

Quaternary Research, v. 47, p. 206-217.

Two profiles, one from the Highland Creek terrace, and one from an alluvial fan within Dry Creek Gully, both in Wind Cave National Park, Black Hills, SD, were studied to demonstrate the utility of complimentary techniques of stable carbon isotope and phytolith analysis. The data collected provides a basis for documentation of the rate and nature of grassland change throughout the Holocene, and an evaluation of temporal sensitivity and spatial limits of the methods used. Interpretations were based in part on modern grassland assemblages.

The first line of evidence, stable carbon isotopes, were used as a proxy for vegetation change in the great plains. Assuming negligible effects of soil formation, 13C values should reflect the balance between C3 and C4 plants. The isotopic ratios measured are therefore a result of either a shift in C3/C4 grassland composition forced by climate, or a result controlled by moisture and fire regimes.

Phytolith assemblages, the second set of data, may be use in accordance with stable carbon isotopes to reconstruct changes in climate and vegetative patterns. Phytolith analysis focused on short-cell forms because the pattern of production and redundancy is better understood, and they are generally assumed to be uniformly preserved and recovered.

This study involved analysis of sediments and soils along two profiles within the Red Valley of the Wind Cave National Park, South Dakota. Samples were taken from a 2.5 m thick cut bank terrace along the Highland Creek, with a basal 14C date of 3,870. Results showed that (a) changes in percent carbon correlated well with the soil and sediment stratigraphy, (b) changes in phytolith recovery and preservation parallel soil stratigraphy, and (c) grass short-cell phytolith assemblages show significant changes with depth. The other sample site, an alluvial fan in Dry Creek Gully measured 6.5 m thick with a basal 14C date of 14,120. An unconformity representing "ca. 8000 to 4500 yr B.P." separated this section, with the upper soils and sediments correlating well with the Highland Creek site. Analysis reveals, (a) "good correspondence" between identified palaeosols and peaks in organic carbon, (b) oldest soils average 13C values of -24.79, indicating almost exclusively C3 biomass, (c) 13C become increasing enriched throughout the early Holocene, and (d) grass phytolith assemblages show significant shifts in the lower portion of the profile. Overall, the authors say their analysis shows a clear change in plants biomass during the early Holocene, and argue that phytolith analysis along with 13C analysis provides a more robust argument for vegetation and climate change.

This is a useful study because it incorporates not only standard carbon isotopic analysis, but also uses phytoliths in order to propose vegetative, and thus climatic changes. I am a bit troubled that both of their profiles were missing almost 5000 years of record. Also, their first profile did not contain the earliest (late Pleistocene) soils and sediments that presumable would have correlated with their other profile. This missing information would have allowed for a better interpretation, and an overall more believable conclusion.

Leuenberger, M., Siegenthaler, U., and Langway, C.C., 1992, Carbon isotope

composition of atmospheric CO2 during the last ice age from an Antarctic ice core:

Nature, v. 357, p. 488-490.

Analysis of CO2 content within bubbles from an ice core drilled at Byrd Station, Antarctica reveals lower atmospheric concentrations of CO2 than the pre-industrial value. This study attempts to attribute the change in concentrations of CO2 to either a change in the "biological pump" of the ocean, or a change in surface alkalinity of the ocean.

The biological pump can be explained by assuming that sinking organic particles continuously export carbon from the surface of the ocean to the depths, causing a depletion of total CO2 in the surface water compared to that of the deep ocean. Since 13C in organic matter is lower than in dissolved inorganic matter, depletion of the surface accompanies an enrichment in the deep sea water. Therefore, a more efficient biological pump (increased upwelling) should produce higher 13C values in surface water and atmospheric CO2. A change in surface alkalinity on the other hand, and therefore a lower pCO2 due to rearrangement of water masses or to changes in carbonate sedimentation by themselves should not affect the 13C values in the atmosphere. Thus the authors say, that isotopic analysis should discriminate between these two mechanisms.

Data from the analysis of air trapped in the ice core revealed 13C values 0.19 ± .18 more negative than in the early Holocene (20-40 kyr BP), and 0.32 ± .17 lower than in the millennium preceding industrialization. This data, according to the authors "might reflect the preferential withdrawal of isotopically light carbon during biomass buildup on land". Analysis of forams and packrat middens from other studies are compatible with lower atmospheric 13C during the last ice age.

By far my least favorite paper thus far. The authors spent way too much time discussing methodology, corrections necessary for gas analysis of CO2, and reasons why the scatter could be disregarded. Just check out some of the disclaimers;

1) "traces of drilling fluid which may have interfered with isotope measurements

(although we have no indication for this)"

2) "we do not consider these old results as reliable, and they are not taken into account in

the mean"

3) "our CO2 values generally agree with earlier measurements obtained by a more precise

method... although some samples deviate..." .

Anyway, the authors do state in their conclusion that the evidence does show that changes in the biological pump alone are not sufficient to account for their values. Overall, a lousy paper that leaves me wondering what the motivation for their study might have been.

Quade, J., Cerling, T.E., and Bowman, J.R., 1989, Development of Asian monsoon

revealed by marked ecological shift during the latest Miocene in northern Pakistan:

Nature, v. 342, p. 163-165.

Well dated sediments in the Potwar Plateau region are abundant within the fine-grained floodplain facies of large river systems. Carbonate nodules within these soils were analyzed with a sampling density averaging 1 palaeosol per 130,000 yrs. This record contains 3 distinct phases with respect to carbon and oxygen isotopes; (1) from 18 - 7.4 to 7.0 Myr almost all values of 13C were -13 to -9 for soil carbonate and up to -15 more negative for organic matter from several samples, (2) from 7.4 - 7.0 to 5 Myr the stable isotopic compositions shift to more positive values, beginning at 7.4 Myr, and (3) from 5 to 0.4 Myr soil carbonate values range from -2 to +2, and organic matter is -14 to -11. This analysis from 52 palaeosols of the Siwalik Group in northern Pakistan indicates a pure or nearly pure C3 biomass dominated floodplain before 7.4- 7.0 Myr.

After 7.4 - 7.0 Myr, C4 grasses become dominate as evidenced by the more enriched values of 13C. This shift in biomass, as compared to modern ecosystems indicated a "closed canopy forest with or without an understorey of C3 shrubs and herbs" before 7.4 Myr, with a transition to C4 dominated grasslands after that time. This ecological shift can be interpreted in two ways according to the authors. A climate change that allowed the invasion of C4 grasses, or an evolutionary 1st appearance of C4 plants unrelated to climate change could account for the isotopic record within these palaeosols.

The origins of the vegetation shift seen in the soil record is explained in terms of the modern monsoonal climate. Modern climate in northern Pakistan is monsoonal, with 70% of the annual 40 cm's of rain falling in the warm summer months. "This moderate rainfall, the strongly seasonal rainfall distribution, and the prevalence of natural fires from summer lightning strikes all favour grasslands over forest." The authors continue to state that if the monsoon system was in place prior to 7.4 Myr, then it experienced a strong intensification at that time. Supporting evidence for this rapid shift include diatom assemblages from the Indian Ocean specific to monsoonal circulation, and a faunal turnover a mammalian browsers to grazers at approximately the same time.

Another article by the group of authors working on the usefulness of carbon isotopes as a proxy for different metabolic plant pathways and climate change. Unlike their previous Nature paper, they do conclude that their evidence indicates a transition from C3 to C4 photosynthesis. One has to wonder why in the previous article they cite this yet unpublished study as inconclusive then months later state their case certainly. Overall I liked this paper, but I was troubled that they related 18O analysis side by side with their carbon isotopes yet did not really use any specifics to support their conclusion.

Smith, B.N., 1972, Natural abundance of the stable isotopes of carbon in biological

systems: BioScience, v. 22, no. 4, p. 226-231.

About 99% of all carbon is the 12C isotope while 1% is 13C. The other isotopes are rare by comparison. Radiocarbon (14C) for instance makes up only 1 atom in every 1012 carbon atoms. The ratios of the stable carbon isotopes 13C/12C vary depending on the material in which they are found. Limestones, atmospheric CO2, marine algae, and land plants all have characteristic ratios. Plants that fix carbon dioxide via the Calvin cycle (C3) have different d13C values than plants that fix carbon dioxide via the C4-dicarboxylic acid pathway. Recent studies have investigated naturally occurring ratios of 13C to 12C of biological materials, which vary due to the different ways of carbon fixation. This paper discusses the methods and assumptions on which a few of these studies have been based.

Various isotopic species of an element have different chemical properties, and thus behave differently. Factor influencing behavior include bond strength, temperature, pressure, chemical composition, etc. which will determine the isotopic fractionation of a given process.

In order to process a sample, the organic materials must be converted to CO2. This is accomplished through combustion @ 800-900 C in an excess of oxygen. The gasses are recycled to ensure total conversion to CO2, then collected in a nitrogen-cooled trap, and subsequently warmed to dry-ice temperatures where the CO2 is collected, presumably free of any water or gas contaminant. The pure sample of CO2 is then isotopically analyzed with a mass spectrometer, and compared to a standard (the fossil carbonate skeleton of Belemnitella americana (PDB)). The function defining the values reported is

d13C per mil = (13C/12C) sample - (13C/12C) standard ¥ 1000

(13C/12C) standard

Therefore, a sample with a value of -10 has a 13C/12C ratio less than the standard by 10 per mil, or 1.0%. Atmospheric CO2 for example is smaller than that of the PDB standard by about 7 per mil, so the d13C per mil of atmospheric CO2 is -7. The precision typical of mass spectrometers is ± 0.1%. Often these values will be reported as being depleted, or enriched (relative to 13C). A depleted sample, with a more negative value means more 12C and less 13C, while an enriched sample would have a more positive value with 13C dominating the ratio.

Different species of organisms (plants) will tend to produce depleted or enriched values of 13C. Marine plants, freshwater aquatics, and some desert plants have relatively more 13C than most terrestrial species. This difference was originally attributed solely to the source carbon being either bicarbonate for marine organisms or atmospheric CO2 for terrestrial organisms. Not until the late 1960's was it discovered that the fractionation of the stable carbon isotopes was largely due to differences in the metabolic pathways during photosynthesis. Plants with enriched values are those with metabolic pathways that fix carbon via phosphoenol pyruvate carboxylase (PEP) while those that tend to be depleted in 13C use ribulose diphosphate carboxylase (Rubisco) This fractionation is only found in the angiosperms, and has led to studies that may allow for climatic and environmental interpretations.

In depth studies have found that the fractionation of carbon may be averaged for a certain species of plant, but is really rather complex. Different chemical fractions of an organism differ from one another isotopically. A common potato tuber for instance, has a tissue value of -25.8, and the lipids = -34.6, while different acids, proteins, and sugars within the tuber range between the other two values.

What can account for the overall fractionation of a given plant species? A model proposed by Park and Epstein (1960) identified 4 major fractionation sites within a plants' system. The first site involves the uptake of CO2 through the cytoplasm boundary due to stomatal resistance to diffusion, and collision frequency of the gas molecule with the mesophyll cell surface. After the CO2 has passed through the membrane, it is partitioned and converted to starch, where some of it is removed through the vascular system for excretion through the roots. The next step, and the most consequential for isotopic fractionation is the actual fixation of carbon (photosynthesis) which is accomplished via Rubisco, or PEP. The last mechanism for possible fractionation includes the conversion of starches to lipids. All of these mechanisms affect the final fractionation that is associated with CO2 fixation, which in principle allows for plants to have the range of 13C values from 0 to -38.

The environmental effects of carbon isotope fractionation have recently been investigated. The isotopic differences between warm water (low latitude) and cold water (high latitude) marine plankton populations implies a certain connection between isotopic ratios and ambient water temperature. However, laboratory studies of plankton growth in which CO2 was controlled (large concentrations) showed no temperature effects. This implies that carbon availability may be more important than temperature in determining fractionation. In general, "natural abundance ratios of isotopes of carbon can be used to study aspects of organism-environment interactions since fractionations of the isotopes will occur at various points in the carbon cycle".

This 1972 article is easily read and understood. The author cites several earlier studies, including Park and Epstein (1960) which are seminal papers on the subject of carbon isotopes on biologic processes. The author encourages others to investigate these phenomenon and use this knowledge to make possible further insight into evolution, etc. It appears that only recently has the scientific community taken heed of his suggestion.

(Park, R., and Epstein, S., 1960, Carbon isotope fractionation during photosynthesis:

Geochim. Cosmochim. Acta., v. 21, p.110-126.)

Stern, A.L., Chamberlain, C.P., Blum, J.D., and Fogel, M.L., 1997, Isotopic lessons in

a beer bottle: Journal of Geoscience Education, v. 45, p. 157-161.

An upper-level undergraduate class entitled "Isotopic Tracers in the Environment", taught at Dartmouth College, brewed an Irish Ale in order to observe the distinct isotopic signatures associated with C3 and C4 plants. Specifically, students observed and interpreted the carbon isotope ratios of the CO2 produced by yeast during the fermentation of sugars. This exercise allowed students to evaluate the mixing of two isotopically distinct reservoirs, observe biologically mediated isotopic fractionation, and acquire and understanding of the isotopic signature of C3 and C4 plants.

The students hypothesized that glucose, primarily from corn sugar (a C4 plant), which ferments easily would ferment first, followed by the fermentation of a mixture of less easily fermentable sugars in malted barley (a C3 plant). Analysis of the CO2 evolved during fermentation should reflect the different isotopic ratios of the source sugars.

Results of the experiment produced enriched 13C values averaging about -16 during the first 30 hours of fermentation. Samples of CO2 after 30 hours showed a marked depletion in 13C, with an average of 22.8. The change in isotopic ratios, specifically between the 30-39 hour interval represented the change in fermentation of source sugars. In other words, the most easily metabolized sugars (sucrose, glucose, fructose) fermented first producing the more positive values of 13C, followed by the less easily metabolized sugars (maltose and maltotriose) producing more negative 13C values.

This experiment assumed no fractionation of carbon during fermentation, so that the isotopic values obtained could be attributed to the fractionation effect of the plant's metabolic pathways during photosynthesis. Students suggested several variations to their experiment such as fermenting a single source sugar to verify no fractionation affects during fermentation, sampling of the fermenting wort along with evolved CO2 to monitor 13C concentrations of sugars and ethanol during fermentation, and a more sophisticated sampling device to minimize atmospheric contamination of CO2 samples.

Well of course I like this paper! I had heard of this experiment for a while before I finally got my hands on the paper. Since I am an avid homebrewer myself, and the topic of C3/C4 plants is germane to our course, this paper precipitated my interest in the whole subject of stable carbon isotopes. As "they" always say, show a student the relevance of what they are learning to their personal life and you will have them hooked. You may look forward to a sample of my own experiment at the end of our course! Cheers?