www.sciencemag.org/cgi/content/full/314/5801/980/DC1

Supporting Online Material for Isotopic Evidence for Dietary Variability in the Early Hominin Paranthropus robustus Matt Sponheimer, Benjamin H. Passey, Darryl J. de Ruiter, Debbie Guatelli-Steinberg, Thure E. Cerling, Julia A. Lee-Thorp Published 10 November 2006, Science 314, 980 (2006) DOI: 10.1126/science.1133827 This PDF file includes: Materials and Methods Figs. S1 and S2 Table S1 References

Supporting Online Material Materials and Methods All specimens analyzed were housed at the Transvaal Museum, Pretoria, South Africa. The Paranthropus teeth temporarily obtained for this study are SK 24605 (RM3), SK 24606 (RM2 or RM3), SKX 5939 (partial molar) and SKW 6427 (partial molar). The Raphicerus sp. specimens obtained are SKX 8494 (RP2), SKX 8535a (RP3), and SKX 14150 (LP3). All specimens were cleaned mechanically with razor blades and acetone was applied using cotton swabs to remove potential contaminants. Individual isotope analyses (continuous-flow GC-IRMS) were of 10-30 nmol of cryofocused CO2 generated by 3 to 8 laser ablation events, using a CO2 laser (10.6 µm) operating at 5-15 W and 8.5 ms pulse duration in a helium atmosphere (S1). Systematic isotope fractionation was monitored by full system analyses of injected CO2 aliquots, with the CO2 calibrated against NBS-19 gas (δ13C = 1.95‰) produced using the sealed vessel phosphoric acid method at 25°C. Fractionation associated with laser ablation production of CO2 was monitored by analyzing a suite of internal tooth enamel standards with carbonate δ13C values calibrated against NBS-19 using the phosphoric acid method. The laser-carbonate isotope fractionation (ε*LASER-carb) for fossil herbivore samples from several South African hominin sites was –1.3 ± 1.5 ‰ (1σ). Paranthropus teeth were sampled at 0.48 mm intervals on average (~6 perikymata high each representing a minimum of 54 days), while Raphicerus teeth were sampled every ~1 mm. We made high-resolution epoxy replicas of each hominin tooth and coated these with a goldpalladium alloy (S2). The perikymata between each sampling track were then counted with the aid of a scanning electron microscope (see Figure 1). Perikymata counts were made under a magnification of 25x, except for regions of the crown in which perikymata were more closely packed (near the cervix), where counts were made at 50x magnification. For intervals in which perikymata were not visible, counts were estimated using the average number of microns between adjacent perikymata for these Paranthropus specimens (75 microns).

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Fig. S1: Plots showing δ13C and δ18O values in Paranthropus robustus. These values are significantly correlated across the data for all four specimens (Fig. S1A; R2=0.50; P<0.001) and within SKX 5939 (Fig. S1B; R2=0.85; P<0.001). We do not observe similar co-variation in fossil Raphicerus, nor in reference enamels and gasses analyzed alongside Paranthropus specimens, so we interpret this as a real ecological signal similar to that found in many South African mammals today (S3).

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Fig. S2: δ13C values of the hair of modern C3 feeders (n=49), C4 feeders (n=126), and Pan troglodytes in the dry (n=21) and wet (n=11) seasons (data from S4). The boxes represent the 25th-75th percentiles (with the medians as horizontal lines) and the whiskers show the 10th-90th percentiles. Analysis of variance of these four groups demonstrates strong differences (F2,203 = 1173.586; P<0.0001), but Fisher’s PLSD test reveals that all significant differences are between the C4 feeders and other groups (P<0.0001). There is no significant difference between dry and wet season chimpanzee hair (P=0.49). Although hair and enamel δ13C values are offset because of fractionation differences, they can be converted for comparison, and when this is done, they are not statistically different in terms of absolute values and magnitudes of variability for individual species (S5). Moreover, hair is not subject to the isotopic overprinting of enamel, and therefore hair δ13C values should be more sensitive to seasonal differences in diet. Thus, the greater amplitude of change observed in Paranthropus enamel compared to Pan hair cannot be ascribed to tissue differences. See S6 for more high-resolution δ13C data for the teeth and hair of primates.

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Table S1: Data used in this study for Paranthropus robustus molars and Raphicerus sp. molars. Sample SKW 6427 SKW 6427 SKW 6427 SKW 6427 SKW 6427 SKW 6427 SKW 6427 SKW 6427 SKW 6427 SKW 6427 SKW 6427

Species Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus

δ13C -7.4 -9.1 -9.0 -8.6 -8.8 -8.6 -9.8 -9.2 -8.2 -8.1 -8.3

δ18O 21.9 20.3 20.7 21.3 20.9 21.8 21.4 21.1 22.2 22.1 21.2

Scan 1 2 3 4 5 6 7 8 9 10 11

SKX 5939 SKX 5939 SKX 5939 SKX 5939 SKX 5939 SKX 5939 SKX 5939 SKX 5939 SKX 5939 SKX 5939 SKX 5939 SKX 5939 SKX 5939 SKX 5939 SKX 5939 SKX 5939 SKX 5939

Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus

-4.1 -3.1 -2.7 -4.9 -3.7 -4.9 -5.0 -5.3 -5.2 -6.7 -5.8 -5.8 -6.9 -7.6 -7.9 -6.9 -6.0

24.0 24.3 24.2 23.8 24.0 24.1 23.6 23.6 23.7 23.6 23.6 23.5 23.4 22.8 22.7 23.1 23.1

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17

0 4 9 13 18 24 30 35 42 49 55 61 67 74 80 86 92

0 0.3 0.7 1.0 1.4 1.8 2.3 2.6 3.2 3.7 4.1 4.6 5.0 5.6 6.0 6.5 6.9

SK 24606 SK 24606 SK 24606 SK 24606 SK 24606 SK 24606 SK 24606 SK 24606 SK 24606 SK 24606

Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus

-7.1 -8.7 -5.1 -4.4 -5.3 -4.7 -6.2 -5.9 -6.6 -7.0

24.8 23.7 24.4 23.9 24.8 24.5 24.4 24.1 24.1 24.6

1 2 3 4 5 6 7 8 9 10

0 6 14 21 31 37 44 52 60 70

0 0.5 1.1 1.6 2.3 2.8 3.3 3.9 4.5 5.3

SK 24605 SK 24605 SK 24605 SK 24605 SK 24605 SK 24605

Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus Paranthropus robustus

-7.4 -7.2 -6.7 -7.5 -6.5 -7.7

24.1 24.1 22.9 23.6 24.2 23.7

1 2 3 4 5 6

0 7 14 21 27 33

0 0.5 1.1 1.6 2.0 2.5

4

Perikymata Millimeters 0 0 9 0.7 16 1.2 21 1.6 26 2.0 33 2.5 39 2.9 44 3.3 50 3.8 57 4.3 62 4.7

SK 24605

Paranthropus robustus

-7.8

23.9

7

SKX 8494 SKX 8494 SKX 8494 SKX 8494 SKX 8494 SKX 8494

Raphicerus sp. Raphicerus sp. Raphicerus sp. Raphicerus sp. Raphicerus sp. Raphicerus sp.

-9.6 -9.8 -9.8 -10.1 -9.7 -10.0

19.5 19.8 19.9 20.1 20.9 21.1

1 2 3 4 5 6

SKX 8535a SKX 8535a SKX 8535a SKX 8535a SKX 8535a

Raphicerus sp. Raphicerus sp. Raphicerus sp. Raphicerus sp. Raphicerus sp.

-8.2 -8.4 -8.4 -8.6 -9.1

23.5 23.8 23.6 23.6 23.6

1 2 3 4 5

SKX 14150 SKX 14150 SKX 14150 SKX 14150 SKX 14150 SKX 14150 SKX 14150 SKX 14150

Raphicerus sp. Raphicerus sp. Raphicerus sp. Raphicerus sp. Raphicerus sp. Raphicerus sp. Raphicerus sp. Raphicerus sp.

-10.0 -10.0 -10.0 -10.2 -9.9 -9.6 -9.8 -10.0

23.3 23.1 23.2 22.5 22.2 22.9 22.2 23.3

1 2 3 4 5 6 7 8

5

39

2.9

References and Notes S1. B. H. Passey, T. E. Cerling, Chem. Geol. doi:10.1016/j.chemgeo.2006.07.002. S2. D. Guatelli-Steinberg, Am. J. Phys. Anthropol. 120, 309 (2003). S3. J. Smith, thesis, University of the Witwatersrand (2005). S4. M. Sponheimer et al., J. Hum. Evol. 51, 128 (2006). S5. M. Sponheimer et al., J. Mammal. 84, 471 (2003). S6. D. M. Codron, University of Cape Town (2003).

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