Learning Outcomes

By the end of this session you should be able to:

• Describe the physical basis of stable isotope fractionation
• Explain how trophic enrichment is quantified
• Discuss tissue-specific isotopic routing
• Apply collagen quality criteria to evaluate specimens
• Articulate the link between dietary isotope mixing models and the Marine Reservoir Effect

What Is a Stable Isotope?

Isotopes are atoms of the same element differing in neutron number and therefore atomic mass.

Stable isotopes do not undergo radioactive decay — unlike ¹⁴C in radiocarbon dating, they are permanent constituents of the biosphere.

The Physics of Fractionation

The Delta (δ) Notation

Absolute heavy-isotope abundances are tiny (~1 %), so ratios are reported as per-mil (‰) deviations from an international standard:

\[ \delta X\ (\text{‰}) = \left[\frac{R_{\text{sample}}}{R_{\text{standard}}} - 1\right] \times 1000 \]

where \(R\) = heavy/light isotope abundance ratio.

Analytical Measurement

Stable isotope ratios are measured by Isotope Ratio Mass Spectrometry (IRMS).

In continuous-flow (CF-IRMS) configurations, sample combustion and MS introduction are integrated:

• As little as 0.3–1.0 mg of collagen required
• Typical precision: ±0.1–0.2 ‰ (1σ) for both δ¹³C and δ¹⁵N
• δ¹³C and δ¹⁵N measured simultaneously on a single run

The δ¹³C / δ¹⁵N Biplot

Why δ¹³C and δ¹⁵N? (and if you have the money δ³⁴S)

Two Types of Fractionation

Equilibrium fractionation

Heavy and light isotopes have slightly different bond energies → temperature-dependent → primary driver of ¹³C partitioning during carbonate precipitation.

Trophic Enrichment Factors (TEFs)

When an organism synthesises tissue from diet, fractionation produces systematic isotopic enrichment. These offsets are TEFs (also: discrimination factors), denoted Δ (capital delta):

\[\Delta^{13}C = \delta^{13}C_{\text{tissue}} - \delta^{13}C_{\text{diet}}\]

\[\Delta^{15}N = \delta^{15}N_{\text{tissue}} - \delta^{15}N_{\text{diet}}\]

When an animal eats food and builds tissue from it, the biochemical processes involved (transamination, deamination, protein synthesis etc.) are not isotopically neutral. Lighter isotopes tend to be preferentially lost or excreted, meaning the tissue ends up slightly enriched in the heavier isotope relative to what was consumed. The TEF (or Δ) simply quantifies that offset.

The Trophic Step Effect on δ¹⁵N

At each trophic level, δ¹⁵N increases ~3–5 ‰ because:

• Animals digest protein → lighter ¹⁴N preferentially lost in urine as urea/ammonia
• Heavier ¹⁵N retained in body tissue
• Effect repeats at every predation event → staircase pattern through the food web

⚠️ Archaeological Confounders of δ¹⁵N

Elevated δ¹⁵N is not always diagnostic of high-status meat consumption:

• Manured soils elevate baseline plant δ¹⁵N, cascading up the food web
• Breastfed neonates show apparent trophic elevation (~+2–3 ‰) relative to their mothers
• Physiological stress (starvation, illness) can increase tissue δ¹⁵N through net protein catabolism
• Arid environments tend to have systematically elevated δ¹⁵N baselines throughout the ecosystem

What Is Isotopic Routing?

Isotopic routing = the differential biochemical fate of dietary macronutrients — proteins, lipids, and carbohydrates — during tissue synthesis.

It has profound implications for interpreting collagen isotope data and for constructing mixing models, because collagen does not reflect bulk diet uniformly across all macronutrient classes.

Carbon Routing to Bone Collagen

Bone collagen is synthesised almost entirely from amino acids, and these are preferentially sourced from dietary protein:

• Dietary protein carbon contributes ~60–80 % of collagen carbon (Ambrose & Norr 1993)
• Dietary carbohydrate and lipid contribute relatively little, even when they dominate caloric intake
• Bone collagen δ¹³C is therefore a better proxy for dietary protein source than for total diet

Historical Development of the Field

Relevance to Radiocarbon Practitioners

Why Quality Assessment Matters

Before any mixing model can be constructed, collagen extract quality must be assessed. Degraded collagen introduces systematic bias because isotope ratios shift with diagenetic alteration.

The DeNiro Acceptance Window