Stable isotopes Colin MacLeod. Using Stable Isotopes (SI) To Investigate Ecological Aspects Background (the boring bit) Nucleus of elements made up of

Stable isotopes

Colin MacLeod

Using Stable Isotopes (SI) To Using Stable Isotopes (SI) To Investigate Ecological AspectsInvestigate Ecological Aspects

Background (the boring bit)• Nucleus of elements made Nucleus of elements made

up of protons and up of protons and neutronsneutrons PP

PP

PP

PP

PP

PP

E.g. Carbon E.g. Carbon NucleusNucleus

NNNN

NN

NN

NN

NN

6 Protons, 6 Protons, 6 6 NeutronsNeutrons

• Some elements have Some elements have different forms with different forms with different numbers of different numbers of neutrons (but same neutrons (but same number of protons)number of protons)

PP

PP

PP

PP

PP

PP

E.g. Carbon E.g. Carbon 1212

NNNN

NN

NN

NN

NN

6 Protons, 6 6 Protons, 6 NeutronsNeutrons

• Nucleus of elements made Nucleus of elements made up of protons and up of protons and neutronsneutrons


Background (the boring bit)



PP

PP

PP

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PP

PP


NNNN

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NN

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NN



PP

PP

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PP


NNNN

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PP

PP

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NNNN

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• Number of neutrons does Number of neutrons does NOTNOT affect chemical affect chemical reactions but affects reactions but affects weightweight

• Some are radio-active: Some are radio-active: C14 C14

• Others are stable: C12, Others are stable: C12, C13C13





• Radioactive isotopes used Radioactive isotopes used for measuring time e.g. for measuring time e.g. radiocarbon dating (C14)radiocarbon dating (C14)

• Stable isotopes used for Stable isotopes used for measuring rates of measuring rates of biological reactions, biological reactions, patterns of energy flow patterns of energy flow etc.etc.

• Usually measured as ratio Usually measured as ratio of two stable isotopes of of two stable isotopes of same element (e.g. same element (e.g. C12:C13 or delta C13 or C12:C13 or delta C13 or ΔC13 or ΔC13 or C13)C13)

PP

PP

PP

PP

PP

PP


NNNN

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• One neutron = One neutron = 1·675 × 101·675 × 10-27-27 kg kg

• Therefore, atoms with more neutrons will be Therefore, atoms with more neutrons will be slightly heavierslightly heavier

• This affects how they ‘behave’: e.g. COThis affects how they ‘behave’: e.g. CO22 in in SolutionSolution

C12C12

C13C13

C12C12

C12C12

C12C12 C12C12

C12C12 C12C12 C12C12

C13C13WaterWater(cold)(cold)

AirAir

Ratio of Ratio of C12:C13C12:C13

1000:121000:12


Why Does This Work ?



C13C13

C12C12

C12C12

C12C12

C12C12

C12C12 C12C12 C12C12

C13C13

AirAir

C12C12




WaterWater(less (less cold)cold)



C13C13

C12C12

C12C12

C12C12

C12C12 C12C12 C12C12

C13C13

AirAir C12C12

C12C12




WaterWater(luke warm)(luke warm)



C13C13

C12C12

C12C12

C12C12

C12C12 C12C12 C12C12

C13C13

AirAir C12C12

C12C12




WaterWater(warm(warm))



C13C13

C12C12

C12C12

C12C12

C12C12 C12C12 C12C12

C13C13AirAir C12C12

C12C12




WaterWater(quite (quite warm)warm)



C13C13

C12C12

C12C12

C12C12

C12C12 C12C12 C12C12

C13C13AirAir C12C12 C12C12




WaterWater(very (very warm)warm)



C13C13

C12C12

C12C12

C12C12C12C12

C12C12 C12C12

C13C13AirAir C12C12 C12C12

Ratio of Ratio of C12:C13C12:C13

1000:201000:20




WaterWater(hot)(hot)


Why Does This Work ?• The same happens in animals as they The same happens in animals as they

metabolise food they have eatenmetabolise food they have eaten• C12 gets preferentially exhaled in C0C12 gets preferentially exhaled in C022 and and

animal becomes relatively enriched with the animal becomes relatively enriched with the heavier C13heavier C13

• Similarly, with isotopes of nitrogen, N14 gets Similarly, with isotopes of nitrogen, N14 gets preferentially excreted in urea etc. and animal preferentially excreted in urea etc. and animal becomes relatively enriched with the heavier becomes relatively enriched with the heavier N15N15

• This happens with each trophic levelThis happens with each trophic level• Therefore, the higher up a food-web the greater Therefore, the higher up a food-web the greater

the enrichment in heavier isotopes of C and Nthe enrichment in heavier isotopes of C and N• In addition, in marine environment, warmer In addition, in marine environment, warmer

waters = greater C13 enrichment at base of waters = greater C13 enrichment at base of food-webfood-web


What Happens in Nature ?

NHNH33++

NHNH33++ NHNH33

++

NHNH33++

SeawaterSeawater PlanktonPlanktonPrimary Primary

CarnivoreCarnivoreSecondary CarnivoreSecondary Carnivore

Tertiary CarnivoreTertiary CarnivoreTop CarnivoreTop Carnivore

(N14:N15)(N14:N15)

1000:31000:3 1000:61000:6 1000:91000:9 1000:121000:12

1000:151000:151000:181000:18

+3‰+3‰ +3‰+3‰ +3‰+3‰

+3‰+3‰

+3‰+3‰


How Can Scientists Use This To Study Ecology ?

What What Trophic Trophic Level Level

Does a Does a Species Species

Feed At ?Feed At ?

What is the What is the Ratio of Ratio of Nitrogen Nitrogen

Isotopes in its Isotopes in its Tissues ?Tissues ?

Rephrase Rephrase Question In Question In

Terms of Terms of IsotopesIsotopes

???????????????????????????? N14:N15??N14:N15??N14:N15??N14:N15??

N

14:N

15?

BlendeBlenderr

ONON

OFFOFF

BlendeBlenderr

ONON

OFFOFF



Mass Mass SpectrometerSpectrometer

(measures weights all (measures weights all atoms of specific atoms of specific

elements in sample)elements in sample)

Ratio of different isotopes Ratio of different isotopes e.g. e.g. 1000:141000:14 N14:N15N14:N15

(reflects trophic level of (reflects trophic level of animal)animal)

PREPARATIOPREPARATIONN

DecalcificatiDecalcificationon

DelipificationDelipificationDehydrationDehydration



NHNH33++

NHNH33++ NHNH33

++

NHNH33++

SeawaterSeawater PlanktonPlanktonPrimary Primary

CarnivoreCarnivoreSecondary CarnivoreSecondary Carnivore

Tertiary CarnivoreTertiary CarnivoreTop CarnivoreTop Carnivore

(N14:N15)(N14:N15)

1000:31000:3 1000:61000:6 1000:91000:91000:121000:12

1000:151000:151000:181000:18

1000:141000:141000:141000:14

Using Stable Isotopes To Using Stable Isotopes To Investigate Beaked Whale Investigate Beaked Whale

EcologyEcology

(the interesting bit!!)• Used data from stranded animals to develop Used data from stranded animals to develop hypotheses on beaked whale ecology in the hypotheses on beaked whale ecology in the North AtlanticNorth Atlantic

• But Strandings data subject to an number of But Strandings data subject to an number of unquantifiable biases - therefore need unquantifiable biases - therefore need independent testing of hypothesesindependent testing of hypotheses

• Used stable isotope analysis of bones from Used stable isotope analysis of bones from stranded animals for testing hypothesesstranded animals for testing hypotheses

• Why bones ? Easy to obtain, reflects long-Why bones ? Easy to obtain, reflects long-term diet...term diet...

...And hard to get a whole whale into a ...And hard to get a whole whale into a blender!blender!

Using Stable Isotopes To Using Stable Isotopes To Investigate Beaked Whale Investigate Beaked Whale

EcologyEcology

(the interesting bit!!)• In total, samples from 150 different individuals In total, samples from 150 different individuals collected between 1800 to 2002 obtained from:collected between 1800 to 2002 obtained from:

• Museums e.g. National Museums of ScotlandMuseums e.g. National Museums of Scotland• Strandings networks e.g. SACStrandings networks e.g. SAC• Researchers and naturalistsResearchers and naturalists

• Represent six different species - all found in North Represent six different species - all found in North AtlanticAtlantic

• Primarily sampled inner posterior mandible - least Primarily sampled inner posterior mandible - least damage to valuable museum specimensdamage to valuable museum specimens

• Samples taken weighed around 0.25gSamples taken weighed around 0.25g• Decalcified to remove inorganic carbon (Soak in 0.5N Decalcified to remove inorganic carbon (Soak in 0.5N

HCl)HCl)• De-lipified to remove fats and lipids (Using De-lipified to remove fats and lipids (Using

Chloroform)Chloroform)• Actual tissue analysed: Bone collagen (Protein Actual tissue analysed: Bone collagen (Protein

component of bones - makes them flexible)component of bones - makes them flexible)• Size of samples analysed in mass spec: 0.7mg Size of samples analysed in mass spec: 0.7mg

Documents

Stable isotopes Colin MacLeod. Using Stable Isotopes (SI) To Investigate Ecological Aspects Background (the boring bit) Nucleus of elements made up of