2 H = ( 2 H / 1 H) Sample vs. ( 2 H / 1 H) Reference 13 C/ 12 C – 15 N/ 14 N – 18 O/ 16 O – 34 S/ 32 S – 37 Cl/ 35 Cl – 81 Br/ 79 Br.

2H = (2H / 1H)Sample vs. (2H / 1H)Reference

13C/12C – 15N/14N – 18O/16O – 34S/32S – 37Cl/35Cl – 81Br/79Br

Malin SödermanJunior Research Engineer

• We develop methods – Visitors come and use them

• Self-cost model

• We are available for

-General isotope advice (planning)

- Collaboration - One-off- Short term- Long term- Proposal assist

… Equilibrium Isotope Effects (EIE) – isotopic phase partitioning… Kinetic Isotope Effects (KIE) – isotopic reaction rates

Zero-point energies…

=(1/2+n)h

(/m)½

Vibrational frequency

Isotope mass

Force constant

Heavy isotopes favour a strong bond

The Equilibrium Isotope Effect

(2H / 1H)Phase X-H*

(2H / 1H)Phase Y-H*

EIE =

solvent analyte

The Kinetic Isotope Effect

krate=Ae-Ea/RT

Fre

e en

ergy

Reaction coordinate

Ea LightEEa HeavyE

KIE = kLight / kHeavy

…KIE = kLight / kHeavy

Holmstrand et al., Chemosphere 69 (2007) 1533–1539

DDT DDE

R(+)or

S(-)

Catalysis by DL-2-haloacid dehalogenase for stereo isomers as

determined from Kinetic Isotope Effects (KIEs)

Papajak et al., Biochemistry 2006, 45, 6012-6017

An Equilibrium Isotope EffectMolecular oxygen adducts in metalloenzymes

– side-on or end-on?

Side-on

End-on

The characterization of intermediates

in oxidation reactions of transition metals – MetalloenzymesSide-on

End-on

The derived oxygen isotopic effects have been used to

(i) differentiate inner-sphere from outer-sphere electron transfer

(ii) identify metal-O2 intermediates

(iii) assess mechanisms of O-O bond-breaking/making reactions

Ashley, Brinkley and Roth, Oxygen Isotope Effects as Structural and Mechanistic Probes in Inorganic Oxidation Chemistry, Inorg. Chem. 2010, 49, 3661–3675

“No deuterium was incorporated… the active site of the enzyme is probably in a closed conformation during catalysis”

Hydrogen exchange labeling to studya reaction mechnism

- choice of labels and analytical technique

2H label

Natural 2H level

Guangqi et al., Biochimie 92 (2010) 1454e1457

Tracers and labels: - Is natural abundance enough? (safety, cost, feasibility)

- Can KIE or EIE help? (additional information)

- Added value of IRMS instruments? (trace weaker signals)

Macronutrient sourcing for biosynthesis using 13C analysis of amino acids

– direct routing from diet to tissue

Filled = diet Open = collagen

Diet compositionDiet Protein EnergyD2A C3 C3

D3G C3 C4

D4H C4 C4

D5I C4 C3

Jim et al., British Journal of Nutrition (2006), 95, 1055–1062

C3

C4

-25‰

-12‰

C3 plants = - 25‰ (e.g. rice)C4 plants = - 12‰ (maize)

?

Chemistry

Labeling

Reactionmechanisms

Reactionmonitoring

Reactionidentification

Linking source &product

Environmentalsciences

Ecology

Plant physiology

Microbiology

Biochemistry

E-mail: [email protected]

Cell phone: 070 760 7229

Website: http://delta.geo.su.se…where you will find this presentation…