NOVEL FLUORESCENT SENSORS FOR THE DETECTION OF ORGANIC MOLECULES IN

EXTRA-TERRESTRIAL SAMPLES Roy C. Adkin, James I. Bruce and Victoria K. Pearson

Twitter: @RCAdkin

Aim of the Research

Development of a fluorescent lanthanide complex which will

interact with meteoritic organic species in situ and/or in

the aqueous phase

Background• Organic material is found in carbonaceous chondrite (CC) meteorites: <5% by

mass (~14000 different molecules (Schmitt-Kopplin et al., 2010))

• Although (most) confirmed as extra-terrestrial in origin due to:– Isotope ratios (H/D, C, N, O)– Structural isomerisation and diversity e.g. racemic ratio, branching– Compounds present in higher concentrations but rare on Earth, e.g.,

isovaline, pseudoleucine etc. (Kvenvolden et al., 1970)

• No defined environment of formation for what is seen in meteorites although several possible cosmological provinces suggested

• BUT, minerals are formed under distinct chemical and physical conditions so can be used as environmental indicators (Velde, 2000)

• Understanding mineral/organic associations more could help clarify organic compound source regions and formation processes

The problem• We know a relationship exists;

– Amount of matrix vs bulk organic material indicated by C and N content (e.g. Anders et al., 1973)

– Removal of minerals by dissolution releases more organic material (e.g. Sephton and Gilmour, 2001)

– Basic labelling reveals organic material predominately associated with matrix (e.g. Pearson et al., 2007)

• Organic molecular inventory and concept of mineral/organic material associations elucidated by destructive analysis of carbonaceous chondrites

Development of a new, non-destructive, in situ analytical tool is required…

Fluorescence - Overview

Emitted light < energy and > λ than the light absorbed

Usually, emission ceases almost instantaneously as irradiation is terminated (ns to μs timescale)

The sensor – Introducing the lanthanides• Lanthanides (Ln) are elements, e.g. europium (Eu) and

terbium (Tb) - amongst the most luminescent elements in the Periodic Table

• Extensively used in biomedical imaging techniques

• Lanthanide metal ion coupled to an organic ligand

• ‘Fingerprint’ emission spectrum consisting of line-like peaks or bands – indicative of the element

• Have long fluorescent lifetimes – the time between termination of irradiation and cessation of emission (ms)

• Ln must be stable, chemically inert yet subject to physical interactions

The sensor – ligand: DOTA

1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraethanoic acid

Commercially available

NNO

O

N

O

ON

OO

O

O

H

H

H

H

The sensor – EuDOTA and TbDOTA

Preliminary researchSources of intrinsic CC fluorescence•Some minerals exhibit fluorescent properties

•Presence of Eu or Tb can activate, enhance or intensify that fluorescence

•Identify organic and inorganic CC components whose excitation and emission λ may be similar to Eu and Tb

• Organic excitation below that of Eu and Tb so not a concern

• Mineral fluorescence, activated and intrinsic, can be background corrected

Preliminary research – DOTA experimentation

1) DOTA was synthesised

2) Suitable analytes were chosen representative of all classes of organic molecules identified in CCs taking into consideration:

– The number and type of reactive sites and functional groups

– Likelihood of interaction with the sensor – structure/size

– Whether they are terrestrially rare or potentially prebiotic

– Solubility in water

Results of DOTA experiments and discussion• 1 mM LnDOTA solution mixed with a range of meteoritic

organic molecules at concentrations expected in CCs

• Spectra showed no peak shifts but a slight, yet trendless, variation in intensity

• Lack of spectral deviation• No interaction with metal centre?• Lanthanide/analyte interaction but fluorescence not altered by

presence of analyte? • Limit of detection?

• Concentrations consistent with chondritic organic matter (µM, 10-6 mol dm-3, to nM, 10-9 mol dm-3) may be too low for detection by this sensor

Fluorimetric analysis – Equimolar (1 mM) EuDOTA/analyte analysis

Fluorimetric analysis – Equimolar (1 mM) TbDOTA/analyte analysis

DOTA Fluorimetric analysis – Conclusion

• Would expect analytes to increase fluorescent intensity due to displacement of water molecules

• No discernible trend regarding analyte structure;• It was expected that conjugated and aromatic analytes could

increase fluorescent intensity by absorption of excitation energy

• Hypothesis: DOTA ligand does not afford interactions• Steric hindrance• Ln atom is too well enveloped• Cannot be sure of limit of detection

• Solution? – Use DO3A ligand…one less pendant arm

The new ligand – DO3A

O

H

ON

NNOH

O

N

O

H

O

H

Fluorimetric analysis – EuDOTA/TbDO3A comparison

Fluorimetric analysis – Eu3+(aq)/EuDOTA/EuDO3A comparison

EuDO3A fluorimetric analysis – EuDO3A and all analytes

EuDO3A fluorimetric analysis – EuDO3A and all analytes

(L)-serine

(L)-tyrosine

(L)-threonine

EuDO3A/EuDOTA fluorimetric analysis - conclusions

• EuDOTA and EuDO3A have shown intensity increase with certain structures or chemical classes only

• Identification of structures or functional groups is feasible

• Individual molecular specificity may not be achievable

Future work

• Produce standards for mixtures of:– Similar compound classes (e.g. all amino acids or all

carboxylic acids etc.)– Similar or analogous structures (e.g. hypoxanthine

and cytosine or adenine and 2,4-diaminopyrimidine etc.)

– Complex mixtures of classes and structures

• Introduce LnDOTA and LnDO3A complexes to these mixtures

• Measure the effects on Ln fluorescent properties

Future work (continued)

• Development of other ligand molecules • change nature of the pendant arms

- facile ligand modifications - broaden the scope of

interactions with analytes- selectivity and sensitivity H

H

H

O

O

N

NNH

NO

O

N

HH

H

N

OO

N

HH

H

• Development of methodology for future solid sample analysis

Thank you for listening.

Any questions?

Twitter: @RCAdkinEmail: Roy.Adkin@open.ac.uk

Fluorimetric analysis – Analyte structures

(L)-serine (L)-ornithine (L)-tyrosine

(L)-threonine (L)-aspartic acid Benzoic acid

Fluorimetric analysis – Analyte structures

Cytosine

Maleic Acid

Adenine Hypoxanthine

Fumaric acid

Itaconic acid N-guanylurea 2,4-diaminopyrimidine