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welcome to this presentation as part of

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astrobiology graduate conference

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organized virtually by elsi in tokyo

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i am thomas lostoler

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a phd candidate

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focused on probiotic chemistry in solid

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state

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and working at roger boschkovic

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first i want to highlight the key

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message

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i want you to take away from this

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presentation and this is that amino

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acids oligomerized to peptides on

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mineral surfaces

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without the need to interact with water

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or aqueous environment

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i want to acknowledge the amazing team

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of scientists with whom i worked on this

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project

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jose gregorio hernandez

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a scientist with world-class talent and

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the originator of this project idea

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kronos lavojarvic who is my phd

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supervisor and my biggest supporter

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the scientist with the right tools and

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solutions who has an incredible passion

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for doing science

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nicola ernest and sasha for offering us

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tremendous support and access

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to analytical techniques that we needed

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to carry out this project

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many thanks to croatian science

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foundation for financing

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to start with the presence of amino

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acids on early earth is consensually

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accepted by scientists in the field

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either stemming from endogenous chemical

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routes or delivered extraterrestrially

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by meteorites

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on the other hand peptides are important

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biological polymers with many catalytic

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functions

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and are regarded to be included in

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ancient molecular symbiosis with nucleic

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acids

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here we have a molecular structure of

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the simplest amino acid called glycine

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generally amino acids have an n-terminus

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with an amino group and a c-terminus

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with a carboxyl group

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peptides on the other hand are chains of

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amino acids

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linked together via peptide bonds which

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is a covalent bond between carbon from a

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carbonyl group and a nitrogen from an nh

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group

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how did nature go from amino acid to

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peptides is a major question in

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probiotic chemistry

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which is chemistry that led to life

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the formation of peptide bonds is an

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example of a condensation reaction where

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two amino acids such as glycine are

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joined together or condensed

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via peptide bond together with the

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elimination of a water molecule

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looking at the basic le chatelier

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principle

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if this reaction is carried out in

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aqueous media

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it is highly thermodynamically

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unfavorable

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because in order for this reaction to

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proceed water molecules need to be

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eliminated

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so it seems that we have a water paradox

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on one hand water is essential for life

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and for biological evolution

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but on the other hand

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it seems that water is not desired for

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the synthesis of life's building blocks

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this is not just related for the

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synthesis of peptides

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but is a broader issue in probiotic

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chemistry

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because there are many condensation

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reactions

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such as the formation of glycosidic

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or phosphodiester bonds

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this has been recognized by more and

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more

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scientists in the field

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and there is a growing realization that

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dry land was critical for concentrating

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organic building blocks

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and enabling their chemical evolution

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there are different literature

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approaches for prebiotic peptide

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synthesis

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firstly thermal polymerization which

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requires high temperatures going back to

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late 50s

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soon after the

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discovery of famous

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military experiment which served as a

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birth of probiotic chemistry

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secondly hypervelocity impact studies

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that uses impacts in the range of

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kilometers per second

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and results in short peptides such as

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dye and tripeptides

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thirdly wet dry cycling which uses

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different approaches

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but always uses water for mixing of the

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reactants and its subsequent evaporation

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in order for peptide bond to form during

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the dry phase

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so we hypothesized what if we avoid the

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use of water and instead turn to

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solid-state reaction environment

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and use only solid mineral surfaces

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our approach was to use mechanical

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energy that could have been provided

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probiotically by impacts erosion

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earthquakes and weathering

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and to use thermal energy

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the availability of dry land is often

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discussed in the field and although its

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availability was only a few percentage

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of today's land mass

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it is considered that there was some

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land available

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near volcanoes and on the emerging

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continents

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we used mechanochemistry as a tool to

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carry out chemical reactions in the

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absence of solvents

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mechanochemistry has been previously

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used in prebiotic chemistry contexts

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photosynthesis of alpha amino nitriles

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and sugars

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we also used temperature controlled ball

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milling setup developed recently by our

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group

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we decided to use

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the simplest amino acid glycine

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due to its probiotic availability

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the presence in meteorites and the

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availability of

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analytical techniques

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needed to characterize reaction products

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reaction mixture was

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closed in a milling jar and was bold

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milled via three stainless steel balls

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the milling jar together with the

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reaction mixture

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was oscillated horizontally

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and

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attached to it was a thermal

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ring element that was connected to a

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temperature controller

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to characterize the reaction products we

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used

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iron pairing high performance liquid

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chromatography

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as we can see on the chromatograms

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raising the reaction temperature going

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from room

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40 degrees 70 100 and 130 degrees

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celsius

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we have observed the formation of longer

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glycine oligomers

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this is

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happening together with the formation of

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higher amounts of

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dicato piperazine a cyclic glycine dimer

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that is sometimes regarded as a dead end

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for probiotic peptide synthesis

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the total yield of all linear oligomers

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was highest when the reaction was

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carried out at 100 degrees celsius and

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in order to better understand the

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dynamics of mechanochemical peptide bone

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synthesis

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we carried out reactions starting from

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dipeptide tripeptide and dicatopirazine

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we can see on the chromatograms

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that for all these cases we observed the

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formation of odd and even number of

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oligomer lengths

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meaning that the mechanochemical peptide

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bond formation is a dynamic process with

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simultaneous

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forming and breaking of peptide bonds

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importantly

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when starting the reaction

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from dicato piperazine

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we have ascertained that it can be a

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productive intermediate

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for the synthesis of peptides as we

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observed the

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formation of

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peptides containing 10 glycine residues

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in order to unambiguously determine the

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presence of

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different oligomer length

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we have developed a method for ultra

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performance liquid chromatography

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coupled with mass spectrometer

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so first

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to separate different oligomers and then

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to detect them using a q-tof detector

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according to their

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molecular mass

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the longest oligomer detected by milling

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glycine with titanium dioxide was a

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peptide with 11 residues of glycine

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this oligomer length

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is long enough for it to establish a

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three-dimensional structure

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which is associated

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with the catalytic properties of

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peptides and proteins

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to summarize our key results

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we have successfully formed peptides

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when also using different mineral

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surfaces

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such as silicon dioxide montmorillonite

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and

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mica mechanochemical peptide bond

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formation tolerates the presence of

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moisture

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as verified by performing

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liquid-assisted grinding experiments

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where we added small amounts of water

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this was to show the robustness of

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mechanochemical route

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that does not require glovebox

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conditions

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that were certainly not available on an

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earlier scenario

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we also tried to use different amino

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acids such as l-alanine and after

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milling it with titanium dioxide we have

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when milling together glycine and

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l-alanine with titanium dioxide

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it resulted in the formation of their

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heteropeptides

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and we have successfully shown

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that it is possible to incorporate other

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again

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i want to highlight a key message for

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you to take away from this presentation

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and this is that amino acids

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oligomerized to peptides on mineral

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surfaces without the need to interact

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with water or aqueous environment

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using only

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the input of mechanical and thermal

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energy

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finally

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what are the implications of

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mechanochemical route towards peptides

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for astrobiology

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can we expect to find peptides in

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asteroid samples

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these are some of the very exciting

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questions please do get in contact with

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us if you have any additional questions