Synthesis and Self-Assembly of Novel ABC Miktoarm Star Terpolymers

Syn hesis and Sel -Assembly o No el ABC Mik oa m S a Te polyme s DISSERTATION zu E langung des akademisches G ades eines Dok o s de Na u wissenscha en (D . e . na .) im Fach Chemie an de Bay eu he G aduie enschule ü Ma hema ik und Na u wissenscha en de Uni e si ä Bay eu h o geleg on And eas Hanisch Gebo en in Kulmbach Bay eu h, 2013 Die o liegende A bei wu de in de Zei on Juli 2008 bis Janua 2013 in Bay eu h am Leh s uhl Mak omolekula e Chemie II un e Be euung on He n P o . D . Axel H. E. Mülle ange e ig . Volls ändige Abd uck de on de Bay eu he G aduie enschule ü Ma hema ik und Na u wissenscha en de Uni e si ä Bay eu h genehmig en Disse a ion zu E langung des akademischen G ades eines Dok o s de Na u wissenscha en (D . e . na .). Disse a ion einge eich am: 25.01.2013 Zulassung du ch die P omo ionskomission: 13.02.2013 Wissenscha liches Kolloquium: 26.04.2013 Am ie ende Dekan: P o . D . Bea e Lohne P ü ungsausschuss: P o . D . Axel H. E. Mülle (E s gu ach e ) P o . D . S ephan Fö s e (Zwei gu ach e ) P o . D . Ma hias Ka g (Vo si z) P o . D . Ka lheinz Sei e “ Li e is like iding a bicycle. To keep you balance you mus keep mo ing. ” Albe Eins ein Meine Familie G e a und Oska Table o Con en s I Table o Con en s Summa y 1 Zusammen assung 3 Glossa y 7 1 – In oduc ion 9 1.1 Solu ion Based Sel -Assembly in Polyme Sys ems 9 1.1.1 Gene al Aspec s o Sel -Assembly 9 1.1.2 Mul icompa men S uc u es om Te na y Sys ems 10 1.1.2.1 Linea T iblock Te polyme s and Di ec ed Sel -Assembly 12 1.1.2.2 Mik oa m S a Te polyme s 16 1.2 Mik oa m S a Polyme s 19 1.2.1 Syn hesis 19 1.2.1.1 ABC Mik oa m S a Te polyme s as Model Sys ems 19 1.2.1.2 O he Mik oa m S a Polyme Sys ems 26 1.2.2 Sel -Assembly in Bulk 27 1.2.3 Applica ion o ABC Mik oa m S a Te polyme s o Func ional Ma e ials 29 1.3 Syn he ic S a egies in Polyme Science 31 1.3.1 DPE Chemis y in Anionic Polyme iza ion 31 1.3.2 Click Chemis y 32 1.4 Aim o his Thesis 34 1.5 Re e ences 35 2 – O e iew o he hesis 41 2.1 Modula Syn hesis o Mik oa m S a Te polyme s 42 2.2 Coun e ion-Media ed Hie a chical Sel -Assembly o an ABC Mik oa m S a Te polyme Con aining a Poly(N-me hyl-2- inylpy idinium iodide) Segmen 45 2.3 Applica ion o he T iiodide-Di ec ed Sel -Assembly o o he ABC and ABA’ Mik oa m S a Polyme s wi h a Poly(N-me hyl-2- inyl- py idinium iodide) Segmen 47 2.4 Indi idual Con ibu ions o Join Publica ions 51 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s ia a New Alkyne-Subs i u ed Diphenyle hylene De i a i e 53 3.1 In oduc ion 54 3.2 Expe imen al Sec ion 57 3.3 Resul s and Discussion 60 3.3.1 Syn hesis o 1-[(4-( e -Bu yldime hylsilyl)e hynyl)-phenyl]-1-phenyl- e hylene (click-DPE) 60 3.3.2 Syn hesis o Alkyne Mid-Func ionalized Diblock Copolyme s 61 3.3.3 Hyd olysis o he Alkyne Mid-Func ionalized Diblock Copolyme s 64 3.3.4 Syn hesis o ω-Azido Homopolyme s 66 Zusammen assung 4 g ega en umlage n. Au g und ih e E scheinung in elek onenmik ospopischen Au nah- men wu de ü diese de Beg i „woodlouse“ („Kelle assel“) einge üh . Die Agg ega e sind nich hohl und bes ehen aus PB/P BMA Lamellen, die übe eine eilweise gequolle- ne P2VPq Phase mi einande e bunden und dadu ch s abilisie we den. Die PB/P BMA Lamellen besi zen dabei eine eilweise en misch e inne e S uk u . Dieses Konzep wu de e olg eich au zwei wei e e Mik oa ms e n-Sys eme ange- wand . Fü eine Se ie on ABC-Mik oa ms e n-Te polyme en aus PS, PB und P2VPq (µ-BVqS) wu de eine Abhängigkei de Mizell o m om hyd ophilen An eil beobach e ; analog zu Diblock-Copolyme en. Eine sei s wu den ü lange P2VPq-Blöcke Agg ega e aus ges apel en Lamellen/Scheiben e hal en, die aus Kugelmizellen als G undbaus einen en s anden sind; ande e sei s wu de ü ku ze P2VPq Blöcke die Ausbildung mul ilamel- la e Vesikel du ch Fusion unilamella e Vesikel beobach e . Ande s als du ch die Polyme a chi ek u e wa e , konn e ü die Vesikelwand keine Kompa men alisie ung beobach e we den, sonde n es wi d eine lamella e Zusammense zung mi einem PB Ke n angenommen, die du ch eine dünne PS-Schich o de P2VPq-Ko ona geschü z wi d. Fü einen de µ-BVqS-Mik oa ms e ne wu de e olg eich eine Nanohyb ids uk u mi Goldnanopa ikeln gebilde . Fü ein zwei es Polyme sys em wu de das elas ische PB-Segmen du ch einen wei e- en glasa igen PS-Block mi un e schiedliche Länge e se z (µ-SVqS‘). Obwohl mi Iodid als Gegenion Vesikel e hal en wu den, bilde en sich bei de T iiodid-induzie en Übe - s uk u bildung aniso ope Agg ega e aus de o mie en Vesikeln. Die Abwesenhei on mul ilamella en Vesikeln wi d dabei au die eduzie e Dynamik de ke nbildenden, glas- a igen PS-Blöcke zu ückge üh . Ähnlich zu den „woodlouse“ Pa ikeln on µ-BVqT, wu den somi ü µ-SVqS‘ längliche Agg ega e mi eine lamella en S uk u e hal en, jedoch übe Vesikel als G undbaus eine. Somi is dieses übe das T iiodid-Ion beein- lussba e Konzep de ges eue en Selbs agg ega ion nich nu au Mik oa ms e n Sys- eme mi einem elas ischen Block besch änk , da die Umo dnungsp ozesse wäh end des Dialyse s a inden, wo das an angs o handene o ganische Lösungsmi el eine aus ei- chende Mobili ä de den Mizellke n bildenden Blöcke zu Folge ha . Zusammen assung 5 Neben eine neuen Syn heses a egie ü Mik oa ms e n-Te polyme e, basie end au einem ielsei igen Alkin-subs i uie en DPE De i a , konn e somi die du ch das T iiodid- Ion induzie e Übe s uk u bildung als neua iges Konzep ü ges eue e Selbs ano d- nungsp ozesse in wäss igen Lösungen au gezeig we den. 6 Glossa y 7 Glossa y 2VP 2- inylpy idine AAO anodic aluminium oxide ATRP a om ans e adical polyme iza ion CoQ10 coenzyme Q10 CRP con olled adical polyme iza ion c yo-TEM c yogenic ansmission elec on mic oscopy CuAAC coppe -assis ed azide-alkyne cycloaddi ion DNA deoxy ibonucleic acid DPE 1,1-diphenyle hylene DVB di inylbenzene w olume ac ion o wa e IPEC in e polyelec oly e complex MCM mul icompa men micelles NRC ni oxide adical coupling SET-LRP single elec on ans e li ing adical polyme iza ion PAA poly(ac ylic acid) PB polybu adiene PBLL poly(ε- e -bu yloxyca bonyl-L-lysine) PCEMA poly(2-cinnamoyloxye hyl me hac yla e) PDMAEA poly(N,N-dime hylaminoe hyl ac yla e) PDMAEMA poly(N,N-dime hylaminoe hyl me hac yla e) PEO poly(e hylene oxide) PEE polye hyle hylene PI polyisop ene PISC poly((sul ama e-ca boxyla e)isop ene) PMA poly(me hyl ac yla e) PMAA poly(me hac ylic acid) PMCL poly(γ-me hyl-ε-cap olac one) PnBu poly(n-bu yl ac yla e) PNiPAAm poly(N-isop opylac ylamide) PS polys y ene PSGA poly(sucinna ed glyce yl monome hac yla e) P BA poly( e -bu yl ac yla e) P BMA poly( e -bu yl me hac yla e) PVBFP poly(pen a lu o phenyl 4- inylbenzyl e he ) P2VP poly(2- inylpy idine) P2VPq poly(N-me hyl-2- inylpy idinium) ROP ing-opening polyme iza ion TEM ansmission elec on mic oscopy THF e ahyd o u an TPPB iphenylphosphonium b omide ΔG agg. Gibbs ee ene gy o micelle o ma ion ΔH agg. en halpy o micelle o ma ion ΔS agg. en opy o micelle o ma ion 8 1 – In oduc ion 9 1 – In oduc ion 1.1 Solu ion Based Sel -Assembly in Polyme Sys ems 1.1.1 Gene al Aspec s o Sel -Assembly Sel -assembly desc ibes he well-de ined o ganiza ion o one species o a se o iden ical molecula building uni s in o highly o de ed agg ega es by non co alen in e ac ions. He e, he complexi y and di e si y achie ed in na u e displays an imp essi e p o o ype o such s uc u es a di e se leng h scales. 1-3 Examples o biological s uc u es o di e - en dimensions, buil up wi h he concep o sel -assembly include e.g. DNA, 4 i uses 5 o spide silk. 6 The enginee ing o na u al building uni s esembles a ema kable pa hway o he p og ammed cons uc ion o nanoscale objec s in a bo om-up app oach. 7 An ou s anding example is DNA-o igami, which is based on he combina ion o a long sca - old s and o i us-DNA wi h small, designed s aple s ands o adequa e base-sequence o ob ain di ec ed olding in o di e se wo-dimensional 8 o h ee-dimensional s uc- u es. 9 Despi e he syn he ic ad ances in polyme chemis y, achie ing such a dis inc com- plexi y wi h polyme sys ems has no been eached. Ne e heless, i ep esen s a desi - able pa adigm o he cons uc ion o complex unc ional s uc u es o di e en dimen- sion in analogy o sys ems in na u e (Figu e 1-1A). 10 In he case o block copolyme s as mos simpli ied polyme ic sys em wo chemically di e en blocks a e co alen ly linked oge he . I he polyme now is exposed o a sol en selec i e o only one block, he indi idual molecules ha e o sel -o ganize o shield he sol ophobic block. The gain in ee ene gy (ΔH agg. < 0) o minimizing he un a o able in e ac ion be ween he sol en and he insoluble block o e comes he loss o en opy (ΔS agg. > 0), leading o an o e all nega i e Gibbs ee ene gy ΔG agg. . ΔG agg. = ΔH agg. - TΔS agg. < 0 Depending on he ac ion o he s abilizing block he polyme s assemble in o sphe i- cal micelles, cylind ical micelles o esicles (Figu e 1-1-B). 11-14 Despi e he s uc u al sim- plici y o hese agg ega es he ailo ed syn hesis o block copolyme s enables cons uc - ing unc ional ma e ial. These micella s uc u es can se e as nanoca ie s 15,16 o 1 - In oduc ion 10 nano eac o s, 17-19 o as sca old o e.g. hyb id ma e ials. 20-22 Vesicles (o also called polyme somes) esemble he syn he ic analogues o liposomes and a e he e o e o in- e es o biological applica ions. 23-25 E en hough his hesis deals wi h he sel - assembly in solu ion i should be men ioned ha he same p inciples also apply o he bulk s a e, p o ided ha he he modynamic incompa ibili y o he wo blocks is su i- cien ly high. The co alen linkage p e en s phase sepa a ion on he mac oscale and he e o e o ces he polyme o align in nanoscopic s uc u es, i.e. sphe ical, cylind ical, gy oid o (pe o a ed) lamella mo phologies. 26,27 Figu e 1-1. (A) Leng h scales o molecula sel -assembled s uc u es in syn he ic and biological sys ems wi h inc easing complexi y. 10 (B) Schema ic ep esen a ion o he s uc u es o AB diblock copolyme s in a sol en selec i e o he B block. Depending on he olume ac ion o he soluble block, B , he polyme sel assembles in o sphe ical micelles, cylind ical micelles o esicles. 11 1.1.2 Mul icompa men S uc u es om Te na y Sys ems Ex ending he unc ionali y o block copolyme sys ems wi h a hi d chemically di e ing block can be accomplished in wo manne s. The linea a achmen leads o iblock e polyme s, whe eas by nonlinea conjunc ion o he hi d block mik oa m s a e polyme s ( om he g eek wo d ικτός o mixed) a e achie ed. 28 The syn hesis o he la e will be discussed in chap e 1.2. In con as o diblock copolyme s di e en block- selec i e sol en s a e possible o hese e na y sys ems, which induce ei he a com- pa men aliza ion o he co e o he co ona. 29 In he ollowing a sho o e iew o possi- ble s uc u es a he example o sphe ical micelles is gi en. I linea e polyme s a e ex- posed o a sol en selec i e o he end-blocks micelles wi h an ei he mixed, pa chy o Janus- ype co ona a e ob ained (Figu e 1-2A). 30 Howe e , accessing he egion o exclu- si ely Janus- ype micelles om ABC iblock e polyme s in endblock-selec i e sol en s 1 – In oduc ion 11 is challenging and mos ly leads o mix u es wi h pa chy micelles. 30,31 Addi ionally, in sol- en s selec i e o he middle and one end-block co e-shell-co ona s uc u es will be ob ained (Figu e 1-2B). 32 In con as o hei linea analogues he in luence o block se- quence is elimina ed o mik oa m s a e polyme s as a consequence o he polyme a chi ec u e. Hence co e-shell-co ona s uc u es a e no possible and o he co ona- compa men alized s uc u es Janus- ype micelles a e a he un a o ed due o he s e ical us a ion o he wo soluble blocks. Figu e 1-2. O e iew o co ona- (A, B) and co e-compa men alized s uc u es (C) ob ained om ABC mik oa m s a e polyme s and linea ABC e polyme s. The s uc u es depic ed a e micelles wi h a mixed, pa chy o Janus- ype co ona (A, om le o igh ), co e-shell-co ona micelles (B) o sol en s selec i e o he end-blocks o linea e polyme s and mul icompa men o “onion”-like micelles (C, om le o igh ). (D) shows a schema ic illus a ion o he chain packing o ABC mik oa m s a and linea e polyme s o ob ain mul icompa men micelles. On he o he hand, sol en s selec i e o only one block lead o a compa men aliza ion o he micella co e (Figu e 1-2C). Inspi ed by biological sys ems, in 1999 Ringsdo in- oduced he concep o mul icompa men micelles (MCM’s) o syn he ic polyme sys- ems. 33 Sepa a ed compa men s wi hin he co e would allow o s o age o wo di e - en payloads wi hin one micelle, while simul aneously main aining access o he su - ounding medium. When iblock e polyme s sel -assemble in sol en s selec i e o one end block, he in e acial ene gies be ween each o he indi idual co e- o ming blocks and he co ona- o ming block de e mine whe he micelles wi h simple concen ic com- pa men s (“onion”-like s uc u es, igh -hand s uc u e in Figu e 1-2C) 34 o micelles 1 - In oduc ion 12 wi h non-concen ic nano-s uc u ed co es (mul icompa men micelles, le -hand s uc- u e in Figu e 1-2C) a e ob ained. 35 Howe e , a ibu ed o hei us a ed a chi ec u e, mik oa m s a e polyme s a e inhe en ly o ced o sel -assemble in s uc u es wi h compa men alized co es upon dissolu ion in a sol en selec i e o one block, dis e- ga ding di e en pai s o in e acial ene gies (Figu e 1-2D). The i s example o isuali- za ion o he indi idual co e-segmen s o mul icompa men micelles and wo ms was gi en by Hillmye , Lodge and co-wo ke s in 2004 o a mik oa m s a e polyme wi h highly incompa ible segmen s, poly(e hyle hylene)-a m-poly(e hylene oxide)-a m- poly(pe luo op opylene oxide), µ-EOF. 36,37 In c yo-TEM he pe luo ina ed segmen s we e in-si u de ec able owing o hei inc eased elec on densi y. In con as o his mik oa m sys em, o linea e polyme s, besides special chemical composi ion, in e polyelec oly e complexa ion, he use o addi i es, sol en mix u es o adequa e assembly pa hways ha e o be applied o guide he sequen ially connec ed blocks o- wa ds mul i-compa men alized s uc u es. In he ollowing wo sec ions I will i s gi e an o e iew o he di ec ed sel -assembly o linea iblock e polyme s in o mul icompa men s uc u es (1.1.2.1) and hen highligh he well-de ined mo phologies ob ained by di e en mik oa m s a e polyme sys ems (1.1.2.2). 1.1.2.1 Linea T iblock Te polyme s and Di ec ed Sel -Assembly In he ield o linea iblock e polyme s Laschewsky e al. p esen ed he i s success ul in-si u isualiza ion o he mul icompa men cha ac e in 2005 o a poly(4-me hyl-4-(4- inylbenzyl)mo pholin-4-ium chlo ide)-block-polys y ene-block-poly(pen a luo ophenyl 4- inylbenzyl e he ) (PVBM-b-PS-b-PVBFP) sys em by c yo-TEM. 38 A e dialysis o wa e he e polyme assembled in o micelles wi h a co e consis ing o sphe ical domains (~3 nm in diame e ) o he pen a luo ophenyl g oup wi hin a hyd oca bon ma ix o med by bo h he polys y ene block and he a oma ic moie y o he luo ina ed block (Figu e 1-3A). The e o e seg ega ion wi hin he PVBFP block ook place. They ex ended his polyme design o o he e polyme s based on ac yla e- ype monome s o ob ain sphe ical compa men alized s uc u es. 39-41 Besides sys ems wi h a sol ophilic endblock, also sys ems wi h a sol ophilic midblock we e shown o sel -assemble in o mul icompa men s uc u es. 40-42 E en hough simula ions o iblock sys ems in mid- 1 – In oduc ion 13 block selec i e sol en s p edic mul ico e micelles, whe e he wo chemically di e en co es a e sepa a ed by he sol ophilic block, 43 he p esence o mul icompa men mi- celles he e migh be a ibu ed o he sho leng hs o he sol ophobic blocks and hei chemical na u e. In e es ingly, ins ead o using iblock e polyme s, mos ecen ly, Langlois and co-wo ke s ollowed ano he app oach u ilizing a s a is ical e polyme based on biocompa ible poly(3-hyd oxyalkanoa es). 44 Again, he seg ega ion is induced by he hyd ophilic, lipophilic and luo ophilic cha ac e o he a ached sidechains. Nanop ecipi a ion in wa e o ced o polyme o o m micelles (18 and 79 nm in diame- e o wo di e en sys ems) wi h dis inc luo ina ed subdomains in he co e. Ano he possibili y o induce phase seg ega ion wi hin he co e is based on segmen s ca ying cha ged unc ions. As a consequence o he cha ge neu ali y o poly((sul ama e-ca boxyla e)isop ene) a low pH, poly((sul ama e-ca boxyla e)isop ene)- block-polys y ene-block-poly(e hylene oxide) (PISC-b-PS-b-PEO) was epo ed o yield micelles wi h a “ aspbe y”-like PISC co e wi h sphe ical PS domains unde acidic condi- ions. 45 The emaining isop ene uni s wi hin he PISC domain and he o ma ion o hy- d ogen-bonds wi h PEO a e supposed o u he educe i s wa e solubili y and addi ion- ally he ansi ion in o micelles wi h a mixed PISC/PEO co ona was demons a ed a in- c eased pH. Schache e al. showed o zwi e ionic polybu adiene-block-poly(N-me hyl- 2- inylpy idinium)-block-poly(me hac ylic acid) (PB-b-P2VPq-b-PMAA) ha in aqueous media pa chy in e micella IPEC domains o P2VPq/PMAA a e o med on he PB co e. 46 Su p insingly, he non-qua e nized and non-hyd olyzed p ecu so polyme also o med mul icompa men micelles wi h a “sphe e on sphe e” mo phology in ace one as selec- i e sol en o he poly( e -bu yl me hac yla e) block. 47 Due o he s ong incompa ibil- i y be ween PB and P2VP, he sys em is assumed o aim su ace minimiza ion o he PB/P2VP in e ace. Apa om he sphe ical mul icompa men micelles discussed so a , Fang e al. demons a ed he hie a chical sel -assembly o p e- o med co ona-compa men alized micelles in o one-dimensional co e-compa men alized s uc u es upon educing he sol en quali y. 48 Depending on he co ona s uc u e (pa chy o Janus- ype) o he mi- celles om poly(4- e -bu oxys y ene)-block-polybu adiene-block-poly( e -bu yl me h- ac yla e) wi h a pe luo o-modi ied midblock insoluble in dioxane, he dialysis in o e ha- 1 - In oduc ion 20 o end- unc ionaliza ion o homopolyme s (1), wi h he s oichome y o adequa e linking eac ions wi h p e- o med polyme ic building blocks o ini ia o molecules (2). E en hough in li e a u e di e en classi ica ions ha e been used, he syn hesis o ABC mik oa m s a e polyme s can be di ided in o ou app oaches o undamen ally di e - ing chemis ies. Fo he i s h ee ypes anionic polyme iza ion is u ilized o he in o- duc ion o special unc ionali ies in de ined posi ions and/o selec i e eac i i y o li ing anionic chain ends (A-C), whe eas he ou h is based on mul i unc ional co e molecules (D). (A) Chlo osilane Me hod Simila o he syn hesis o egula s a s by eac ion o li ing anionic polyme chains wi h chlo osilanes, 85,86 hese compounds can be used as linking agen s o he cons uc ion o ABC mik oa m s a e polyme s. The e o e, li ing anionic chain ends o di e en eac i i- y owa d chlo ine-silicon bonds ha e o be used o allow o s ep-wise subs i u ion. Ia ou e al. syn hesized a mik oa m s a wi h polyisop ene, polys y ene and polybu adiene segmen s wi h ichlo ome hylsilane as i unc ional linking agen . 87 Howe e , he syn hesis can only be conduc ed in he speci ic sequence PS > PI > PB, due o he eac i i y o he li ing anion owa d he chlo osilane unc ionali y. The less eac- i e and mos s e ically hinde ed polyme anion has o be added i s , whe eas he less s e ically hinde ed and mos eac i e polyme anion has o be added a he end o gua - an ee ull con e sion. E en wi h his sequence (Scheme 1-2) he las s ep is ime- consuming wi h eac ion imes o up o 4 weeks and o each s ep he s oichome y is o ou e mos impo ance. Simila ly, an ABCD mik oa m s a qua e polyme was syn he- sized by expanding he sys em wi h poly(4-me hyl s y ene) and using e achlo osilane. 88 The applica ion o his app oach o o he less eac i e polyme anions like poly(me hyl me hac yla e) o poly(2- inylpy idine) equi es a pos - o p e-modi ica ion o he chlo osilane compound, espec i ely. 89,90 1 – In oduc ion 21 Scheme 1-2. Syn hesis o an ABC mik oa m s a e polyme by he successi e eac ion o li ing polyme anions wi h ichlo osilane as linking agen . (B) Mac omonome Me hod Apa om he special chemis y o chlo osilanes, diphenyle hylene (DPE) and i s dou- ble-diphenyle hylene de i a i es ep esen a powe ul class o compounds, as hei ina- bili y o homopolyme ize pe mi s selec i e unc ionaliza ion o diblock copolyme s. 91 P e o med mac omonome s wi h a e minal DPE unc ionali y can be applied o he sequen ial li ing anionic polyme iza ion o diblock copolyme s. By sequen ial addi ion o he mac omonome a e polyme iza ion o he i s block, ABC mik oa m s a e polyme s a e accessible unde con ol o he s oichome y (Scheme 1-3A). Fo his pu pose Qui k e al. used 1,4-bis(1-phenyl-e henyl)benzene (Scheme 1-3B) o syn hesize A 2 B and ABC mik oa m s a polyme s. 92 When he DPE de i a i e was used in wo- old excess o he linking eac ion wi h polys y yl-li hium p ima ily monoaddi ion ook place o gene a e he polys y ene mac omonome . Simila ly, Hücks äd e al. demons a ed 1- (4-b omome hylphenyl)-1-phenyle hylene (Scheme 1-3B) o be a sui able e mina ion agen o li ing anions o polybu adiene 93 and polys y ene 94 . These se ed as mac omonome s o he syn hesis o a polybu adiene-a m-polys y ene-a m-poly(me hyl me hac yla e) mik oa m s a e polyme and se ies o polys y ene-a m-polybua diene- a m-poly(2- inylpy idine) mik oa m s a e polyme s, espec i ely. Howe e , also in he case o he b omo- unc ionalized DPE, o ma ion o dime ic mac omonome s can occu unde inadequa e eac ion condi ions as a esul o Wu z-analogous side eac ions. Be- sides he use o DPE de i a i es as e mina ion agen s o he mac omonome syn hesis, Qui k and co-wo ke s p o ed 1-(4-hyd oxyp opylphenyl)-1-phenyle hylene o be a sui - able ini a o o he syn hesis o poly(e hylene oxide) mac omonome s a e dep o ona ion wi h iphenylme hylpo assium (Scheme 1-3B). 95 In his way a polys y- ene-a m-poly(e hylene oxide)-a m-poly( e -bu yl me hac yla e) mik oa m s a e polyme was success ully syn hesized. In a simila s a egy a polys y ene-a m- poly(dime hylsiloxane)-a m-poly( e -bu yl me hac yla e) mik oa m s a e polyme was 1 - In oduc ion 22 accessible by ing opening polyme iza ion o hexame hylcyclo isiloxane wi h li hia ed pa a-(dime hylhyd oxy)silyl-α-phenyls y ene as ini ia io . 96 Fo he syn hesis in ol ing mac omonome s he s oichome y o he endcapping eac ion has o be conside ed o allow o simple pu i ica ion o possible side p oduc s. Fu he mo e, he choice o mon- ome sequence is limi ed o he di e en eac i i ies o li ing anionic polyme chains. Scheme 1-3. (A) Schema ic ep esen a ion o he syn hesis o ABC mik oa m s a e polyme s ia sequen- ial anionic polyme iza ion u ilizing mac omonome s. (B) DPE-de i a i es used o he syn hesis o mac omonome s: 1,4-bis(1-phenyl-e henyl)benzene, 92 1-(4-b omome hylphenyl)-1-phenyle hylene, 93,94 1- (4-hyd oxyp opylphenyl)-1-phenyle hylene dep o ona ed wi h ime hylphenylpo assium 95 ( om le o igh ) (C) Mid-Func ionalized Diblock Copolyme s Ano he possibili y o cons uc ing ABC mik oa m s a e polyme s is he syn hesis o mid- unc ional diblock copolyme s. The hi d a m hen is a ached by adequa e eac- ions wi h he unc ional g oup (Scheme 1-4). Again, DPE chemis y is ad an ageous wi hin his con ex , as homopolyme iza ion is excluded and he e o e mono unc ionaliza ion is gua an eed unde app op ia e eac ion condi ions and polyme iza ions sequences. Exclusi ely hyd oxyl- unc ionalized DPE’s in hei p o ec ed o m ha e been used so a . 97,98 Lambe e al. syn hesized mid- unc ional polys y ene- block-poly(e hylene oxide) and polys y ene-block-poly(me hyl me hac yla e) by sequen- ial anionic polyme iza ion wi h 1-[4-(2- e -bu yldime hylsiloxy)e hyl]phenyl-1- phenyle hylene (Scheme 1-4A). A e dep o ec ion and dep o ona ion he hyd oxyl- unc ion se ed as ini ia o o he anionic ing opening polyme iza ion o ɛ- cap olac one 97 o L-lac ide 98 in he case o polys y ene-block-poly(e hylene oxide) as diblock copolyme , o o e hylene oxide 98 in he case o polys y ene-block-poly(me hyl 1 – In oduc ion 23 me hac yla e). Using a simila DPE de i a i e, Hi ao and co-wo ke s syn hesized hyd oxy mid- unc ionalized polys y ene-block-poly(2-(pe luo ooc yl)-e hyl me hac yla e) diblock copolyme s (Scheme 1-4B). 99 Howe e , a e ans o ma ion o he silyl-p o ec ed hy- d oxyl unc ion in o benzyl b omide, li ing anionic polyme s o 2- inylpy idine and me- hyl me hac yla e we e g a ed o he diblock copolyme s by anionic coupling eac ions. Modi ica ion o he syn he ic s a egy and using a dual hyd oxy- unc ionalized DPE u - he yielded A 3 B, ABC 2 and ABCD mik oa m s a polyme s. Ano he elegan way o in- oduc ion o a hyd oxyl unc ion a he bo de o wo blocks is he use o 2- me hoxyme hoxyme hyloxi ane o he endcapping o li ing polyme anions (Scheme 1- 4C). 62,64,100 The hyd oxy unc ion inhe en ly gene a ed du ing he coupling eac ion wi h he endcappe is used o he ing opening polyme iza ion o e hylene oxide as second block. A e dep o ec ion he second hyd oxy unc ion was used o a ach a ca boxyl- e mina ed poly(pe luo op opylene oxide) ia es e i ica ion 100 o “g a ing- om” o γ- me hyl-ɛ-cap olac one 62 o N,N-dime hylaminoe hyl ac yla e 64 a e pos -modi ica ion o he alcohol o ob ain amphiphilic ABC mik oa m s a e polyme s. In a simila manne polys y ene-block-poly(e hylene oxide) diblock copolyme s bea ing an p ima y amino- 101 o allyl- unc ion 102 a he block bo de we e syn hesized by F ey and co-wo ke s om he co esponding unc ionalized glycidyl e he s. Due o hei de ined mid- unc ionali y, hese a e o possible u u e in e es o he cons uc ion o mik oa m s a polyme s. Howe e , o such glycidyl compounds as endcapping agen s, he polyme iza ion me- hod o he second block is es ic ed o anionic ing-opening polyme iza ion. Also o he app oaches u ilizing DPE o gene a e he mid- unc ionali y he choice o monome sequence is dependen on he eac i i y o he monome s. 91 1 - In oduc ion 24 Scheme 1-4. O e iew o mid- unc ional diblock copolyme s o he cons uc ion o ABC mik oa m s a e polyme s. U ilizing DPE chemis y (A,B) 97-99 o a glycidyl e he (C) 62,64,100 hyd oxyl mid- unc ionalized diblock copolyme s we e syn hesized which se e as p ecu so o he a achmen o he hi d block ia “g a ing- om” o “g a ing- o” app oaches. (D) He e o unc ional Co e Molecules All he examples discussed up o now ake ad an age o anionic polyme iza ions s eps o gene a e well-de ined and unc ionalized polyme s as building blocks o he mik oa m s a e polyme syn hesis. Howe e , in he pas decade, an inc easing numbe o eac- ions ul illing he c i e ia o click chemis y we e u ilized in polyme chemis y o he cons uc ion o a ious polyme a chi ec u es in combina ion wi h con olled adical polyme iza ion me hods. 103 He ein, he e o unc ional co e molecules can se e as com- mon junc ion poin o he “g a ing- om” and “g a ing- o” o di e en polyme seg- men s by a combina ion o such click eac ions wi h s anda d polyme iza ion me hods o cons uc mik oa m s a e polyme s. Fo example Zhang e al. syn hesized a i unc ional co e molecule bea ing an alkyne-, hyd oxyl- and b omine- unc ion. 104 Due o he compa ibili y and ole ance o he eac ion condi ions simul aneous azide-alkyne click chemis y, ing-opening polyme iza ion and ATRP was possible o syn hesize poly- s y ene-a m-poly(ɛ-cap olac one)-a m-poly(N,N-dime ylaminoe hyl me hac yla e) o poly(e hylene oxide)-a m-poly(ɛ-cap olac one)-a m-poly(N,N-dime ylaminoe hyl me h- ac yla e) in a one-po eac ion (Scheme 1-5A). Based on he same p inciples, di e se o he s a egies a e epo ed in li e a u e whe e consecu i e ATRP, ing-opening polyme iza ion, click eac ions like azide-alkyne o hiol-ene click chemis y, es e i ica- ion and ans o ma ion eac ions we e combined o cons uc ABC mik oa m s a e polyme s. 66-68,77,105-110 Fu he mo e, Tunca and co-wo ke s p esen ed an app oach u ilizing h ee o hogonal click eac ions o he cons uc ion o a poly(e hylene oxide)- a m-poly(ɛ-cap olac one)-a m-poly(N-bu yl oxano boneneimide) mik oa m s a 1 – In oduc ion 25 (Scheme 1-5B). 111 Fi s he poly(e hylene oxide) segmen was a ached o he co e mole- cule by a Diels-Alde click eac ion, ollowed by he simul aneous liga ion o he o he wo blocks by azide-alkyne cycloaddi ion and ni oxide adical coupling click eac ion. Huan e . al epo ed a acile s a egy o he cons uc ion o sup amolecula ABC mik oa m s a e polyme s using β-cyclodex in as co e molecule. 112 Recen ly, he g oup o Li demons a ed he Passe ini h ee-componen eac ion o be a powe ul me hod o simul aneously in oduce an ATRP ini a o and an alkyne- unc ion o aldehyde end- unc ionalized poly(e hylene oxide). 113 S a ing wi h his dual- unc ionalized PEO di e se ABC mik oa m s a e polyme s we e accessible ei he by consecu i e ATRP and azide- alkyne cycloaddi ion o simul aneous SET-LRP and click eac ion (Scheme 1-5C). Addi- ionally, in li e a u e di e en o he mik oa m s a a chi ec u es like ABCD, 114 s a - 115 and H-shaped ABCDE mik oa m s a s 116 and he i s ABC mik oa m s a e polyme wi h cyclic a ms 117 a e epo ed by modi ica ions o hese s a egies wi h he e o unc ional co e molecules. Scheme 1-5. Syn he ic s a egies o he cons uc ion o ABC mik oa m s a e polyme s wi h he e o unc ional co e molecules ia simul aneous ATRP, ROP and CuAAC (A), 104 consecu i e iple click eac ions (B) 111 o h ee-componen Passe ini eac ion (C). 113 1 - In oduc ion 26 1.2.1.2 O he Mik oa m S a Polyme Sys ems Hi ao and co-wo ke s designed special eac ion sequences which enable he i e a i e syn hesis o asymme ic s a b anched polyme s by epea ing he sequence. 76 These indi idual s eps in ol e he linking eac ion o a li ing polyme anion wi h a polyme con aining a sui able unc ional g oup (1), which is ollowed by he egene a ion o he unc ional g oup a he eac ion si e (2). Based on he silyl-p o ec ed hyd oxyl- unc ionalized DPE shown in Scheme 1-4B such an i e a i e me hodology is possible, s a ing wi h a li ing polyme anion endcapped wi h his DPE de i a i e. A e ans o - ma ion o he p o ec ed hyd oxy g oup in o a benzyl b omide unc ion a second li ing anion endcapped wi h his DPE is eac ed wi h he b omide, which esembles s ep 1. Regene a ion o he b omide unc ion a he co e om he newly in oduced silyl- p o ec ed hyd oxyl- unc ion esembles s ep 2, which allows o epea ing he cycle (Scheme 1-6). In his manne an ABCD mik oa m s a polyme was syn hesized. 118 How- e e , as a p e equisi e o he e-in oduc ion o he unc ional g oup he li ing polyme anion used in he las s ep has o be nucleophilic enough o copolyme ize wi h DPE. The e o e, o less nucleophilic li ing anions o polyme s like poly(me hyl me hac yla e), a ein oduc ion o he unc ional g oup is no possible and hey ha e o be a ached in he las s ep. This me hodology was also applied o b omide- unc ionalized DPE 119 o b omide- unc ionalized 1,3-bu adiene, 120 which a e used o e-in oduce a non- homopolyme izable DPE o bu adiene end- unc ionali y. Repea ing hese s eps, s a s con aining up o se en di e en a ms could be achie ed. Ano he unc ionali y capable o such linking eac ions wi hou unde going homopolyme iza ion is he α-phenyl ac y- la e, which can also copolyme ize wi h less eac i e monome s like e -bu yl me hac y- la e. Using a dual hyd oxy- unc ionalized DPE de i a i e, which p o ec ing g oups can be selec i ely clea ed o es e i y hem wi h he α-phenylac ylic acid in sepa a e s eps, ABCDE mik oa m s a s we e syn hesized wi h an i e a i e me hodology in he g oup o Hi ao. 121 Fo all hese s a egies addi ional ac iona ion is necessa y o ob ain he pu e mik oa m s a polyme s as he linking eac ions a e conduc ed wi h excess o he li ing anionic polyme chain o gua an ee 100% con e sion. 1 – In oduc ion 27 Scheme 1-6. I e a i e me hodology o he syn hesis o s a b anched polyme s ia eac ion o li ing pol- yme anions wi h a benzyl b omide unc ion. 118 Ano he widely used me hod is he di inylbenzene (DVB) me hod, based on he abili y o p e o med polys y ene a ms o anionically copolyme ize wi h di inylbenzene o o m a co e wi h ac i e eac ion si es. 122 Addi ion o a second monome leads o polyme chains g owing om his co e. The e o e, wi h his “in-ou ” me hod, s a sys ems o he ype A n B n wi h symme ical composi ion and di e en second blocks we e easily accessible as shown by Tsi silianis and co-wo ke s. 123-126 By he addi ion o a hi d monome u he - mo e A n (B-block-C) n mik oa m s a sys ems wi h a polys y ene and poly(2- inylpy idine)- block-poly( e -bu yl ac yla e) segmen s we e syn hesized. 127 Howe e he numbe o a ms ep esen s an a e age alue, which is only oughly adjus able by he amoun o DVB and molecula weigh o he s a ing polyme . 1.2.2 Sel -Assembly in Bulk In addi ion o he sel -assembled s uc u es o mik oa m s a e polyme s in solu ion (see 1.1.2.2), hei mo phology in he bulk s a e ep esen s an unique p ope y. Al eady linea iblock e polyme s we e shown o p oduce a wide a ie y o in e es ing mo - phologies due o he linea connec ion o h ee immiscible blocks wi h h ee di e en se s o in e ac ion pa ame e s. 27,71,128-131 Howe e , o mik oa m s a e polyme s o su icien incompa ibili y he cons i u ion o he h ee polyme segmen s a a common 1 - In oduc ion 28 poin o ces hese o align in a one-dimensional ashion. In con as o ha he wo di - e en junc ion poin s in linea ABC iblock e polyme s a e loca ed a wo-dimensional polyme in e aces. Consequen ly, comple ely seg ega ing mik oa m s a e polyme s a e supposed o assemble in o columna bulk mo phologies (Figu e 1-6A). 132 The wo- dimensional c oss-sec ions o he spa ial a angemen o hese cylinde s can be de- sc ibed by A chimedian iling pa e ns. 132-134 Due o he challenging syn hesis o mik oa m s a e polyme s sys ema ic in es iga- ions o he composi ional in luence on he mo phology a e qui e a e. Besides ea lie mo phological s udies, 135,136 Thomas and co-wo ke s p o ed he alignmen o he junc- ion poin s in a one-dimensional ashion o a sys em consis ing o polys y ene, polyiso- p ene and poly(me hyl me hac yla e). 137 Hücks äd e al. ga e he i s de ailed mo pho- logical s udy on a polys y ene-a m-polybu adiene-a m-poly(2- inylpy idine) sys em o a ious composi ions. 94 Fo ano he sys em con aining polys y ene, polyisop ene and poly(2- inylpy idine) a ms he a ia ion o he olume ac ions yielded mani old s uc- u es o complex iling pa e ns as exempla ily depic ed in Figu e 1-6B. 138-141 When mik oa m s a s o mo e asymme ic composi ions we e in es iga ed, ansi ions o high- ly pe iodic, subs uc u ed lamella mo phologies could be obse ed. 142 Simila ly, Ikkala and co-wo ke s epo ed he hie a chical smec ic sel -assembly o a mik oa m s a e polyme con aining a α-helical poly(ε- e -bu yloxyca bonyl-L-lysine) (PBLL) seg- men . 143 The mo phology composed o an o e all lamella s uc u e wi h one ype o lamella o packed helices o PBLL and a second ype o med om al e na ing ec angula cylinde s o polys y ene and polyisop ene (Figu e 1-6C). In addi ion Abe z e al. demon- s a ed ha blending o polys y ene-a m-polybu adiene-a m-poly(2- inylpy idine) mik oa m s a s wi h diblock copolyme s can be u ilized o une he ob ained mo pholo- gy wi hin ce ain limi s. 144 1 – In oduc ion 29 Figu e 1-6. (A) Schema ic illus a ion o he one-dimensional alignmen o junc ion poin s and he esul ing columna a angemen o he indi idual blocks o ABC mik oa m s a e polyme s. 132 (B) TEM mic og aphs o ou polyisop ene-a m-polys y ene-a m-poly(2 inylpy idine) (µ-ISV) mik oa m s a e polyme s ( op ow) and he co esponding schema ic iling pa e ns (bo om ow): (a) µ-I 1.0 S 1.8 I 1.0 , (b) µ-I 1.0 S 1.8 I 1.6 , (c) µ- I 1.0 S 1.8 I 2.0 , (d) µ-I 1.0 S 1.8 I 2.9 ( he subsc ip s deno e he co esponding olume a ios). The da k, ligh and g ay domains co espond o PI, PS, and P2VP phases. 141 (C) TEM mic og aph and schema ic illus a ion o he bulk mo phology o a µ-(PS)(PI)(PBLL) mik oa m s a . The sample is s ained wi h OsO 4 (PI da k, PS and PBLL ligh ). 143 1.2.3 Applica ion o ABC Mik oa m S a Te polyme s o Func ional Ma e- ials Due o he b oad a ie y o hie a chies ob ained bo h in solu ion and bulk (see 1.1.2.2 and 1.2.2), mik oa m s a e polyme s a e o special in e es o he design o no el ma- e ials wi h compa men alized s uc u es. Up o now only a limi ed numbe o publica- ions has been dealing wi h he applica ion o mik oa m s a e polyme s in ma e ials esea ch, mos p obably due o he complica ed syn hesis and accessibili y o la ge amoun s o ma e ial. Howe e , his si ua ion will change wi h he ad ances ob ained in mik oa m s a syn hesis du ing he las decade. 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The e o e, a new app oach o he syn hesis o a ious ABC mik oa m s a e polyme s was de eloped. Mid-alkyne unc ionalized diblock copolyme s, which we e syn hesized by anionic polyme iza ion u ilizing 1,1-diphenyle hylene (DPE) chemis y, o m he key building blocks. These allowed modula liga ion wi h azido- unc ionalized homopolyme s ia azide-alkyne Huisgen cycloaddi ion, which is p esen ed in Chap e 3. Fo a mik oa m s a e polyme con aining a poly(2- inylpy idine block) (P2VP) he mechanism o hie a chical supe s uc u e o ma ion was in es iga ed. The polyme a chi ec u e, qua e niza ion o P2VP and iiodide as coun e ion we e ound o be p e equisi es o he di ec ed sel -assembly in o complex “woodlouse”-shaped agg ega es. The o ma ion o cylind ical s uc u es om sphe ical micelles, he meande - like a angemen o hese cylinde s and, inally, he usion in o lamella agg ega es o ba el-like shape was shown o be he unde lying mechanism. Chap e 4 includes he esul s o his coun e ion-media ed hie a chical supe s uc u e o ma ion. Wi h he knowledge o he necessa y pa ame e s o such a di ec ed sel -assembly, his s udy was u he expanded o o he ABC mik oa m s a e polyme s and an ABA’ mik oa m s a copolyme . Enabled by ou modula app oach he in luence o molecula and chemical composi ion on he ob ained supe s uc u e was e alua ed (Chap e 5). He ein, s uc u es simila o he “woodlouse” agg ega es we e obse ed, howe e om a di e en mechanism s a ing wi h esicles as p ima y building uni s ins ead o micelles. Below, an o e iew o e he mos impo an esul s o each o he h ee ollowing chap e s is gi en. The eade is e e ed o he co esponding chap e o an ex ensi e 2 – O e iew o he Thesis 42 discussion on he pa icula opic o syn hesis and sel -assembly o he mik oa m s a e polyme s. 2.1 Modula Syn hesis o Mik oa m S a Te polyme s Besides ou es elying exclusi ely on anionic polyme iza ion, he syn hesis o mik oa m s a e polyme s is ypically accomplished by a combina ion o di e en polyme iza ion me hods. The e o e, specially designed linking eac ions in combina ion wi h p ecisely unc ionalized diblock copolyme s and pos unc ionaliza ion/ ans o ma ion eac ions ha e o be conduc ed. In his chap e we p esen a mo e gene alized me hod, which can be applied o a b oad a ie y o ( unc ional) monome s. The syn he ic s eps in ol e sequen ial anionic polyme iza ion o alkyne mid- unc ionalized diblock copolyme s and subsequen liga ion wi h azido- unc ionalized diblock copolyme s (Scheme 2-1). Scheme 2-1. Syn he ic ou e o ABC mik oa m s a e polyme s by he combina ion o anionic polyme iza ion in THF wi h DPE chemis y and click chemis y. Fo he p epa a ion o he alkyne mid- unc ionalized diblock copolyme we u ilized DPE chemis y, as i allows exac inco po a ion o exac ly one DPE uni a he block bo de unde app op ia e choice o monome sequence. Fo his pu pose, we syn hesized a no el DPE de i a i e ca ying a p o ec ed alkyne- unc ion (click-DPE, 1-[(4-( e - bu yldime hylsilyl)-e hynyl)phenyl]-1-phenyle hylene). Due o he conjuga ion o he π- sys em wi h he alkyne- unc ion in pa a-posi ion he co esponding li ing anion exhibi s a ba hoc omic shi in he UV-spec um as compa ed o unsubs i u ed DPE (Figu e 2-1A). 2 – O e iew o he Thesis 43 As a consequence o his al e ed elec onic con igu a ion click-DPE was shown o copolyme ize wi h li ing anions o poly(2- inylpy idine) (P2VP) by MALDI-ToF MS in es iga ion o an app op ia e es eac ion (Figu e 2-1B), in con as o o he DPE de i a i es epo ed in li e a u e. Hence, he li ing anion o click-DPE can p incipally bo h be used o s a he anionic polyme iza ion 2- inylpy idine (2VP) and o end unc ionaliza ion o li ing anions o P2VP. Figu e 2-1. (A) UV- is abso p ion spec um o he adduc o 1,1-diphenyle hylene (do ed line) and 1-[(4- ( e -bu yldime hylsilyl)e hynyl)phenyl]-1-phenyle hylene (solid line) wi h sec-BuLi in THF. The inse shows he pic u e o he cu e e con aining he li ing anion o he unsubs i u ed DPE ( op) and he click-DPE (bo om). (B) MALDI-ToF MS spec a o poly(2- inylpy idine) be o e (black line) and a e addi ion o click- DPE (cDPE, ed line), eco ded in e lec on mode (B) wi h AgTFA as ioniza ion agen .  He e, we p epa ed di e en alkyne mid- unc ionalized diblock copolyme s wi h polybu adiene as i s block and poly(2- inylpy idine), poly( e -bu yl me hac yla e) (P BMA), o poly(N,N-dime hylaminoe hyl me hac yla e) (PDMAEMA) as second block. Excep hyd olysis no u he ans o ma ion eac ions a e necessa y o gene a e he alkyne- unc ion. We de e mined he deg ee o alkyne- unc ionaliza ion by a es click eac ion wi h an azido- unc ional pe ylene bisimide as ch omopho e in combina ion wi h UV measu emen s. Fo he diblock copolyme s wi h P2VP as second block only a sligh o e -inco po a ion o he click-DPE could be achie ed unde app op ia e eac ion condi ions (~120% alkyne- unc ionaliza ion). This is due o he addi ion o a second DPE uni o some chains. In con as , o P BMA as second block nea -quan i a i e unc ionaliza ion was p o en (~93% alkyne unc ionaliza ion). λ max = 549 nm λ max = 500 nm 4750 4800 4850 4900 4950 VP45 VP42cDPE1 VP41cDPE1 VP44 m/z 4780 4800 4820 VP 44 VP41cDPE1 ~ 4.5 g/mol A B 400 450 500 550 600 650 700 abso bance λ [nm] 2 – O e iew o he Thesis 44 By a modula combina ion o hese mid- unc ional diblock copolyme s wi h azido- unc ionalized homopolyme s we syn hesized di e en no el ABC mik oa m s a e polyme s ia coppe -ca alyzed azide-alkyne Huisgen cycloaddi ion. Fo he example o a PB-b-P BMA diblock copolyme (cBT2) he mik oa m s a e polyme s ob ained a e click eac ion wi h azido- unc ional polys y ene (PS), poly(e hylene oxide) (PEO) and PDMAEMA homopolyme s a e lis ed in Table 2-1. The co esponding SEC eluog ams a e depic ed in Figu e 2-2. In all cases na owly dis ibu ed mik oa m s a e polyme s we e ob ained. We u he showed o click- eac ions wi h PEO and PDMAEMA homolopolyme s ha a eac ion pa hway wi h an excess o he homopolyme was possible and monomodal mik oa m s a e polyme s we e achie ed a e app op ia e pu i ica ion p ocedu es, like e.g. dialysis (Figu e 2-2B). Table 2-1. Molecula Cha ac e iza ion o ABC Mik oa m S a Te polyme s Ob ained om Click Reac ions wi h Alkyne Mid-Func ionalized PB 111 -b-P BMA 42 Diblock Copolyme cBT2 (Subsc ip s Deno e he Co esponding Deg ees o Polyme iza ion) sample ID a DP(3 d block) b M n, h. c [kg/mol] M n,app d [kg/mol] Ɖ d µ-BT2S1, (µ-B 38 T 38 S 23 15.9 ) 36 15.9 20.5 1.03 µ-BT2S2, (µ-B 33 T 33 S 3318.1 ) 57 18.1 22.4 1.03 µ-BT2E, (µ-B 32 T 32 E 3518.8 ) 150 18.8 25.4 1.03 µ-BT2D, (µ-B 34 T 34 D 3117.6 ) 34 17.6 20.8 1.11 a The supe sc ip deno es he heo e ical numbe a e age molecula weigh o he ABC mik oa m s a e polyme , as calcula ed om he espec i e alues o he diblock copolyme and homopolyme and he indices he heo e ical weigh ac ion o he espec i e blocks. b Calcula ed om he molecula weigh o he 3 d block. c Theo e ical molecula weigh . d Appa en molecula weigh and dispe si y de e mined by SEC wi h polys y ene calib a ion. Fo he mik oa m s a e polyme s con aining PDMAEMA THF-SEC wi h addi ional 0.25 w % e abu ylammonium b omide (TBAB) was used. 2 – O e iew o he Thesis 45 Figu e 2-2. SEC aces o mik oa m s a e polyme s om click eac ions o a polybu adiene-block- poly( e -bu yl me hac yla e) diblock copolyme (cBT2) wi h (A) wo polys y ene homopolyme s (PS1-N 3 , PS2-N 3 ) and a poly(e hylene oxide) homopolyme (PEO-N 3 ) and (B) a poly(N,N-dime hylaminoe hyl me hac yla e) homopolyme (PDMAEMA-N 3 ). Fo (B) THF-SEC wi h addi ional sal was used and as he click eac ion was conduc ed wi h 2 equi o he PDMAEMA-N 3 , excess homopolyme was emo ed ia dialysis. Wi h his modula app oach a a ie y o unc ional mik oa m s a e polyme s a e accessible. Fu he mo e, click-DPE o e s no el syn he ic ad an ages in he cons uc ion o polyme ic a chi ec u es ia a combina ion o anionic polyme iza ion and click- chemis y. The mik oa m s a e polyme s syn hesized he ein all ca y a polybu adiene block, which can be used o u he modi ica ion and unc ionaliza ion ia hiol-ene chemis y. Mo eo e new amphiphilic mik oa m s a e polyme s we e syn hesized, which ca y hyd ophilic (PEO, PDMAEMA) and s imuli- esponsi e a ms (PDMAEMA, P BMA a e hyd olysis). 2.2 Coun e ion-Media ed Hie a chichal Sel -Assembly o an ABC Mik oa m S a Te polyme Con aining a Poly(N-me hyl-2- inylpy idinium iodide) Segmen Fo a mik oa m s a e polyme con aining a polybu adiene, poly( e -bu yl me hac yla e) and poly(2- inylpy idine) segmen (µ-BVT), qua e niza ion wi h me hyl iodide and ans e in o aqueous solu ion was obse ed o yield wo s uc u ally comple ely di e ing limi ing cases o agg ega ion o ms: sphe ical micelles (d micelle = 24.5 ± 2.0 nm) and pa icles wi h a complex lamella in e io (200-500 nm in longi udinal axis). The concen a ion o iiodide as pola izable coun e ion o he qua e nized P2VP 26 28 30 32 34 36 38 0.0 0.2 0.4 0.6 0.8 1.0 cBT2 PS1$N 3 PS2$N 3 PEO$N 3  $BT2S1  $BT2S2  $BT2E no malized RI signal V e [mL] 14 15 16 17 18 19 0.0 0.2 0.4 0.6 0.8 1.0 V e [mL] cBT2 PDMAEMA$N 3 . $BT2D, undial. . $BT2D, dial. A B 2 – O e iew o he Thesis 52 Felix H. Schache , Hi oshi Jinnai and Axel H. E. Mülle we e in ol ed in scien i ic discussions and co ec ion o he manusc ip . Chap e 5 This wo k is published in Polyme 2013, 54, 4528-4537 unde he i le: “Hie a chical Sel -Assembly o Mik oa m S a Polyme s Con aining a Polyca ionic Segmen : A Gene al Concep ” by And eas Hanisch, And é H. G öschel, Melanie Fö sch, Tina I. Löbling, Felix H. Schache ,* and Axel H. E. Mülle * I w o e he publica ion and conduc ed mos o he expe imen s. Excep ions a e s a ed below: And é H. G öschel was in ol ed in scien i ic discussions. Melanie Fö sch pe o med almos all TEM and c yo-TEM measu emen s. Tina I. Löbling eco ded some o he TEM mic og aphs. Felix H. Schache and Axel H. E. Mülle we e in ol ed in scien i ic discussions and co ec ion o he manusc ip . 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 53 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s ia a New Alkyne-Subs i u ed Diphenyle hylene De i a i e And eas Hanisch, Holge Schmalz, and Axel H. E. Mülle ABSTRACT: We in oduce a modula ou e o he syn hesis o well-de ined ABC mik oa m s a e polyme s. To his aim, he syn hesis o a 1,1-diphenyle hylene de i a- i e bea ing a p o ec ed alkyne unc ion (1-[(4-( e -bu yldime hylsilyl)e hynyl)phenyl]- 1-phenyle hylene) was de eloped. This compound was o he i s ime employed in sequen ial anionic polyme iza ion o eadily p epa e alkyne mid- unc ionalized diblock copolyme s wi h polybu adiene as i s and a poly(alkyl me hac yla e) (poly( e -bu yl me hac yla e), poly(N,N-dime hylaminoe hyl me hac yla e)) as second block. Fo he hi d a m con olled adical polyme iza ion me hods (polys y ene, poly( e -bu yl me h- ac yla e), poly(N,N-dime hylaminoe hyl me hac yla e)) and anionic ing-opening polyme iza ion (poly(e hylene oxide)) we e used o sepa a ely p epa e homopolyme s wi h an azido unc ion. A e wa d, azide-alkyne Huisgen cycloaddi ion was success ully employed o syn hesize a lib a y o ABC mik oa m s a e polyme s wi h di e en mo- lecula weigh s and chemical composi ions ia modula combina ion o he unc ional- ized diblock copolyme s and homopolyme s. The esul ing new ABC mik oa m s a e polyme s showed na ow, monomodal molecula weigh dis ibu ions wi h dispe si ies ypically below 1.10, as de e mined by size exclusion ch oma og aphy. 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 54 3.1 In oduc ion Polyme a chi ec u e and composi ion is a decisi e ac o o con olling s uc u e o - ma ion bo h in bulk and in solu ion. Besides he choice o monome and he numbe and sequence o polyme ic building blocks he way o molecula conjunc ion o hese blocks is o ou e mos impo ance. Compa ed o hei linea analogues, mik oa m s a e polyme s, whe e he polyme chains a e connec ed a one common junc ion poin ,1 p esen a much mo e complex sys em. A a ie y o unique bulk mo phologies we e ound,2-4 which a e inaccessible by linea iblock e polyme s. This is a di ec conse- quence o he con ined geome y o he polyme chains, which o ces he common junc- ion poin s o he h ee di e en blocks o be aligned in a 1-dimensional ashion. These mik oa m s a e polyme mo phologies we e also u ilized o gene a ing cylind ical, compa men alized pa icles by selec i e c osslinking o one block.5 In addi ion, heo e - ical s udies sugges ha e en mo e complex mo phologies a e expec ed unde cylind i- cal con inemen o ABC mik oa m s a e polyme s.6 Depending on he leng h a io o he espec i e a ms, Hillmye , Lodge and co-wo ke s7,8 ob ained di e en mul icompa men s uc u es in aqueous solu ion om ABC mik oa m s a e polyme s wi h a luo ina ed segmen . These anged om hambu ge micelles o segmen ed wo mlike micelles and nanos uc u ed esicles. The conjunc ion o h ee chains a one common linking poin leads o special e- qui emen s conce ning he syn he ic s a egy. The main di icul ies a e (1) he exac unc ionaliza ion o a diblock copolyme wi h a unc ional g oup o polyme block a he junc ion o he wo blocks and (2) he s oichome y o he linking eac ion. Fou di e - en syn he ic s a egies ha e been epo ed so a o mik oa m s a e polyme s. One app oach uses he di e en eac i i y o li ing anionic polyme chains owa d he chlo ine-silicon g oups o ichlo osilanes. Using me hyl ichlo osilane as linking agen , Hadjich is idis and cowo ke s syn hesized a mik oa m s a e polyme consis ing o poly- isop ene, polys y ene, and polybu adiene.9 Howe e , he sequence o in oduc ion o a ms is s ongly limi ed o he eac i i y and s e ic demand o he polyme ic anions, and adap ion o o he monome s equi es a modi ica ion o he me hod.10,11 The second anionic app oach u ilizes mac omonome s wi h nonhomopolyme izable endg oups 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 55 which we e di ec ly sequen ially copolyme ized wi h wo so s o monome s.2,12-14 In ano he app oach, i s diblock copolyme s we e syn hesized ca ying unc ional g oups a he junc ion poin o he wo blocks. Again, nonhomopolyme izable compounds o special polyme iza ion s a egies ha e o be employed o assu e ha only one unc ional g oup is loca ed exac ly in be ween he wo polyme ic building blocks. Up o now, main- ly hyd oxyl- unc ions we e used o conjuga e he hi d block, which we e in oduced ia a 1,1-diphenyle hylene (DPE) de i a i e15,16 o 2-me hoxyme hyloxyme hyloxi ane17-19 as end-capping molecule. This hyd oxyl unc ion se es as ancho ing poin o he hi d block, which can be “g a ed om” by anionic ing opening polyme iza ion (AROP)15-17 o con olled adical polyme iza ion (CRP) a e app op ia e modi ica ion19 o “g a ed o” ia di ec es e i ica ion.18 Hi ao and co-wo ke s ecen ly used in-chain-benzyl b omide- unc ionalized diblock copolyme s o he mik oa m s a e polyme syn hesis in ol ing specially designed anionic linking eac ions.20 The possibili y o cons uc ing complex polyme ic a chi ec u es wi h con olled adi- cal polyme iza ion me hods inc eased wi hin he las decade.21,22 This suppo ed he de elopmen o new s a egies o he syn hesis o ABC mik oa m s a e polyme s whe e anionic polyme iza ion is no in ol ed. In addi ion, click eac ions in gene al, o coppe (I)-ca alyzed azide-alkyne cycloaddi ion (CuAAC) in pa icula ha e become pow- e ul ools o highly e icien linking o polyme ic building blocks.23-25 These no el syn- he ic ou es ake ad an age o he o hogonali y o he di e en polyme iza- ion/liga ion me hods, which is a p e equisi e o he syn hesis o well-de ined ABC mik oa m s a e polyme s. By designing special co e molecules wi h h ee di e en unc ionali ies, he a ms can be “g a ed on o” o “g a ed om” in di e en s eps.26-31 He ein, we p esen a mo e gene al me hod by combining anionic polyme iza ion, employed o he syn hesis o eadily alkyne mid- unc ionalized diblock copolyme s, wi h o he polyme iza ion echniques ia azide-alkyne Huisgen cycloaddi ion (Scheme 3- 1). Ba ne -Kowollik and co-wo ke s al eady epo ed he liga ion o alkyne mid- unc ionalized homopolyme s wi h azide-bea ing homopolyme s.32 Howe e , only A2B mik oa m s a e polyme s we e accessible. The syn hesis o an alkyne mid- unc ionalized diblock copolyme consis ing o polys y ene and poly( e -bu yl ac yla e) was also epo ed.33,34 Ne e heless, mul iple polyme analogous eac ions had o be 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 56 conduc ed and/o swi ching he polyme iza ion sys em was necessa y. Thus, we syn he- sized 1-[(4-( e -bu yldime hylsilyl)e hynyl)phenyl]-1-phenyle hylene o di ec ly p epa e mid-chain alkyne-subs i u ed diblock copolyme s ia sequen ial anionic polyme iza ion. Because o he di ec in oduc ion o he alkyne-g oup in o he diblock copolyme by he use o DPE chemis y, a wide a ie y o unc ional diblock copolyme s a e accessible, and excep hyd olysis no edious ans o ma ion eac ions ha e o be conduc ed. The ob- ained diblock copolyme s we e a e wa d liga ed wi h di e en azide-bea ing homopolyme s p epa ed by a om ans e adical polyme iza ion (ATRP), e e sible ad- di ion- agmen a ion chain ans e (RAFT) polyme iza ion, and anionic ing opening polyme iza ion ia azide-alkyne click chemis y. Di e en no el ABC mik oa m s a e polyme s we e syn hesized u ilizing his modula combina ion. Scheme 3-1. Syn he ic ou e o he syn hesis o ABC mik oa m s a e polyme s o he example o a s a consis ing o a ms o polybu adiene, poly( e -bu yl me hac yla e), and polys y ene 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 57 3.2 Expe imen al Sec ion Ma e ials. Fo he pu i ica ion o monome s, eagen s and sol en and he syn hesis o used compounds see he Suppo ing In o ma ion. Syn hesis o 1-[(4-( e -Bu yldime hylsilyl)e hynyl)phenyl]-1-phenyle hylene (Click- DPE). The s a ing molecule 1-(4-b omophenyl)-1-phenyle hylene was syn hesized as desc ibed elsewhe e.35 The syn he ic p o ocol o he coupling eac ion wi h he p o- ec ed alkyne de i a i e was adop ed om he li e a u e.36 Fi s , 1-(4-b omophenyl)-1- phenyle hylene was dissol ed in pipe idine (~50 mg/mL) and bis( iphenylphosphine)palladium(II) dichlo ide (0.03 equi ) and CuI (0.004 equi ) we e added unde a ni ogen a mosphe e. A e degassing wi h ni ogen o 30 minu es, e - bu yldime hylsilylace ylene (1.2 equi ) was added d opwise a 50 °C, and he solu ion was s i ed o e nigh . The eac ion mix u e was il e ed, he sol en e apo a ed, and he c ude p oduc was dissol ed in THF. A e addi ion o wa e , he p oduc was ex ac ed wi h hexane o h ee imes. The o ganic ac ions we e d ied o e sodium sul a e, and he sol en was e apo a ed o ob ain a b own oil. This was u he pu i ied by column ch oma og aphy wi h hexane as sol en o ob ain a clea oil. Dis illa ion o he p oduc was no possible e en unde high acuum condi ions. Be o e he use in anionic polyme - iza ion, he compound was eeze-d ied om benzene solu ion on a high acuum line and subsequen ly dissol ed in d y THF o ob ain a s ock solu ion o low iscosi y. sec- BuLi was added d opwise ( ypically 1-2 d ops) unde ni ogen low un il a deeply iole colo was ob ained. Besides p o ing he absence o impu i ies, he pe sis en colo also se ed as indica ion o he pu i y o he s ock solu ion. 1H NMR (300 MHz, CDCl3): δ = 7.50-7.27 (m, 9H, A ), 5.50 (s, 2H, -C=CH2), 1.03 (s, 9H, SiC(CH3)3), 0.22 (s, 6H, SiCH3) Polyme iza ions. Sequen ial Li ing Anionic Polyme iza ion in THF. All polyme iza ions we e ca ied ou a low empe a u es in a he mos a ed labo a o y au ocla e (Büchi) unde d y ni ogen a mosphe e using THF as sol en . The day be o e polyme iza ion, eshly dis illed THF (~250 mL o 15 g o polyme ) was ea ed wi h 1 mL o sec-BuLi pe 100 mL o sol en a -20 °C, ollowed by s i ing o e nigh o o m li hium alkoxides. These exhibi s abilizing e ec s on he li ing chain end o polybu adiene and u he - mo e in he case o me hac yla e- ype monome s no addi ion o LiCl is necessa y o ob- 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 58 ain well-de ined polyme s.37 Fo he diblock copolyme s, i s bu adiene was ini ia ed wi h sec-BuLi a -70 °C and hen polyme ized a -50 o -15 °C depending on he block leng h o he di e en polyme iza ions. To ensu e comple e consump ion o he bu adi- ene he polyme iza ion was ollowed by in-line NIR ibe -op ic spec oscopy. The li ing polybu adienyl anion was end-capped wi h 1-[(4-( e - bu yldime hylsilyl)e hynyl)phenyl]-1-phenyle hylene (1.1-1.5 equi ) a -50 °C o 2 h. Be o e and a e he addi ion o he click-DPE samples o SEC and MALDI-ToF MS we e wi hd awn. In he case o poly(2- inylpy idine) (P2VP) as second block he monome was added a -70 °C and polyme ized o 5 minu es. A e e mina ion wi h degassed me ha- nol he polyme (cBV) was isola ed by p ecipi a ion in wa e . Fo he diblock copolyme s wi h e -bu yl me hac yla e ( BMA) he monome addi ion was conduc ed a -70 °C and he polyme iza ion ook place a -45 °C o 1 h be o e i was e mina ed wi h degassed me hanol. A e he addi ion o he BMA he iole colo o he DPE end-capped li ing anion slowly aded away. The inal polyme (cBT) was p ecipi a ed in a mix u e o iso- p opanol/wa e 3/1 ( / ). Fo he hi d ype o diblock he N,N-dime hylaminoe hyl me hac yla e (DMAEMA) was injec ed in o he eac o a -70 °C and polyme ized o 1 h. Immedia ely a e addi ion o he monome he colo o he DPE end-capped polybu adienyl anion anished comple ely. A e e mina ion wi h degassed me hanol, he esul ing polyme (cBD) was dialyzed agains THF and inally eeze-d ied om dioxane. The molecula weigh s o he diblock copolyme s wi h P2VP o PDMAEMA as second block we e calcula ed by a combina ion o MALD-ToF MS and 1H NMR. The e- o e, he Mn o he polybu adiene (PB) p ecu so polyme was de e mined by mass spec- ome y. By he ela i e mola a ios o he cha ac e is ic signals o PB (5.10-5.60 ppm, 2H 1,2-PB; 5.70-5.15 ppm, 1H 1,2-PB and 2H 1,4-PB) and P2VP (8.50-8.00 ppm, 1H) o PDMAEMA (4.30-3.90 ppm, 2H) he o e all molecula weigh was calcula ed using he PB p ecu so molecula weigh as e e ence. The Mn o he PB-b-P BMA diblock copoly- me s was di ec ly measu ed by MALDI-ToF MS. All he da a o he diblock cha ac e iza- ion a e summa ized in Table 3-1. Alkyne Dep o ec ion. Fo he dep o ec ion p ocedu e, he espec i e polyme was dis- sol ed in THF (~0.1 g/mL) and degassed o 30 min. Then, 10 equi o e abu ylammonium luo ide (1 M solu ion in THF) ela i e o alkyne unc ions we e 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 59 added a 0 °C and s i ed a his empe a u e o 2 h. A e wa ming up o oom empe - a u e he solu ion was s i ed o e nigh . The polyme was p ecipi a ed in pu e wa e o isop opanol/wa e 3/1 ( / ) o PB-b-P2VP and PB-b-P BMA, espec i ely. A e wa d, he polyme was dissol ed in THF and dialyzed agains THF (MWCO 1 000 g/mol) o emo e impu i ies. Finally, he polyme s we e eeze-d ied om dioxane. In he case o he diblock copolyme wi h DMAEMA he eac ion mix u e was di ec ly dialyzed agains THF p io o eeze-d ying. Click Reac ions. Fo he de e mina ion o he deg ee o unc ionaliza ion wi h alkyne- subs i u ed DPE, es click eac ions wi h N-(1-hep yloc yl)-N′-(hexyl-6′-azido)-pe ylene- 3,4,9,10- e acaboxylic acid bisimide we e conduc ed. In a ypical un, 100 mg o he dep o ec ed diblock copolyme we e dissol ed in 5 mL THF in a sc ew-cap glass. Then, 2 equi o he azido- unc ionalized pe ylene bisimide ela i e o he alkyne unc ion was added. A e pu ging wi h ni ogen o 10 min, 1 equi o CuB was added, ollowed by u he degassing o 10 min. By addi ion o 1 equi o N,N,N′,N′,N′′- pen ame hyldie hylene iamine (PMDETA) as ligand, he click eac ion was s a ed and s i ed a oom empe a u e o 3 days. A e e mina ion by exposu e o ai , he e- maining coppe was emo ed by il a ion o e a sho silica gel column. P epa a i e SEC wi h THF as eluen was employed o emo e excess pe ylene. An in ense ed-colo ed polyme was ob ained a e eeze-d ying om dioxane. The click eac ions o he cons uc ion o he mik oa m s a e polyme s we e con- duc ed in a simila manne . Typically, he mola a ios o alkyne unc ion:azido unc- ion:CuB :PMDETA was se o 1:1:1:1 (i no s a ed elsewhe e), and he polyme concen- a ion was ~20 mg/mL in THF. The eac ions we e conduc ed a oom empe a u e and ollowed by wi hd awing samples o SEC measu emen s. Finally, when no u he changes in he SEC eluog amms we e obse ed, he esul ing mik oa m s a e polyme was pu i ied by passing h ough a small silica gel column o emo e coppe . The poly- me s we e ob ained as whi e powde s a e eeze-d ying om dioxane. 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 60 3.3 Resul s and Discussion 3.3.1 Syn hesis o 1-[(4-( e -Bu yldime hylsilyl)e hynyl)phenyl]-1- phenyle hylene (Click-DPE) The key compound o his modula ou e owa d ABC mik oa m s a e polyme s (Scheme 3-1) was di ec ly syn hesized by he palladium-ca alyzed Sonogashi a coupling eac ion be ween 1-(4-b omophenyl)-1-phenyle hylene and e -bu yldime hylsilyl p o- ec ed ace ylene. Simila compounds, like 4-( ime hylsilyl)e hynyls y ene36 and me hac- yla e de i a i es as 3- ime hylsilyl-2-p opynyl me hac yla e,38 ha e al eady been syn- hesized and used in anionic polyme iza ion o ob ain polyme s wi h p edic able mo- lecula weigh s and na ow dis ibu ions. As known o DPE and i s de i a i es homopolyme iza ion o such compounds is no possible due o s e ic hind ance.39 In con as o me hods epo ed in li e a u e,33,34 his syn he ic ad an age enables us o use one sequen ial polyme iza ion p ocess o selec i ely inco po a e exac ly one alkyne unc ion be ween he wo polyme ic blocks unde adequa e choice o monome s. The click-DPE was pu i ied ia column ch oma og aphy o gi e a clea iscous oil. The chemi- cal s uc u e and he pu i y we e con i med by 1H NMR spec oscopy (Figu e 3-1a). Figu e 3-1. 1H NMR spec a o (a) 1-[(4-( e -bu yldime hylsilyl)e hynyl)phenyl]-1-phenyle hylene (click- DPE), (b) he p o ec ed alkyne- unc ionalized diblock cBT2, and (c) he co esponding diblock a e hyd o- lysis o he silyl-p o ec ing g oup (sol en signals a e s iked ou ). 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 61 The UV- is spec um o he deeply iole solu ion o he li ing anion (Figu e 3-2) gene - a ed by eac ion o 1-[(4-( e -bu yldime hylsilyl)e hynyl)phenyl]-1-phenyle hylene wi h sec-BuLi shows a maximum a 549 nm. In con as , he li ing anion o unsubs i u ed 1,1- diphenyle hylene has a deep ed colo , and we de e mined he abso p ion maximum a a ound 500 nm unde he same condi ions.40 This clea ba hoc omic shi o igina es om he conjuga ion o he π-sys em wi h he p o ec ed alkyne g oup. Simila ly, Tsuda e al. epo ed a b ownish- ed colo in he anionic polyme iza ion o 4- ( im hylsilyl)e hylens y ene,36 in con as o he o ange colo o polys y yl anions.41 Figu e 3-2. UV- is abso p ion spec um o he adduc o 1,1-diphenyle hylene (do ed line) and 1-[(4-( e - bu yldime hylsilyl)e hynyl)phenyl]-1-phenyle hylene (solid line) wi h sec-BuLi in THF. The inse shows he pic u e o he cu e e con aining he li ing anion o he unsubs i u ed DPE ( op) and he click-DPE (bo - om). 3.3.2 Syn hesis o Alkyne Mid-Func ionalized Diblock Copolyme s Bu adiene was chosen o he i s block and o he second block 2- inylpy idine (2VP) o me hac yla e ype monome s as e -bu yl me hac yla e ( BMA) and N,N- dime hylaminoe hyl me hac yla e (DMAEMA). The anionic polyme iza ion o bu adiene was conduc ed in THF a -50 o -15 °C (depending on he espec i e block leng h) using sec-BuLi as ini ia o . A e comple e con e sion – as ollowed in si u wi h ibe -op ics NIR spec oscopy – 1.1 o 1.5 equi o he click-DPE s ock solu ion was added ia sy inge a -50 °C. The addi ion o he diphenyle hylene de i a i e was clea ly isible by he imme- 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 68 weigh s close o 100 000 g/mol we e p oduced ia CuAAC,49,50 we suppose ha in ou case he alkyne unc ion a he block junc ion is s e ically less amenable han an alkyne unc ion in α- o ω-posi ion. Ou i s ials wi h highe molecula weigh polyme s (ex- ceeding 40 000-50 000 g/mol o he indi idual compounds) we e no success ul. The e- o e, we chose mode a e molecula weigh s o he indi idual compounds, so ha he o e all molecula weigh did no exceed 25 000 g/mol and he unc ional g oups we e no oo dilu ed. The cou se o he click eac ions wi h cBT diblock copolyme s was ol- lowed by SEC. The eac ions we e conduc ed unde equimola condi ions. A e 24 h he molecula weigh dis ibu ion o he diblock copolyme s cBT1 and cBT2 shi ed com- ple ely (see Figu e 3-3 and Figu e 3-S4), esembling he success ul gene a ion o he de- si ed mik oa m s a e polyme . Howe e , despi e equimola eac ion condi ions in bo h cases he e was s ill a mino amoun o un eac ed homopolyme le . Fu he SEC meas- u emen s showed ha he peak o he PS1-N3 al eady disappea ed nea ly comple ely a e 11.5 h (Figu e 3-S5) o he click eac ion wi h cBT2. Inc easing he eac ion ime did no lead o any change in he eluog amm. The e o e, we addi ionally ca ied ou a eac ion be ween cBT1 and only 0.8 equi o PS1-N3. Unde he same eac ion condi- ions, again, a compa able amoun o homopolyme was le ( esul s no shown). We assume ha du ing polyme iza ion and wo k-up o he homopolyme , a mino pa o he b omine g oup is elimina ed, as al eady epo ed in li e a u e.51 This leads o a small amoun o non-azido- unc ionalized homopolyme which canno ake pa in he click eac ion (see discussion o MALDI-TOF spec um in he Suppo ing In o ma ion). This was also con i med by he click eac ion o cBT2 wi h PS3-N3 (Figu e 3-S6). Unde equimola eac ion condi ions no u he liga ion ook place a e 15 h, leading o a non- negligible amoun o un eac ed diblock and homopolyme . In con as o he click eac- ions wi h lowe molecula weigh polys y enes he con e sion o he click eac ion wi h PS3-N3 was much lowe a e 3.25 h (whe e he eac ion was mo e o less inished in he o he cases). Fu he addi ion o 0.5 equi o homopolyme esul ed in a comple e shi o he diblock due o comple e click e iciency as depic ed in Figu e 3-S6. Thus, he de- g ee o un unc ionalized polys y ene inc eased wi h inc easing molecula weigh . 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 69 26 28 30 32 34 36 0.0 0.2 0.4 0.6 0.8 1.0 no malized RI signal V e [mL] cBT1 PS1-N3 PS2-N3 µ -BT1S1 µ -BT1S2 Figu e 3-3. THF-SEC (RI signal) o alkyne- unc ionalized diblock cBT1, azido- unc ionalized PS (PS1-N3, PS2- N3) and he co esponding ABC mik oa m s a e polyme s (µ-BT1S1 and µ-BT1S2) ob ained a e equimola click eac ion o 24 h. Ne e heless, comple e shi s o he molecula weigh dis ibu ions o he mik oa m s a e polyme s owa d lowe elu ion olumes compa ed o he p is ine diblock copolyme s we e de ec ed in all cases. This demons a es he success ul o ma ion o ABC mik oa m s a e polyme s. Addi ionally, he o e all shi s o he esul ing ABC mik oa m s a e polyme s we e no oo dis inc . This is in acco dance wi h heo y, whe e he hyd o- dynamic adius o s a polyme s is expec ed o be smalle han he hyd odynamic adius o a linea polyme wi h he same composi ion and molecula weigh due o he highe segmen al densi y o s a polyme s.52 All syn hesized µ-BTS s a e polyme s exhibi ed symme ical peaks wi h a na ow molecula weigh dis ibu ion. The heo e ical molecu- la weigh s we e calcula ed om he known molecula weigh s o he liga ed diblock copolyme s. These a e shown oge he wi h he appa en molecula weigh s and he espec i e dispe si ies in Table 3-3. Fu he mo e an exempla ily 1H NMR o µ-BT1S1 is shown in Figu e 3-S7. All cha ac e is ic signals o he polyme ic building blocks a e p e- sen in he ob ained mik oa m s a e polyme . 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 70 Table 3-3. Molecula Cha ac e iza ion o ABC Mik oa m S a Te polyme s Ob ained om Click Reac ions wi h Alkyne Mid-Func ionalized Diblock Copolyme s sample IDa DP(PB)b DP(2nd block)c DP(3 d block)d Mn, h. e [kg/mol] Mn,app [kg/mol] Ɖ µ- BT1S1 (µ-B 63 T 18 S 18 20.7) 242 27 36 20.7 28.0 1.03 µ- BT1S2 (µ-B 57 T 17 S 26 22.9) 242 27 57 22.9 30.0 1.03 µ- BT1E (µ-B 55 T 16 E 28 23.6) 242 27 150 23.6 32.7 1.04 µ- BT1D (µ-B 58 T 17 D 24 22.4) 242 27 34 22.4 31.5 1.10 µ- BT2S1 (µ-B 38 T 38 S 23 15.9) 111 42 36 15.9 20.5 1.03 µ- BT2S2 (µ-B 33 T 33 S 33 18.1) 111 42 57 18.1 22.4 1.03 µ-BT2S3 g (µ-B 29 T 29 S 40 20.5) 111 42 80 20.5 25.0 1.03 µ- BT2E (µ-B 32 T 32 E 35 18.8) 111 42 150 18.8 25.4 1.03 µ- BT2D (µ-B 3 4 T 34 D 31 17.6) 111 42 34 17.6 20.8 1.11 µ-BDT g (µ-B 40 D 28 T 31 24.5) 181 44 53 24.5 26.3 1.08 μ-BDE g (µ-B 42 D 29 E 28 23.5) 181 44 150 23.5 32.5 1.06 aThe numbe -a e age molecula weigh o he ABC mik oa m s a e polyme was calcula ed om he espec i e alues o diblock copolyme s and he homopolyme s. The e o e, he supe sc ip deno es he heo e ical numbe -a e age molecula weigh o he ABC mik oa m s a e polyme and he indices he heo e ical weigh ac ion o he espec i e blocks. bDeg ee o polyme iza ion (DP) de e mined om MALDI-ToF MS o he PB-p ecu so . cCalcula ed by he di e ence in Mn de e mined by he MALDI-ToF MS spec a o diblock and p ecu so . dCalcula ed om he molecula weigh o he 3 d block. eTheo e ical mo- lecula weigh . Appa en molecula weigh and dispe si y de e mined by SEC wi h polys y ene calib a ion. Fo he mik oa m s a e polyme s con aining PDMAEMA THF-SEC wi h addi ional 0.25 w % e abu ylammonium b omide (TBAB) was used. gIn hese cases he appa en molecula weigh and dispe is y we e de e mined excluding he sepa a ed homopolyme peak. Click Reac ions wi h ω-Azido-Func ionalized Poly(e hylene oxide). These i s success ul es eac ions wi h azido- unc ionalized polys y ene p o e he easibili y o ou ap- p oach. To p epa e amphiphilic mik oa m s a e polyme s we conduc ed click eac ions wi h PEO-N3. Fi s a emp s unde equimola eac ion condi ions led o a non-negligible amoun o un eac ed diblock copolyme . The e o e, he click eac ions we e ca ied ou wi h an excess o he unc ionalized PEO-N3. By s epwise addi ion o PEO-N3 o he eac- ion mix u e, he equi alen s necessa y o a comple e click eac ion we e de e mined 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 71 (Figu e 3-S8). The e o e, in a i s s ep, he eac ion wi h 1 equi o PEO-N3 was ollowed by SEC. As a e 24 h no u he change in he eluog am was de ec ed, and s ill a signi i- can amoun o un eac ed diblock copolyme was le o e , 0.5 equi o PEO-N3 was added subsequen ly. Finally, a e 3.5 h eac ion ime, u he 0.2 equi o PEO-N3 was added o he click eac ion con aining al eady 1.5 equi . As his did no esul in a change compa ed o he eluog am a e eac ion wi h 1.5 equi o PEO-N3, he equi a- len s necessa y o comple e liga ion we e de e mined o be 1.5 equi . The eason o he use o such a huge excess o PEO-N3 s ill emains unclea . F om he amoun o coupled p oduc (~6 w % om SEC) only a sligh excess o he azido- unc ionalized compound would be easonable. Howe e , maybe pa ial elimina ion o he azido g oup o PEO o uncomple e unc ionaliza ion due o side eac ion du ing he end-capping eac ion could be a easible explana ion (see discussion o MALDI-ToF spec- um in he Suppo ing In o ma ion). The espec i e SEC aces o he click eac ions wi h cBT1, cBT2, and cBD (whe e he a io o cBX:PEO-N3 was minimum 1:1.5) a e shown in Figu e 3-4. In case o he diblock copolyme s wi h P BMA as second block he excess PEO-N3 was easily emo ed du ing pu i ica ion om coppe wi h a sho column o silica due o in e ac ions wi h he column. Using PEO-N3 in excess and subsequen emo al o un eac ed homopolyme gua an eed he comple e con e sion o he alkyne unc ion. Howe e , in he case o he diblock cBD con aining DMAEMA a highe amoun o PEO homopolyme was le o e a e wo k-up. Fu he pu i ica ion wi h a column, c ys alli- za ion om cold THF, o dialysis in aqueous media did no educe he amoun o he undesi ed PEO homopolyme . A possible explana ion could be ha PEO o ms hyd ogen bonds wi h PDMAEMA, which he e o e p e en he comple e emo al o excess PEO homopolyme . He e, one has o no ice ha o u he applica ions, whe e he PEO se es as co ona in aqueous solu ions, his mino amoun o PEO homopolyme is no p oblema ic. In con as o he p e ious click eac ions, dis inc shi s o he molecula weigh dis- ibu ions o he ABC mik oa m s a e polyme s compa ed o he co esponding diblock p ecu so s we e de ec ed. The absence o a peak om esidual diblock also gi es e i- dence ha all diblock copolyme chains ca y an alkyne unc ion. Like o he click eac- 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 72 A B 14 15 16 17 18 0.0 0.2 0.4 0.6 0.8 1.0 cBD PEO-N3 µ -BDE, dial. V e [mL] 26 28 30 32 34 0.0 0.2 0.4 0.6 0.8 1.0 no malized RI signal V e [mL] cBT1 cBT2 PEO-N3  BT1E  BT2E ions wi h PS-N3 low dispe si ies o he esul ing ABC mik oa m s a e polyme s we e de ec ed (Table 3-3). Figu e 3-4. THF-SEC (A) o alkyne- unc ionalized diblock copolyme s cBT1, cBT2, he azido- unc ionalized PEO-N3, and he co esponding ABC mik oa m s a e polyme s (µ-BT1E and µ-BT2E) ob ained a e azide- alkyne click coupling and sal -THF-SEC aces (B) o cBD, PEO-N3, and he esul ing ABC mik oa m s a e polyme (µ-BDE) a e dialysis. In all cases he RI-signals a e shown. Click Reac ions wi h ω-Azido-Func ionalized Poly(N,N-dime hylaminoe hyl me hac yla e). To ob ain PDMAEMA as a wa e -soluble polyme , which esponds bo h o pH and em- pe a u e,53 we used an azido-subs i u ed CTA.44 As he gene al applicabili y o ou ap- p oach unde equimola condi ions was p o en we conduc ed he click eac ions wi h a 2- old excess o PDMAEMA-N3 o 5 days, wi hou op imizing he eac ion ime. Wi h his eac ion pa hway we we e also able o gua an ee 100% con e sion o he alkyne- compound. The SEC aces o he aw p oduc o he click eac ions o he azido- unc ionalized PDMAEMA-N3 wi h he wo cBT diblock copolyme s a e shown in Figu e 3-5A and 3-5B, espec i ely. In bo h cases he molecula weigh dis ibu ions o he mik oa m s a e polyme s shi ed comple ely in he co esponding SEC- aces com- pa ed o hei diblock copolyme p ecu so s. The excess PDMAEMA-N3 was emo ed by dialysis in a mix u e o me hanol/isop opanol (2/1 / ), whe e he mik oa m s a e polyme o ms micelles wi h a polybu adiene co e. The e o e, a dialysis memb ane wi h a cu -o (50 000 g/mol) much highe han he molecula weigh o he homopolyme was used. The SEC aces o he ob ained mik oa m s a e polyme s a e plo ed in Figu e 3-5A and B. In he case o µ-BT1D he iny coupling shoulde a lowe elu ion olume became mo e p onounced a e dialysis. The e o e, we assume ha his 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 73 A B 14 15 16 17 18 19 0.0 0.2 0.4 0.6 0.8 1.0 V e [mL] cBT2 PDMAEMA-N3 µ -BT2D, undial. µ -BT2D, dial. 14 15 16 17 18 19 0.0 0.2 0.4 0.6 0.8 1.0 no malized RI signal V e [mL] cBT1 PDMAEMA-N3 µ -BT1D, undial. µ -BT1D, dial. shoulde migh be he esul o some agg ega ion e ec s, which occu ed o his eac- ion. Ano he explana ion could be he oxida ion o he amine, which leads o he o - ma ion o amine oxide. Fo he dialysis o µ-BT2D e en unde ex ensi e exchange o he sol en , he small esidual peak om un eac ed PDMAEMA-N3 could no be emo ed comple ely. Howe e , compa ed o he p oduc peak his peak is negligible. The molecu- la cha ac e iza ion o he µ-BTD mik oa m s a e polyme s is lis ed in Table 3-3. In a simila way a µ-BDT mik oa m s a e polyme consis ing o he same monome uni s was syn hesized by liga ion o cBD wi h P BMA-N3 (see Suppo ing In o ma ion and Ta- ble 3-3). Figu e 3-5. Sal -THF-SEC (RI signal) o µ-BTD mik oa m s a e polyme s ob ained by click coupling o he alkyne- unc ionalized diblock copolyme s (A) cBT1 and (B) cBT2 wi h he azido- unc ionalized PDMAEMA- N3. As in bo h cases 2 equi o PDMAEMA we e used wi hin he click eac ion, SEC aces o he aw p o- duc and he mik oa m s a e polyme a e dialysis a e shown. 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 74 3.4 Conclusions This gene al modula ou e o he syn hesis o ABC mik oa m s a e polyme s com- bines anionic polyme iza ion echniques wi h CRP me hods and anionic ing opening polyme iza ion using azide-alkyne Huisgen cycloaddi ion as liga ion me hod. Fo his pu pose, we success ully employed ou alkyne-modi ied DPE de i a i e in sequen ial anionic polyme iza ion. In he case o me hac yla e- ype monome s well-de ined alkyne mid unc ional diblock copolyme s consis ing o a i s block o PB and a second block o ei he P BMA o PDMAEMA we e syn hesized. Click eac ions wi h azido- unc ionalized pe ylene bisimide e i ied he success ul inco po a ion o only one DPE uni a he block junc ion. In con as o o he DPE de i a i es, we obse ed ha click-DPE can copoly- me ize wi h 2VP, which o e s he ad an age o p oducing alkyne mid- unc ionalized diblock copolyme s wi h P2VP as i s block and a me hac yla e as second block. The e- o e, 2VP and bu adiene and s y ene and hei de i a i es can be used as i s blocks and me hac yla e ype monome s as second block o ob ain well de ined-diblock copolyme s bea ing an alkyne unc ion a he block junc ion. By using a oolbox o di e en azido- unc ionalized homopolyme s, we demons a e ha a a ie y o well-de ined ABC mik o a m s a e polyme s is accessible ia his modula app oach. These exhibi ed monomodal molecula weigh dis ibu ions wi h small dispe is ies close o 1.10 o e en lowe . The ob ained mik oa m s a e polyme s a e in e es ing new polyme s ega ding hei bulk mo phologies and solu ion-based sel -assembly. Resul s om he sel - assembly o hese ABC mik oa m s a e polyme s will be epo ed in subsequen publi- ca ions. A u he ad an age is ha due o he use o anionic polyme iza ion mik oa m s a e polyme s wi h a polybu adiene block a e accessible, which can a e wa d be unc ionalized h ough hiol-ene click chemis y54 o used o selec i e c osslinking.5,55 Fu he mo e, he ABC mik oa m s a e polyme con aining a ms o polybu adiene, poly(N,N-dime hylaminoe hyl me hac yla e) and poly(e hylene oxide) is a p omising candida e in he ield o bio echnological applica ions like gene deli e y.56-58 Ou ap- p oach enables he upscaling o he syn hesis o ABC mik oa m s a e polyme s o mo e han one g am. Bo h syn he ic s eps o diblock and homopolyme syn hesis a e well- es ablished polyme iza ion me hods, and no u he complex syn he ic p ocedu es a e 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 75 necessa y, as compa ed o o he me hods. Because o he s ong e sa ili y o click chemis y and he inc easing amoun o publica ions dealing wi h i , we expec ou click- DPE o o e a a ie y o new possibili ies in he design o no el cus om polyme a chi- ec u es and ma e ials. Suppo ing In o ma ion. Ma e ials sec ion, polyme iza ion p ocedu es, cha ac e iza ion sec ion, cha ac e iza ion o ω-azido homopolyme s, click eac ion wi h poly( e -bu yl me hac yla e) and discussion o he eac i i y o click-DPE owa d li ing P2VP-Li; UV- is spec a o pe ylene-labeled diblock copolyme s, SEC aces o polybu adiene p ecu so s, poly(e hylene oxide) and click eac ions, MALDI-ToF mass spec a o polybu adiene p e- cu so s and an exempla ily 1H NMR spec um o µ-BT1S1. This In o ma ion is a ailable ee o cha ge ia he In e ne a h p://pubs.acs.o g/. Acknowledgmen s. This wo k was suppo ed by he Deu sche Fo schungsgemeinscha wi hin SPP 1165 (Mu896/22). We hank Ch is o B. Ts e ano and Ma kus Müllne o ui ul discussion. Melanie Fö sch, Annika P a enbe ge , and Ch is ophe V. Syna schke a e acknowledged o pe o ming MALDI-ToF MS measu emen s and Ma ie - a Böhm, Ka ha ina Neumann, and Robin Pe au o conduc ing SEC measu emen s. We u he hank And é H. G öschel o p o iding one o he polys y ene homopolyme s, S e an Reinicke o he syn hesis o he azidoace yl chlo ide, and Julia Ewe o RAFT polyme iza ion o he PDMAEMA homopolyme . And eas S. Lang is acknowledged o p o iding he azido- unc ionalized pe ylene bisimide. 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 76 3.5 Re e ences (1) Hadjich is idis, N. J. Polym. Sci., Pa A: Polym. Chem. 1999, 37, 857-871. (2) Hücks äd , H.; Göp e , A.; Abe z, V. Mac omol. Chem. Phys. 2000, 201, 296-307. (3) Junnila, S.; Houbeno , N.; Hanski, S.; Ia ou, H.; Hi ao, A.; Hadjich is idis, N.; Ikkala, O. Mac omol- ecules 2010, 43, 9071-9076. (4) Ma sushi a, Y.; Hayashida, K.; Takano, A. Mac omol. Rapid Commun. 2010, 31, 1579-1587. 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Biomac omolecules 2012, 13, 857-866. 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 84 26 28 30 32 34 36 38 0.0 0.2 0.4 0.6 0.8 1.0 no malied RI signal V e [mL] cBT2 PS1-N3 click: 3.25 h click: 11.5 h and 48 h Figu e 3-S5. THF-SEC (RI-signal) o samples wi hd awn du ing equimola click eac ion be ween cBT2 and PS1-N3 a e 3.25 h (-), 11.5 h and 48 h (···). 26 28 30 32 34 36 0.0 0.2 0.4 0.6 0.8 1.0 cBT2 PS3-N3 click: 1 eq./3.25 h click: 1 eq./48 h click: 1.5 eq./25 h no malized RI signal V e [mL] Figu e 3-S6. THF-SEC (RI signal) o click eac ion be ween cBT2 and PS3-N3 wi h u he addi ion o 0.5 equi PS3-N3 48 h a e s a o he click eac ion. 25 h a e addi ion o he second po ion o PS3-N3 a inal sample was aken o SEC. 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 85 Figu e 3-S7. 1H NMR spec um o µ-BT1S1 in CDCl3. The cha ac e is ic signals o he espec i e polyme blocks a e highligh ed. 26 28 30 32 34 36 0.0 0.2 0.4 0.6 0.8 1.0 no malized RI signal V e [mL] cBT2 PEO-N3 click: 1 eq./1.75 h click: 1. eq./24 h click: 1.5 eq./3.5 h and click: 1.7 eq./1.75 h Figu e 3-S8. THF-SEC (RI signal) o click eac ion be ween cBT2 and PEO-N3 wi h s epwise addi ion o PEO- N3. Fu he 0.5 equi PEO-N3 we e added 24 h a e s a o he click eac ion and 3.5 h la e u he 0.2 equi PEO-N3 we e added. The co esponding eac ion imes a e ela i e o he poin o addi ion o a u - he PEO-N3 po ion. The excess o PEO-N3 is no isible as he PEO is adso bed a he silica column used o emo ing coppe ca alys . 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 86 3.6.3 Tes Polyme iza ion wi h P2VP and Click-DPE In con as o polybu adienyl and polys y yl li hium, he li ing anion o poly(2- inylpy idine) is less nucleophilic and hence, he ini ial assump ion was ha i is no able o a ack excess click-DPE. This would lead o he desi ed diblock copolyme s wi h only one click-DPE a he block junc ion. I he 2VP would inco po a e mo e uni s o click-DPE he diblock would ca y mo e han on alkyne unc ion, which would esul in s a e polyme s wi h mo e han h ee a ms. Hogen-Esch e al. epo ed ha he endcapping o P2VPLi wi h unsubs i u ed DPE is an equilib ium eac ion9 and ha e en he addi ion o a la ge excess o 1,1-diphenyle hylene o li ing poly(2- inylpy idine) does no lead o a signi ican endcapping o he anion.10 Hi ao e al. showed ha silyl-p o ec ed hyd oxyl- unc ionalized DPE, 1-(3- e -bu yldime hylsilyloxyme hylphenyl)-1-phenyle hylene did no unde go copolyme iza ion wi h li ing poly(2- inylpy idine) anions.11 Also, in p e ious es s on he endcapping o poly(2- inylpy idine) wi h DPE unc ionalized polybu adiene mac omonome s, we ound no signi ican coupling be ween he wo polyme ic chains. The e o e, a es eac ion was conduc ed o e i y whe he he alkyne-subs i u ed click- DPE can be a acked by a li ing P2VPLi as a esul o he al e ed elec onic con igu a ion o he DPE de i a i e. Fo his pu pose, 2- inylpy idine was polyme ized in THF a -70 °C wi h sec-BuLi as ini ia o , yielding a molecula weigh o 5 000 g/mol. Then, a 2- old ex- cess o he alkyne- unc ionalized click-DPE was added a he same empe a u e. Di ec ly a e injec ion, he colo o he P2VPLi anion changed om ed o iole gi ing a i s quali a i e indica ion o he addi ion o he click-DPE. 1H NMR de e mined he deg ee o alkyne unc ionaliza ion as a ound 98%. MALDI-ToF MS suppo ed a quan i a i e end- unc ionaliza ion (Figu e 3-S9A). Be o e addi ion he Mn o he P2VP was 4 750 g/mol. A e 1 h eac ion wi h click-DPE he Mn inc eased o 5 230 g/mol. These alues indica e ha click-DPE addi ion ook place quan- i a i ely, conside ing he accu acy o he de e mina ion me hod. Mo eo e , he mo- lecula weigh dis ibu ion shi ed comple ely and in he case o he e lec on mode a dis inc shi o he indi idual peaks was no iceable (Figu e 3-S9B). This p o ed he p es- ence o only one species a e addi ion o click-DPE. The shi o he peaks wi h a di e - ence o a ound 4.5 g/mol is in good ag eemen wi h he expec ed 3 g/mol di e ence (M(2VP) = 105.14 g/mol, M(click-DPE) = 318.53 g/mol; Δ = 318.53 g/mol - 3 x 105.14 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 87 g/mol = 3.11 g/mol). Fo example, he peak a m/z = 4791.6 in he P2VP p ecu so , i.e. be o e addi ion o click-DPE, co esponds o 44 epea ing uni s o 2VP wi h a sec-bu yl esidue and Ag+ as coun e -ion. This is in acco dance wi h he heo e ical m/z alue o 4792.0. In he spec um a e click-DPE addi ion, he modi ied P2VP wi h he same num- be o epea ing uni s and an addi ional click-DPE was de ec ed a m/z = 5111.1. Also he e, his alue is consis en wi h he expec ed 5110.5 (= 4792.0 + 318.5). Hence, click- DPE adds o li ing poly(2- inylpy idine) anions quan i a i ely, in con as o o he DPE de i a i es epo ed in li e a u e. Figu e 3-S9. MALDI-ToF MS spec a o poly(2- inylpy idine) be o e (black line) and a e addi ion o click- DPE (cDPE, ed line), eco ded in linea (A) and e lec on mode (B) wi h AgTFA as ioniza ion agen . In summa y, he anion o he alkyne- unc ionalized DPE has a nucleophilici y compa able o P2VPLi, whe eas he anion o unsubs i u ed DPE is mo e nucleophilic han P2VPLi. One eason o his migh be he elec on wi hd awing e ec o he e - bu yldime hylsilyl g oup which leads o a educ ion o he elec on densi y in he double bond and hus dec eases he nucleophilici y o he click-DPE as compa ed o he unsubs i u ed DPE.12 As he li ing anion o he click-DPE is able o s a he polyme iza- ion o P2VP and click-DPE also adds o P2VPLi, i is possible o copolyme ize P2VP and click-DPE. The e o e, i he click-DPE is used in a sligh excess and no unde equimola condi ions, he DPE de i a i e migh also be inco po a ed in o mos p obably a e con- sump ion o all 2VP monome a he end o he P2VP block o he aimed PB-b-P2VP diblock copolyme s. In con as o bu adiene and s y ene, me hac yla es a e known o be no nucleophilic enough o a ack diphenyle hylene and i s de i a i i es.13,14 Conse- 4780 4800 4820 VP44 VP41cDPE1 ~ 4.5 g/mol 4750 4800 4850 4900 4950 VP45 VP42cDPE1 VP41cDPE1 VP44 m/z 2000 4000 6000 8000 P2VP-p ec P2VP-DPE i P2VP-p ec i P2VP-DPE m/z 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 88 quen ly, monome s like BMA and DMAEMA can be polyme ized as second block o ob- ain well-de ined diblock copolyme s wi h exac ly one alkyne unc ion a he block junc- ion. Due o he al e ed eac i i y o he click-DPE owa d li ing P2VP-Li compa ed o o he DPE de i a i es bo h α- and ω-alkyne modi ied P2VP homopolyme s a e accessible. In addi ion, alkyne mid- unc ionalized diblock copolyme s wi h poly(2- inylpy idine) as i s and a me hac yla e- ype monome as second block can be syn hesized. 3.6.4 Cha ac e iza ion o ω-Azido Homopolyme s Besides he quali a i e p oo o he p esence o an azido unc ion ia IR (Figu e 3-S10) he deg ee o azide- unc ionaliza ion was addi ionally ied o be quan i ied by MALDI- ToF MS o 1H NMR. 4000 3500 3000 2500 2000 1500 1000 P BMA-N3 PDMAEMA-N3 PEO-N3 ela i e ansmission wa enumbe cm-1 PS-1-N3 Figu e 3-S10. IR spec a o he co esponding homopolyme s show he cha ac e is ic s e ching ib a ion o he azido g oup a ~ 2098 cm-1. In case o PS1-N3 and PEO-N3 mass spec ome y in e lec on mode was possible. The espec i e spec a a e shown in Figu e 3-S11 and 3-S12. In he case o he azido- e mina ed PS1-N3 only a small peak esembling he desi ed unc ional species is p e- sen . As al eady epo ed in li e a u e o ma ion o pos sou ce me as able ions om azido- unc ionalized polyme s is possible unde high lase powe s in e lec on mode.15 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 89 The e o e, by compa ing he ela i e in ensi ies o he azido- unc ionalized species and he co esponding me as able ion wi h he in ensi y o he un unc ionalized species, as main popula ion he azido- e mina ed polys y ene is clea ly concluded. We ha e o men ion he e ha he peak a he le side o he p o on- e mina ed polys y ene could no be assigned clea ly. 3320 3360 3400 3440 3480 [PS29H+Ag]+[PS30H+Ag]+ [PS31N3-N2+Ag]+ me as able [PS30N3-N2+Ag]+ me as able [PS31N3+Ag]+ O O N3 x m/z = PSxN3 [PS30N3+Ag]+  ~ 104 Figu e 3-S11. MALDI-ToF MS spec um o PS1-N3 eco ded in e lec on mode wi h AgTFA as ioniza ion agen . In he MALDI spec um o PEO-N3 (Figu e 3-S12) wo se ies o peaks we e p esen . He e, he main popula ion esembles he azido- e mina ed PEO. The second popula ion could be a ibu ed o azido- e mina ed PEO wi h a p o on as coun e ion. Ano he possible explana ion could be deduced om SEC measu emen s, whe e a small second peak a lowe elu ion olume was de ec ed (a ound 6 w .%, see Figu e 3-S13). This o igina es om minimal esidual amoun s o hionyl chlo ide which was used o he syn hesis o he azidoace yl chlo ide and is ha d o emo e comple ely despi e ho ough pu i ica- ion.1 This popula ion canno ake pa in he click eac ion. Fo he molecula cha ac e - iza ion, only he majo peak was aken in o conside a ion. The molecula weigh de e - mined by MALDI-ToF MS was consis en wi h he alues om SEC. The e o e, he second popula ion in MALDI can be in e p e ed as he lowe molecula weigh side o he cou- pling p oduc wi h hionyl chlo ide. E en hough he main peak co esponds o he de- 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 90 si ed unc ionalized PEO, one has o no ice ha chains whe e HN3 was elimina ed (Δ ~43 g/mol) and chains which we e e mina ed wi h a p o on ins ead o he acid chlo ide (Δ ~5 g/mol) would no be dis inguishable and could possibly be p esen . 6180 6200 6220 6240 6260 [PEO126SO+Ag]+[PEO127SO+Ag]+ = PEOx+ySO OS O O x y [PEO133N3+H]+[PEO134N3+H]+  ~ 44 = PEOxN3 O O N3 x [PEO131N3+Ag]+ m/z [PEO132N3+Ag]+ Figu e 3-S12. MALDI-ToF MS spec um o PEO-N3 eco ded in e lec on mode wi h AgTFA as ioniza ion agen . 27 28 29 30 31 32 33 34 0.0 0.2 0.4 0.6 0.8 1.0 no malized RI signal V e [mL] PEO-N3 Figu e 3-S13. THF-SEC (RI signal) o azido- unc ionalized poly(e hylene oxide). Bo h in case o P BMA-N3 and PDMAEMA-N3 endg oup de e mina ion was no possible ia MALDI-ToF MS. In con as o P BMA-N3 o he DMAEMA homopolyme calcula ion 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 91 o he deg ee o azido- unc ionaliza ion was possible om he cha ac e is ic signals in he 1H NMR spec um (Figu e 3-S14). Acco ding o he calcula ed alue o app oxima ely 96 %, quan i a i e azido- unc ionaliza ion o he polyme was assumed. Figu e 3-S14. 1H NMR spec um o PDMAEMA-N3 in CDCl3. The cha ac e is ic signals o he monome uni and chain ans e agen a e highligh ed. 3.6.5 Click Reac ion wi h ω-Azido-Func ionalized Poly( e -bu yl me hac y- la e) Since we also syn hesized an alkyne- unc ionalized diblock wi h DMAEMA as second block (cBD), a µ-BDT mik oa m s a e polyme was achie able by click eac ion wi h he azido- e mina ed P BMA. Simila o he click eac ions wi h PDMAEMA-N3, he eac ion condi ions we e no op imized. The liga ion was conduc ed o 3 days wi h 1.3 equi o azido- unc ionalized homopolyme o gua an ee 100 % conjuga ion o he diblock. The SEC o he click-p oduc a e dialysis in me hanol/isop opanol (2/1 / ) is shown in Fig- u e 3-S15 and a comple e shi o he molecula weigh dis ibu ion o he mik oa m s a e polyme compa ed o he diblock e polyme was de ec ed. This demons a es he e iciency o ou app oach. E en hough we we e no able o emo e all excess homopolyme , we showed ha o ma ion o a µ-BDT mik oa m s a e polyme is possi- 3 – A Modula Rou e o he Syn hesis o ABC Mik oa m S a Te polyme s 92 ble s a ing om wo di e en diblock copolyme s. The molecula pa ame e s o he ob ained ABC mik oa m s a e polyme a e summa ized in Table 3-3. 14 15 16 17 18 19 0.0 0.2 0.4 0.6 0.8 1.0 cBD P BMA-N3 µ -BDT, dial. no malized RI signal Ve [mL] Figu e 3-S15. Sal -THF-SEC (RI-signal) o he µ-BDT mik oa m s a e polyme s ob ained a e click eac ion o alkyne- unc ionalized diblock cBD wi h he azido unc ionalized P BMA-N3. 3.6.6 Re e ences (1) Reinicke, S.; Schmalz, H. Colloid Polym. Sci. 2011, 289, 497-512. (2) Gondi, S. R.; Vog , A. 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(13) Hsieh, H. L.; Qui k, R. P. In Anionic Polyme iza ion: P inciples and P ac ical Applica ions; Ma cel Dekke : New Yo k, 1996. (14) Qui k, R.; Yoo, T.; Lee, Y.; Kim, J.; Lee, B. Ad . Polym. Sci. 2000, 153, 67-162. (15) Li, Y.; Hoskins, J. N.; S ee ama, S. G.; G ayson, S. M. Mac omolecules 2010, 43, 6225-6228. 4 – “Woodlouse” S uc u es om µ-BVqT 93 4 – Coun e ion-Media ed Hie a chical Sel -Assembly o an ABC Mik oa m S a Te polyme And eas Hanisch, And é H. G öschel, Melanie Fö sch, Ma kus D echsle , Hi oshi Jinnai, Thomas M. Ruhland, Felix H. Schache , and Axel H. E. Mülle ABSTRACT: Di ec ed sel -assembly p ocesses o polyme ic sys ems ep esen a powe ul app oach o he gene a ion o s uc u al hie a chy in analogy o biological sys ems. He ein, we u ilize iiodide as a s ongly pola izable coun e ion o induce hie a chical sel -assembly o an ABC mik oa m s a e polyme comp ising a polybu adiene (PB), a poly( e -bu yl me hac yla e) (P BMA), and a poly(N-me hyl-2- inylpy idinium) (P2VPq) segmen . He eby, he mik oa m a chi ec u e in conjunc ion wi h an inc easing a io o iiodide e sus iodide coun e ions allows o a s epwise assembly o sphe ical micelles as ini ial building blocks in o cylind ical s uc u es and supe s uc u es he eo . Finally, mic ome e -sized mul icompa men pa icles wi h a pe iodic lamella ine s uc u e a e obse ed, o which we in oduce he e m “woodlouse”. The coun e ion-media ed de- c ease in hyd ophilici y o he co ona- o ming P2VPq block is he unde lying igge o induce his hie a chical s uc u e o ma ion. All indi idual s eps and he co esponding in e media es owa d hese well-de ined supe s uc u es we e in ensi ely s udied by sca e ing and elec on mic oscopic echniques, including ansmission elec on mic o omog aphy. Keywo ds: ABC Mik oa m S a Te polyme , Hie a chical Sel -Assembly, Polyelec oly es, Mul icompa men Micelles, Poly(2- inylpy idine) 4 – “Woodlouse” S uc u es om µ-BVqT 100 Be o e s udying he sel -assembly mechanism in de ail, we we e in e es ed in he in lu- ence o he polyme s uc u e i sel on he ob ained agg ega es. The e o e, ma e ials o simila chemical composi ion bu di e en a chi ec u e we e used, mo e p ecisely a diblock copolyme (B 109 V 81 ) and a linea iblock e polyme (B 1108 V 142 T 93 ). Howe e , only ill-de ined agg ega es we e ob ained om hese linea polyme s a e qua e niza ion and compa able p epa a ion condi ions (see ep esen a i e mic og aphs o he iblock e polyme in Figu e 4-S4). These indings demons a e ha he mik oa m a chi ec u e i sel is an impo an c i e ion o he hie a chical sel -assembly in o subs uc u ed pa i- cles. Hadjich is idis and co-wo ke s al eady showed bo h expe imen ally and by heo e - ical in es iga ions ha he micelliza ion beha iou o A 2 B mik oa m sys ems is di e en compa ed o linea AB diblock copolyme s due o opological di e ences a he co e- co ona in e ace. 45 4.2.2.1 Hie a chical Supe s uc u e Fo ma ion Acco ding o c yo-TEM (Figu e 4-1D), he “woodlouse” agg ega es can be p elimina ily desc ibed as ei he supe s uc u es om cylind ical micelles o lamellae. Howe e , as he pe iodici y o hese pa icles is in he size ange o he diame e obse ed o he sphe ical micelles in Figu e 4-1B, we assume hese o ac as p ima y building blocks. The di e en s uc u es obse ed would hen simply co espond o di e en le els o supe - s uc u e o ma ion. Rega ding he o e all mo phology, simila s uc u es, such as la e - ally s uc u ed esicles o laye ed s uc u es, we e al eady p edic ed by simula ions o ABC mik oa m s a e polyme s wi h wo sol ophobic blocks. 46,47 Mo e de ailed in o ma- ion abou he supe s uc u e o ma ion was gained om in es iga ions o he in e me- dia e s uc u es ob ained unde compa able p epa a ion condi ions (Figu e 4-2A). He e, he agg ega es clea ly consis o se e al in e wined and pa ially w apped-up cylind ical micelles. Again, he dimensions o he cylinde s a e in good ag eemen wi h he diame- e o he sphe ical micelles (d cylinde = 27.5 ± 2.5 nm, as compa ed o d micelle = 24.5 ± 2.0 nm), which co obo a es he assump ion ha sphe ical micelles a e indeed he unde ly- ing building blocks. S aining wi h OsO 4 e eals a PB (o mixed PB/P BMA) co e o he cylinde s (Figu e 4-2B). Subsequen ly, hese cylinde s align in a pa allel, ibbon-like ash- ion, which we ega d as ano he in e media e le el on he way owa d highly pe iodic 4 – “Woodlouse” S uc u es om µ-BVqT 101 lamella “woodlice”. Ou assump ion o a mixed PB/P BMA co e is u he suppo ed by he calcula ed phase seg ega ion pa ame e , o PB/P BMA, χN ~1.1. 48 Acco ding o he heo y o phase sepa a ion o diblock copolyme s, his is in he diso de ed egime due o he small deg ees o polyme iza ion o ou sys em. 49,50 Fu he mo e, cas ilms o PB- b-P BMA diblock copolyme s wi h compa able molecula weigh s did no show any phase sepa a ion in he bulk ( esul s no shown). C yo-TEM shows highly in e wined and en angled cylind ical micelles (Figu e 4-2C). A su p isingly highly egula packing o indi idual cylinde s can be obse ed in some cases, highligh ed in he inse s and g ay scale analysis o Figu e 4-2C. Rema kably, some o he s uc u es show long- ange o de and dimensions o up o 1 µm in leng h.  Figu e 4-2. TEM (A,B) and c yo-TEM mic og aphs (C) o in e media e micella s uc u es om µ-BVqT in aqueous solu ion. The inal polyme concen a ion was 0.2 g/L o TEM and 0.4 g/L o c yo-TEM. Whe eas o (A) no s aining was pe o med, he sample in (B) was s ained wi h OsO 4 . He e, he g ay scale analysis o he highligh ed a ea e eals a ibbon-like a angemen o cylinde s. Wi h he example o hese in e media e s uc u es, we u he in es iga ed he in lu- ence o wo o he pa ame e s du ing he p ocess o s uc u e o ma ion ( o de ails see Suppo ing In o ma ion): (i) he solu ions we e he mally annealed, and (ii) he ime in be ween qua e niza ion in dioxane and dialysis o wa e was inc eased o up o 1 mon h. Whe eas he mal annealing did no esul in highe s uc u al pe ec ion a all, he s uc u es we e sligh ly mo e de eloped a e 1 mon h in dioxane p io o dialysis (Figu e 4-S5). None heless, om all he TEM in es iga ions discussed so a , di e en 4 – “Woodlouse” S uc u es om µ-BVqT 102 le els o hie a chy om app oxima ely 20 nm o 1 µm we e de ec ed o µ-BVqT, s a - ing om sphe ical micelles (le el 1), which agg ega e in o cylinde s (le el 2), and inally in o mul ilamella supe s uc u es (le el 3). 4.2.2.2 Impo ance o he Na u e o he Coun e ion I is known ha me hyl iodide can unde go pho odecomposi ion o o m ee iodine. 51 Based on his ac , one hypo hesis o he obse ed s uc u al di e ences was he p es- ence o a ying amoun s o elemen a y iodine in he espec i e dioxane solu ions. In combina ion wi h he iodide coun e ion, his hen o ms iiodide, I 3- , which is a s ongly pola izable coun e ion, 52,53 al eady desc ibed o qua e nized poly(4- inylpy idine) (P4VPq). 54 The inc eased hyd ophobici y o P2VPq wi h iiodide as coun e ion is clea ly demons a ed by DLS measu emen s o a P2VPq homopolyme in wa e (Figu e 4-S8). Highe amoun s o added iodine led o an inc ease o he hyd odynamic adii, which we assign o hyd ophobic in e ac ions. In con as , chlo ide o me hyl sul a e coun e ions did no lead o he o ma ion o any hie a chically s uc u ed agg ega es o he µ-BVqT sys em (see Suppo ing In o ma ion). Howe e , addi ion o supplemen a y iodine o an aqueous micella solu ion o µ- BVqT (Figu e 4-1B) did no induce signi ican s uc u al changes. We a ibu e his o wo di e en easons: i s , owing o he educed co e dynamics, ea angemen p ocesses a e supp essed. Second and mos impo an , iodine i sel is no soluble in wa e bu is solubilized by he o ma ion o iiodide. Howe e , he iodide coun e ions a e mainly loca ed wi hin he micella co ona – app oxima ely 90% acco ding o in es iga ions o qua e nized poly(N,N-dime hylaminoe hyl me hac yla e) s a s 55 – leading o e y slow exchange p ocesses. Consequen ly, we added di e en amoun s o iodine o he µ-BVqT solu ion in dioxane p io o dialysis. The samples we e allowed o equilib a e o 2 h o gua an ee he con e sion o iiodide and hen dialyzed o wa e . Al eady he addi ion o 0.08 equi o I 2 induced d as ic s uc u al changes (Figu e 4-3A), as wo m-like micelles and elonga ed supe s uc u es we e obse ed in con as o sphe ical micelles in he absence o iodine (Figu e 4-1B). Again, he diame e o hese cylind ical agg ega es e- e s o he ini ially obse ed sphe ical micelles (d micelle = 24.5 ± 2.0 nm and d cylinde = 25.0 ± 2.0 nm). The co ona- o ming P2VPq block can be clea ly dis inguished in c yo-TEM 4 – “Woodlouse” S uc u es om µ-BVqT 103 (inse in Figu e 4-3A and Figu e 4-S9A). Fo 0.25 equi , almos exclusi ely supe s uc- u es om agg ega ed and in e wined cylind ical micelles a e ound. In addi ion, ewe p o usions and an inc eased endency o he cylinde s o o m meande -like s uc u es we e obse ed. The inse in Figu e 4-3B depic s an a ea whe e he cylinde s o m p e- s ages o ibbons. When 0.42 equi o I 2 was added, “woodlouse” agg ega es wi h a pe- iodic, mul ilaye ed s uc u e we e ound (d lam = 19.5 ± 1.0 nm, Figu e 4-3C). This pa e n is also clea ly isible in c yo-TEM, accompanied by a eas whe e he lamellae a e less densely packed (highligh ed a ea in Figu e 4-S9B) o pa icles which seem o be apped as cylind ical supe s uc u es (inse in Figu e 4-S9B). A mino ac ion o micelles and cylinde s was ound, as well.  Figu e 4-3. TEM mic og aphs om 0.2 g/L aqueous micella solu ions o µ-BVqT a e dialysis in he p es- ence o di e en amoun s o iodine. The solu ions we e p epa ed wi h 0.08 (A), 0.25 (B) and 0.42 (C) equi o iodine wi h espec o P2VPq monome uni s. The inse in (A) shows he co esponding c yo-TEM mi- c og aph. The schema ic illus a ions ep esen he dominan agg ega e mo phology. Fo he espec i e sample wi hou addi ional iodine, see Figu e 4-1B. To e alua e his lamella packing wi hin he pa icles in mo e de ail, addi ional SAXS measu emen s we e conduc ed om eeze-d ied powde s o he sample depic ed in Figu e 4-3C. The SAXS pa e n (Figu e 4-4A) shows peaks wi h a q a io o 1:2:3, ep e- sen ing he [100], [200], and [300] e lec ions, hus con i ming he lamella s uc u e. The long pe iod was calcula ed o d lam = 20.0 ± 1.5 nm and is in pe ec ag eemen wi h he alues obse ed in TEM (d lam = 19.5 ± 1.0 nm). In he SAXS pa e n o he in e medi- a e s uc u e (as shown in Figu e 4-S5), he e lec ions we e less p onounced (Figu e 4- 4B). This was al eady expec ed om he TEM images (Figu e 4-S5), which e eal a less egula a angemen . Howe e , he assump ion o an o e all lamella mo phology and he p esence o he [100] and [200] e lec ions allow he calcula ion o d lam = 22.5 ± 2.0 4 – “Woodlouse” S uc u es om µ-BVqT 104 nm. Compa able long pe iods om “woodlice” and agg ega ed cylinde s u he suppo ou p oposed mechanism o supe s uc u e o ma ion. Figu e 4-4. SAXS pa e n o a eeze-d ied powde om (A) he “woodlouse” s uc u e (Figu e 4-3C) and (B) he in e media e s uc u e (Figu e 4-S5). The in ege numbe s indica e he ela i e e lex posi ions, and he inse in A depic s he sca e ing pa e n obse ed a he 2D de ec o . These di e en le els o hie a chical sel -assembly o µ-BVqT in o sphe ical micelles (le el 1), ollowed by cylinde s (le el 2), supe s uc u es he eo and, inally, he s acking and back- olding o lamellae in o compa men alized mic ome e -sized polyme pa i- cles (le el 3) can be a ibu ed o su ace minimiza ion due o dec easing hyd ophilici y o he P2VPq co ona igge ed by he p esence o iiodide coun e ions. He eby, he su- pe s uc u e o ma ion o lamellae ia olding (o s acking) is suppo ed by simula ions on he sphe e-cylinde -lamellae ansi ion in diblock copolyme sys ems. 56 As we a e mos p obably dealing wi h non-equilib ium s uc u es o med du ing dialysis, he old- ed cylinde s a e supposed o be ansien in e media e s uc u es on he way owa d lamellae. Simila s uc u es we e al eady epo ed o linea ABC iblock e polyme s, whe e ei he he addi ion o a diamine in he case o poly(ac ylic acid) as solubilizing block 31-33 o changes in sol en quali y o he P2VP co ona 23 igge ed he o ma ion o supe s uc u es. Howe e , in ou case, his is clea ly caused by changes in he pola izabili y o he coun e ion, igge ed by he addi ion o iodine. The amoun o iiodide ep esen s he essen ial igge he e. Bo h he assembly pa hway and a delica e balance o iodide/ iiodide egula e s uc u al p ecision and he o e all colloidal s abili y o he subs uc u ed pa icles. Fu he mo e, he e, he well-de ined compac s uc u es 4 – “Woodlouse” S uc u es om µ-BVqT 105 clea ly e ol e h ough a complex agg ega ion and usion ia cylind ical building uni s in con as o he s acking o disk-like s uc u es as epo ed in li e a u e. 23,31-33 The sys em p esen ed he e is ela i e simple as no sophis ica ed p epa a ion pa hways u ilizing bi unc ional addi i es ha e o be applied, and he s uc u es a e ob ained in aqueous solu ion a he han in sol en mix u es. Also, he di ec ed sel -assembly is induced by he mono alen coun e ion, and he s uc u al in eg i y is a he una ec ed by he pH o he solu ion (see agg ega es wi hin acidic media o pH 3 as depic ed in Figu e 4-S10) 4.2.3 S uc u al Cha ac e iza ion o “Woodlouse” Agg ega es So a , we ha e desc ibed he iiodide-media ed hie a chical sel -assembly o µ-BVqT in o compa men alized pa icles. The highly pe iodic in e nal ine s uc u e is con i med by g ay-scale analysis o he TEM mic og aphs o indi idual pa icles (Figu e 4-5A). He e, he da ke domains co espond o he P2VPq phase con aining iodide/ iiodide coun e ions (d 1 = 8 nm), whe eas he con as is in e ed when s aining wi h OsO 4 was pe o med (selec i e o he PB phase, d 2 = 11 nm Figu e 4-5B). The wide lamellae p e- sumably consis o a mixed P BMA/PB phase as al eady discussed abo e. In c yo-TEM, e en h ee di e en epea ing dis ances a e isible (Figu e 4-5C). He e, he obse ed pe iodici ies can be explained in a simila way. The 8 nm o d 2 ep esen a mixed phase o PB and P BMA, which is sligh ly b oadened in TEM (d 2 in Figu e 4-5B), as he lamella is la ened and collapsed on o he ca bon ilm in he d ied s a e. On he o he hand, bo h d 1A and d 1B ep esen P2VPq, se ing as he co ona o he mixed PB/P BMA domains. A he in e ace (d 1A ), he densi y o P2VPq chains is highe as com- pa ed o he pe iphe y (d 1B ), leading o an inc eased elec on densi y. The p oposed chain packing and he co esponding dimensions a e illus a ed in Figu e 4-5D. He eby, he indi idual lamella shee s o he supe s uc u es can ei he s ack (whi e a ow in Figu e 4-5C) o back- old (black a ows in Figu e 4-5C), leading o “open” o “closed” s uc u es a he edge o he pa icles. 4 – “Woodlouse” S uc u es om µ-BVqT 106 Figu e 4-5. TEM mic og aphs o “woodlouse” agg ega es o µ-BVqT ob ained ia dialysis om dioxane in o wa e (A,B). The concen a ion was 0.1 g/L. In (A), he con as eme ges solely om he iodide coun e ion o he P2VPq phase, whe eas (B) was s ained wi h OsO 4 . In c yo-TEM (C), a egula pa e n o h ee dis- ances is isible. The concen a ion was 0.6 g/L. The co esponding g ay scale analyses a e shown below he mic og aphs. The p oposed a angemen o he mik oa m s a e polyme s is illus a ed in (D). We addi ionally pe o med c yo-TEM a di e en il angles (Figu e 4-S11 and ideo 4- S3). The eby, he in luence o he woodlouse o ien a ion wi hin he i i ied ilm was examined, and om he p ojec ions a di e en il angles, a ci cula c oss sec ion o he agg ega es can be clea ly deduced. Again, he p esence o la wo-dimensional assem- blies can be excluded, which is addi ionally suppo ed by SEM o he d ied pa icles (Fig- u e 4-S12). In he c yo-TEM il images, he in e nal ine s uc u e is only isible i he pa icles a e o ien ed pe pendicula o he beam di ec ion. I he s age is il ed u he , he s uc u al ea u es blu and, inally, disappea comple ely. The ac ha such s uc- u al ea u es a e only isible unde speci ic iewing angles has al eady been obse ed. 57 C oss-sec ional analysis o he “woodlice” wi hin hin ilm cu s o epoxy esin embed- ded pa icles (Figu e 4-6A and 4-S13) clea ly showed a pe iodic ine s uc u e, u he con i ming ou assump ion ha he pa icles a e no hollow. When he sample was ea ed wi h OsO 4 (s aining o PB, Figu e 4-6A), undula ed lamellae a e isualized as al- 4 – “Woodlouse” S uc u es om µ-BVqT 107 eady discussed o he in e media e s uc u es. The a he undula ed shape o he PB/P BMA lamella migh be ela ed o pa ial demixing o PB and P BMA, despi e he a he low χN. This could be a consequence o he longe DP o he PB block (109) as compa ed o P BMA (DP = 53), as well as di e en χ o he PB/P2VP and he P BMA/P2VP in e ac ions, leading o minimiza ion o he PB/P2VP in e ace. 48,58 Addi- ionally, om DSC measu emen s o he mik oa m s a e polyme , he p esence o a T g a -2 °C also hin s owa d he p esence o a sepa a ed PB phase in he bulk s a e (Figu e 4-S14). This leads o a di ec P BMA/P2VPq in e ace, accompanied by an in e ace o P2VPq wi h he adjacen mixed PB/P BMA phase and, inally, a pu e PB domain wi hou a PB/P2VPq in e ace. The p oposed a angemen o he cons i u ing segmen s is illus- a ed in Figu e 4-6B. S aining wi h OsO 4 enhances con as mainly in he PB phase, bu also he mixed phase appea s da ke (Figu e 4-6A), whe eas he pu e P BMA domains appea b igh e . Simila ly, he TEM mic og aphs o he cylind ical in e media e agg e- ga es unde he same sample p epa a ion me hod also hin owa d an undula ed phase bounda y (Figu e 4-S6B). Finally, he olume mo phology o he “woodlice” was in es iga ed using TEM omo- g aphy (TEMT). We he e o e p epa ed hicke slices (~150 nm) o he esin-embedded sample and pe o med s aining wi h OsO 4 o p o ide maximum con as . In Figu e 4-6C, a h ee-dimensional econs uc ion o a slice o a woodlouse pa icle wi h iew in o he lamella bulk mo phology is shown. Addi ionally, Figu e 4-6D shows h ee di e en slices o he same econs uc ion o a single pa icle. Bo h h ee-dimensional econs uc ions clea ly con i m he p esence o lamellae wi h an undula ed su ace h oughou he en- i e sample (also see ideo 4-S4). He eby, he o ma ion o well-de ined lamella s uc- u es as compa ed o micella clus e s (obse ed o linea polyme s; see Figu e 4-S4) is p obably a di ec consequence o he mik oa m a chi ec u e. In acco dance wi h bo h heo y and expe imen al wo k, 59 he inc eased segmen al densi y has a dis inc i e e ec on he su ace cu a u e and, in he case o ou sys em, acili a es he o ma ion o la- mellae wi h low su ace cu a u e. 4 – “Woodlouse” S uc u es om µ-BVqT 108 Figu e 4-6. (A) TEM mic og aph o 50 nm hick cu s om eeze-d ied and embedded samples o µ-BVqT agg ega es, s ained wi h OsO 4 . (B) Schema ic illus a ion o he block a angemen wi hin he “woodlouse” s uc u e is depic ed, wi h he wo possibili ies o ben (uppe pa ) and s acked lamellae (lowe pa ). The g ay a eas esemble he P2VPq phase; P BMA is ed, and PB is blue. The iole domains ep esen a mixed PB/P BMA phase. TEM omog aphy 3D econs uc ions o a slice o he “woodlouse” s uc u e (C) and c oss-sec ional analysis o a single pa icle (D). The omog aphy images we e ob ained om a 150 nm hick cu , which was addi ionally ea ed wi h OsO 4 o selec i ely s ain he mixed PB/P BMA phase (plo - ed in g een). The app oxima e leng h o he long ma ked edge o he econs uc ions is 280 nm in (C) and 220 nm in (D). 4 – “Woodlouse” S uc u es om µ-BVqT 109 4.3 Conclusions We ha e demons a ed he hie a chical sel -assembly o an ABC mik oa m s a e polyme in o subs uc u ed pa icles o up o 1µm in size, which we e m “wood- louse” pa icles. Sphe ical micelles wi h a mixed PB/P BMA co e and a P2VPq co ona ac as he basic building blocks (le el 1). S epwise agg ega ion o hese esul s in cylind ical micelles (le el 2), ollowed by supe s uc u es he eo , and inally compa men alized pa icles o up o 1 µm in size. These pa icles ea u e a highly pe iodic, lamella ine s uc u e (le el 3). The p esence and amoun o iiodide as a highly pola izable coun e ion o he P2VPq co ona is an essen ial igge o induce his supe s uc u e o ma ion in o di e en le els o hie a chy (Figu e 4-7). All in e media es o inc easing hie a chy in ol ed on he way o he inal “woodlice” a e isualized by TEM and c yo- TEM. The iodide/ iiodide sys em is an elegan app oach o di ec ing he hie a chical sel -assembly o such ma e ials. The na u e o he coun e ion, he mik oa m s a a chi- ec u e, and he assembly pa hway a e essen ial pa ame e s and in luence s uc u al pe ec ion and he inal mo phology. To ecapi ula e, a de ailed unde s anding o he sel -assembly mechanism in o complex supe s uc u es was ob ained, which bea s uc- u al simila i ies o biological sys ems such as mi ochond ia. Applying hese esul s o o he mik oa m s a e polyme sys ems wi h highe segmen al incompa ibili y migh lead o comple ely phase-sepa a ed co es and ep esen s an in e es ing app oach o co e-compa men alized s uc u es o complex shape. Fu he unc ionaliza ion and modi ica ion o hese s uc u es will enable he p epa a ion o a a ie y o s imuli- esponsi e highly s uc u ed ma e ials wi h de ined in e nal pe iodici y. The ques ion a ises whe he his app oach is applicable o o he mik oa m s a sys- ems con aining polyca ionic segmen s in gene al o whe he he combina ion o P2VPq and a “dynamic”, low T g segmen like PB is a p e equisi e. This will be he subjec o u- u e wo k. The use o iodide/ iiodide as a se sc ew o di ec he sel -assembly o di e - en ma e ials in o well-de ined hie a chical supe s uc u es would be ad an ageous and desi able. 4 – “Woodlouse” S uc u es om µ-BVqT 116 4.5 Re e ences (1) Whi esides, G. M.; G zybowski, B. Sel -Assembly a All Scales. Science 2002, 295, 2418-2421. (2) Zhang, S. Eme ging Biological Ma e ials Th ough Molecula Sel -Assembly. Bio echnol. Ad . 2002, 20, 321-339. (3) Naga ajan, R.; Ruckens ein, E. Theo y o Su ac an Sel -Assembly: a P edic i e Molecula The - modynamic App oach. 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(63) C ow he , R. A.; DeRosie , D. J.; Klug, A. The Recons uc ion o a Th ee-Dimensional S uc u e om P ojec ions and i s Applica ion o Elec on Mic oscopy. P oc. R. Soc. London A 1970, 317, 319-340. 4 – “Woodlouse” S uc u es om µ-BVqT 119 4.6 Suppo ing In o ma ion 4.6.1 Addi ional Expe imen al Sec ion Syn hesis Qua e niza ion o 2-Vinylpy idine Homopolyme s (P2VP) and P epa a ion o Aqueous Solu ions The qua e niza ion p ocedu e was simila o he me hod desc ibed o he mik oa m s a e polyme wi h sligh modi ica ions. A e h ee days o eac ion wi h me hyl iodide 10 ol% o wa e we e added o he pa ially p ecipi a ed eac ion mix u e in dioxane and i was allowed o s i o an addi ional day. A e wa d, i was i s dialyzed o a mix- u e o dioxane:wa e (90:10) and hen he sol en composi ion was g adually inc eased o (80:20). The ob ained s ock solu ion in he dioxane:wa e mix u e was a e wa d dia- lyzed o wa e . The iiodide complexes we e p epa ed in he same manne as desc ibed in he manusc ip . P epa a ion o µ-BVqT wi h Di e en Coun e ions The exchange o he iodide coun e ion wi h chlo ide was achie ed by dialysis o he dioxane s ock solu ion o 50 mM solu ion o LiCl in THF. A e wa d he solu ion was dia- lyzed agains pu e THF o emo e excess sal . Be o e dialysis o wa e , he sol en was again changed o dioxane h ough dialysis. The qua e niza ions wi h dime hyl sul a e we e conduc ed ei he in THF o dioxane. A polyme solu ion was p epa ed wi h a concen a ion o 2 g/L and hen degassed o 15 minu es. A e wa d, 10 equi o dime hyl sul a e ega ding 2VP uni s we e added and he eac ion mix u e was allowed o s i a 40 °C o 3 days. Finally he solu ion was pu i- ied om he excess qua e niza ion agen by dialysis wi h he espec i e eac ion me- dium. A e dilu ion o a concen a ion o 1 g/L he solu ions we e dialyzed o wa e . Cha ac e iza ion Size Exclusion Ch oma og aphy (SEC) SEC measu emen s we e pe o med on a se o 30 cm SDV-gel columns (5 µm bead size, wi h po e sizes o 10 5 , 10 4 , 10 3 and 10 2 Å) using e ac i e index and UV (λ = 254 nm) 4 – “Woodlouse” S uc u es om µ-BVqT 120 de ec ion. THF was used as eluen a a low a e o 1 mL/min. Toluene was used as in- e nal s anda d and he sys em was calib a ed wi h PS and 1,4-PB s anda ds. 1 H NMR Spec oscopy 1 H NMR spec a we e eco ded on a B uke Ul ashield 300 spec ome e a an ope a - ing equency o 300 MHz. CDCl 3 was used as sol en and e ame hylsilane as in e nal s anda d. Ma ix-Assi ed Lase Deso p ion Ioniza ion Time-o -Fligh Mass Spec ome y (MALDI- ToF MS) MALDI-ToF MS analysis was pe o med on a B uke -Re lex III appa a us equipped wi h a N 2 lase (λ = 337 nm) a an accele a ion ol age o 20 kV. ans-2-[3-(4- e -Bu ylphenyl)- 2-me hyl-2-p openyliden]maloni ile (DCTB, Fluka, 99,0 %) was used as ma ix and sil e i luo oace a e (AgTFA, Sigma-Ald ich, 99.99%) as ioniza ion agen . Samples we e p e- pa ed om THF solu ion by mixing ma ix, polyme and sal in a a io o 20/5/1 ( / ). Scanning Elec on Mic oscopy (SEM) The pa icles we e analyzed by ield emission scanning elec on mic oscopy on a Zeiss LEO 1530 Gemini mic oscope equipped wi h a ield emission ca hode ope a ing a 0.5-5 kV. Specimen p epa a ion was accomplished as ollows: silicon wa e s we e cleaned us- ing isop opanol and ace one in a s anda d p ocedu e. To ob ain a eas o di e en con- cen a ion o pa icles, he wa e was i s dip-coa ed om 0.02 g/L solu ion and hen a u he d op o he solu ion was deposi ed and allowed o d y. The specimens we e coa ed wi h a hin pla inum laye using a spu e coa e (C essing on 208HR) o ende he specimen conduc i e. Dynamic Ligh Sca e ing (DLS) DLS measu emen s we e pe o med on an ALV DLS/SLS-SP 5022F compac goniome e sys em wi h an ALV 5000/E c oss co ela o and a He-Ne lase (λ = 632.8 nm). The meas- u emen s we e ca ied ou in cylind ical sca e ing cells (d = 10 mm) a an angle o 90° and a empe a u e o 20 °C. P io o he ligh sca e ing measu emen s, he sample solu- ions we e il e ed using nylon il e s (Magna, Ro h) wi h a po e size o 5 µm. The CON- 4 – “Woodlouse” S uc u es om µ-BVqT 121 TIN algo i hm was applied o analyze he ob ained co ela ion unc ions. Appa en hy- d odynamic adii we e calcula ed acco ding o he S okes-Eins ein equa ion. DSC The mal analysis and de e mina ion o he glass ansi ion empe a u es was pe o med on a Pe kin–Elme Diamond DSC a a hea ing a e o 20 K/min. 4.6.2 Syn hesis and Cha ac e iza ion o PB-a m-P2VP-a m-P BMA Mik- oa m S a Te polyme The syn hesis o he PB-a m-P2VP-a m-P BMA mik oa m s a e polyme was accom- plished by combining sequen ial anionic polyme iza ion wi h azide-alkyne Huisgen cycloaddi ion. The e o e, an alkyne- unc ionalized polybu adiene-b-poly(2- inylpy idine) (PB-b-P2VP) diblock copolyme was di ec ly p epa ed by anionic polyme iza ion using an alkyne- unc ionalized DPE de i a i e (click-DPE). The molecula cha ac e iza ion o he diblock copolyme is lis ed in Table 4-S1. As al eady epo ed, in he case o he click- DPE, inco po a ion in, o mos p obably a he end o he poly(2- inylpy idine) (P2VP) block akes place. This leads o diblock copolyme s which bea addi ional alkyne- unc ions in o a he end o he P2VP block, as he DPE de i a i e is used in excess. Ne - e heless, unde app op ia e eac ion condi ions, i.e. using only a sligh excess o click- DPE, low empe a u es and sho eac ion imes an o e all deg ee o alkyne- unc ionaliza ion o 116 % was achie ed. 1 Taking in o accoun he e o s o he de e mi- na ion me hod his esembles only a sligh o e -inco po a ion o click-DPE in o he ma- e ial. The alkyne g oup is hen used o a ach he hi d block, poly( e -bu yl me hac yla e) (P BMA), ia azide-alkyne Huisgen cycloaddi ion. Thus, in he ollowing click eac ion an excess o 10% o azido- unc ionalized homopolyme chains ela i e o he diblock chains was used o ensu e ha all diblock chains unde go conjuga ion. As a di ec consequence he ob ained mik oa m s a e polyme con ains a mino ac ion wi h mo e han one chain o P BMA. Figu e 4-S1 displays he SEC aces o he p ecu so homopolyme and diblock copolyme , and he ob ained mik oa m s a e polyme . Table 4-S1 lis s he co - esponding cha ac e iza ion da a. Due o he use o an azido- unc ionalized ATRP ini ia- 4 – “Woodlouse” S uc u es om µ-BVqT 122 o o he P BMA polyme iza ion in case o ecombina ion polyme s bea ing wo azido end-g oups a e o med. The e o e, a small pe cen age o α,ω-azido-bi unc ionalized homopolyme could lead o H-shaped mik oa m s a e polyme s and explain he ob- se ed highe molecula weigh shoulde . F om es click eac ions wi h PS homopolyme s, whe e he b omo- unc ion was ans o med in o an azide a e polyme - iza ion, no such coupling shoulde was de ec ed, suppo ing ou assump ion. Table 4-S1. Molecula Cha ac e is ics o Poly( e -bu yl me hac yla e) (T), Polybu adi- ene-block-poly(2- inylpy idine) (BV), and µ-BVT Polyme a M n b / kg/mol PDI c T 53 7.6 1.17 B 109 V 81 14.7 1.02 µ-B 109 V 81 T 53 22.2 1.07 a The subsc ip s deno e he deg ees o polyme iza ion o he co esponding blocks as calcula ed om he espec i e numbe a e age molecula weigh s. b Fo he P BMA homopolyme he numbe a e age molec- ula weigh was de e mined by SEC using a P BMA calib a ion. Fo he diblock copolyme he molecula weigh was calcula ed om 1 H NMR using he molecula weigh o he PB as e e ence. This was measu ed di ec ly ia MALDI-ToF MS. Fo he ABC mik oa m s a e polyme he o e all molecula weigh was calcu- la ed om he co esponding p ecu so polyme s. c De e mined by SEC in THF calib a ed wi h PS s anda ds o in he case o P BMA homopolyme wi h P BMA s anda ds. Figu e 4-S1. SEC eluog ams o he PB-a m-P2VP-a m-P BMA mik oa m s a e polyme (µ-BVT), he p e- cu so diblock copolyme (PB-b-P2VP), and he homopolyme (P BMA). In all cases he RI signal is shown.  26 28 30 32 34 36 38 0.0 0.2 0.4 0.6 0.8 1.0 no malized RI de ec ion V e [mL] PB b P2VP P BMA " BVT 4 – “Woodlouse” S uc u es om µ-BVqT 123 4.6.3 Sel -Assembly o µ-BVqT Figu e 4-S2. In ensi y-weigh ed DLS CONTIN plo (A) o qua e nized µ-BVT (µ-BVqT) in 1 g/L dioxane solu- ion, R h,app = 12.5 nm.  Figu e 4-S3. O e iew o a ious c yo-TEM images o di e en “woodlouse” agg ega ion o ms obse ed in ~0.6 g/L aqueous solu ions o µ-BVqT. 4 – “Woodlouse” S uc u es om µ-BVqT 124  Figu e 4-S4. (A) TEM and (B) c yo-TEM mic og aphs om aqueous solu ions o linea B 1108 Vq 142 T 93 a 0.2 and 0.65 g/L, espec i ely. A e dialysis o wa e p ecipi a ion occu ed g adually. Whe eas he image in (A) is uns ained, he inse in (A) shows an enla ged a ea a e s aining wi h OsO 4 . In bo h cases micella clus e s can be seen, wi hou any in e nal ine s uc u e (see s aining in he inse in (A) o he b igh e a eas in he c yo-TEM in (B)). Please no e ha he molecula weigh o he iblock e polyme was much highe and also he mola ac ions we e di e en o he s udied mik oa m s a e polyme µ-B 101 V 81 T 53 . Addi ionally, he sequence was PB-b-P2VP-b-P BMA due o he es ic ions o anionic polyme iza ion and he e o e he middle block o ms he co ona. This leads o a olding o he polyme chain o shield he hyd ophobic end blocks. 4.6.3.1 In e media e S uc u es o “Woodlouse” Agg ega es The dioxane solu ion o µ-BVqT, which yielded he in e media e s uc u es (Figu e 4-2), was allowed o age o an addi ional mon h be o e dialysis o wa e . TEM analysis showed ha he ob ained agg ega es we e o sligh ly highe s uc u al o de and he in e nal ine s uc u e o he “woodlouse” was al eady isible o some objec s (Figu e 4- S5A). Ne e heless, he s uc u es seem o be kine ically apped du ing he p ocess, as u he ageing o aqueous solu ions did no lead o any inc ease in o de . As obse ed by c yo-TEM he supe s uc u es we e mo e densely packed (Figu e 4-S5B). Less cylind ical p o usions emana e om he cen e o he agg ega es, and he g ay-scale analysis con- i ms h ee pe iodic dis ances (d 1 = 4.5 ± 1.0 nm, d 2 = 4.5 ± 1.0 nm, d 3 = 9.0 ± 1.0 nm. The s uc u e was u he isualized using di e en il angles in c yo-TEM (Suppo ing ide- os 4-S1 and 4-S2), con i ming a 3-dimensional a ay o he cylinde s.  4 – “Woodlouse” S uc u es om µ-BVqT 125  Figu e 4-S5. TEM (A) and c yo-TEM mic og aphs (B) o in e media e micella s uc u es o µ-BVqT ob- ained by dialysis o wa e a e ageing o he dioxane solu ion o one mon h. The inal polyme concen- a ion was 0.2 g/L o TEM and 0.4 g/L in case o c yo-TEM. The uppe inse in (B) displays a g ay-scale analysis o he highligh ed a ea.  We u he p epa ed hin ilm cu s o his s uc u al in e media e by eeze-d ying o he solu ion and embedding he pa icles in o an epoxy esin. The co esponding TEM mi- c og aphs om hin slices (wi h and wi hou addi ional OsO 4 s aining) a e shown in Fig- u e 4-S6 and an in e nal lamella s uc u e can be obse ed (black a ows in Figu e 4-S6). Addi ionally, OsO 4 s aining e ealed an undula ed shape o he lamellae, which can also be in e p e ed as a dense packing o sphe es (Figu e 4-S6B). The same obse a ion was made o some o he cylind ical s uc u es (whi e a ows in Figu e 4-S6B). As indica ed by he whi e a ows o he non-s ained mic og aphs in Figu e 4-S6A (only P2VPq is isi- ble), cylind ical p o usions wi h a ubula na u e we e also iden i ied. These indings con i m ou ini ial assump ion o cylind ical micelles as in e media es. Fu he mo e, he absence o s uc u es wi h a di use co e egion suppo s he assump ion o compac pa icles. The p esence o bo h cylind ical p o usions and elonga ed pa icles o a he compac shape wi h a sphe ical c oss sec ion we e u he con i med ia SEM measu e- men s (Figu e 4-S7). 132 5 – T iiodide-Di ec ed Sel -Assembly o Di e en Mik oa m S a Polyme s 133 5 – Hie a chical Sel -Assembly o Mik oa m S a Polyme Sys ems Con aining a Polyca ionic Segmen : A Gene al Concep And eas Hanisch, And é H. G öschel, Melanie Fö sch, Tina I. Löbling, Felix H. Schache , and Axel H. E. Mülle ABSTRACT: We ecen ly in oduced a concep o he coun e ion-media ed hie a chical sel -assembly o amphiphilic mik oa m s a e polyme s in aqueous media in o mic om- e e -sized compa men alized pa icles wi h a highly pe iodic lamella ine s uc u e (“woodlice”). He ein, we ex end his concep o di e en mik oa m s a polyme sys- ems con aining a polyca ionic segmen . The p esence o a poly(N-me hyl-2- inylpy idinium) (P2VPq) block and i s in e ac ion wi h iodide/ iiodide coun e ions is c ucial. In analogy o linea diblock copolyme sys ems he hyd ophilic/hyd ophobic bal- ance o polybu adiene-a m-poly(N-me hyl-2- inylpy idinium iodide)-a m-polys y ene mik oa m s a e polyme s de e mines he mo phology o he p ima y building blocks (sphe ical micelles and cylind ical micelles/ esicles) and he ob ained supe s uc u es (s acked lamella s uc u es and mul ilamella esicles) du ing his hie a chical p ocess. When an ABA’ mik oa m s a copolyme (polys y ene-a m-poly(N-me hyl-2- inylpy idinium iodide)-a m-polys y ene) wi hou a dynamic co e- oming block was in es iga ed, a di e en mechanism in o “woodlouse”-s uc u ed agg ega es ia agg e- ga ion and de o ma ion o in e media e esicles was ound. The indi idual s eps o he 5 – T iiodide-Di ec ed Sel -Assembly o Di e en Mik oa m S a Polyme s 134 di e en sel -assembly p ocesses we e in es iga ed by ansmission elec on mic oscopy and addi ionally suppo ed by dynamic ligh sca e ing, di e en ial scanning calo ime y, and small-angle X- ay sca e ing. Keywo ds: Mik oa m S a Polyme , Polyelec oly e, Hie a chical Sel -Assembly 5 – T iiodide-Di ec ed Sel -Assembly o Di e en Mik oa m S a Polyme s 135 5.1 In oduc ion One s aigh o wa d me hod o con ol he mo phology o sel -assembled s uc u es om block copolyme s is changing he olume ac ion o he cons i u ing segmen s. This led o a di e si y o mo phologies in he bulk 1-4 and in solu ion. 5-10 In he la e case, p edominan ly sphe ical micelles a e o med, bu also cylind ical o esicula s uc- u es, as well as he co esponding in e media es ha e been epo ed. 11 Despi e he ple ho a o nanos uc u es being accessible oday, he s uc u al di e si y and complexi y eached in biological sys ems s ill ep esen s he ul ima e benchma k. He e, sel -o ganiza ion o di e se biomac omolecules induces hie a chy o e se e al leng h scales h ough in a- and in e molecula in e ac ions by encoding u mos unc- ionali y ia he indi idual building blocks. Such p ocesses can be conside ed as con e - gen since he coope a i i y o a mul i ude o (di e en ) weak in e ac ions leads o he o ma ion o p ima y building blocks, which u he assemble in o s uc u es o in- c eased complexi y a a he modynamic minimum. The accumula ion o weak in e ac- ions wi hin and be ween he building blocks compensa es o he loss in en opy. 12,13 Fo example, p o eins old in o complex h ee-dimensional s uc u es guided by he in- o ma ion encoded wi hin he p ima y s uc u e o he amino acid sequence. 14 Com- monly, hese s uc u es a e monodispe se, as shown o he obacco mosaic i us (TMV), whe e 2130 iden ical p o ein subuni s a ange a ound a single s and o RNA in a helical ashion o p oduce a igid od o 18 x 300 nm. 15 E en hough he dimensions ob ained in na u al sys ems a e di icul o each by di- ec ed sel -assembly o syn he ic mac omolecules in gene al, his s ill p o ides a s aigh o wa d app oach o he design o well-de ined unc ional ma e ials. Inc easing he ansc ip ed chemical in o ma ion o he sys em by an addi ional block (AB o ABC) al eady leads o a d as ic inc ease o he me e numbe o mo phologies being accessi- ble. 16 This is a consequence o h ee unlike polyme -polyme in e ac ion pa ame e s and he possibili y o sol en s being selec i e o one o wo segmen s. Depending on he indi idual solubili y, his enables he p epa a ion o co ona- o co e-compa men alized s uc u es. 17-19 In his con ex Hillmye , Lodge and co-wo ke s ga e a de ailed in es iga- ion o he compa men aliza ion in micella sys ems by using mik oa m s a 5 – T iiodide-Di ec ed Sel -Assembly o Di e en Mik oa m S a Polyme s 136 e polyme s. Th ough his unique mac omolecula a chi ec u e “hambu ge ” micelles, segmen ed wo m-like micelles, and subs uc u ed esicles could be ob ained as co e- compa men alized analogs o sphe ical, wo m-like and esicula s uc u es. 20-22 Mo e- o e , in he ield o iblock e polyme s i is well demons a ed ha he complexa ion wi h single s anded DNA 23 o he use o diamines as addi i es 24,25 display addi ional igge s o hie a chical s uc u e o ma ion. Especially “kine ic con ol” allowed o he s epwise sel -assembly o poly(ac ylic acid)-block-poly(me hyl ac yla e)-block- polys y ene iblock e polyme s in o o oids, s acks o disk-like micelles, and lamella phase sepa a ed d ople s. He eby, he s uc u es we e in luenced by he polyme se- quence, composi ion, he na u e o he di alen coun e ion, he sol en mix u e, and he p epa a ion pa hway and he con ol o hese pa ame e s allowed o manipula ion o he ob ained agg ega ion o ms. 25-30 Recen ly, we epo ed he coun e ion-di ec ed sel -assembly o a polybu adiene-a m- poly(N-me hyl-2- inylpy idinium iodide)-a m-poly( e -bu yl me hac yla e) (µ-BVqT) mik oa m s a e polyme in o mul ilaye ed “woodlouse” agg ega es, aking ad an age o bo h he polyme a chi ec u e and he unabili y o he coun e ion. 31 The p esence o iodide as coun e ion o he qua e nized poly(2- inylpy idine) segmen was c ucial o achie e supe s uc u e o ma ion. Iodine as addi i e led o he o ma ion o iiodide, a highly pola izable coun e ion, which induced hie a chical sel -o ganiza ion om sphe i- cal micelles in o cylind ical s uc u es and well-de ined supe s uc u es he eo . Besides he na u e o he coun e ion and he mik oa m s a a chi ec u e also he p epa a ion pa hway was essen ial o ob ain well-de ined agg ega es. He e, we y o ob ain a mo e de ailed insigh in o he unde lying mechanism o his p ocess by expanding i o o he mik oa m s a polyme sys ems. We ake ad an age o ou newly de eloped modula app oach o he s aigh o wa d syn hesis o such ma e ials. 32 Fi s , he in luence o he molecula composi ion on he ob ained building uni s om h ee di e en polybu adiene-a m-poly(N-me hyl-2- inylpy idinium iodide)-a m-polys y ene mik oa m s a e polyme s (µ-BVqS) and he esul ing coun e ion-di ec ed supe s uc u es will be elabo a ed. In his con ex , he P2VPq co ona was addi ionally u ilized o he gene a- ion and hos ing o Au nanopa icles. Second, he iiodide-di ec ed sel -assembly o a 5 – T iiodide-Di ec ed Sel -Assembly o Di e en Mik oa m S a Polyme s 137 polys y ene-a m-poly(N-me hyl-2- inylpy idinium iodide)-a m-polys y ene mik oa m s a copolyme (µ-SVqS’) as a model sys em wi hou a low T g segmen is in es iga ed. 5 – T iiodide-Di ec ed Sel -Assembly o Di e en Mik oa m S a Polyme s 138 5.2 Expe imen al Pa Syn hesis Ma e ials Bu adiene (Rießne -Gase) was passed h ough columns illed wi h molecula sie es (4 Å) and basic aluminium oxide and s o ed o e dibu ylmagnesium (1 M solu ion in hep ane, Ald ich). 2-Vinylpy idine (2-VP, Ald ich) was degassed, s i ed wi h ie hylaluminium (1 M solu ion in hexanes, Ald ich) and condensed on a high acuum line. S y ene (BASF) was pu i ied in a simila manne , excep ha i was s i ed wi h dibu ylmagnesium (1 M solu ion in hep ane, Ald ich) ins ead o ie hylaluminium. THF (Sigma-Ald ich) was dis illed om CaH 2 and Na/K alloy. sec-Bu ylli hium (Ac os, 1.3 M in cyclohexane/hexane: 92/8) was used wi hou u he pu ifica ion. e -Bu yl me hac yla e ( BMA, Sigma-Ald ich) and s y ene (Sigma-Ald ich) o ATRP we e il e ed o e basic aluminium oxide be o e use. N,N,N′,N′,N′′- Pen ame hyldie hylene iamine (PMDETA) and CuB we e pu chased om Ald ich and dis- illed and degassed o ea ed wi h pu e ace ic acid and il e ed, espec i ely. 1-(4- e -bu yl- dime hylsilyl)e hynylphenyl)-1-phenyle hylene (click-DPE) was syn hesized om 1-(4- b omophenyl)-1-phenyle hylene as al eady epo ed 32 . The aqueous solu ions we e p e- pa ed wi h dis illed desalina ed wa e . All o he chemicals we e o analy ical g ade and used as ecei ed. The dialysis memb ane used o all s eps was pu chased om Ro h (Spec a Po ), wi h a molecula weigh cu -o (MWCO) o 1 000 g/mol. Syn hesis and Qua e niza ion o Mik oa m S a Te - and Copolyme s The de ailed p ocedu es o he syn hesis o he alkyne mid- unc ional diblock copolyme s (polybu adiene-block-poly(2- inylpy idine)) and azido- unc ionalized homopolyme s (poly- s y ene and poly( e -bu yl me hac yla e)) ha e al eady been epo ed elsewhe e. 32 Simila - ly, ano he alkyne mid- unc ionalized diblock copolyme , polys y ene-block-poly(2- inylpy idine) (PS-b-P2VP), was also syn hesized ia anionic polyme iza ion in THF u ilizing he alkyne-subs i u ed diphenyle hylene (click-DPE) de i a i e. The e o e, s y ene was ini i- a ed wi h sec-BuLi a -70 °C and allowed o polyme ize a his empe a u e o 25 minu es be o e he click-DPE was added (1.1 equi ela i e o li ing chains). A e s i ing a -50 °C o 2 h he empe a u e was se o -70 °C and 2VP was added and allowed o polyme ize o 5 5 – T iiodide-Di ec ed Sel -Assembly o Di e en Mik oa m S a Polyme s 139 minu es be o e e mina ion wi h degassed isop opanol. The polyme was pu i ied by p ecipi- a ion in wa e . Finally, he liga ions o he alkyne- unc ionalized diblock copolyme s wi h he azido- unc ionalized homopolyme s we e conduc ed by azide-alkyne Huisgen cycloaddi ion. 33 The e o e, a mix u e o he diblock copolyme and 1.1 equi o he espec i e homopolyme we e dissol ed in THF a a concen a ion o ~20 g/L and degassed o 10 min. A e addi ion o 1 equi CuB , he solu ion was degassed o u he 15 min. PMDETA (1 equi ) was added o complex he coppe and s a he eac ion, which was ollowed by SEC. A e 2 days, he esul ing mik oa m s a e polyme s we e pu i ied by passing he solu ion h ough a small column wi h basic alumina o emo e coppe and inally eeze-d ied om dioxane. P epa a ion o Aqueous Micella Solu ions The ans o ma ion o he P2VP compa men in o a s ong ca ionic polyelec oly e (P2VPq) was conduc ed using me hyl iodide as qua e niza ion agen . The de ailed p ocedu e is gi en in he li e a u e. 31 To simpli y ma e s, in case o all qua e nized solu ions he gi en concen- a ions esemble he concen a ion o he p is ine mik oa m s a e polyme be o e he modi ica ion, hus neglec ing he inc ease o mass due o qua e niza ion. Fo he p epa a ion o he iiodide complexes an iodine s ock solu ion was p epa ed wi h dioxane as sol en a 15 g/L. This iodine solu ion was hen added o he dioxane solu ion o he qua e nized s a e polyme s un il he desi ed a io o iodine o amino- unc ion was achie ed. A e wa ds, he solu ions we e s i ed o 2 h and hen ea ed wi h ul asound o 15 min. Subsequen ly, he solu ions we e dialyzed agains wa e , as desc ibed abo e and ob ained wi h concen a ions anging om 0.3 o 0.7 g/L. Some o he solu ions we e col- loidally s able o e mon hs, whe eas o he s sedimen ed wi h ime. Gene a ion o Au Nanopa icles To 1.5 mL o an aqueous solu ion o µ-BVq1S, which was ob ained om dialysis o he co e- sponding dioxane solu ion wi h addi ional 0.25 equi I 2 ( ega ding P2VPq uni s), was added a highly concen a ed aqueous solu ion o HAuCl 4 un il an Au:N a io o 0.6 was eached. Di- ec ly a e addi ion, p ecipi a ion occu ed, and he e o e he sample was ea ed wi h ul- asound o 30 minu es. A e s i ing o 90 minu es he solu ion was subjec ed o cen i u- ga ion. The supe na an liquid was decan ed and he solid con en was edispe sed in 1.5 mL 5 – T iiodide-Di ec ed Sel -Assembly o Di e en Mik oa m S a Polyme s 140 o wa e . A eshly p epa ed solu ion o NaBH 4 was added (5 M equi wi h espec o Au) and hen he solu ion was s i ed o e nigh . A e cen i uga ion o emo e excess eagen s, he solid con en was again edispe sed in 1.5 mL o wa e o ob ain a solu ion o cha ac e is ic ed- iole colo . Cha ac e iza ion T ansmission Elec on Mic oscopy (c yo-TEM and TEM) TEM mic og aphs we e aken wi h a Zeiss CEM 902 o 922 OMEGA elec on mic oscope op- e a ed a 80 kV o 200 kV, espec i ely. Bo h machines we e equipped wi h an in column ene gy il e . Fo sample p epa a ion 2 µL o he solu ion ( ypically 0.1-0.2 g/L) we e depos- i ed on a hyd ophilized TEM g id (coppe , 200 mesh). A e wa ds, he emaining sol en was emo ed wi h a blo ing pape . In ano he p epa a ion pa hway a d op o he solu ion was deposi ed on a TEM g id on a piece o pa a ilm, immedia ely ozen wi h liquid ni ogen and a e wa ds eeze-d ied. Fo in es iga ion o he pa icle ilm he eeze-d ied polyme was embedded in o a esin (EpoTek 301). 50 nm hin cu s we e p epa ed wi h a Leica EM UC7 mic o ome equipped wi h a diamond kni e and deposi ed on o TEM g ids (coppe , 200 mesh). Fo c yo-TEM s udies, a d op (~2 mL) o he aqueous micella solu ion (c ~0.4-0.7 g/L) was placed on a lacey ca bon-coa ed coppe TEM g id (200 mesh, Science Se ices), whe e mos o he liquid was emo ed wi h blo ing pape , lea ing a hin film s e ched o e he g id holes. The specimens we e shock i ified by apid imme sion in o liquid e hane in a empe a u e-con olled eezing uni (Zeiss C yobox, Zeiss NTS GmbH) and cooled o ap- p oxima ely 90 K. The empe a u e was moni o ed and kep cons an in he chambe du ing all o he p epa a ion s eps. A e eezing he specimens, hey we e inse ed in o a c yo- ans e holde (CT3500, Ga an) and ans e ed o a Zeiss EM922 OMEGA EFTEM ins u- men . Examina ions we e ca ied ou a empe a u es a ound 90 K. The mic oscope was op- e a ed a an accele a ion ol age o 200 kV. Ze o-loss fil e ed images (ΔE = 0 eV) we e aken unde educed dose condi ions. All images we e egis e ed digi ally by a bo om-moun ed CCD came a sys em (Ul ascan 1000, Ga an), combined, and p ocessed wi h a digi al imaging p ocessing sys em (Ga an Digi al Mic og aph 3.9 o GMS 1.4). E alua ion o he espec i e leng h scales o he s uc u es was achie ed by measu ing 50-100 di e en spo s wi hin he sample wi h he UTHSCSA ImageTool V. 3.00. 5 – T iiodide-Di ec ed Sel -Assembly o Di e en Mik oa m S a Polyme s 141 UV-Vis Spec oscopy UV- is spec a we e eco ded on a Hi achi 3000 spec opho ome e . Dynamic Ligh Sca e ing (DLS) DLS measu emen s we e pe o med on an ALV DLS/SLS-SP5022F compac goniome e sys- em equipped wi h an ALV 5000/E c oss co ela o and a He-Ne lase (λ = 632.8 nm). The measu emen s we e ca ied ou in cylind ical sca e ing cells (d = 10 mm) a an angle o 90 ° and a empe a u e o 20 °C. The solu ions we e measu ed wi hou il a ion. The CONTIN algo i hm was applied o analyze he ob ained co ela ion unc ions. Appa en hyd ody- namic adii we e calcula ed acco ding o he S okes-Eins ein equa ion. Small-Angle X-Ray Sca e ing (SAXS) SAXS measu emen s o he eeze-d ied powde s we e pe o med on a B uke AXS Nanos a (B uke , Ka ls uhe, Ge many), equipped wi h a mic o ocus X- ay sou ce (Incoa ec IµS Cu E025, Incoa ec Gees hach , Ge many), ope a ing a λ = 1.54 Å. A pinhole se up wi h 750, 400, and 1000 µm (in he o de om sou ce o sample) was used and he sample- o-de ec o dis ance was 107 cm. Samples we e moun ed on a me al ack and ixed using ape. The sca e ing pa e ns we e co ec ed o he beam s op and he backg ound (Sco ch ape) p io o e alua ions. The measu emen ime was 12 h. Di e en ial Scanning Calo ime y (DSC) The mal analyses and de e mina ion o he glass ansi ion empe a u es we e pe o med on a Pe kin-Elme Diamond DSC a a hea ing a e o 20 K/min. The desc ip ion o he SEC, NMR and MALDI-ToF expe imen s is gi en in he Suppo ing In- o ma ion