Experimental Chemistry II
Experimental Chemistry II CHEM C1260
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Date Created: 09/19/15
lnline Monitoring of Crystallization Processes Lynne S Taylor Hakan Wikstrc39jm Alan Gift Yuerong Hu U I I V I I I I 39I39I Outline Crystallization from solution Solution concentration Polymorphic form Transition temperature Recrystallization during wet granulation Kinetics Effect of excipients PURDUE UIIVIISI39I39Y Primary Instrumentation Control Electronics ProbeHeat Kaiser Optical Systems Inc Rxn1 Raman system operating at 785 nm PURDUE 3 UNIVERSITY Crystallization from Solution Goals Determine how much information can be extracted from Raman data Solution concentration Initial polymorphic form Conversion kinetics Correlate Raman data with light scattering data FBRM to verify signal interpretation PURDUE 4 UIIVIISI39I39Y Model System Fufenamic Acid 42 C Form lt gt Form Form Iwhite For I yellow 5 Raman Spectra Ethanol l 1680 cm 1 i Solute Form v 1 200 400 600 800 1000 1200 1400 1600 Raman Shift cm391 PURDUE UIIVIISITY 1800 Raman Monitoring of Desuperaturation During Crystallization IE 00 04 E E 03 3 02 01 D 1000 1000 1000 1020 1040 1000 1000 1000 1020 Raman Shift 11390ij Normalized to ethanol peak at 881 cm i PURDUE 7 UIIVIISITY Feasibility quantitative measurement of solution concentration correlation between Raman and UV data 190 0029 7 170 7 150 0024 7 E Q 7 130 a 2 E a i l C c 0019 7 o 7 7 110 5 g E E E m w a 7 7 90 g 0014 7 o 0 70 0009 7 39 7 50 0004 i i 30 0 20 40 60 80 100 Time minute P E UIIVIIIITY 8 Multistage Crystallization solution data 250 j 200 E m E E 150 39 Raman 2 Temperature 5 g 100 39 39 I 39 S 2 W 39 39 o W 50 o 39 u I c39 Solubility of Form Ill 34 4 mgml 39 Solubility from Raman data 3414 mgml 0 I I I I l I 0 100 200 300 1200 1300 1400 1500 Time minutes PURDUE UIIVIIII39I39 70 60 50 40 30 20 Temperatu r C Raman Spectra Ethanol l 1680 cm 1 i Solute Form Form 200 400 600 800 1000 1200 1400 1600 1800 Raman Shift cm391 PURDUE 10 UIIVIISITY Form wlw Multistage Crystallization Form 100 8 I 80 a 39 39 0 39 39 f t o 9 60 i o 39 Form seeds 339 4o c39 f 20 O w I I III I I I 0 50 100 1300 1400 1500 TIme minutes 11 UIIVIIII39I39Y Concentration mglmL Multistage Crystallization All Mass Balance 250 39 I i We 200 o 39 A 1 a 39 Mm WX 39 t e 0 i 150 1 I A i Solute 3 Formlll 100 s 4 A Forml 05 0quot i 3 39 2 W M f 3 50 39 39 a 1 O I I I v I I 0 100 200 300 1500 1300 1400 1500 Time minutes 12 UIIVIIII39I39Y Total Particle lsecond Correlation Between Spectroscopic Data and FBRM 10000 7 7 240 8000 7 Particle Count 180 Soute 6000 Temperature Onset of Nucleation a 120 4000 7 60 2000 7 0 0 0 20 40 60 80 100 120 Time minutes PURDUE 13 UIIVIIII39I39Y Concentration mglmL Total Particle lsecond Correlation Between Spectroscopic Data and FBRM Concentration of Form mglmL 10000 7 8000 7 180 Lasentec Form 6000 Temperature 7 120 4000 7 60 2000 7 0 0 0 20 40 60 80 100 120 Time minutes PURDUE 14 UIIVIIII39I39Y Summary Linear relationships were observed between Raman signal and both solution concentration and polymorphic ratios Solution concentration could be monitored during crystallization Polymorphic form and kinetics of conversion could be readily monitored PURDUE 15 UIIVIISI39I39Y Monitoring of Crystal Form Changes During Wet Granulation U I I V I I I I TY Statement of Problem Water plays an integral role throughout the pharmaceutical manufacturing chain Crystallization Wet granulation Coating Performance testing eg dissolution 13 of all pharmaceutical drugs can form hydrates1 Lower solubility Inferior particle properties 1 KR Morris in Polymorphism in Pharmaceutical Solids Ed HG Brittain 1999 17 UIIVIISI39I39Y SolventMediated Transformation Three stages2 Metastable phase dissolves Stable phase nucleates Growth of the stable phase Described by Cardew and Davey using mechanistic model without nucleation3 2 E Shefter et al J Pharm Sci 52981 1963 3 PT Cardew et al Proc R Soc Lond 398415 1985 18 UIIVIIII39I39Y nLine Monitoring of High Shear Wet Granulation Hydrate formation can occur during wet granulation mm mm WEIEI man man iEAEI mm D Ramaquot SM icm 19 Ill39llllTY Hydrate Formation 105 90 75 60 5 lgt 45 30 15 0 i i i i i i i i i i i i i i i i i i i i O 20 4O 6O 80 100 Time min Typical transformation profile for hydrate formation obtained using Raman spectroscopy PURDUE 2o UIIVIIII39I39Y Hydrate Formation 105 a 90 V do 7 A 75 kDA SA CA 3 607 Hydrate formation will occur E 45 through a solventmediated transformation SMT 30 Dissolution ofthe metastable 15 phase 05 min i gt 0 i i i i i i i i i i i i i i i i i 0 20 40 60 80 100 V v Time min 39 39 M I I A l 39 In TM 39i i a f f A I a 5 AA Noyes eta J Am Chem Soc 1912930 1897 Dissolution of the metastable phase continues until all have been dissolved 21 A Metastable phase anhydrate V I l V l I l l 39I39 Y Hydrate Formation 105 a 7 ENB 90 75 a 0 Hydrate formation will occur g 60 39 through a solventmediated 45 a transformation SMT 30 Dissolution ofthe metastable phase 05 min 15 7 Nucleation of the stable phase 5 min 0 l l l 0 80 100 39 39 I w ne min r In TM 12 0 g a am i t V f A I a 6 D Turnbull eta J Chem Phys 1717 l 1949 Dissolution of the metastable phase continues until all have been dissolved 22 B Stable phase hydrate I l l V l I l l 139 Y Hydrate Formation Hydrate formation will occur through a solventmediated transformation SMT Dissolution ofthe metastable phase 05 min Nucleation of the stable phase 5 min Growth ofthe stable phase 575 min 80 100 Time min 39 u 30 min 90 min EM is L umquot M a 1 7 N RodriguezHornedo et a Pharm Res 85643 1991 23 Dissolution of the metastable phase continues until all have been dissolved II I l V I I I I 39I39 Y Sampling Methodology Evaluation Sampling techniques for monitoring A flat face immersion probe 60 pm spot size Kaiser RXN1 spectrometer 25 noncontact optics 150 pm spot size Kaiser RXN1 spectrometer 10 cm noncontact optics 3 mm spot size Modified RXN1 PhAT System PURDUE 24 UIIVIIII39I39Y PURDUE 25 Sampling Volume Determination Spot size diameter provided by manufacturer Penetration depth determined by analyzing 13 mm diameter AT compacts PhAT Standard Immersion System 25quot NCO optics mm Spot size 3000 150 60 um Pen depth 2000 600 300 um Vsampling 14100 106 085 n1 Index 16588 125 1 AT Theophylline anhydrous NCO Noncontact optics u I I v I I s I 39I39 Y 26 Effect of Sampling Volume Calibration comparison between small and large spot sizes Calibration of monohydrate nitrofurantoin 5 min collection time compared to 10 s 100 100 yx410396 yx210 6 R2 09732 R2 09992 80 80 8 6O 8 60 a a D D 3 3 o 40 O 40 2o 20 O l m l l O l l l o 50 100 o 50 100 Predicted Predicted PURDUE 27 UIIVIIII39I39 Intensity IU 14000 12000 10000 8000 6000 4000 2000 Effect of Sampling Distance The granulation bed moved 34 cm in height per mixing revolution PhAT System 90 SNR throughout the range 25 NCO as low as 20 SNR PhAT System never less than 35 SNR Immersion optics 10000 Standard NCO 8000 1100 PhAT System 900 E 6000 E gt gt a a 700 5 5 g 4000 g 500 2000 T f 0 00 05 10 15 20 25 00 20 40 60 80 100 120 140 160 00 50 100 150 200 Sampling Distance cm Sampling Distance cm Sampling Di ssss ce cm SNR Signaltonoise ratio 28 UIIVIIII39I39Y 250 MT Effect of Sticking immersion probe sticking problems poo I I mmersion Time min 29 PURDUE VII39IIII39I39Y MT Theophylline monohydrate Summary Raman spectroscopy is suitable for monitoring hydrate formation in aqueous environments Noncontacts optics is preferred over immersion optics due to sample stickiness to the end of the probe Large spot size system results in more representative sampling Faster sampling rates Less susceptible to subsampling PURDUE 30 UIIVIISI39I39Y Hydrate Formation Five model compounds selected Material Polymorphic Form mp CAS Molecular Structure C CDC Ref Code 0C pKa log P O A tricljnic Bphase form caffeine N N NIWFEE ocform CAF 238 104 007 583908392 N o monocljnic 45 hydrate form N CAFIN39EOI monocljnic 10W temp form carbamazepine CBMZPNI 0 CBZ N 193 1391 245 298464 Monoclinic dihydrate O NHZ FENOTOZ o monoclinic B form nitrofurantoin O l LABJON NF N O N N NH 268a 72 047 67209 I orthorhombic form H O o HAXBUD H d if monoclinic form H su aguam me ZZZAYP03 SFG HZN lt gt N 113 122 57670 O gt NHz monoclmlc HN SOGUANZO O i orthorhombic 10W temp form theophylljne N N BAPLOTOI 270 TP 274 86 002 58554 HN o monoclinic form zN THEOPHOI 353225 PURDUE 31 bCalculated UIIVIISI39I39 Hydrate Formation Three different experiments Slurry experiments 2 9 drug 40 ml water Seeded slurries as above but 10 seeds Granulations 4 9 drug 2 ml water Compound Transformation Time Slurry Seeded Granulation sulfaguanidine 03 03 03 theophylline 30 30 30 oarbamazepine 100 60 80 caffeine 125 90 70 nitrofurantoin 180 60 50 PURDUE 32 UIIVIISI39I39Y Effect of Processing Small scale granulation and unseeded slurry experiments comparable for TP Small scale granulation and seeded slurry experiments similar for NF 250 Time min Time min MN Nitrofurantoin monohydrate 33 UIIVIISI39I39Y Manipulating Hydrate Formation U I I V I I I 3 TY MT Slurry Experiments Effect of excipients on transformation kinetics 100 85 70 55 40 25 Water PVP HPC HPMC E4M HPMC K4M HPMC F4M MC A4M Time min PVP Polyvinyl pyrrolidone HPC Hydroxypropyl cellulose PURDUE 35 HPMC Hydroxypropyl methylcellulose u I I v I I 3 I 1 1 MC Methylcellulose Growth Rate Determination Result from growth rage dether ination rowt ra e Polymer HmS None 059 PVP 045 HPC 009 HPMC 007 MC 003 PURDUE 36 UIIVIISI39I39Y x In 1 y 100 f a Q l quot wig r l 397 39 I I 51 50 min t 1 In f PURDUE 37 W MT Granulation Experiments Similar trend to slurry experiments Excipients affect transformation rate and onset time Methylcellulose the most effective UIIVIIII39I39 quotV 9 80 21 PVP K2932 HPC 70 o5 HPMC E4M 13 o5 HPMC K4M o5 HPMC F4M 60 f o5 MC A4M 15 o5 MC A15 i50 39 C i 3 40 12 1 3 3 3 g 30 3 09 20 i i 06 10 i i I i i I I I I 0 03 Water PVP K HPC HPMC HPMC HPMC MC MC A15 05 2 3395 5 65 2932 E4M K4M F4M A4M Time min PURDUE 38 Granulation Experiments SEM pictures of granules Morphology changed when using additives SEM Scanning electron microscopy P a Water as granulating liquid 39 b MC A15 solution as granulating liquid LabScale Granulation Individual effects of excipients in small scale granulations 100 Water PVP K 2932 85 39 MCC Mannitol 70 39 HPMC E5 55 E o 40 25 10 5 I 1 0 1 2 3 4 5 6 Time min MCC Microscrystalline cellulose 40 UIIVIIII39I39 LabScale Granulation Standard formulation 110 90 70 I E 507 No HPMC1 No HPMCZ No HPMC 3 30 10 10 w 0 2 4 6 8 10 12 14 Time min PURDUE 41 UIIVIIII39I39Y LabScale Granulation Standard formulation compared with 03 ww HPMC E5 added 110 90 7 7o 7 No HPMC1 2 50 No HPMC 2 No HPMC 3 O 30 7 034 HPMC 10 a 10 r o 2 4 6 8 1o 12 14 Time min PURDUE 42 UIIVIIII39I39Y Caffeine Water 062 d 47 HPMC 067 j 40 CAF HPC 022 l 05 Hydrate CMC 025 j 12 PAA No Growth PVP 040 j 25 PEG 068 j 27 0 20 40Timemin60 80 100 No Growth 3000 7 2500 7 2000 7 g g 1500 7 O 1000 7 Caffeine 0 200 400 600 800 1000 PURDUE 43 UIIVIIII39I39 CBZ Hydrate 100 Growth Rate 80 7 Polymers pmsec Water 49 i 16 60 PAA 44 i 14 CMC 25 i 13 40 7 water HPC HF C 008 i 06 20 7 HPMC PVP 004 i 02 0 w I I HPMC 0003 t 001 0 10 20 30 40 Time min Carbamazepina 0 100 200 300 400 500 600 Time min PURDUE 44 W Summary Hydrate formation can be manipulated by commonly used excipients The difference between experimental types lies in the nucleation kinetics Excipients can have a cooperating effect during granulation experiments leading to complete inhibition The major mechanism of action seems to be specific adsorption to crystal surfaces PURDUE 45 UIIVIISI39I39Y Conclusions Raman spectroscopy is suitable for monitoring hydrate formation in aqueous environments Hydrate formation kinetics is dependent on compound specific properties as well as extrinsic factors Commonly used pharmaceutical excipients have a profound influence on the transformation kinetics PURDUE 46 UIIVIISI39I39Y Acknowledgements Particle Technology and Crystallization Consortium AstraZeneca Pfizer Kaiser Optical Systems Inc Steven Byrn Allan Myerson Steef Boerrigter Jessica Liang PURDUE 47 UIIVIIII39I39Y s Plasmonic Nanostructures in Sensing Molecules g Presentation Outline You a re here Introduction Surface Plasmon Resonance SPR Localized SPR Nanoshells Surface Enhanced Raman Scattering SERS Challenges Conclusions Summary Introduction Variety Precise Control a QMquot SlD lAu nanushell Antibody Easy binding to Bio Sensing Cancer Therapy Molecules Promise of more and more advanced applications than conventional techniques Fluorescence Chromatography Mass Spectrometry Introduction Motivation ENERGY 0V 25 2 30 1 390 L15 V B G Y 0 R T GOLD e RUBY 0 RED l E Q 0 SILVER 2 YELLOW 900 560 660 73 WAVELENGTH nm Plasmon resonances give specific More recently plasmon I metallic nanoparticles a strong and Resonant nanoparticles g well defined color This effect was have been used as Bulk Gold Yellow already used in the Middle Ages to biomarkers fabricate stained glass windows Nanogold Red Proc Nat Ac Sci vol 97 pp 9961001 2000 Plasma Resonance Plasma resonances for various geometries Single atom Mi KC 9 fJCMV Nucleus COD plasma frequenc 39y restoring force spring constant sk mix O P iNex E747239P 8 Material Resonance Resonance condition Frequency Bulk Metal gaff 0 mp Planar Surface 5quot p geff 8d 2 S here a p Self 725 1 7 L EIIIpSOId Serf LM V came Optical Properties of Metal Clusters 95 Kreibig and Vollmer19 Types of Plasmonic Structures Surface Plasmon Resonance SPR StrUCtured LSPR or SERS lms plastnon metal lm slide quot250 mu Nanoshells LSPR lt5 Presentation Outline You a re here Introduction Surface Plasmon Resonance SPR Localized SPR Nanoshells SERS Challenges Conclusions Summary First Publication of SPR Effects 1902 The phenomenon of anomalous diffraction on diffraction gratings due to the excitation of surface plasma waves was first described by Wood in 1 902 XXV On a Rerazarkable Case of fazeven Distribution qf Light in a Di v action Grating Spectruuz By R VV WOOD Prqfessar O f b39svperiwnentai Physics Johns Ioplcins University 9 IT is a well known fact that in the spectra fanned by a dif raction grating the light is unevenly distributed that is the total light in any one speetruul will not recombine to forln white light I have been examining a 11051 remarkable grating recently ruled on one of the Rowland dividingengines in which this uneven distribution is carried to a degree 31111051 incoznpre hepsible If the spectra of an incandescent lamp are viewed 9 Read June 20 190 V What is a surface plasmon polariton Plasmon Surface plasmon E Z Electron densi wave Dielectric Strong local eld ty A 8d excuted by photons of light Metal 1 9 H 8I39T I rmn gpm mm 7 evanescent wave air 12 Occurs on the metal dielectric surface glass En InCPdBifII light 1 t n varying n 12 8 8 a Wave vectors must satisfy ksp k m d 8d Slng 8m 8d C SPR Introduction Plasmonpolariton excitation produces absorbance peak at specific frequency Shift in the absorbance spectrum indicates presence of analyte molecules change in dielectric refractive index First experiments on biosensing using SPR method min 9pm mm evanesvcem wave air HI quot REFLECTANCE glass rm 1 V ll llCHllEflt llght u ax var1an 2 9 1b ANGLE 0F INCIDENCE degrees Liedberg et al 1983 10 b SPR Excitation Techniques Optical wave Prism coupler Mefol layer Me AnalMe i g 5 Es Grating ozp cci wove Guided optical wave I Waveguide Meicl lover uidlng Kretsch ma nn Geometry The periodic structure translates to k5 Ksp im2 j m012 a The sensor structure is irradiated from waveguide 11 SPR Applications Chemical Biological 1 Sensing 1 Real time sensing 2 Reaction kinetics 2 DEtECtiOIquot 0f binding 3 Concentration reaCtlons measurement 3 Proteomics Mass Spectrometry 4 Plasma membrane studies 5 Equilibrium properties 5 Drug denvery teCl ll39liClU39EES and many more and many more O BIACORE 399 TEXAS INSTRUMENTS wwwblacovecom 12 lt5 Presentation Outline You a re here Introduction Surface Plasmon Resonance SPR Localized SPR Nanoshells SERS Challenges Conclusions Summary 13 Localized Surface Plasmon Resonance from propagating to localized plasmons propagating localized mm 11cm Ilm eevanescem wave air n21 glass n1l III llgl lL re ecmghr gt1 vE TryIrg n I J 0P R t f 8 8 a a esonan requency i d i ksp k m i 5394 111 9 res J3 geometry conSIderatIons gm 84 c In both cases resonance depends on dielectric permittivity 14 Localized Surface Resonance Plasmons LSPR 7 1 l 2 I A 5565 5684 5 015 7 y gt ShapeSize Rayleigh scattering Ag nanoparticles Spheres Prisms I disks rods 39 39 39 Av Diameter 35nm g an n A if I I 30 6M TIN InnaImam mi 39 5880 6008 51on mj l l39m EnVIronment Ag nanoparticle in different solvents kmax shifts to larger k as ed increases Wequot 250 nm 250 um Binding of antibody 4 5 Wai enm Df nm 65 m to Ag nanosensor Calculated Extinction spectra in various solvent environments Molecule sensmg nitrogen methanol propanol 32nm snlft n resonance peak chloroform benzene Mie theory Nano Lett Vol 4 N0 6 2004 Nano Lett Vol 3 N0 8 2003 Van Duyne Group Northwestern Univ 15 Nanoshells one con guration for LSPR 20mm 113nm r141 r251 nm g 86 99 a E a m w u u 1 rm WU g wmmimi 39 131 149 so rm amaze onm gm 6 2mm 9 5M 5 7 Optical resonances of gold silica a nu core nanoshells as a function Calculated CII lt3eS of their coreshell ratio EXPeI Imental SOIId solid gold spheres clashed More sensitive than simple nanoparticles gain r2r1 Resonance frequency is a strong function of geometry More resonant frequencies possible Technology 397 Cancer Research amp 77 eatment 3 1 2004 Nano Lett 3 10 1411 1414 2003 16 Preparation of LSPRNanoshells One process to make a localized surface plasmon resonance structure Van Duyne A 11 Clean Substrate Building a nanoshell Halas 0quot to 55 mo Nanoparn lc summrcmnmmd uizrxmau uz nm Adduiunal Gold Punquot Core IOD nm withAn lnes Gold calloudAumhed um Goldshdl ls Cumplece Nanoshell fabrication nanoscale metallic overlayer grown on top of silica core Nano Lett 3 7 939 943 2003 17 lt5 Nanoshells in bio applications Properties Optical activity in bio compatible wavelengths Strong tunable absorption in NIR region 700 1300 nm maximum light penetration through tissue in NIR Easy conjugation with speci c proteins Chemical Photochemical stability Biocompatible non toxic to tissue gold 18 NIR Photothermal Tumor Therapy NIR radiation is now regularly used in the treatment of tumors Radiation heats the tumor cells causing cell damage or destruction AT for radiation therapy is on the order of 10 C not very powerful Is there a way to make this procedure more efficient and powerful using plasmonics YES We can prepare special nanoshells with a resonance peak at the required frequency act as heat delivery system Nanoshell preparation Nanoshells used 128 nm core diameter 14 nm old shell peak absorbance at 820 nm max penetration of light through tissue Passivation Minimize aggregation to physiological environment saline Conjugate the nanoshell with a tumor specific protein antibodies against oncoproteins PEG to block non specific absorption sites and enhance biocompatibility Hirsch et al 2003 Summer Bioengineering Conference Florida Hirsch et al Technology in Cancer Research amp Treatment 3 1 2004 19 Nanoshell NIR Photothermal Tumor Therapy on Mice n x a 25 E r 15 L 9 quotJ i z 4 8 1a Tern erature vs time Cells treated with laser only vs Nanosge treatment Red 3 Nanoshell NIR both nanoshells and laser in Vitro t39ssue amage area I Blue NIR only Nanoshells injected 2 5 mm beneath the tumor surface and bind to tumor proteins Nanoshells preferably accumulated at the tumor site due to enhanced permeability and retention EPR leaky tumor vessels with wide junctions incomplete membranes channels Death of cells at 820 nm 35Wcm2 A T 374 C within 4 6 min NIR only A Tlt 10 C No healthy tissue damage observed no nanoshell leakage outside of tumor Technology in Cancer Research amp Treatment 3 1 2004 Proc Nat Ac Sci 10023 13549 13554 2003 20 Photothermally triggered E J BSA releaeetl lmglg dw Time mini Release of BSA from Non irradiated diamond Irradiated hydrogels triangle Irradiated nanoshells square Conjugate nanoshell with biomolecules 5 95A nelsaeecl mgg dwl drug system l 56 sh 7390 11mg Izmir Release of BSA from nanoshell composite hydrogels in response to irradiation Strong absorbance at a specific frequency will force release of the biomolecule 800 1200 nm Small increase in T doesn t cause tissue damage West JL and Halas NJ Current Opinion in Biotechnology 11 215 217 2000 21 Sensing using Nanoshells General idea Change in resonance when there is change in the dielectric constant of the environment Change due to Change in host material Molecule binding Dimer formation 1 J E E E E o 5 1 aon sun awn 1 W avelength nm Single nanoshell with a 96 nm diameter core and a 22 nm thick shell solid line and a dimer system composed of uch nanoshells whose surfaces are separated by 10 and 40 nmA LR Hirsch JB Jackson A Lee NJ Halas JL West Anal Chem 75 2377 2381 2003 22 lt5 Whole Blood Immunoassay Using Nanoshells 1 8 S C A 1 e s Coat the nanoshells with the antigen 1 J 939 391 The two nanoshells get connected with E08 quotquot target molecule form dimers g mquot UY m 52 A r 7 L r Antigen induced aggregation detected via 039 4 500 SCPD 700 BBC EIOO 1 CBC red shifting of the spectrum 720nm and Waveengmmm reduction of the amplitude of the spectrum Disperse nanoshells solid Nanoshellantibody conjugates dotted Rapid detection of specific analyte in complex biological media Detection of immunoglobulins in saline serum and whole blood Detection limits in the order of 100pgmL 44ngmL in 10 30min Stable dimers still sensitive after a week LR Hirsch JB Jackson A Lee NJ Halas JL West Anal Chem 75 2377 2381 2003 5 Presentation Outline You a re here Introduction Surface Plasmon Resonance SPR Localized SPR Nanoshells SERS Challenges Conclusions Summary 24 Raman Scattering a primer Sample Ra man Spectra Diamond 30000 25000 Incident photons M0ecue WI Virtual state u n 8 a 15000 lnlemlly mu 1 none 2cm 7cm 1 mm om w inunsnylam Raman Scattering Energy Diagrams Stokes Scattering Diamond 39 o CC lattice 0 Only one bond type Vibrational Flnal 0 One Raman band States Initial 00 Van 2200 2700 Virtual state 1k Raman Shit lnv cm Cequotquot39 se AntiStokes Scattering Cellulose C6H1005 0 Organic molecule 0 Many bond types Vibrational Initial 0 Several Raman bands states Final 391on 27cc 32cc 3700 Rarnzn shin nv cm 25 Ra ma n Scattering Raman scattering is a weak effect Raman scattering cross section 10393O cm2 If we can increase the local elds we can obtain a larger effective scattering cross section More Raman scattering higher signal Lower detection limits How can we increase the local elds Plasmon excitation produces induced elds Fields can be localized and intense Raman scattering scales with E4 Surface enhanced Raman scattering SERS 26 Field Enhancement in Metal Nanostructures Structured Arrays MBA yisu I z 1 xl 051102 1 a 393 391 b C Periodical array of metal nanoparticles a and calculated local Raman enhancement in such system at A 06 um 17 and A 1 pm 6 The sizes are given in nm Genov AK Sarychev VM Shalaev and A Wei Nano Letters 41 2004 Fractal Aggregates Higher local field intensity Higher Raman signal for molecules experiencing those larger fields Raman enhancement scales as the 4th power of E leading to local enhancement as high as 1 O1 1 VM Shalaev Nonnear Opt5 OfRandom Med22 Springer Verlag Berlin 2000 27 Adaptive Silver Film ASF as SERS substrate Protein Sensing with ASFs Outside spot Disintegrated particles High filling fraction 65 Very little clustering RMS roughness 65 nm 1 Metal nanoparticles on glass substrate 2 Deposition of a protein droplet 3 Metal particles and proteins form closelyspaced structures Inside spot Clustered particles Lower filling fraction 50 RMS roughness 46 nm 4 Meta paniclesthatare notattachedto Smallergaps result in larger field enhancement the protein are removed by washing H Proteins stabilize films washable Protein functionality preserved SERS Protein Studies with ASFS Insulin Analog Detection Millions of diabetics many who must use insulin daily Insulin Humulin is slow acting While Humalog is fast acting Humulin and Humalog are very hard to distinguish from each other Protein Binding Detection Binding responsible for many processes in the body Key for diagnostics Drug development research 29 Insulin Analog Detection N Terminus Gly lle Val 39 L11 Chum A 1 cyscy mGlnl m A 39n n ms Len Cy Gly Set His Len Val Glu Ala Len 1y L39cu Val 6111 Am Chain B Val Pb N Terminus Humltlog yn xl39n Humulin 0 tends to form dimers and hexamers in the presence of Zinc ions oslow acting in bloodstream Humalog o lysine Lys and proline Pro are reversed 0 same empirical formula 0 same molecular weight 0 minimizes the tendency to form dimers and hexamers fast acting Monomer Hexamer a SERS Spectra of Humulin amp Humalog SERS of Humulin and Humalog SERS at sub monolayer prOtein Coverage Average enhancement factor 25x106 to Rama on quartz 3x106 to Rama in liquid D Ferrari J R Diers D F Bocian N C Kaarsholm and M F Dunn Biopolymers Biospectroscopy 62 249 2001 35 3 a A i 325 i W 3 2 7 g E 39 1 i 015 C C 7 r g 1 L l 1 W 05 a D 500 700 500 1100 1300 1500 Wavenumbercm1 200 100 L 391 g 0 1000 1500 Ram an shift Inv cm VP Conventional Rama n spectr urn for human insulin Drachev MD Thoreson EN Khaliullin VJ Davisson VM Shalaev J Phys Chem 2004 31 SERS for Monolayer Protein Binding Detection STE PS Ab Y Y Y lmnz39lobilization of antibody SERS monoclonal Ab 05uM 2ul on ASF surface ASF restructuring SERSactive substrate ASF Ag Incubation with anti 39 39 gen solutlon 9 O Q O 90 9 0 30 min Flagtagged protein FTP BAP bacterial alkaline phosphatase c E 5 y W vrr Wash way unbound proteins SERS only specific binding remains 32 Protein Binding on ASFS Ab and Ag Binding Ab Ag incubation Blue Antibody Ab Red Ab inc C Antigen C Ag Green Ab inc UnAg Substrates are bio array compatible and exhibit monolayer sensitivity sun auu luau lzluu lqr Raman Shift cm 1 Region of interest 33 Other SERS biosensing applications Phosphorylation disease detection Glycosylation Bioagent detection 2 B CO ZIWMM lniens ily 500 000 700 500 900 1000 1100 1200 1300 1400 1500 1000 1700 Invelse em 1 03 gm 0 R Ebola Virus Sena A Z g I 02 I MN em 00 70 500 000 1500 100 Iuvelse 0m TP Goyani Purdue Masters Thesis 2004 34 5 Presentation Outline You a re here Introduction Surface Plasmon Resonance SPR Localized SPR Nanoshells SERS Challenges Conclusions Summary 35 Plasmonic Biosensor Challenges Reusabilib A 020 7 r Anti Blotln 0 1 8 Adsorption 016 1 73 3 014 LLl 012 r 6 0 O 010 x 500 600 700 800 9 0 Wavelength nm E 03920 AmiBiotin o 1 8 2quot esorption C 39 2 I 1 2 0316 r E 439 012 20 2a 010 r v 500 600 700 800 9 0 Wavelength nm Journal of Fluorescence Vol 14 No 4 July 2004 Van Duyne LSPR Ag nanoparticles 1 2 Ag nanoparticles Anti biotin added 1 Ag with Anti biotin 2 Biotin added Peak 6711 nm 6818 nm Peak 6818 nm 6702 nm 36 Plasmonic Biosensor Challenges Fabrication Lithography Integration in larger units High yield Stability Can it be used over and over again even after long time Specificity Recognizable sensing Multifunctional capability use of shapesize Big issue in high protein concentration environ ments Chemistry Biocompatibility Understanding of protein surface interactions 37 a Summary Conclusions Ease of fabrication advanced interest in nanomaterials and applications Plasmonic nanostructures can be used in a variety of applications sensing bio environmental Structures SPR Nanoparticles Nanoshells SERS Plasmonic Nanostructures are a very exciting and promising eld that might give solutions to problems recent technology cannot solve 38
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