Ultrathin layer chromatography using
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1 Ultrathin layer chromatography using electrospun nanofibers SusanV. Olesik Department of Chemistry Ohio State University International Symposium for High-Performance Thin-Layer Chromatography Basel July 6, 2011
2 Thin Layer Chromatography Ultra Thin Layer Chromatography h (UTLC) Uses thin stationary phase (~10 μm) in comparison to HPTLC (~ 200 μm) Non traditional stationary phase structures Silica Monoliths and Nanostructures Improve sensitivity while reducing analysis time and amountof of consumables required Lower sample capacity than HPTLC
3 Electrospinning Apparatus Silicon (Collector Plate) Syringe Pump High Voltage Source
4 Electrospinning Electrospinning is a process in which a polymer solution is used to make ultrafine fibers Electric field is applied to the polymer solution Charge repulsion causes formation of Taylor cone Taylor Cone
5 Electrospinning Any polymer that can be electrospun can be used as the stationary phase 2 polymer systems: Polyacrylonitrile (PAN) Initial studies SU 8 Photoresist HEAT Glassy Carbon
6 Spinnability of a Polymer The ability of a polymer solution to form uniform fibers is dependent upon many parameters: Solution properties Polymer molecular weight Viscosity Conductivity Surface tension Electric field Applied voltage Distance from tip to collector Solution flow rate Temperature Humidity
7 Effect of Concentration 75% 70% 20 µm 20 µm 50% 25% 20 µm 20 µm
8 UTLC -- Electrospun Polyacrylonitrile Fibers 10% PAN AIR PAN Al Foil Thickn ness (microns) SEM micrographs of the electrospun stationary phase used for UTLC Time (minutes)
9 Variation in Retention Factor Commercial Phase R f % Acetone (Acetone:H 2 O) E-ULTC R f 0.5 androsterone, cholesterol and cortisone. (n=5) % Acetone (Acetone:H 2 O)
10 High Efficiency Nano Sil CN PAN UTLC Steroid Analysis How? Effic ciency, N Z N 16 S w Androsterone Cholesterol Cortisone J.E. Clark and S.V. Olesik Anal. Chem. 81(10), (2009).
11 Ultra-thin layer chromatography using electrospun fibers N 16 2 Z S w CN-Modified TLC Spot Width (mm) Electrospun PAN UTLC Spot Width (mm) Androsterone Cholesterol Cortisone
12 High Efficiency 50 Electrospun 10% PAN Commercial Si 40 Nano Sil CN Silica Dist tance (m mm) N 16 Z S w Square Root of Time (sec 0.5 )
13 High Efficiency 50 Electrospun 10% PAN Commercial Si 40 Nano Sil CN Silica Dist tance (m mm) Square Root of Time (sec 0.5 )
14 E-ULTC: Polyacrylonitrile Fibers 400 nm fibrous stationary phase E-ULTC requires less time and therefore less solvent than typical TLC plates Efficiency of the separations substantially improved compared to that determined using commercial phases. Separations used minimal materials (1 ml polymer) and solvent (< 5 ml) Mat thickness impacts efficiency. Thicker mat improved efficiency J. Clark, S. Olesik, Anal. Chem. 81 (10) (2009).
15 Carbon Ultra Thin Layer Chromatography SU 8 carbon 7 cm Glassy Carbon Substrate
16 Devices Used for Carbon TLC Comparison Device Mat Thickness (μm) Avg. Fiber Diameter (nm) PAN 24 ± ± 55 UTLC 600 C 16 ± ± C 10 ± ± C 13 ± ± 70 Silica Gel 200 N/A
17 Variation in Migration Distance with Time Mig gration Distance (mm) Silica PAN UTLC 600 C glassy carbon UTLC Time (sec) 0.5
18 Migration Distance as a Function of Fiber Diameter
19 Z Lucas-Washburn Model Predicting Solvent Travel Behavior 2 f Rt cos 2 Device Mat Thickness (μm) Avg. Fiber Diameter (nm) Effective Pore Radius, R = surface tension PAN 24 ± ± ± 10 nm = solution viscosity = contact angle 600 C 16 ± ± ± 35 nm R = effective pore radius t = time 800 C 10 ± ± ± 15 nm 1000 C 13 ± ± ± 25 nm Silica m 345 ± 25 nm Gel diameter
20 S640 Study of Separation of Laser Dyes Rh590Cl Rhodamine 101 Rh610P & Rh610Cl Kiton Red P597
21 Retardation Factors of Laser Dyes Left rhodamine 610 perchlorate, rhodamine 610 chloride, kiton red Right pyrromethene 597, rhodamine 101, sulforhodamine 640 Mobile phase: 2-propanol.
22 Laser Dye Separation 250 Rh 610 P 200 S640 Intensity P Distance, mm Jonathan E. Clark, S.V. Olesik, J. Chromatogr. A, 1217 (27) (2010).
23 Efficiency Comparison Numbe er, N Plate Commercial PAN UTLC 1000 C Carbon 0 Rh610P Rh610Cl KR
24 Efficiency Comparison 8x10 4 PAN UTLC 600 C 800 C 1000 C, N Number Plate 6x10 4 4x10 4 2x S640 P597 Rh101
25 High Resolution 60 PAN UTLC Plate Laser Dye Analysis 40 Res solution S640/Rh610P S640/P597 Rh610P/P597
26 Study of Separation at of Essential Amino Acids Phenylalanine Lysine Threonine
27 Tunable Retention Migration Order: 600 C: Thr Phe Lys y 800 C: Phe Thr Lys
28 Essential Amino Acid Analysis 250 Lys 200 Inte ensity Phen Thr mm development distance 600 C Max. Sample capacity 10-5 M Distance, mm
29 Efficiency Comparison Plate Number, N Compound 600 C 800 C 1000 C Cellulose* Lysine Threonine Phenylalanine 37,500± ± ± ,000± ,400± ± ,000± ,600± ±30 N/A *S.A. Nabi, M.A. Khan, Acta Chromatogr. 13,161(2003).
30 6 High Resolution Amino Acid Analysis 600 C 800 C C Mixed Cellulose Resolutio on Lys/Thr Lys/Phe Thr/Phe
31 Variation of Plate Number with Development Distance Plate Height (micron) Development Lysine Threonine Phenylalanine Distance (cm)
32 Biodegradeable Polymers: Electrospun Polyvinyl l Alcohol l 190±50 nm
33 Importance of Cross-linking No crosslinking Crosslinked and soaked water Crosslinked and soaked in optimized solvent ( th (ethanol/butanol/water) l/b t l/ t )
34 Thickness of Mat with Each Additional Layer
35 selectiv vity α 2,4 2,2 2 1,8 1,6 his/cys PVA Selectivity Mobile phase:methanol/butanol/water (7:5:1, v:v:v) 1,4 1,2 phe/asp met/gln tyr/thr ala/trp arg/asp 2,4 1 2,2 2 Silica Selectivity Mobile phase:ethanol/butanol/water (5:7:0.5) sele ectivity/α 1,8 16 1,6 1,4 phe/asp try/thr 1,2 1 cys/his met/gln trp/ala arg/asp
36 Optimization of Capillary Diameter Used for Application plate heigh ht (μm) phe thr trp tyr 250 um 100 um 50 um 20 micron thick mat with 190 nm fibers
37 Optimization i of Iinjection volume d 30 ht (μm) plate heig thr trp try 2 nl 6 nl
38 Concentration Range Appropriate Using 50 micron Capillary Pla ate height (μ μm) thr trp tyr 14 mm 7 mm 4 mm 3 mm
39 Travel distance as a function of Development Time 3,5 3 2,5 (cm) Distance 2 1,5 1 0, Square Root of Time (sec 0.5 )
40 Efficiency Comparison to Commercial Phase
41 Summary E-UTLC provides Lower mobile phase use than other TLC separations Higher speed separations Improved efficiency Devices are chemically and mechanically robust Future: Much to be studied d on exactly how improved efficiency i is gain further work on improving precision of retention factors underway
42 Acknowledgments Funding: NSF- Chemistry Division; NSF-Engineering g Division, NSEC; NSF- Engineering - Fluid Dynamics Division Funding for Education Efforts: NSF-DGE, NSF-DUE, Ohio Board of Regents
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