Evaluation of Materials Using Single Molecules as Nanoscale Probes

1. Evaluation of Materials Using Single Molecules as Nanoscale Probes Miyasaka lab. Yamada Keisuke

2. Contents I. IntroductionThe significance of measuring single molecules in evaluating microscopic properties of materials II. Single molecule imaging (SMI) methodsWide-field microscopy and its related techniques III. Evaluation of mesoporous materials with SMIPrevious works: mesoporous silica My current research topic: mesoporous organosilica IV. Summary

3. Microscopic heterogeneity in condensed matter Ex. Solids of polymer Polymer chains Packed area Loose area Inhomogeneity in nanometer scale Free volume distribution, permittivity, density, viscosity, elasticity, etc.

4. The result of ensemble measurement Emission spectrum of a single molecule Each guest molecule is in different environment. Different electronic state → Position-dependent spectrum and lifetime Heterogeneity in structure, viscosity, etc.→ Position-dependent diffusion coefficient of guest By measuring each guest molecule, nanoscale property changes in materials can be evaluated.

5. The motions of individual guest molecules strongly depend on microenvironment. Tracking the motion of guests can provide information on microscopic properties of materials: ・ Nanostructures ・ Mobility of guests (Free volume in polymer, Host-Guest Interaction, etc. ) ・ Spatial heterogeneity ....

6. Laser Wide-field Microscope objective lens DM Band-pass filter PC Back illumination CCD Camera

7. Defocus Imaging Method J. Opt. Soc. Am. B, 2003, 20, 554-559 The direction of the emitting dipole 8 mm Defocus by~ 1 mm

8. Single Molecule Tracking (SMT) Biophysical J. (2001) 81, 2378. Poly(2-hydroxyethylacrylate) X0 = 274.03 +/- 0.0339 pix. Y0 = 148.17 +/- 0.0351 pix. 330 nm Exp. Time: 500 ms 10 frame/s, 32.2 nm/pixel Actual precision of tracking ~ 10 nm in our experimental set up

9. 2 nm 50 nm Microporous Mesoporous Macroporous Application of SMT to the evaluation of mesoporous silica What is mesoporous silica ? Applications ・ Adsorption system ・ Molecular sensor ・ Drag delivery C.T. Kresge,et al. Nature359 (1992), p. 710. hexagonal cubic Lamellar ・ Periodic nanometer-scale pores ・ Large surface area ・ Easy-controlled structure and pore size

10. How to synthesize the mesoporous material Reagent ; water, ethanol,surfactant, silane monomer, catalyst C.T. Kresge,et al. Nature359 (1992), p. 710.

11. Evaluation TEM (Transmission Electron Microscope) XRD (X-Ray Diffraction) Kresge, C. T et al. ,Nature 1992, 359, 710~712. ・ Space resolution in nano meter ・ Measure diameters of pore and observe the structure directly Measure the distance between pores SMT (Single Molecule Tracking) ・ High time and space resolution ・ Evaluation mobility of guest molecules in pores 11

12. Previous Work CCD Mesoporous Silica (silica framework) ・record emission of fluorescent guest molecule in mesoporous ・ diffusion behavior show the surrounding structure Kirstein, J. et al. Nat. Mater.2007, 6 (4), 303-310. Trajectory of guest molecule （show mesoporous strucuture）

13. Mesoporous Organosilica Organic groups: ethane, benzene, biphenyl, naphthalene, ... Inagaki, S. et al. Nature2002, 416, 304-307. Fujita, S.; Inagaki, S.Chem. Mater.2008, 20, 891-908. (Toyota Central R&D Laboratories)

14. Organic Silane monomer: BTEB Surfactant: Brij76 Fluorescent dye BP-PBI / Brij76,Exp. time: 250 ms BP-PBI TDI (Playing at 30 fps)

15. BP-PBI / Brij76 4.2 × 10-3 μm2s-1 (1D diffusion) TDI / Brij76 2.0 × 10-4 μm2s-1 (1D diffusion) Diffusion coefficient is strongly dependent on guests.

16. My Work Dual color SMT System 325,442,488,532 nm 633,660,690 nm DM CCD2 CCD1 Optical filter Two kinds of guest molecules are incorporated into the same mesoporous structure. Host-guest interaction can be precisely evaluated. BP-PBI TDI 16

17. Summary • I have shown the significance of measuring single molecules in evaluating microscopic properties of materials. • Wide-field microscopy and its related techniques permitted the tracking of translational and rotational motions of individual molecules. • Tracking single guest molecules in mesoporous silica visualized its mesostructures, and detail analysis of the trajectories evaluated the host-guest interaction. 17

18. Diffusion behavior dependence on guest molecule

19. Fluorescence Imaging

20. Analytical method Analysis Fluorescence Imaging per 1 frame and determination position coordinate of molecule Analysis formula 2-Dimension Gaussian Function Tracking translational diffusion of each molecule

21. Tracking translational motion

22. One-dimensional and Two-dimensional 1D diffusion 2D diffusion translation diffusion in mesoporous surface diffusion Diffusion in structural defect region

23. I0 I Principle Absorption I0 I0 I I Fluorescence Sensitive detector Single fluorescence molecules in dark space can be detected optically.

24. Characteristics 同じ資料に異なるゲスト分子をいれて、違いを評価することで材料のロットによるばらつきを抑えて議論できる。 そのための装置開発（２色SMT装置）もテーマであることを述べる The result of ensemble measurement • Each molecule’s fluctuations • High time resolution • Evaluation mobility of guest molecules in pores • The difference of microscopic environment Ensemble Single molecule Emission spectrum of a single molecule

25. Till the 90s… In the 90s … History There are some problems of mesoporous material ・ irregularly space ・ broadly distribution in size ・ too small pore size to apply 1990 T.Yanagisawa et al. (Waseda Univ.) 1992 C.T. Kresge et al. (Mobil) ・ Control pore size of mesoporous silica ・ Build up the synthesis based on sol-gel process

26. Characteristics hexagonal cubic Lamellar C.T. Kresge,et al. Nature359 (1992), p. 710. ・ Periodical meso structure of nano meter scale ・ Large surface area ・ Control the structure and pore size