1. 1 Aqueous Cholesteric Liquid Crystals Using Uncharged Rodlike Polypeptides Enrico G. Bellomo, Patrick Davidson, Marianne Impéror-Clerc, and Timothy J. Deming J. Am. Chem. Soc., 126 (29), 9101 -9105, 2004 Presented by: Erick Soto April 28, 2006

2. 2 Overview: Introduction Background Experimental and Results Discussion Conclusions Epilogue

3. 3 Solid Liquid Gas Liquid Crystal Introduction States of Matter Adapted from: http://www.chem.purdue.edu/gchelp/atoms/states.html

4. 4 Introduction States of Matter Solid Liquid Gas Energy per particle increases Order increases Liquid Crystal From: http://www.chem.purdue.edu/gchelp/atoms/states.html

5. 5 Applications of liquid crystals From: http://www.beyondconnectedhome.com/aboutus/press/downloads.html

6. 6 From: http://www.petsolutions.com/Images/200/15522455.jpg http://www.goratec.com/applications.php?ApplicationID=12&language=en Liquid Crystal Thermometers

7. 7 From the physicist’s perspective: 50-1000 Å 5 Å

8. 8 Characteristics of molecules capable of forming liquid crystals: Rod-like molecular structure Rigidness of the long axis Strong dipoles and/or easily polarizable susbtituents Examples: adapted from: http://liq-xtal.case.edu/lcdemo.htm#Diagrams p-decyloxybenzylidene p'-amino 2-methylbutylcinnamate ("DOBAMBC") Methoxybenzylidene Butylanaline (“MBBA”)

9. 9 1888 Friedrich Reinitzer discovered LC’s The crystals transformed at 145.5 º C into a cloudy fluid which suddenly clarified only on heating at 178.5 ºC. He observed colors. Cholesteryl benzoate (604-32-0)

10. 10 The distinguishing characteristic of the liquid crystalline state is the tendency of the molecules (mesogens) to point along a common axis, called the director. To quantify how much order is present in a material, an order parameter (S) is defined From: http://plc.cwru.edu/tutorial/enhanced/files/lc/intro.htm  = 0 º S=1  = 54.7 º S=0

11. 11 Liquid Crystal Phases: The nematic liquid crystal phase is characterized by molecules that have no positional order but tend to point in the same direction (along the director). From: http://www.nat.vu.nl/~fcm/ComplexFluids/ComplexFluids.html http://www.nat.vu.nl/~fcm/ComplexFluids/4a.gif

12. 12 Smectic A phase Smectic C phase From: http://portellen.phycmt.dur.ac.uk/sjc/thesis_dlc/node15.html http://www.warwick.ac.uk/fac/sci/Chemistry/jpr/lq/liqcry.html Smectic phases:

13. 13 Birefringence: Liquid crystals are birefringent. This is due to the fact that they are anisotropic (directionally dependent) isotropicanisotropic polarizer analyzer

14. 14 Polypeptide liquid crystals are valuable as chiral NMR solvents for enantiomeric resolution.  New, and Accurate Method To Determine the Enantiomeric Purity of Amino Acids Based on Deuterium NMR in a Cholesteric Lyotropic Liquid Crystal J. Am. Chem. Soc.; 1994; 116(5); 2155-2156  Two-Dimensional Deuterium NMR Spectroscopy of Chiral Molecules Oriented in a Polypeptide Liquid Crystal: Applications for the Enantiomeric Analysis through Natural Abundance Deuterium NMR J. Am. Chem. Soc.; 1999; 121(22); 5249-5258

15. 15 Probably the best studied example of polypeptide liquid crystal is PBLG Repeating unit of poly(  -benzyl-L-glutamate) (PBLG)

16. 16  -helical structure of PBLG H bonding holds it From: http://pszichologusboy.freeblog.hu/Files/Alfa%20hélix.jpg

17. 17 PBLG exhibits liquid crystal behavior in several organic solvents. Unfortunately, there is no aqueous phase polypeptide analogue of PBLG. PBLG liquid crystal in pyridine

18. 18 There are some water soluble polypeptides. For example poly(glutamic acid) and polylysine. Problem: Charge repulsion Poly (glutamic acid) sodium salt Polylysine·HBr

19. 19 There was considerable interest in development of non-ionic, water-soluble, conformationally regular polypeptides derived from chemically modified aminoacids. The best materials resulting from this work are poly(N-hydroxyalkyl- L-glutamines). (a) Lupu-Lotan, N.; Yaron, A.; Berger, A.; Sela, M. Biopolymers 1965, 3, 625-655. (b) Lupu-Lotan, N.; Yaron, A.; Berger, A. Biopolymers 1966, 4, 365-368. (c) Okita, K.; Teramoto, A.; Fujita, H. Biopolymers 1970, 9, 717-738

20. 20 The best example, poly(N-hydroxybutyl-L-glutamine), PHBG, is ca. 65% helical in neutral water at 20 º C Nearly all the side chains could be functionalized. The resulting polymers were found to be water soluble.  The aminolysis reaction resulted in significant backbone cleavage  Studies on their helical structure were limited because they contained substantial random coil content when dissolved in water. poly(N-hydroxyalkyl-L-glutamines)

21. 21 Biocompatibility: An important feature of  -helical water soluble polypeptides would be biocompatibility. This is for biomedical applications. Poly(N-hydroxyalkyl-L-glutamines) are recognized are foreign and rapidly degraded in vivo. Most biocompatability strategies employ polyethylene glycol (PEG), which is typically grafted onto other polymers.

22. 22 PEG  Water soluble  Non ionic  Not recognized by immune systems They wanted to incorporate these attractive properties of PEG into polypeptides. 

23. 23 Strategy: (1) (2) (3)

24. 24 Polymer synthesis pre-monomer PEG (short) PEG (short) * pre-monomer– PEG (short)

25. 25 F1 polymer pre-monomer– PEG (short) monomer– PEG (short)

26. 26 Structure and schematic representation of polymer L-1

27. 27 The magenta vector can be decomposed in two green vectors of equal intensity. The change in intensity of the magenta electric field vector causes these two vectors to rotate oppositely. From: http://www.imb-jena.de/ImgLibDoc/cd/tut1a.html Circular Dichroism

28. 28 [  ] at 222 nm as a function of temperature for 1 (M n = 93100) and PHBG ( M n = 105000) in H 2 O ([polymers] = 0.5 mg/mL, pH 7). Circular Dichroism

29. 29 LC I + LC I  polymer Adapted from: Miller et al., 1974.

30. 30 From: http://www.microscopyu.com/articles/polarized/polarizedintro.html Birefringence:

31. 31 Test tubes filled with increasing weight fractions of L-1 in deionized water. Samples were imaged between crossed polarizers. (A) (Mw = 62 kDa): i = 32.7%; ii = 39.3%; iii = 41.9%; iv = 43.5%; v = 45.4%; vi = 47.9% Polymer solutions observed between cross polarizers

32. 32 B) (Mw = 120 kDa): i = 15.4%; ii = 18.7%; iii = 23.8%; iv = 25.2%; v = 38.5%; vi = 39.8%. All sample compositions are in weight percent Polymer solutions observed between cross polarizers

33. 33 Between the concentrated and the dilute regime, a biphasic domain is found where samples display macroscopic phase separation. These observations show the existence of a first-order phase transition between the isotropic and liquid crystalline phases. 32.7%39.3%41.9%43.5%45.4%47.9% 15.4%18.7%23.8%25.2%38.5%39.8% M w = 62 kDa M w = 120 kDa

34. 34 From: http://www.microscopyu.com/articles/polarized/polarizedintro.html Optical Microscopy

35. 35  The textures of the liquid-crystalline samples held in flat glass capillaries were clear enough to allow mesophase identification.  Biphasic samples displayed banded birefringent liquid spherulites floating in a dark isotropic liquid.  Fully liquid-crystalline samples displayed large dark homeotropic regions separated by bright regions containing fingerprint patterns.  All of these features are typical of the cholesteric (chiral nematic) phase.

36. 36

37. 37 The fingerprint patterns are caused by the cholesteric pitch. half pitch distance

38. 38 From: “Ordered phases of filamentous viruses”, (to be published in Current opinion in Colloid and Interface Science) Cholesteric pitch

39. 39  From a practical standpoint the two major challenges in cholesterics are adjusting and controlling the pitch.  The pitch depends on solvent and temperature but it also varies with the optical purity of the sample.  If a cholesteric liquid-crystalline compound is mixed with its opposite enantiomer (i.e. L and D) the pitch should increase and approach infinity as the mixture become racemic, and then the liquid crystalline phase will become nematic.

40. 40 Cholesteric pitch tuning:

41. 41 Polarized optical micrographs showing the dependence of the cholesteric pitch on the enantiomeric composition of 1. All samples were prepared at 60 wt % in deionized water and are defined as the mol % of L-1 in a L-1 + D-1 mixture. (A) = 0%; (B) = 15%; (C) = 30%; (D) = 40%; (E) = 60%; (F) = 70%; (G) = 85%; (H) = 100%.

42. 42 Cholesteric pitch (  m), measured from optical micrographs, as a function of the enantiomeric composition of 1 (mol % of L-1 in a L-1 + D-1 mixture). The solid line is a fit of the data using a hyperbolic divergence at 50 mol %.

43. 43 Sample Alignment: In order to gain more information on the structure and stability of these cholesteric phases, they sought to untwist the pitch by aligning the rodlike polypeptide chains. It is known that magnetic fields can align  -helical polypeptides parallel to the direction of the applied magnetic field. In preliminary studies they found that a moderate magnetic field (1.7 T) was not sufficient to align optically pure polypeptide samples, or even the weakest cholesteric samples (i.e. near the equimolar L + D). A stronger magnetic field (magnet of an NMR spectrometer ~8T) was able to align the polypeptide molecules into a nematic phase.

44. 44 100 mol % L polymer 55 mol % L polymer, aligned by magnetic field CCD camera and rotating anode X-ray source were used to record the scattering patterns.  

45. 45 Be the judge PBLG liquid crystal Conmar Robinson 1957 Polylysine-PEG Tim Deming 2004

46. 46 Magnetic field aligned (8 T) F1 55 mol % of L-polypeptide (60 wt %) I versus  I versus  Maximum at 2.6 nm -1 which corresponds to the average distance between the rods of 2.4 nm Order parameter, S = 0.85 ± 0.05. It’s large but it’s typical of lyotropic nematic phases.

47. 47 Shear alignment: The inability to record structural information in situ during magnetic field alignment, a Couette cell setup was used. Application of a shear flow is a very powerful way of aligning viscous mesophases and has been successfully applied to liquid crystal phases of PBLG in m-cresol.

48. 48 100 mol % L polymer, high shear55 mol % L polymer, low shear I versus  I versus  Radial X-ray diffraction Tangential X-ray diffraction showed no anisotropy.

49. 49 55 mol % of L-polypeptide (60 wt %) Maximum at 2.6 nm -1 which is the same as obtained by magnetic field alignment. Order parameter, S = 0.88 ± 0.05. Which is comparable to the obtained by magnetic alignment.

50. 50 Discussion:  System behaved as predicted by the Onsager model. (Strong first order isotropic/nematic phase transition, with phase coexistence)  The Onsager model predicts the volume fractions  n = 4.2 D/L and  i = 3.3 D/L for the isotropic and nematic phase transition.  For a short L- polymer (M w = 62 kDa; L= 32.3 nm and D= 2.2 nm) the values should be  n = 29% and  i = 23%. These results are in fair agreement with the experimental values of 47% and 38% respectively. (  n /  i ~same) 32.7%39.3%41.9%43.5%45.4%47.9% M w = 62 kDa

51. 51  For a longer L- polymer (Mw= 120 kDa; L= 62.6 nm and D= 2.2 nm) the values should be  n = 15% and  i = 12%. These results are in fair agreement with the experimental values of 39% and 17% respectively. This deviation could be due to chain flexibility and polydispersity.  The electrostatic interactions are negligible in this system because similar properties were observed when samples were prepared in 100 mM NaCl. 15.4%18.7%23.8%25.2%38.5%39.8% M w = 120 kDa

52. 52 Conclusions:  A new aqueous cholesteric liquid crystal has been reported which is formed by uncharged rodlike polypeptide molecules.  The mentioned system undergoes isotropic/nematic phase transition.  The cholesteric pitch was found to be easily tuned.  Important structural information was obtained by aligning the rodlike polypeptide molecules by shear or magnetic field. (S and d)  The system was found to behave as predicted by the Onsager model.

53. 53 Epilogue: This publication has been cited 3 times to date. Research ideas: Fundamental To investigate the persistence length vs. molecular weight behavior. To investigate D vs. concentration. To observe their interaction with spheres. New phases may arise.

54. 54 Applied To use a more rigid polypeptide backbone. (glutamate) Potential candidates for drug delivery

55. 55 Thanks for your attention

56. 56 Macromolecules, 32, 4500 back

57. 57 back

58. 58 Macromolecules, 32, 4500

59. 59 Radial (I versus q) X-ray scattering pattern of a 55 mol % L-1 sample (>70 wt % in deionized water) showing hexagonal symmetry of mesophase of 1 with diffraction lines in the ratio 1, 31/2, 2, 71/2 (labeled as the 10, 11, 20, and 21 reflections, respectively). DH = diffuse halo of scattering from ethylene glycol side-chains