1. SERS characterization of self-assembling oligopeptides for tissue engineering
M. Di Foggia1, A. Torreggiani2, P. Taddei1, M. Dettin3, S. Sanchez-Cortes4, A. Tinti1 1 Dip. Scienze Biomediche e Neuromotorie, Università di Bologna, Italy. 2 ISOF-CNR, Bologna, Italy. 3 Dip. di Processi Chimici dell’Ingegneria, Università di Padova, Italy. 4 Dep. de Espectroscopia Vibracional y Procesos Multifotonicos, Madrid, Spain.

2. 1st Faculty of Chemistry
Bologna docet
1929
1st University
1st Faculty of Chemistry
1st Raman Spectrometer
1st SERS?

3. BIOMEDICAL MATERIALS SURFACE PROPERTIES
Growth and support of damaged tissues
Secret of success…
SURFACE PROPERTIES
improving the morphology of the material itself (porosity, synthesis, roughness…)
inserting something that cells love!

4. SELF-ASSEMBLING PEPTIDES
Biomimetic coatings for orthopaedics and dentistry
Biocompatible and biodegradable
Form membranes and stable fibrillar aggregates
Many biomedical and pharmaceutical applications:
drug-release systems
matrices supporting cellular growth
(neurite growth and synapsis formation)
LEGO® peptides

5. leukocytes excrete high levels of ·OH: O2·- + H2O2 → ·OH + OH- + O2
The peptides “hide” the prosthetic device to the immunitary system, but the surgical procedure triggers the beginning of an inflammatory process:
leukocytes excrete high levels of ·OH:
O2·- + H2O2 → ·OH + OH- + O2
g-radiation of aqueous solutions of peptides in protected atmosphere (NO2), produces ·OH at physiological concentrations:
H2O + hν → ·OH + H·
Can radical attack alter peptide structure, thus hampering biomimetic coating functionality? 5

6. PEPTIDES
EAK-161 + -
b-sheet structure (best interaction surface-peptide-cells):
electrostatic interactions between charged side chains (COO-/NH3+)
hydrophobic interactions between aliphatic side chains
p-p interaction between aromatic side chains
1 Zhang S et al, PNAS, 1993

7. PEPTIDES

8. STRUCTURE SENSITIVE BANDS
Amides bands depends on H-bonds strenght: highly sensitive structural bands.
Amide II
Amide III
Amide I
Raman

9. METHODS
Structural changes caused by radical attack were evaluated by FT-Raman spectroscopy and SERS technique on Ag colloids (≈ physiological conditions).
SERS can be used to establish the relative importance of specific functional groups in controlling peptide adsorption (very debated problem!).
Hydroxylamine hydrochloride
Cl-

10. Radiation treatment: peptides 1, 3, 5, 7, 8
non-irradiated peptide
difference spectrum

11. Peptides 2, 4: Asp  Glu non-irradiated peptide difference spectrum
Structural variations on backbone chain
Structural variations on aliphatic side-chains
b-sheet
decrease
a-elix/random
increase
non-irradiated peptide
difference spectrum

12. Structural variation in Tyr
Peptide 6: Tyr
Structural variation in Tyr
side chains
non-irradiated peptide
difference spectrum

13. CONCLUSIONS (part 1)
Peptides 2 and 4 showed severe structural variations: Asp decarboxylation
Peptides 1, 3, 5, 7 and 8 were resistant to radical attack (1, 5 and 7 were found to stimulate osteoblasts proliferation in cell culture studies) 13

14. SERS spectra: peptides 1, 2, 7
Ag-COO- interaction
ns COO-
Am. III
nas COO-
b-sheet
n C-COO-
_ SERS
_ FT-Raman
Am. IV
r CO

15. SERS spectra: peptides 3, 4 Mainly Orn (NH3+…Cl-) interaction
r CH2
def as NH3+
t NH3+
Am. IV
n COO-
def NH3+
n C-COO-
r NH3+
_ SERS
_ FT-Raman Ag
Cl-
Cl-

16. Both COO- and NH3+ interaction
SERS spectra: peptide 5
Both COO- and NH3+ interaction
n COO-
def as NH3+
n C-COO-
_ SERS
_ FT-Raman
b-sheet
Am. III

17. Both COO- and amide bond interaction
SERS spectra: peptide 8
Both COO- and amide bond interaction
Am. I
n COO-
n C-COO-
Am. III
_ SERS
_ FT-Raman
Am. IV

18. Both Tyr- and COO- interaction
SERS spectra: peptide 6
Both Tyr- and COO- interaction
1605
Tyr-
n COO-
Tyr-
Tyr-
_ SERS
_ FT-Raman
Tyr-
1616
n C-COO-

19. Small changes in the SERS spectra
SERS spectra: peptide 1, g-irradiation
Small changes in the SERS spectra
SERS
Difference spectrum
ns COO-
Am. IV
r CO
n C-COO-

20. Major changes in the SERS spectra
SERS spectra: peptide 4, g-irradiation
Major changes in the SERS spectra
ns COO-
SERS
Am. I
Am. III
n C-COO-
def NH3+
Am. IV
Difference spectrum
def as NH3+
r CH2
t NH3+

21. CONCLUSIONS (part 2)
SERS spectra evidence different peptide – colloid interactions as a function of a.a. substitution.
SERS spectra highlights major changes on the peptide – colloid interaction for g-degraded peptides (i.e. pept 2, 4 and 6).
Bibliography:
[2] A. Tinti, M. Di Foggia, P. Taddei, A. Torreggiani, M. Dettin, C. Fagnano, J. Raman Spetrosc. 39 (2008) 250.
[3] M. Di Foggia, P. Taddei, C. Fagnano, A. Torreggiani, M. Dettin, S. Sanchez-Cortes, A. Tinti, J. Mol. Struct (2009), 120.
[4] M. Di Foggia, P. Taddei, A. Torreggiani, M. Dettin, A. Tinti, J. Raman Spetrosc. 42 (2011) 276.
[5] M. Di Foggia, P. Taddei, A. Torreggiani, M. Dettin, A. Tinti, J. Raman Spetrosc. (2013) DOI /jrs.4271., 21

22. Thank you for your kind attention!
Ab-initio calculations:
attributions
interaction type
Thank you for your kind attention! 22