1. INTERCALATION COMPOUNDS OF LAYERED DOUBLE HYDROXIDES WITH ANTIMICROBIAL BENZOATE ANIONS AS FILLER OF BIODEGRADABILE POLYMERIC MATRICES FOR NEW PACKAGING SYSTEMS Loredana Tammaro Department of Chemical and Food Engineering, University of Salerno, Italy Sezione Tematica INSTM: 6. Materiali polimerici funzionali e strutturali VI Convegno Nazionale sulla Scienza e Tecnologia dei Materiali (INSTM 2007) Perugia, 12-15 Giugno 2007

2. Outline Structural and physical characterization Study of thermal properties Preparation and characterization of composites based on hydrotalcite as model systems for “active food packaging” Aim Loredana TammaroUniversity of Salerno Preparation of nano-hybrids: Intercalation of active molecules in layered double hydroxides (LDH) Preparation of nano-bio-hybrids composites: polymeric matrix: poly(  -caprolactone) (PCL) inorganic fillers: modified hydrotalcites

3. Advantages of “active packaging materials” Controlled release systems are becoming part of a wide category of new food packaging concepts known as ‘‘active packaging materials’’ which have the aim of extending the shelf life of the packaged foodstuff, inhibiting the microbial growth and preserving its sensory properties An eco-friendly solution is introducing the use of polymers either biodegradable or from renewable sources. However some properties of biodegradable polymers are not suitable for the applications in the food packaging field. Therefore many efforts are necessary to produce biodegradable materials with mechanical and barrier properties suitable for specific necessities. A good way to improve the performance of these systems is the incorporation of inorganic materials, with lamellar structure, into polymeric matrix at nanometric level Loredana TammaroUniversity of Salerno

4. The idea! The innovative idea of our research is based on the fact that it is possible to fix by ionic bonds, on the inorganic lamellae of anionic clay, active molecules (antimicrobial or antioxidants). These molecules not only can improve the compatibility with the polymer matrix, but can be released with controlled kinetics in particular environments, too Loredana TammaroUniversity of Salerno

5. Biopolymers Layered double hydroxides Active molecules films Nano-hybrid composite polymeric material Nano-hybrid composite membraneshydrogelsfibers Scaffolds tissue engineering Drug release Active packaging Chemical propertiesmorphological and structural propertiesphysical properties Mechanical properties (tensile,toughness, hardness, impact, fatigue properties.and creep properties)barrier and mechanical properties biodegradability biocompatibility release kinetics Suture threads Loredana TammaroUniversity of Salerno

6. Why PCL? PCL is a biodegradable polymer with good processing properties M. Vert, J. Feijen, A. C. Albertsson, G. Scott, E. Chiellini (eds.), Biodegradable Polymers and Plastics, Royal Society of Chemistry, London, 1992 "Layered Double Hydroxides: present and future" Vicente Rives Editor, Nova Science Publisher, INC. New York, 2001. They can be prepared with simple procedures, at high level of purity, are cheap and eco-compatible and can be organically modified with a variety of organic anions, generally much more numerous than organic cations. This latter characteristic will make these layered compounds compatible with a large variety of polymers and it is easy to foresee the possibility to prepare interesting new hybrid polymeric materials Why Layered Double Hydroxide? Loredana TammaroUniversity of Salerno

7. Chemical Structure of LDH Hydrotalcite: [M II 1-x M III x (OH) 2 ] x+ [A n-. nH 2 O] inter Structure of brucite Ions of metals M(II) and M(III) six times coordinate with ions OH - identifying octraedrals that form innumerable packed layers joined by ionic bonds and/or Van der Waals forces. The whole structure is constituted by the stacking of such layers, intercalating charge- balancing anionic species and water molecules Loredana TammaroUniversity of Salerno

8. ISOMORPHOUS REPLACEMENT Replacement of bivalent cations such as Mg 2+ e Fe 2+ present in the lattice with trivalent cations such as Al 3+ creates positive charges balanced by the presence of counter-anions such as CO 3 2-, Cl -, NO 3 - located into the interlamellar region MODIFIED HYDROTALCITES Although the hydrophilic nature of the clay minerals hinders their homogeneous dispersion into the polymer it has been possible to render the clays more compatible with the polymeric matrix Loredana TammaroUniversity of Salerno GENERALLY By replacing the charge balancing anions (ex. NO 3 - ) with organofilic groups, such as long chain of carboxylic acid salts. In this way the miscibility of the oxide layers in the polymer matrix can be enhanced LDHs a unique class of layered solids to be used as host of polymers, molecules bearing a negative charge and/or polymers copolymerized with a small amount of a negative charged monomer

9. Intercalation of active molecules in hydrotalcite- like compounds Structural formula of: benzoate (Bz) (a), 2,4-dichlorobenzoate (DCB) (b), o- hydroxybenzoate (o-OHBz) (c), p-hydroxybenzoate (p-OHBz) (d) anions (a) (b) (c)(d) Loredana TammaroUniversity of Salerno

10. A) Preparation of nano-hybrids Preparation of ZnAl-2,4dichlorobenzoate (ZnAl-DCB) and ZnAl-p hydroxybenzoate (ZnAl-p-OHBz) The intercalation of the benzoate derivatives were performed equilibrating 1 g of ZnAl-NO 3 in 15 cm 3 of solution water/acetone (1/1, v/v) 0.5 M of the anions obtained by titrating the corresponding acid forms with NaOH 1 M until the hydrolysis pH 7. The suspensions were stirred for 3 days. The obtained intercalation compounds were washed with CO 2 -free de-ionized water and dried at R. H.=75% Preparation of ZnAl-Benzoate (ZnAl-Bz) and ZnAl-salycilate (ZnAl-o-OHBz) The intercalation of benzoate and salicylate anions was achieved by equilibrating the nitrate form of hydrotalcite with an aqueous solution of the anion 0.5 mol/dm 3 (molar ratio organic anions/ NO 3 - = 3) for 24 hours at room temperature. The recovered solids were three times washed with CO 2 -free de-ionized water and dried at R.H.=75% Loredana TammaroUniversity of Salerno

11. DENOMINATION Acronyms of molecular ions intercalated into ZnAl-LDH, interlayer distance and composition of the intercalation compounds Aniond (Å)Composition Bz15,5[Zn 0.65 Al 0.35 (OH) 2 ]Bz 0.35 x1H 2 O o-OHBz15,5[Zn 0.65 Al 0.35 (OH) 2 ]o-OHBz 0.27 (NO 3 ) 0.08 x1H 2 O p-OHBz15,3[Zn 0.65 Al 0.35 (OH) 2 ]p-OHBz 0.2 (NO 3 ) 0.15 x0.66H 2 O DCB16,8[Zn 0.65 Al 0.35 (OH) 2 ]DCB 0.32 (NO 3 ) 0.03 x1H 2 O Loredana TammaroUniversity of Salerno

12. Structural analysis XRPD of intercalation compounds dried over saturated NaCl solution (R.H.=75%): (a) ZnAl- NO 3 ; (b) ZnAl-p-OHBz; (c) ZnAl-o-OHBz, (d) ZnAl-Bz and (e) ZnAl-DCB Loredana TammaroUniversity of Salerno

13. Structural Model Structural models of the intercalation compounds is proposed taking into account their chemical composition, interlayer distance, van der Waals dimension of the guests and considering that the intercalation process does not alter appreciably the structure of the layers. Computer generated models showing the most probable arrangement of (a) o-OHBz; (b) p-OHBz; (c) DCB and (d) Bz anions between the LDH layers (d) Loredana TammaroUniversity of Salerno

14. Thermogravimetric analysis Weight losses: 1) 80°-300°C: co-intercalated water and dehydroxylation of the inorganic layers 2) 300°C -600°C: decomposition of organic guests 3) > 600°C: formation of ZnO and ZnAl 2 O 4 TGA and DTA curves of the sample ZnAl-Bz. Operative conditions: air flow; heating rate: 5°C/min Loredana TammaroUniversity of Salerno

15. Infrared Analysis FT-IR spectra of ZnAl-Bz: (a) room temperature; (b) after heating at 300°C for 24 hours Spectrum (a) 1) 3000-3750 cm -1 : lamellae OH stretching involved in H- bonds with hydration water and carboxylic groups 2) 1537 cm -1 and 1397 cm -1 : asymmetric and symmetric stretching vibrations of the carbon-oxygen bonds of COO - group 3) 1595 cm -1 : in-plane skeletal vibration of monosubstituted aromatic ring 4) 719 cm -1 and 689 cm -1 : bending of the five adjacent hydrogen atoms of the ring Spectrum (b) The broad band centred at 3420 cm-1 disappear because of the removal of hydration water and the condensation of OH group of the lamellae, while the typical –COO- stretching are still present Loredana TammaroUniversity of Salerno

16. High Energy Ball Milling Use of mechanical energy to blend different materials Advantages: No solvents No high temperatures Possibility to prepare composites using natural polymers (i.e. pectins, starch...) that don’t melt, but degrade with temperature Possibility to incorporate into polymers, at low temperatures, inorganic solids with organic molecules thermally sensible Loredana TammaroUniversity of Salerno B) Nano-bio-hybrid incorporation in PCL Operating conditions: Cylindrical steel jar = 50 cm 3 Steel balls (10 mm diameter)= 5 Rotation speed = 580 r.p.m. Milling time = 1 hour Temperature= 25°C Different percentages (wt/wt) PCL-LDHs

17. Characterization of the composites: X-Ray ZnAl-Bz (a) and the composites PCL/ZnAl-Bz3 (b), PCL/ZnAl-Bz6 (c), PCL/ZnAl-Bz11 (d) ZnAl-DCB (a) and the composites PCL/ZnAl-DCB3 (b), PCL/ZnAl-DCB6 (c), PCL/ZnAl-DCB9 (d) ZnAl-o-OHBz (a) and the composites PCL/ZnAl-o-OHBz4 (b),PCL/ZnAl-o-OHBz6 (c), PCL/ZnAl-o-OHBz11 (d) ZnAl-p-OH (a) and the composites PCL/ZnAl-p-OHBz4 (b), PCL/ZnAl-p-OHBz7 (c), PCL/ZnAl-p-OHBz10 (d) Loredana TammaroUniversity of Salerno

18. Characterization of the composites: FT-IR 1540 cm- 1 : COO - antisymmetric stretching 1595 cm- 1 : C=C stretching vibration PCL (a), ZnAl-Bz (b) and the composites PCL/ZnAl-Bz3 (c), PCL/ZnAl-Bz6 (d), PCL/ZnAl-Bz11 (e) Relationship between the infrared absorbance intensity of the band at 1540 cm-1 and the inorganic content of the nanohybrid ZnAl-Bz in the relative composites Loredana TammaroUniversity of Salerno

19. Characterization of the composites: TGA PCL (a), PCL/ZnAl-Bz3 (b), PCL/ZnAl-Bz6 (c), PCL/ZnAl-Bz11 (d), ZnAl-Bz (e). Heating rate 5°C/min, in air flow PCL displays one main degradation step with a Td midpoint value of 402°C, followed by a small tail at about 450°C. Incorporation of the nano-hybrid ZnAl-Bz within PCL anticipates the midpoint of thermal degradation, due to the lower degradation point of the ZnAl-Bz. Degradation temperature slightly decreases on increasing the ZnAl-Bz content. Loredana TammaroUniversity of Salerno

20. Conclusion Layered double hydroxides intercalated with anionic active molecules based on benzoic acid and derivatives have been prepared and characterized The nano-bio-hybrids, having antimicrobic properties, dispersed into PCL have been prepared and physically characterized They can constitute model systems for “active food packaging” applications Loredana TammaroUniversity of Salerno

21. Work in progress…. Analysis of mechanical properties Analysis of mechanical properties Analysis of controlled release through UV spectroscopy of the active molecules Analysis of controlled release through UV spectroscopy of the active molecules References Petersen, K., Nielsen, P. V., Bertelsen, G., Lawther, M., Olsen, M. B., Nilssonk N. H. and Mortenseny G. Trends in Food Science & Technology;1999, 10: 52-68. M. Chasin, R. Langer (Eds), Biodegradable polymers as drug delivery systems. Marcel Decker, New York, 1990. Loredana TammaroUniversity of Salerno

22. Thanks to: Dr. Francesca Montanari, Dr. Morena Nocchetti, Prof. Umberto Costantino Department of Chemistry, University of Perugia, Italy This work was supported by PRISMA 2005 project entitled “Nanobiohybrids in polymeric matrices as controlled delivery of active molecules” My scientific group: Dr. Valeria Bugatti, Dr. Giuliana Gorrasi, Prof. Vittoria Vittoria Department of Chemical and Food Engineering, University of Salerno, Italy Loredana TammaroUniversity of Salerno

23. Synthesis of the carbonate form: MgAlCO 3 The method used for the preparation of the carbonate form was the hydrolysis of urea in the presence of mixture of M(II) (Mg 2+ ) and M(III) (Al 3+ ) “Costantino et al. (1998a)”. Solid urea was added to 0.5 mol/dm 3 metal chloride solutions (MgCl 2 and AlCl 3 ), having molar fraction Al(III)/Al(III)+Mg(II) equal to 0.33, until the molar ratio of urea/Mg(II)+Al(III) reached the value 3.3.The clear solution was heated, under stirring, at temperature between 60 and 100°C for 72 hours. The solid, separated from the solution, was washed with distilled water, then dried at 80°C and stored in a desiccator with P 4 O 10 at room temperature. Synthesis of the chloride form: MgAlCl This form has been obtained by titrating at room temperature the carbonate form, dispersed in 0.1 mol/dm 3 NaCl aqueous solution with a 0.1 mol/dm 3 HCl solution by means of aRadiometer automatic titrator operating at pHstat mode and pH=5. After titration the solid waswashed with CO 2 free deionized water and finally dried over phosphorus pentoxide.