Philippe Evon
email: philippe.evon@ensiacetfr

Virginie Vandenbossche

Pierre-Yves Pontalier

Luc Rigal

Pierre-Yves Pontaliera

New thermal insolation fiberboards from cake generated during biorefinery of sunflower who le plant in a twin-screw extruder

Keywords:

teaching and research institutions in France or abroad, or from public or private research centers.

New thermal insulation fiberboards from cake generated during biorefinery of sunflower whole plant in a twin-screw extruder

Introduction ABSTRACT

The objective of this study was to manufacture new thermal insulation fiberboards by thermo-pressing. The starting material was a slightly deoiled cake (17.6% oil content), generated during the biorefinery of sunflower (Helianthus annuus L) whole plant in a co-rotating (Clextral BC 45, France) twin-screw extruder. Ali fiberboards produced were cohesive mixtures of proteins and lignocellulosic fibers, acting respectively as binder and reinforcing fillers in what could be considered as a natural composite. The molding experiments were conducted using a 400 ton capacity heated hydraulic press (Pinette Emidecau Industries, France). The influence of mol ding conditions on board density, mechanical properties and heat insulation properties was examined. Mol ding conditions included mold temperature ( 140-200 oc), pressure a pp lied ( 150-250 kgffcm 2 ) and molding time ( 40-76 s ), and these great! y affected board density and thus the mechanical and heat insulation properties. Board density increased with increasingly extreme molding conditions, rising from 500 to 858 kgfm 3 • The mechanical properties increased at the same time (from 52 to 660 kPa for flexural strength at break, from 5.9 to 49.4 MPa for elastic modulus, from 0.5 to 7.7k]/m 2 for Charpy impact strength, and from 19.2 to 47.1" for Shore D surface hardness). Conversely, heat insulation properties improved with decreasing board density, and the lowest thermal conductivity (88.5 mW fm Kat 25 oc) was obtained with the !east dense fiberboard. The latter was produced with a 140 oc mold temperature, a 150 kgffcm 2 pressure a pp lied and a 40 s mol ding time. A medium mold temperature (160°C) was needed to obtain a good compromise between mechanical properties (272 kPa for flexural strength at break, 26.3 MPa for elastic modulus, 3.2 k]/m 2 for Charpy impact strength, and 37.3° for Shore D surface hardness), and heat insulation properties (99.5 mWfmK for thermal conductivity). The corresponding board density was medium (687 kgfm 3 ). Because of their promising heat insulation properties, these new fiberboards could be positioned on walls and ceilings for thermal insulation of buildings. The bulk cake also revealed very low thermal conductivity properties (only 65.6mWfmK at 25 oq due to its very low bulk density (204 kgfm 3 ). It could be used as loose till in the attics of houses. carcinogenicity [START_REF] Galvin | Toxicity data for commercial hexane and hexane isomers[END_REF]. Consequently, numerous solvents have been considered, including water [START_REF] Rosenthal | Aqueous and enzymatic processes for edible oil extraction[END_REF]. Sunflower (Helianthus annuus L) is cultivated for the high oil content of its seeds. Oil represents up to 80% of its economie value. The industrial process for oil production consists of four successive stages: trituration, pressing, extraction of the residual oil using hexane, and refining [START_REF] Isobe | A new twin-screw press design for oil extraction of dehulled sunflower seed[END_REF][START_REF] Rosenthal | Aqueous and enzymatic processes for edible oil extraction[END_REF]. Extraction yields are close to 100% with very good oil quality. However, the use of hexane for oil production is an increasingly controversial issue and could be prohibited due to its Severa! re se archers have studied the aqueous extraction of sunflower oil [START_REF] Evon | Direct extraction of oil from sunflower seeds by twin-screw extruder according to an aqueous extraction process: feasibility study and influence of operating conditions[END_REF][START_REF] Evon | Aqueous extraction of residual oil from sunflower press cake using a twin-screw extruder: feasibility study[END_REF][START_REF] Hagenmaier | Aqueous processing of full-fat sunflower seeds: yields of oil and protein[END_REF][START_REF] Southwell | Extraction of oil from oilseeds using the hot water flotation method[END_REF] that is an environmentally friendly alternative to solvent extraction. 1t can be conducted using whole seeds [START_REF] Evon | Direct extraction of oil from sunflower seeds by twin-screw extruder according to an aqueous extraction process: feasibility study and influence of operating conditions[END_REF] or from a press cake [START_REF] Evon | Aqueous extraction of residual oil from sunflower press cake using a twin-screw extruder: feasibility study[END_REF] in a Clextral BC 45 (France) co-penetra ting and co-rota ting twin-screw extruder that enables efficient mechanicallysis of the cens. Three essential unit operations are carried out in a single step and in continuous mode: conditioning and grinding of the initial material, liquidfsolid extraction, and liquidjsolid separation. A fil ter section is positioned on the barrel to collect an extract (filtrate) and a raffinate (cake), separately. However, the introduction of a lignocellulosic residue upstream from the filtration module is essential to enable liquid/solid separation.

When applied to the whole plant, aqueous fractionation in a twin-screw extruder does not require the addition of a lignocellulosic residue (Evon et al., 2010a) due to the natural abundance of fibers in sunflower stalk [START_REF] Maréchal | Characterization of by-products of sunflower culture: commercial applications for stalks and heads[END_REF], and twinscrew extrusion technology thus appears to be an original and powerful solution for the biorefinery of sunflower whole plant. Under optimal operating conditions, oil extraction yield reaches 57%, and residual oil content in cake is 14.3% of the dry matter. These conditions lead to the co-extraction of proteins but also pectins and hemicelluloses. The corresponding protein extraction yield is 44%, and residual protein content in cake is 7.3% of the dry matter.

Cake moisture content is relatively high (at !east 62%), and soit is first dried to facilitate conservation. ft has a porous structure, and is main! y composed of lignocellulosic fi bers ( around 58% of the dry matter), but also cell debris from the kernel breakdown process. Actually, it is lixiviated matter where soluble molecules (proteins, pectins ... ) and lipids are partly removed, although plant structural molecules are not extracted. The cake is thus suitable for use in animal feeds and for energy production in pellet burning furnaces. Nevertheless, newvalorizations of the cake as a mixture ofproteins and lignocellulosic fibers can be also considered [START_REF] Orliac | Effects of additives on the mechanical properties, hydrophobicity and water uptake of thermo-moulded films produced from sunflower protein isolate[END_REF][START_REF] Orliac | Effects of various plasticizers on the mechanical properties, water resistance and aging ofthermo-moulded films made from sunflower proteins[END_REF][START_REF] Rouilly | DSC study on the thermal properties of sunflower proteins according to their water content[END_REF][START_REF] Rouilly | Thermal denaturation of sunflower globulins in low moisture conditions[END_REF]Rouilly et al., , 2006a,b),b).

The cake's thermo-mechanical behavior has been previously studied (Evon et al.,201 Ob). DSC (differentiai scanning calorimetry) measurements indicate that denaturation of cake proteins is al most complete, and the DMTA (dynamic mechanical thermal analysis) spectrum obtained from dried and ground sam pies reveals a significant peak at high temperature (around 180°C) which, as already observed with industrial sunflower cake [START_REF] Geneau | Manufacturing Process of Agromaterial Natural Composite by Twin-screw Extrusion and Injection Moulding from Sunflower Cake[END_REF], is associated with the glass transition of proteins. Because the cake is a mixture of proteins and lignocellulosic fibers, it can be considered as a natural composite, and can thus be successfully processed into cohesive, biodegradable and value-added fiberboards by thermopressing, proteins and lignocellulosic fibers acting respectively as internai binder and reinforcing fillers (Evon et al., 2010b(Evon et al., , 2012a)).

The mechanical properties for bending of fiberboards, increase with temperature, pressure and length oftime ofthermo-pressing (Evon et al., 2010b(Evon et al., , 2012a)). The highest flexural strength at break (11.5 MPa) and the highest elastic modulus (2.2 GPa) are obtained from a cake with a dry matter residual oil content of 14.5%, and under the following molding conditions: 500mgfcm 2 for cake quantity, 200 oc for the temperature of the two aluminium plates of the heated hydraulic press, 320 kgffcm 2 for pressure applied, and 60s for molding time (Evon et al., 2010b). The fiberboard thickness is only 3.9mm, its density is quite high (1035kgfm 3 ) and DMTA analysis reveals a significant vibratory oscillation peak at low temperature (between -20 and -14 °C), which is attributed to the

セ M エ イ 。 ョ ウ ゥ エ ゥ ッ ョ @
of proteins (glass transition of their side chains) (Rouilly et al., 2006b;[START_REF] Zhang | Mechanical and thermal properties of extruded soy protein sheets[END_REF]. No significant transition is observed between 0 oc and 200 oc, meaning that proteins ensure the agromaterial's cohesion without any phase change in this temperature range. Finally, lignocellulosic fibers' entanglement also act as reinforcement. Because of its promising flexural properties, su ch a fiberboard would be potentially usable as inter-layer sheet for pallets in the handling and storage industry or for the manufacture of biodegradable, multi-board containers, e.g. composters, crates for vegetable gardening (Evon et al., 2010b).

Another industrial application offiberboards made from renewable resources is he at insulation of buildings ( walls and ceilings ), where the main advantages ofvegetable fibers are abundance, low cost (the majority are agricultural residues), minimal environmental impact, independence from fossil resources, and their natural capacity for thermal insulation (Saiah et al., 2010). Insulation boards can be made from maize husks and cobs [START_REF] Paiva | A contribution to the thermal insulation performance characterization of corn cob particleboards[END_REF][START_REF] Pinto | Corn's cob as a potential ecological thermal insulation material[END_REF][START_REF] Sampathrajan | Mechanical and thermal properties ofparticle boards made from farm residues[END_REF], a mixture of durian peels and coconut coir fibers [START_REF] Khedari | New insulating parti de boards from durian peel and coconut coir[END_REF][START_REF] Khedari | New low-cost insulation particleboards from mixture of durian peel and coconut coir[END_REF], cellulose [START_REF] Nicolajsen | Thermal transmittance of a cellulose loose-fill insulation material[END_REF], wastes from tissue paper manufacturing and corn peel [START_REF] Lertsutthiwong | New insulating particleboards prepared from mixture of solid wastes from tissue paper manufacturing and corn peel[END_REF], kenaf fi bers [START_REF] Ardente | Buildingenergy performance: a LCA case study ofkenaf-fibres insulation board[END_REF], flax and hemp fibers [START_REF] Benfratello | Thermal and structural properties of a hemp-lime biocomposite[END_REF][START_REF] Korjenic | Development and performance evaluation of natural thermal-insulation materials composed of renewable resources[END_REF]Kymalainen and Sjoberg, 2008), cotton stalk fibers (Zhou et al.,201 0), jute fibers [START_REF] Korjenic | Development and performance evaluation of natural thermal-insulation materials composed of renewable resources[END_REF], cocon ut fibers [START_REF] Alavez-Ramirez | Thermal conductivity of coconut fibre filled ferrocement sandwich panels[END_REF][START_REF] Panyakaew | New thermal insulation boards made from coconut husk and bagasse[END_REF], sunflower pith [START_REF] Vandenbossche | New agro-materials with thermal insulation properties[END_REF], date palm fibers [START_REF] Chikhi | Experimental investigation of new biocomposite with low cost for thermal insulation[END_REF], etc.

The thermal conductivity of insulation boards is often influenced by their densities [START_REF] Benfratello | Thermal and structural properties of a hemp-lime biocomposite[END_REF][START_REF] Chikhi | Experimental investigation of new biocomposite with low cost for thermal insulation[END_REF][START_REF] Khedari | New insulating parti de boards from durian peel and coconut coir[END_REF][START_REF] Khedari | New low-cost insulation particleboards from mixture of durian peel and coconut coir[END_REF][START_REF] Lertsutthiwong | New insulating particleboards prepared from mixture of solid wastes from tissue paper manufacturing and corn peel[END_REF][START_REF] Panyakaew | New thermal insulation boards made from coconut husk and bagasse[END_REF][START_REF] Vandenbossche | New agro-materials with thermal insulation properties[END_REF][START_REF] Zhou | An environment-friendly thermal insulation material from cotton stalk libers[END_REF], and low-density materials have the lowest thermal conductivities. As an example, the thermal conductivity of an insulation board from sunflower pith is only 38.5mWfmK at 25°C with a board density of 36kgfm 3 [START_REF] Vandenbossche | New agro-materials with thermal insulation properties[END_REF]. It is comparable to that of conventional insulation materials like expanded polystyrene (37.4mWfmK with a board density of 50kgfm 3 ), rock wool (35.6 mW fm K with a board density of 115 kgfm 3 ), and glass wool (35.4mWfmK with a board density of 26kgfm 3 ). Thermal conductivity is higher with medium-density materials: 46-68 mW fm K at room temperature for cocon ut husk insulation boards with board densities of 250-350 kgfm 3 [START_REF] Panyakaew | New thermal insulation boards made from coconut husk and bagasse[END_REF], 81.5mWfmK for a cotton stalk fibers insulation board with a board density of 450 kgfm 3 [START_REF] Zhou | An environment-friendly thermal insulation material from cotton stalk libers[END_REF], 89.9-107.9mWfmK for hemp fibers insulation boards with board densities of 369-475 kgfm 3 [START_REF] Benfratello | Thermal and structural properties of a hemp-lime biocomposite[END_REF], 103.6mWfmK for a coconut coir insulation board with a board density of 540kgfm 3 [START_REF] Khedari | New insulating parti de boards from durian peel and coconut coir[END_REF], and 150mWfmK for a date palm fibers insulation board with a board density of754 kgfm 3 [START_REF] Chikhi | Experimental investigation of new biocomposite with low cost for thermal insulation[END_REF]. Nevertheless, such boards are viable options for use in building insulation (walls and ceilings).

Heat insulation properties, of fiberboards from cake generated during the biorefinery of sunflower whole plant in a twin-screw extruder are also promising, even if the corresponding board densities are quite high (904-966kgfm 3 ) (Evon et al., 2012b). Indeed, thermal conductivity at 25 oc is rather low for the three fiberboards tested, and it decreases from 135.7 to 103.5 mW fm K with an increase in board thickness from 5.4 to 1 0.2 mm. Th us, the thickest fiberboard ( density 917 kgfm 3 ) gives the best thermal insulation. Fiberboards from such a cake with lower density and higher thickness would perhaps produce a significant improvement in their heat insulation properties.

This study aimed to manufacture by thermo-pressing, new thermal insulation fiberboards with medium density (from 500 to 900kgfm 3 ), and high thickness (more than 10mm and up to 20 mm), from cake generated during the biorefinery of sunflower whole plant in a twin-screw extruder, and to evaluate the influence of molding conditions (temperature, pressure, and time) on their mechanical (flexural properties, Charpy impact strength, and Shore D surface hardness) and heat insulation properties.

Materials and methods

Material

Thermo-mechanical fractionation in the twin-screw extruder was carried out using a batch of sunflower (Helianthus annuus L) whole plant of the oleic type (La Toulousaine de Céréales, France) (Table 1), harvested in September, i.e. at plant maturity. Whole plant was previously dried in a ventilated oven (50°C, 48 h) and crushed using a hammer mill (Electra VS 1, France) fitted with a 15 mm screen. The moisture content of the powdered plant (batch of around 250kg) was 7.2±0.1% (French standard NF V 03-903 ).

Tablel Chemical composition of the sunflower whole plant used for the experiment and of the cake obtained, after its thermo-mechanical fractionation in the Clextral BC 45 twin-screw extruder(% of the dry matter).

Mate rial

Minerais

Filter セ @ 1--___L_ _ ____, r-' ---' -------' b セ オ イ ・ ャ @
A\:is a nd sc rew profil e Filtra te

Twin-screw extruder

Thermo-mechanical fractionation was conducted with a Clextral BC 45 (France) co-rotating twin-screw extruder. The extruder had seven modular barrels, each 200 mm in length, and different twin-screws which had segmentai screw elements each 50 and 100mm long (Fig. 1). Four modules (modules 3, 4, 5 and 7) were heated to 80 ac by thermal induction and cooled by water circulation. A fil ter section consisting of six hemispherical dishes with 1 mm diameter perforations, was positioned on module 6 to enable the filtrate to be collected. Screw rotation speed (Ss). sunflower whole plant feed rate (Qs), and barrel temperature (Oc) were monitored from a control panel.

Thenno-mechanicalfractionation of sunjlower whole plant in the twin-screw extruder

Sunflower whole plant was fed into the extruder inlet port by a Clextral 40 (France) volumetrie screw feeder on the first module. Water was injected using a Clextral DKM K20-2-P32 (France) piston pump at the beginning of module 4 (Fig. 1 ). The screw profile chosen in this study (Fig. 2) had already been used successfully for the aqueous extraction of oil from sunflower whole plant (Evon et al., 2010a). The trituration zone was located in modules 2 and 3, and consisted of a succession of 10 mono lobe paddles and 5 bilobe paddles, 5 cm apart. The extraction zone, situated in modules 4 and 5, was composed of a second series of 5 bilobe paddles. The reverse pitch screws were positioned in module 7, immediately Twin-screw extrusion was performed for 30 min before any sampling to ensure stabilization of operating conditions. Recorded operating conditions included feed rates of sunflower whole plant and water, temperature, plus motor current. On stabilization, the filtrate and the cake were immediately collected for a period of 90 min to avoid any variation in outlet flow rates. Sample collection time was determined with a stopwatch, and carried out once. The filtra te and the cake were then weighed.

The cake was dried in a ventilated oven ( 60 ac, 24 h) immediately after collection. Particle size distribution inside the cake was then determined using a Re tseh AS 300 (Germany) vibra tory si eve shaker with a 500 g test sample mass. The sieve acceleration was 1.5xg, and sieving time 10 min.

Oil extraction yield in the twin-screw extruder was calculated according to the following formula:

(1

)
where RL is the oil extraction yield based on the residual oil content in the cake (%), Qs the inlet flow rate of the sunflower whole plant (kg/h), Qc the flow rate of the cake (kg/h), Ls the oil content in the sunflower whole plant (%), and Le the oil content in the cake (%).

The protein extraction yield in the twin-screw extruder was calculated according to the following formula:

Rp = (Qs x ー セ @ セ @ セ セ @ x Pc) x 100 (2)
where Rp is the protein extraction yield based on the residual protein content in the cake(%), Ps the protein content in the sunflower whole plant (%), and Pc the protein content in the cake (%).

The energy consumed by the motor was determined according to the following formulae:

Ss P=Uxlxcoscpx Smax (3)
where P is the electric power supplied by the motor (W), U the motor's operating voltage (U=460V), I the current feeding the motor (A), cos 'P the theoretical yield of the extruder motor (cos 'P = 0.95), and Ss and Smax the test speed and maximum speed (600 rpm) of the rota ting screws (rpm), respectively. where SME is the specifie mechanical energy consumed by the mo tor per unit weight of sunflower who le plant (Wh/kg).

Thenno-pressing

The cake was molded by thermo-pressing inside an aluminium mold, using a 400 ton capacity Pinette Emidecau Industries (France) heated hydraulic press, producing 150 mm x 150 mm square fiberboards. Cake quantity for ali experiments was 258 g (i.e. 1147 mgfcm 2 ). Two fiberboards were manufactured for ali the b llf is the flow rate of the filtra te.

Qr." (kgfh) Qc(kgfh) 20.2 12.1 thermo-pressing conditions tested (including temperature of the aluminium mold, pressure applied, and molding time). Their thickness was measured immediately after thermo-pressing at twelve points, with a 0.01 mm resolution electronic digital sliding caliper, and the mean value (to) recorded. Then, fiberboards were equilibrated in a climatic chamber (60% RH, 25 oc) for three weeks before any analyses. A first fiberboard was used to assess mechanical properties for ben ding, and a second one for measuring: he at insulation properties, Shore D surface hardness, and finally Charpy impact strength.

Analytical methods

The moisture contents were determined according to French standard NF V 03-903. The mineral contents were determined according to French standard NF V 03-322. The oil contents were determined according to French standard NF V 03-908. The protein contents were determined according to French standard NF V 18-100. An estimation of the three parietal constituents (cellulose, hemicelluloses, and lignins) was made using the ADF-NDF method of Van Soest andWine (1967, 1968). An estimation of the water-soluble components was made by measuring the mass Joss of the test sam pie after 1 h in boiling water. Ali determinations were carried out in duplicate.

Mechanical properties for bending

AJFC 5-kN H5KT (France) uni versai testing machine fitted with a 1 00 N Joad ce li was used to assess the flexural properties orthe test specimens according to French standard NF EN 31 O. Such properties included energy-to-break (E), breaking Joad (F), flexural strength at break (a't), and elastic modulus (Er). The energy-to-break for each specimen, was estimated by the area under the Joad deformation curve from zero to rupture, calculated using the trapezium method. After fiberboard equilibration in the climatic chamber, test specimens were eut 30 mm wide and their thickness was measured at three points and their length at two points, with a 0.01 mm resolution electronic digital sliding caliper. The mean values ofthickness (t) and length (1) were recorded to calculate the volume of specimens, which were ali weighed to calculate their mean apparent density (d). The test speed was 3 mmjmin and the grip separation was 120 mm. Ali determinations were carried out four times.

7. Charpy impact strength

A Testwell Wolpert 0-40 daN cm (France) Charpy machine was used to assess the impact strength of the unnotched test specimens according to the French standard NF EN ISO 179. It included absorbed energy (W), and resilience (K). The test specimens, 60 mm long and 15 mm wide, were eut after fiberboard equilibration in the climatic chamber. Their thickness was measured at three points with a 0.01 mm resolution electronic digital sliding caliper, and the mean value (t) was recorded to calculate their section. Impact strength measurements were made at 23 oc according to the three point bending technique, and grip separation was 25 mm. Ali determinations were carried out twelve times. A Bareiss (Germany) durometer was used to assess the Shore D surface hardness of the fiberboards, according to the French standard NF EN ISO 868. Ali determinations were carried out 48 times (24 times for each side of the fiberboard).

Heat insulation properties

Thermal conductivity (À) and thermal resistance (R) of fiberboards were determined at three temperatures (10°C, 25°C, and 40 oc) according to the ISO 8302 08-91 standard, using a Lambda-MeBtechnik GmbH Dresden EP 500 (Germany) À-Meter hot plate apparatus. The area measured was 150 mm x 150 mm, and because of swelling in the elima tic cham ber, fiberboards had to be eut bef ore measuring to obtain the required dimensions. The difference of temperature between the two plates was 5 K. Measurements were also made with the bulk cake using a polycarbonate box 1 mm thick and 50 mm high. Here, the solid, cake particles were evenly distributed in the box, and a 'box effect' correction made on the thermal conductivity measurements. The cake was equilibrated in the climatic chamber (60% RH, 25 oc) for three weeks before being tested.

Results and discussion

Cake production by twin-screw extrusion

Thermo-mechanical fractionation of whole plant and aqueous extraction of sunflower oil were conducted simultaneously in the Clextral BC 45 twin-screw extruder using a screw configuration previously optimized (Evon et al.,201 Oa). A filtra te and a cake were collected continuously from the filter section positioned in the barrel on module 6 (Fig. 1). Lipids and water-soluble components, mainly proteins but also pectins and hemicelluloses, were partly extracted during the process. Operating conditions used for cake production and results of the thermo-mechanical fractionation are given in Table 2. Cake moisture content was 67.8 ± 0.2%, and it was dried in a ventilated oven immediately after production to facilitate conservation. 1t was a powder consisting of inhomogeneous partides (Figs. 3 and4), with chemical composition shown in Table 1. lt was richer in oil (17.6% of the dry matter for oil content) compared with other cakes described in previous studies (from 13.1 to 14.5%) (Evon et al., 2010a,b), leading to an oil extraction yield (RL, yield based on the residual oil content in the cake) of 45.9 ± 0.2%, and a corresponding protein extraction yield (Rp, yield based on the residual protein content in the cake) of 40.4 ± 1.2%.

The low efficiency of aqueous, whole plant extraction of sunflower oil could be explained by the relatively low (3.6) ratio of water to solid (Cà/Qs) in the twin-screw extruder compared to the ratio used for more efficient experiments (up to 4.1) (Evon et al., 2010a,b). 1t was also due to a low extruder filling ratio (0.09 kgjh rpm), defined as the input flow rate of sunflower whole plant to screw rotation speed ratio (Qs/Ss). resulting in an unsatisfactory liquidjsolid separation of filtrate and cake in the reverse pitch screws. Optimal efficiency of sunflower oil extraction with water (57%), was obtained in a previous study with an extruder Table3 Thermo-pressing conditions for the manufacture of the five fiberboards. during molding, corresponded to that giving the best mechanical properties for ben ding in a previous study (Evon et al.,201 Ob ). The lowest temperature (140 oq conversely, was chosen to be near the protein glass transition temperature. A previous study using the DSC technique, showed that this temperature for sunflower proteins was clearly influenced by their water content [START_REF] Rouilly | DSC study on the thermal properties of sunflower proteins according to their water content[END_REF], with a large drop (from 181 to soC) observed as this protein water content increased from 0 to 26.1% of the dry matter, confirming that water acted as a plasticizer for the proteins. Cake moisture was 2.6±0.1% at molding, and the glass transition temperature of proteins in this case was therefore estimated to be close to 140 oc. Moreover, no temperature higher than 200 oc was tested because thermal degradation of most of the organic compounds in the cake (i.e.lipids, proteins, hemicelluloses, and cellulose) occurs in the 200-37S°C temperature range (Evon et al., 2012b). More specifically, another study indicated that thermal degradation of sunflower proteins from an industrial cake occurred above 2SO oc and below 3S0°C [START_REF] Geneau | Manufacturing Process of Agromaterial Natural Composite by Twin-screw Extrusion and Injection Moulding from Sunflower Cake[END_REF]. A second thermal degradation stage in the cake from whole plant, situated at around 42S oc, is also observed. This corresponds to the thermal degradation of lignins only (Evon et al., 2012b).

The pressure applied varied from 1SO to 2SOkgffcm 2 . It was lower than for other studies: up to 320 kgffcm 2 (Evon et al.,201 Ob), from 320 to 360kgffcm 2 (Evon et al., 2012b), and from 2SO to SOO kgffcm 2 (Evon et al., 2012a). The objective for such a choice was to minimize the density of the materials obtained, in order to manufacture fiberboards with medium-density. Molding time varied from 40 to 76 s, which was quite similar to values used in a previous study (30 s and 60s) (Evon et al.,201 Ob), and was sufficient for sunflower proteins to reach a rubbery state during molding. In addition, for the fiberboard molded at 200 oc (trialS), the low molding time (76 s) was also a guarantee against initiation of thermal cake degradation during molding.

The low cake mois ture at mol ding (on! y 2.6%) was chosen to minimize vapor generation during thermo-pressing and so restrict the risk of defects like blisters inside the fiberboards. Cake quantity for ali the experiments was 1147 mgfcm 2 , and this led to the manufacture of thicker fiberboards compared with other materials from sunflower cake described in previous studies (Evon et al., 201 Ob, 2012a,b): at !east 10.3 mm thick (and up to 1S.2 mm) immediately Table4 after thermo-pressing, and at !east 13.0 mm (and up to 20.2 mm) after elima tic cham ber conditioning (60% RH, 2S oq for three weeks (Table 4).

Ali the fiberboards were cohesive. As previously observed (Evon et al., 201 Oa,b, 2012a,b ), proteins acted as an internai binder inside fiberboards, and they contributed to ensure cohesion of the agromaterial, in addition to the entanglement of lignocellulosic fibers also acting as reinforcement. Conditioning in the climatic chamber was conducted immediately after thermo-pressing in order to assess the mechanical and heat insulation properties offiberboards from equilibrated materials. lt resulted in water sorption, and this increased with a decrease of temperature, pressure, and molding time (Table 4). Indeed, the moisture content offiberboard from trial 1 was 8.3 ± 0.2% after conditioning, instead of only 6.8 ± 0.1% for the fiberboard from trial S. This led to the five fiberboards swelling, and the relative increase in their dimensions tended to increase further with decreasing thermo-pressing conditions: + 32.S% for trial 1 instead of +27.0% for trial S for thickness, and +4.2% for trial 1 instead of +2.6% for trialS for length (Table 4).

The density of the equilibrated fiberboards was clearly influenced by the thermo-pressing conditions used, increasing with increasing conditions (Table 4). lt varied from SOO to 8S8kgfm 3 , and the fiberboard with the lowest density (SOOkgfm 3 ) was Iogically that from trial 1 with the lowest values for mold temperature, pressure applied, and molding time: 140°C, 1S0kgf/cm 2 , and 40 s, respectively. This meant that a decrease in the fiberboard density favored water sorption during climatic chamber conditioning, due to its higher porosity. The mechanical properties of fiberboards were also clearly influenced by the thermo-pressing conditions, and could be correlated to their densities. Indeed, the higher the fiberboard density, the higher its mechanical properties (Table S). Regarding flexural properties, the energy-to-break, the breaking Joad, the flexural strength at break and the elastic modulus increased progressively from 3.2 to 44.4 rn], from 3.5 to 18.7 N, from S1.8 to 6S9.9 kPa, and from S.9 to 49.4 MPa, respectively, with increasing thermo-pressing temperature, pressure, and molding time. Similarly, for Charpy impact strength, the absorbed energy and the resilience increased from 0.1S to l.S1J, and from O.S to 7.7 kjfm 2 , respectively. Finally, the Shore D surface hardness increased from 19.2 to 4 7.1 o. Th us, the most fragile fiberboard was also the !east dense board and was from trial 1.

Since the temperature of the aluminium mold was at !east equal to the proteins' glass transition temperature (around 140°C), this change occurred systematically during molding and they were thus always in a rubbery state. However, because the proteinbased resin became more and more viscous with decreasing mold temperature, this meant that the fiber wetting became progressively worse, which could explain why the mechanical properties Swelling, water sorption and density offiberboards after conditioning in the climatic chamber (60% RH, 25 'C) for three weeks. The percentages in parentheses refer to the relative increase in the fiberboard dimensions (thickness and length) after conditioning in climatic chamber.

• lo is the length of fiberboards immediately after thermo-pressing.

b Measurements were made on specimens prepared for bending tests. c Mean apparent density of fiberboards was calculated after weighing of specimens prepared for bending tests. d Moisture contents were determined after crushing of the equilibrated fiberboards using a Foss Cyclotec 1093 (Denmark) mill fitted with a 1 mm screen. Trial ('C) of fiberboards diminished as the mold temperature decreased. Thus, when the mold temperature was only 140°C (trials 1 and 2), proteins became a Jess effective binder within the material due to insufficient wetting of the fibers, logically leading to the most fragile fiberboards of the study (Table S ). Conversely, fi ber wetting was much improved at a 200 oc molding temperature (trialS). Such thermo-pressing conditions were also associated with the highest values for pressure applied and molding time (2SO kgffcm 2 and 76 s, respectively), leading logically to the densest fiberboard of the study and so to the most mechanically resistant insulation board.

Influence of thermo-pressing conditions on heat insulation properties of jiberboards

The heat insulation properties of the five fiberboards were measured at three temperatures: 10°C, 2S°C, and 40°C. As generally observed for thermal insulation solids, the thermal conductivity of the five fiberboards tested increased with increasing temperature, with the thermal resistance logically and simultaneously decreasing (Table 6). The same tendencywas also observed for measurements made on the bulk cake. Consequently, the capacity for thermal insulation of ali the mate rials tested, decreased noticeably with increasing temperature. The thermal conductivity of the bulk cake and of the five fiberboards was also clearly influenced by their density, and it increased linearly with increasing density (Fig. Sa). Similar characteristics have been observed on severa! occasions, in particular for insulation boards made from durian peel and coconut coir [START_REF] Khedari | New insulating parti de boards from durian peel and coconut coir[END_REF][START_REF] Khedari | New low-cost insulation particleboards from mixture of durian peel and coconut coir[END_REF], wastes from tissue paper manufacturing and corn peel [START_REF] Lertsutthiwong | New insulating particleboards prepared from mixture of solid wastes from tissue paper manufacturing and corn peel[END_REF], cotton stalk fi bers (Zhou et al.,201 0), cocon ut husk and bagasse [START_REF] Panyakaew | New thermal insulation boards made from coconut husk and bagasse[END_REF], sunflower pith [START_REF] Vandenbossche | New agro-materials with thermal insulation properties[END_REF], hemp fibers [START_REF] Benfratello | Thermal and structural properties of a hemp-lime biocomposite[END_REF], and date palm fibers [START_REF] Chikhi | Experimental investigation of new biocomposite with low cost for thermal insulation[END_REF]. At the same time, the thermal resistance logically decreased (Fig. Sb), and these trends were observed at ali three temperatures.

However, here we will focus on the 2S oc results, considered as ambient temperature. Thermal conductivity was rather low for the five fiberboards (Table 6), due to their medium densities (from SOO to 8S8 kgfm 3 ) contributing to better heat insulation properties compared with previous results obtained with denser materials (Evan et al., 2012b ). Moreover, as already mentioned, it decreased with the decrease in board density. 1t was 110.5mW/mK for the most dense (8S8kgfm 3 ) fiberboard and only 88.SmW/mK for the !east dense one (SOO kgfm 3 ). The increase in porosity within the boards thus improved their thermal insulation capacity, and such values were in line with the thermal conductivities of other experimental medium-density insulation boards, made from cotton stalk fibers [START_REF] Zhou | An environment-friendly thermal insulation material from cotton stalk libers[END_REF], hemp fibers [START_REF] Benfratello | Thermal and structural properties of a hemp-lime biocomposite[END_REF] or coconut coir [START_REF] Khedari | New insulating parti de boards from durian peel and coconut coir[END_REF], and considered as viable options for use in building insulation: 81.SmW/mK (4SOkgfm 3 for board density), 89.9-107.9 mW fm K (369-47S kgfm 3 for board densities) and 103.6mW/mK (S40kgfm 3 for board density), respectively. At the same time, the thermal resistance varied from 0.118 to 0.228 m 2 K/W (Table 6). In addition, the adsorption of water during climatic chamber conditioning was greater for the Jess dense fiberboards (Table 4) due to their higher porosity, and thus the heat insulation properties also depended on the moisture content of the equilibrated boards. Indeed, the thermal conductivity decreased with increasing moisture content (Fig. 6a), and the thermal resistance rose at the same time (Fig. 6b).

Because the heat insulation properties of the fiberboards improved with decreasing density (Fig. S), the fiberboard with the weakest thermal conductivity (trial 1) also corresponded to the most fragile insulation board (Table S ). Conversely, the most conductive fiberboard (trialS) was also the most mechanically resistant insulation board. Consequently, the medium density (687 kgfm 3 ) fiberboard from trial3 (160°C for mold temperature, 2SOkgffcm 2 for pressure a pp lied, and 76 s for mol ding time) was a good compromise between mechanical properties (272 kPa for flexural strength at break, 26.3 MPa for elastic modulus, 3.2 kJ/m 2 for Charpy impact strength, and 37.3° for Shore D surface hardness) and those for he at insulation (99.S mW fm K for thermal conductivity at 2S oc, and 0.161 m 2 K/W for the corresponding thermal resistance). Fitted on walls and ceilings, it could be used for the thermal insulation of buildings. Measurements made on the bulk cake indicated that it was an even better insulation material (only 6S.6mW/mK for thermal conductivity at 2S oc, and O. 792m 2 K/W for thermal resistance), which was certainly due toits very low bulk density (204kgfm 3 ) and toits porous structure (Table 6). It also would be suitable for the thermal insulation of houses as loose till in attic spa ces.

To conclude, the originality of this study derives from two distinct points. Firstly, the optimal fiberboard (trial3) was a selfbonded composite materiallike the four others, meaning that there was no necessity to add any external binder to obtain a panel with good cohesion. Secondly, the reinforcing fibers inside these panels originated essentially from sunflower stalks, and these fibers are not now commercially available. Indeed, the sunflower harvest concerns only the seeds and not the by-products of its culture like the stalks and head s. However, the ir promising ability for both reinforcement of composite materials and thermal insulation could, in the future, justify their harvest in the field at the same time as the seeds. Which in turn could genera te an additional source of in come for farmers.

Conclusion

New thermal insulation fiberboards were manufactured using a heated hydraulic press, from a cake generated during the biorefinery of sunflower whole plant in a twin-screw extruder. Ali fiberboards were cohesive. Proteins acted as an internai binder, and entanglement of lignocellulosic fibers also acted as reinforcement. The thermo-pressing conditions had an important influence on board density and on mechanical and heat insulation properties. The density of the insulation materials varied from 500 to 858 kgjm 3 , and the !east dense fiberboard was produced un der the lowest values of mold temperature (140aC), pressure applied (150kgfjcm 2 ), and molding time {40s). The heat insulation properties improved with decreasing fiberboard density, and thermal conductivity of the !east dense fiberboard was only 88.5mW/mK at 25 ac. However, such a board was also the most fragile. A 160 ac mold temperature gave a good compromise between mechanical properties (272 kPa for flexural strength at break, 26.3 MPa for elastic modulus, 3.2 k]jm 2 for Charpy impact strength, and 37.3a for Shore D surface hardness) and heat insulation properties (99.5mW/mK for thermal conductivity). Positioned on walls and ceilings, it could be used for the thermal insulation of buildings. The heat insulation capacity of the bulk cake was even better (only 65.6 mW fm K). It also would be sui table for the thermal insulation of houses wh en used as loose fil! in attic spaces.

Fig. 1 .

 1 Fig. 1. Schematic modular barrel of the Clextral BC 45 twin-screw extruder used for the thermo-mechanical fractionation of sunflower whole plant (Oc= 80'C).

  filtration module, to press the liquidfsolid mixture.

Fig. 2 .(

 2 Fig. 2. Screw configuration of the Clextral BC 45 twin-screw extruder used for the thermo-mechanical fractionation of sunflower who le plant. T2F, trapezoïdal double-thread screw; C2F, conveying double-thread screw; ClF, conveying simple screw; DM, monolobe paddle-screw; BB, bilobe paddle-screw; CFtC, reverse screw; the numbers following the type of the screw indicate the pitch ofT2F, C2F, Cl F and CFtC screws, and the length of the DM and BB screws.

  Moisture content (%)d 8.33 (±0.17) 7.70 (±0.10) Water absorbed (%dry matter) 9.08 (±0.20) 8.34 (±0.11(±0.12) 7.31 (±0.13)

  À) and thermal resistance (R) at three temperatures (10'C, 25 'C, and 40 'C) of the bulk cake and of the five fiberboards manufactured by thermo-pressing.

  Fig. S. Thermal conductivity (a) and thermal resistance (b) of the bulk cake and of the five fiberboards manufactured by thermo-pressing, as a function oftheir density at three temperatures (10"C, 25 "C, and 40"C).

Fig. 6 .

 6 Fig. 6. Thermal conductivity (a) and thermal resistance (b) of the five fiberboards manufactured by thermo-pressing, as a function of their moisture content at three temperatures (10"C, 25 "C, and 40"C).


Professor Dominique Crozat

Tourism development in UNESCO biosphere reserves: a typology of action

Keywords: Tourism development, UNESCO biosphere reserves, legitimation of action

Diversity of forms and governance logics in most UNESCO biosphere reserves reveals that a strong pragmatism leads to value natural and, more-over, non-natural (culture, heritage,…) local assets. This study tries to build a typology of strategies and governance logics used to process tourism development with seven study cases in France, Germany and Chili.

A-Biosphere Reserves

Biosphere reserves are areas of terrestrial and coastal ecosystems which are solutions to reconcile the conservation of biodiversity and its sustainable use.

With today more than 500 biosphere reserves in one hundred countries, they are "Living laboratories" (Mousli, 2008) to develop integrated management of land, water and biodiversity.

Their goals are the conservation of biodiversity and sustainable development

The MAB program (Man and Biosphere) of the UNESCO has the aim to implement :

-the Agenda 21 -the Convention on Biological Diversity (especially the ecosystem approach) -realize the objectives of the "Millennium Development Goals" (such as environmental sustainability) and the Decade "United Nations Education for Sustainable Development"

More widely, we need to question his goal of preserve and improve the environment.

In fact, UNESCO biosphere reserve responds to three basic functions:

-Conservation -Sustainable development -Logistical assistance to scientific investigation and education They include three types of spaces:

 A central area: long-term protected, with the aim to conserve biodiversity, protection of landscapes, ecosystems and species it contains.

 A transition area surrounding the central area is designed to protect it from the disturbance. This sparingly populated area is designed to greet traditional activities or activities with low impact (hunting, fishing, sports, tourism, forestry, teaching and research). Then, this decision initiates a public debate which requires that local actors legitimate this option to their citizens to justify these constraints.

C-Legitimate the action

The theory of "justification" by Boltanski and Thévenot (1991) gives elements to analyse; they say that public actors justify it by one of 8 models 1 to legitimate it

D-Models of action

1 -The civic and anthropocentric model concerns great projects (often regional) where only prevails general (often national) interest; local environment and population are forgotten. The France of the 60' and 70', Tunisia of the 80' are good models. If, sometimes, nature exists, it's for the benefit of the full nation not for local population.

-Technocratic and normative model: it has the same logic but two differences:

a-the political agent refers to an expert (often international) to justify his action.

b-Nature become a material to legitimate other interests: it's the case of the Unesco biosphere reserve of Mont Ventoux (real estate at regional scale).

-Civic

6-The local development model shows or how local powers can value all opportunities and

the context. This model permits to describe the opportunist ability to adapt to spatial contexts.

The studied cases are peri-urban biosphere reserves; they are «rich», because many urban opportunities were long time developed here. Here, in the Unesco biosphere reserves of Gatinais Fontainebleau, near Paris or the Campana-Peñuelas reserve, in the mid-road from Santiago to Valparaiso, the two main chilian cities.

The use of UNESCO biosphere reserve or other label offers new opportunities: Fontainebleau detains also Unesco World heritage and 3 major French labels.

Here, the protection of nature is a hazard: it's already impossible to develop any activities on these very well protected spaces… Fontainbleau is one of the oldest protected natural spaces of the world since the 16 th century. But the biosphere label attracts thousands tourists more.

7-« Deep ecology » model reveals an ideological relation between society and nature: it must seek the perfection, the "pure" immersion in a totally preserve nature. Every dimension of life has to respect the fusion between man and nature. If it's impossible, the man must disappear from this space to let nature preserved.

It could be the hope of some members of the team in the biosphere reserve of Camargue but they need to accept a reality imposed, mainly economical, from outside of the reserve that limits this utopist dream.

Models of justification Study cases

Civic models: the civic and anthropocentric model

No study case here (ex. touristic planning of Languedoc French Coast during the 60')

Civic models: the technocratic and normative model

UBR of Mont Ventoux

Civic models: the model of the civic educator

UBR of Camargue

The marketing services model UBR of Spreewald (Germany)

Real protective action Incident protective action

To conclude, the UNESCO biosphere reserves have the goal of preserve and improve the environment and nature. But, we can see that this purpose is rarely a priority: except on the Camargue biosphere reserve most, the studied cases aren't concerned by this aim, or it's an indirect action. The economic logic is dominant and often a real success with only one exception: the Camargue biosphere reserve.

Beyond the apparent rigidity of the rules governing UNESCO biosphere reserves, touristic valorizations reveal a real freedom but, also, a lot of "greenwashing"… A full paper with complete description of each study case is in progress; publication by the end of June

Bibliography and documentation



  A transition zone where live most of the population and with major economic activities B-Complex social uses and action A Biosphere reserve is a complex space because there's: a) A dual process of territorial ecological improvement based on both the spectacle of nature and also tourists' relationship with the place. b) The ambiguity of a situation in which an economical process combines diversified ethics and plural rationalities c) The environmental labeling means that quite large spaces are enacted in a foreclosure situation exclusive of any other uses (central area) or restricted (transition area; transition zone)

  business model with productive logic 6 -The traditionalist and conservative model 7 -The local development model 8 -The model based on "deep ecology" I've studied the politics in 7 cases of touristic development: 6 UNESCO biosphere reserves in France, Germany and Chili and 2 UNESCO World heritage; in fact, Fontainebleau has the two labels. Most of the Boltanski and Thévenot categories are represented.

  educator model is very different with an ideological relation with nature; the goal of the action becomes to learn a new relation with nature: it's only the case of the Unesco 1 Adapted from Corneloup J., Bouhaouala M., Vachée C., Soulé B., (2004) Formes de développement et positionnement touristique des espaces sportifs de nature, revue Loisir et société, vol. 24, n° 1, pp. 21-46 biosphere reserve of Camargue. Yet, it perfectly corresponds to the goals of the biosphere reserves. 4-In the marketing services model, connected here to business model with productive logic, the reserve of biosphere is an economic resource mixed with others. The marketing logic is dominant; the biosphere preservation theme's has a low capacity to orient the politics, except about economical valorization. Near Berlin (UBR of Spreewald) the joint use of the Biosphere Reserve and a close Malaysian funding Tropical Island Resort is mainly a true business model. 5-The traditionalist and conservative model reveals an anti-action: the UNESCO label is used to help local powers to boost out of age social organizations. But different cases reveal two conservative use: -In the franco-german biosphere reserve of Vosges du Nord-Pfälzerwald, this approach has the function to permit the adaptation of fragile spaces to a changing world. -Close to Spain frontier, the Upper-Vallespir valley is a part of the Canigou mountain UNESCO world heritage; in this a social group try to stop the erosion of its domination. Both cases reveal an indirect action of nature protection by «freezing» spaces (in the Vosges du Nord-Pfälzerwald reserve the landscape is promoted as historical heritage with a really sustainable use; in the Upper-Vallespir valley, hoteliers have politic powers and stop the development of other activities than spas.



  Barrère, C , 2007, Vers une théorie substantiviste du patrimoine, « Economie du patrimoine », N° spécial Economie appliquée Tome LX, N° 3, septembre 2007  Bateson, G. (1977) Vers une écologie de l'esprit, Paris, Gallimard  Boltanski, L. & Thévenot, L. (1991). De la justification. Métaillé, Paris  Corneloup J., Bouhaouala M., Vachée C., Soulé B., (2004) Formes de développement et positionnement touristique des espaces sportifs de nature, revue Loisir et société, vol. 24, n° 1, pp. 21-46.  Equipe MIT (2002) Tourisme 1. Lieux communs, Paris, Belin  Equipe MIT (2011) Tourismes 3 La révolution durable, Paris, Belin, 332p.  Glacken, J. (1967) Traces on the Rhodian Shore, Berkeley, Univ. of Califonia Press.  Kalaora, B. (1986). La forêt pacifiée. L'Harmattan, Paris  Lafaye, C. & Thévenot, L. (1993). Une justification écologique. Revue Française de Sociologie, 34, 495-524  Leriche, F. et al, L'économie culturelle et ses territoires, Ed. Presses universitaire du Mirail, Toulouse, 2008  Michaud, D. (1989). L'avenir de la société alternative. Logiques sociales, Paris.  Mousli, S. (2008) UNESCO. FAQ -Réserve de biosphère ; Unesco, doc. n° D:20081124171833  Pecqueur B., Peyrache-GadeauV., 2002, Les ressources patrimoniales : une modalité de valorisation par les milieux innovateurs de ressources spécifiques latentes ou existantes, Colloque du GREMI 5, Neuchâtel, 26-27 avril  Planque B. & Gaussier N., 1998, Milieux innovateurs et développement durable, des mésologies complémentaires, Aix-en-Provence, Centre d'Economie Régionale, 1998, Notes de recherche n°207  Raffestin, C. (1992) Géographie et écologie humaine, in Bailly, A., Ferras, R. et Pumain, D. (dir.) Encyclopédie de Géographie, Paris, Economica, pp. 23-36.  Rossi, G. (2001) L'ingérence écologique. Environnement et développement rural du Nord au Sud. Paris, CNRS ed., 254p.  Valette E., 2003, Pour une approche géographique de l'innovation sociale. L'exemple des territoires ruraux périurbains de la garrigue nord-montpelliéraine, Paris, Thèse de doctorat en géographie, Université Denis Diderot Paris 7.

Personal presentation


Olivier Mongin

Muniappan Sankar

Marina Charlot

Youssef Mir

Mireille Blanchard-Desce
email: m.blanchard-desce@ism.u-bordeaux1.fr

Strong enhancement of two-photon absorption properties in synergic

Keywords:

Photodynamic therapy (PDT) is a medical technique used in oncology for the treatment of several tumours as well as in ophthalmology especially in the treatment of age-related macular degeneration (ARMD). 1,2 The use of a two-photon (2P) excitation offers new perspectives for PDT, especially for the treatment of small areas, such as small solid tumours. Indeed two-photon absorption (2PA) offers several advantages, which includes the ability for highly selective excitation in biological media and intrinsic three-dimensional resolution as well as increased penetration depth in tissues. 3 This however calls for the use of optimized 2P photosensitizers that should combine large 2PA cross-sections in the biological spectral window (700-1100 nm) and high singlet oxygen quantum yields (or ROS production). Most of the photosensitizers currently used clinically or in clinical trials are porphyrin derivatives, such as porfimer sodium (Photofrin) and verteporfin (Visudyne). These compounds exhibit high singlet oxygen quantum yields but low 2PA cross sections (10 GM and 50 GM, respectively) in the NIR, 4 as model tetraphenylporphine (TPP, 12 GM). 5 _ENREF_8 Dramatic enhancement of the 2PA crosssections can be achieved in porphyrinoids, 6 or expanded porphyrins, 7 in conjugated porphyrin dimers, trimers and oligomers, 8 planarized fused or bridged porphyrins 9 or supramolecular assemblies. 8a, 10 The extension of the -conjugated system involves the porphyrin or porphyrinoid macrocycles thus leading to major modification of their photophysical properties: a marked broadening and red-shift of the Q-bands is observed resulting in residual one-photon absorption (1PA) overlapping with the 2PA band located in the NIR region. This promotes resonance enhancement of the 2PA band leading to giant 2PA cross-sections. Other consequences are however possible loss of 3D resolution (due to concomitant 1PA) and weakening of fluorescence and photosensitizing properties. Nevertheless biphotonic photosensitizers that maintain significant fluorescence are highly desirable to allow in vivo monitoring and subsequent localized irradiation.

Here we describe our efforts towards the design of porphyrin-based 2P photosensitizers exhibiting large 2PA in the NIR -with no 1PA -and retaining the fluorescence as well as the excellent photosensitization properties of the porphyrin moiety. Dendritic antenna systems based on resonant energy transfer (FRET) from peripheral 2P absorbers towards a single porphyrin core have been previously designed. 11 In contrast, our strategy is based on multiporphyrin assemblies with a weak conjugation between a central 2PA unit and peripheral porphyrins. A smooth electronic disconnection between the porphyrin moieties and the conjugated -system is ensured by deviation from planarity (Figure 1), so as to retain some of the photochemical features of native porphyrins (in particular singlet oxygen production and fluorescence).

To achieve this goal, we chose meso-tetraarylporphyrin derivatives, where a phenyl ring in meso position prevents porphyrins to be coplanar with the 2PA unit, due to steric effects [START_REF] Silvers | TPP for example, the dihedral angle between the phenyl rings and the macrocycle is more than 60[END_REF] (Figure 1). Four multiporphyrin assemblies (1a, 1b, 2, 3) were synthesized. Inspired by our seminal work on the design of conjugated quadrupolar, octupolar chromophores for 2PA, 13 we selected fluorene and triphenylamine as cores in dumbbell-like or three-branched derivatives, and phenyleneethynylene or fluorenylene-ethynylene as connectors to the peripheral porphyrin units (Figure 1). Derivatives 2 and 3 bear pyridyl and triethyleneglycol substituents which should improve their hydrophilicity and biocompatibility. It should be stressed here that pyridines might be further quaternarized.

1a : M = 2 H 1b : M = Zn 2 3 N N N N N N N N N N N N N N H H H H O O O O O O N N N N N N N N H H O O O O O O N N N N N N N H H N N N N N N N N N NN N N M M M O O O O O O N N N N N N N O H H O O O O O Figure 1.
Chemical structures of "electronically disconnected" porphyrin dimers and trimers.

Porphyrin trimer 1a was obtained from the threefold Pd-catalyzed cross-coupling reaction between ethynyl porphyrin 4 14 and triodotriphenylamine 5. 13a Its metalation with zinc acetate afforded zinc porphyrin trimer 1b (Scheme 1). Porphyrin dimer 2 was synthesized in a multistep sequence from 2,7-diiodofluorene 6b. 15 Dialkylation of fluorene 6b with [2-[2-(2-methoxyethoxy)ethoxy]ethoxy]p-toluenesulfonate in the presence of NaOH and tetrabutylammonium bromide afforded 7a. The double Sonogashira coupling of diiodo derivative 7a with 2-methyl-3-butyn-2-ol afforded 7c, which was deprotected with NaOH to afford diethynylfluorene 7d. The double coupling of the latter with bromoporphyrin derivative 8 16 gave porphyrin dimer 2 (Scheme 2). The synthesis of extended porphyrin trimer 3 started also from diiodofluorene derivatives 6b and 7a. The latter compound was desymmetrized by reaction with a single equivalent of 2-methyl-3-butyn-2-ol, affording monoiodo derivative 7b. The Sonogashira reaction of triethynyltriphenylamine core 9 13a with three equivalents of 7b led to extended core 10a, the protecting groups of which were cleaved with NaOH, giving triethynyl derivative 10b. Extended porphyrin trimer 3 was finally obtained from the threefold palladium(0) catalyzed reaction of 10b with three equivalents of bromoporphyrin 8 (Scheme 2).

The photophysical characteristics (including one-photon absorption, luminescence, singlet oxygen generation and two-photon absorption) of the new porphyrin assemblies are gathered in Table 1. As shown in Figures 2 and3, all compounds exhibit intense absorption bands in the UV and visible regions. Absorption spectra of the free-base multiporphyrin assemblies (1a, 2, 3) closely resemble that of TPP, 17 the characteristic Q bands and Soret band being unaffected. This reveals that the lower excited states are preserved and localized on the porphyrin peripheral units, suggesting that no major electronic coupling between the conjugated core and the peripheral porphyrins occurs in these states (Figure 2, left). The absorption spectrum of trimer 1b is also typical of zinc tetraarylporphyrin complexes (See Supporting Information, Figure S1). No bathochromic shift of the Q bands is observed thus preserving full transparency in the NIR. In toluene. b In THF. c Fluorescence quantum yield determined relative to TPP in toluene. d Singlet oxygen quantum yield determined in toluene relative to TPP in toluene. e Ratio between the maximum 2PA cross section of each compound (normalized by the number of porphyrins in the assembly) and the 2PA cross section of TPP at 790 nm. f Data from lit. 17 g Data from lit. 18 h Data from lit. 19 i Data from lit. 5 In contrast, the presence of the conjugated core moiety promotes the rise of a new absorption band in the near-UV region to the blue side of the Soret band (Table 1), whose intensity increases with increasing size of the core (Figure 1). This band reveals the emergence of higher excited states that modify the excitation behavior of the assembly as compared to TPP. The photochemical properties of multiporphyrin assemblies 1-3 are similar to that of TPP, in agreement with a lower excited state mostly localized on peripheral porphyrin units. As a result, fluorescence and singlet oxygen quantum yields are close to that of TPP (Table 1). We note however that the presence of the core moiety slightly affects the lower excited state as evidenced from the emission spectrum of 1-3 which are slightly modified: the intensity of the emission sub-band at 717-720 nm increases with respect to that of isolated porphyrin moieties (Figure 2, right). Hence a slight electronic coupling is occurring in the excited state. This reveals that the different moieties in the multiporphyrin assemblies (i.e. conjugated core and porphyrins) do not behave as fully independent units but slightly interact.

 max (M -1 .cm -1 )  abs (Q bands) (nm)  em max (nm)  F c   d  2PA max (nm)  2 max (GM)
The 2PA properties of porphyrin assemblies 1-3 were further examined. Taking advantage of the fluorescence properties, they could be determined in the NIR range by investigating their two-photon excited fluorescence (2PEF), following the experimental protocol described by Xu and Webb. 20 Below 740 nm, giant 2PEF signal increases (and concomitant slight deviation from quadraticity) are observed due to the onset of 1PA resonance (leading to apparent 2PA cross-sections larger than 10 5 GM for compound 3 at 700 nm). We thus restrict further discussion to the spectral range where a fully quadratic dependence of the fluorescence intensity on the excitation power is observed and 2PA responses are fully non-resonant ( ≥ 790 nm). Quite interestingly, in this spectral range, a marked increase of the 2PA responses compared to that of model TPP (12 GM at 790 nm), up to 1360 GM for trimer 3 at the same wavelength is observed (Figure 3). Tripodal triphenylamine cored compounds 1a-b, reveal an enhancement factor of more than 10 while a larger value of 20 is obtained with dumbbell-like fluorene derivative 2 (Table 1). Finally, a ~40-fold enhancement is obtained with the extended tripodal compound 3. These striking enhancements provide evidence that the conjugated cores generate higher excited states that are responsible for enhanced 2PA, while allowing fast return (in agreement with Kasha rule) to the lower-excited state responsible for fluorescence (as demonstrated by 2PEF experiments) and singlet oxygen generation. This confirms that the different moieties in 1-3 do not behave as fully independent chromophoric units but slightly interact in the higher excited states. The large (non-resonant) 2PA responses are reminiscent of octupolar (1a-b, 3) or quadrupolar (2) schemes. 13 In addition, extension of the conjugated branches leads to an increase of the 2PA cross sections and of the enhancement factor of 3 (compared with 1a-b), as expected from an octupolar scheme. 13 This suggests that the porphyrin end-groups are indeed contributing to the 2PA response by acting as electro-active end-groups (probably in relation with field rather than resonance effects) in the higher excited states.

In conclusion, by engineering a smooth electronic disconnection between 2PA conjugated systems and peripheral porphyrin emitters/photosensitizers, we have achieved striking (nonresonant) 2PA enhancement in the NIR while retaining excellent fluorescence and photosensitization properties. This "semi-disconnection" strategy, which is different in essence from the strategies based on the full extension of conjugation in porphyrinoids or (supramolecular) assemblies and from FRET based covalent assemblies, opens a promising route for combined two-photon imaging and photodynamic therapy.

Scheme 1 .

 1 Scheme 1. Reagents and conditions (a) Pd 2 dba 3 , AsPh 3 , DMF/Et 3 N, 40 °C, 15 h (47%). (b) Zn(OAc) 2 . 2 H 2 O, CHCl 3 /MeOH, reflux, 1h (82%).

Scheme 2 .

 2 Scheme 2. Reagents and conditions (a) I 2 , H 5 IO 6 , AcOH, H 2 SO 4 , H 2 O, 75 °C, 2 h (68%). (b) CH 3 (OCH 2 CH 2 ) 3 OTs, NBu 4 Br, KOH, 75 °C, 8h (59%). (c) 2-methyl-3-butyn-2-ol (1 equiv), Pd(PPh 3 ) 2 Cl 2 , CuI, toluene/Et 3 N, 40 °C, 16 h (29%). (d) 2-methyl-3-butyn-2-ol (3 equiv), Pd(PPh 3 ) 2 Cl 2 , CuI, toluene/Et 3 N, 40 °C, 18 h (94%). (e) NaOH, toluene/iPrOH, reflux, 1.5 h (58%). (f) 7d (1 equiv), 8 (2 equiv), Pd 2 dba 3 , PtBu 3 , DMF/Et 3 N, 90 °C, 5 h (27%). (g) 7b (4 equiv), 9 (1 equiv), Pd(PPh 3 ) 2 Cl 2 , CuI, toluene/Et 3 N, 40 °C, 60 h (43%). (h) NaOH, toluene/iPrOH, reflux, 1 h (51%). (i) 8 (3 equiv), 10b (1 equiv), Pd 2 dba 3 , PtBu 3 , DMF/Et 3 N, 90 °C, 5 h (14%).

Figure 2 .

 2 Figure 2. Absorption (left) and emission (right) spectra of free-base porphyrin derivatives 1a, 2 and 3 and comparison with TPP.

Figure 3 .

 3 Figure 3. 2PA spectra of TPP 5 and "electronically semi-disconnected" porphyrin assemblies.

Table 1

 1 Photophysical data of porphyrin assemblies and model TPP.

	Compd	 abs (core) (nm)	 abs (Soret) (nm)

Acknowledgments

ANR PNANO (07-102) is gratefully acknowledged for postdoctoral fellowships to Y. M. and M. S. CNRS is also acknowledged for a grant to M. C. We wish to thank Vincent Hugues and Marine Baudequin for contribution in 2PEF and synthesis, respectively.

Supplementary data

Supplementary data (synthetic procedures and spectroscopic data) associated with this article can be found, in the online version, at http://dx.doi.org/***.