RAFFINERIA PETROLCHIMICA BIORAFFINERIA RAFFINERIA PETROLCHIMICA BIORAFFINERIA PETROLIO Composizione complessa Non rinnovabile Estrazione lunga, laboriosa e costosa Costo elevato (70-80$/barile, circa 370 €/ton) Giacimenti limitati in particolari aree del Mondo Carburanti, energia, calore, chemicals Processi inquinanti BIOMASSE (in particolare scarti agro-forestali lignocellulosici). Composizione complessa Rinnovabile Raccolta semplice e veloce Basso costo (10-150 €/ton) Disponibile in qualsiasi Paese Biocarburanti, energia, calore, chemicals Processi non inquinanti e rispettosi dell’ambiente (la CO2 prodotta durante il processo viene rapidamente fissata dalle biomasse) SCOPO BIORAFFINERIA: Valorizzare in toto la biomassa di partenza Bioraffineria Valutazione della composizione della biomassa Pretrattamento (fisico, chimico o biologico) per separare le diverse componenti della biomassa Separazione delle diverse frazioni ottenute dai pretrattamenti Utilizzo delle molecole contenute nelle diverse frazioni per la realizzazione dei prodotti Separazione, purificazione e formulazione dei prodotti ottenuti

SCHEME OF BIOREFINERY PLANT lignocellulose pretreatment to fractionate the recalcitrant lignocellulose structure; enzymatic hydrolysis of the isolated cellulose moiety, by which cellulases hydrolyze reactive intermediates to fermentable sugars; and fermentation, which produces cellulosic ethanol or other bio-based chemicals

STRUTTURA DELLA CELLULA VEGETALE La parete cellulare ha una struttura molto complessa e altamente organizzata essa è composta da diversi strati concentrici che si depositano durante l’accrescimento della cellula. Rappresentazione schematica di cellule vegetali e dell’organizzazione della parete cellulare. Lamella mediana (ML), parete primaria (P), strato esterno della parete secondaria (S1), strato intermedio della parete secondaria (S2), strato interno della parete secondaria (S3), membrana plasmatica (W).

CELLULOSE Glucose polymerization (β-1,4 glycosidic linkages) Chains assembled into sheets (intramolecular + intermolecular H-bonds) Sheets stacking (Van der Waals) Spin of the sheets into highly organized fibrils (crystalline and amorphous domains) Incredibly strong fibers, resistant to the action of enzymes (cellulases) that can crack them back into their simple-sugar components Mechanical (mill)/ chemical (ionic liquid) pretreatment required to break cellulose for the production of fuel alternatives

Aggregazioni della cellulosa in microfibrille A cellulose chain may be 5-7 μm long, but a fibril can be much longer, probably at least 40 μm due to several chains overlapping each other. Each cellulose microfibril has approximately 36 glucose chains, and every elementary fibrils are further associated into larger units, called fibrils aggregates, by means of non-cellulosic polymers

HEMICELLULOSE (ARABINOXYLANS, XYLANS ) Shorter, branched chains: 500-3000 sugar units (cellulose: 7000-15000) that cross-link together cellulose fibrils and covalently bind lignin via ester and ether linkages (Lignin-Carbohydrate Complexes, LCC) Hemicellulose is composed of carbohydrates based on pentose sugars, mainly xylose and arabinose, as well as hexose sugars, such as glucose and mannose. Mannuronic and galacturonic acid are often present Abundant in crop residues, great potential in the production of chemicals

Oxidative coupling of mesomeric phenoxy radicals LIGNIN Tridimensional, racemic, non-crystalline, hydrophobic polymer that embeds together the lignocellulosic components. Plays a crucial role in providing mechanical support and in the plant natural defense against degradation. β-O-4 β-β β-5 5-5’-O-4 Paracoumaryl alcohol (P-OH) Coniferyl alcohol (G-OH) Sinapyl alcohol (S-OH) Oxidative coupling of mesomeric phenoxy radicals LIGNIN Hardwood: Softwood: Herbaceous: G-OH + S-OH G-OH P-OH + G-OH + S-OH

LIGNIN-CARBOHYDRATE COMPLEXES (LCCs) In herbaceous plants, lignin and hemicellulose are connected through a phenolic bridge. Ferulic and p-coumaric acids are esterified to hemicelluloses and lignin, respectively. In wood, LCCs mainly consist of ester and ether linkages connecting sugar hydroxyls of hemicellulose to the a-carbanol of phenylpropane subunits in lignin. Crestini C.; Argyropoulos D.S. Structural Analysis of Wheat Straw Lignin by Quantitative 31P and 2D NMR Spectroscopy. The Occurrence of Ester Bonds and β-O-4 Substructures. J. Agric. Food Chem. 1997, 45, 1212-1219

Lignocellulose biomass pretreatment The objective of pretreating lignocellulosics is to alter the structure of biomass and to make the cellulose and hemicelluloses more accessibile and ame:nable to hydrolytic enzymes following these criteria: minimization of hemicellulose degradation products limiting the formation of by-products that inhibit ethanol fermentation reducing energy/water use and lowering environmental impacts, capital and operating costs

Lignocellulose biomass pretreatment: Steam Explosion Steam explosion involves rapidly heating biomass with steam at elevated temperatures (190-240 °C) with residence times of 3-8 minutes followed by explosive decompression. This treatment promotes hemicellulose hydrolysis and opens up the plant cell structure, although enhanced digestibility of cellulose is only weakly correlated with the physical effects

Lignocellulose biomass pretreatment: Acid and base pretreatment Dilute acid pretreatment has been extensively studied and typically employs 0.4-2% H2SO4 at temperatures of 160–220 °C to remove hemicelluloses and enhance cellulase digestion of cellulose Aqueous lime or NaOH pretreatment has been shown to be effective for wheat straw and sugarcane bagasse at lower temperatures than acid treatments; however, the treatment times are in some cases on the order of hours (20). The use of an alkaline treatment also incurs additional capital cost, as the recovery of salts requires a lime kiln to regenerate the base.

Lignocellulose biomass pretreatment: Ammonia pretreatment Ammonia pretreatment involves pretreating biomass with an aqueous ammonia solution causing depolymerization and cleavage of lignin–carbohydrate bonds. Agricultural residues and herbaceous plants treated in this manner exhibit an excellent response to cellulase. However, woody biomass is often not efficiently treated by this technology, and in all cases, ammonia recovery is an additional cost and an important consideration

Lignocellulose biomass pretreatment: organosolv pretreatment Organosolv pretreatment of biomass resides on the use of an organic solvent system (23-26) with enhanced lignin solubilizing properties. Usually, the resultant cellulosic fraction is highly susceptible to enzymatic hydrolysis, generating very high yields of glucose that can be readily converted to ethanol

Lignocellulose biomass pretreatment: ionic liquid Once the ionic liquid has dissolved the lignocellulose biomass into its components, the subsequent addition of an anti-solvent, such as water or ethanol, results in the sugars being precipitated out while a fairly large fraction of the lignin remains in solution

dissolvere il materiale nelle sue componenti in solventi tradizionali Il network tridimensionale che lega lignina e polisaccaridi rende molto complicato dissolvere il materiale nelle sue componenti in solventi tradizionali LIQUIDO IONICO (sale costituito da grande catione organico e piccolo anione inorganico) Alto bollente Non tossico Biodegradabile Riciclabile Non infiammabile Chimicamente stabile Alta conduttività termica Composto verde 1-allyl-3-metilimidazolo cloruro- [Amim]Cl Catione organico: interazione π-π con lignina; Anione inorganico: rottura legami-H inter-intra molecolari della cellulosa

Meccanismi di idrolisi Doppia possibilità di meccanismo: “Endo” “Eso”

Depolimerizzazione cellulosa; Trattamento CON CELLULASI MECCANISMO Trattamento cellulolitico: i campioni vengono messi in bagno termostatico per due giorni a 40 °C con tampone acetico e cellulasi, un’enzima secreto dal fungo Trichoderma reseei capace di idrolizzare la cellulosa a glucosio Meccanismo di azione dell’enzima