**9. The SCF application as dispersion for biocatalysis**

The use of the SCF as a dispersion for biocatalysis was described in 1985 and there is now a growing trend in using the SCF as a reaction media for enzymes. The advantage of using enzymes in the SCF include; synthesis reactions in which water is a product can be driven to completion, the increased solubilities of hydrophobic materials, greater thermostability of biomolecules in SCF, readily solvent recycling, integrated biochemical reactions and separations. CO2 is the most widely used SCF, however, there is a growing interest in using other SCFs (e.g., ethylene, fluoroform, ethane, sulfur hexafluoride and near critical propane).

Enzymes such as alpha amylase, glucose oxidase, lipase and catalase retained their activities in the solution of high pressure CO2 in water. Among the enzymatic reactions in SCF, the use of lipase shows most commercial promise. A SC-CO2/H2O mixture may be used as a reaction medium for either hydrolytic or synthetic reactions catalyzed by lipase and other

Supercritical Fluid Application in Food and Bioprocess Technology 563

Waste treatment is one of the most important and urgent problems in environmental management around the world. SC-water oxidation has attracted attention for the treatment of industrial waste, especially toxic and refractory waste. In a study, SC-water oxidation with H2O2 was applied as the oxidant to the treatment of a model municipal solid waste containing proteins, fats, vitamins, fiber, and inorganic minerals. The effects of temperature, oxidant concentration, and reaction time on the decomposition of solid waste were investigated in a batch reactor with hydrogen peroxide over the temperature range of 673-823 K. (Mizuno et al. 2000). SC-water is very reactive, corrosive, and miscible with air and oxygen. An industrial process was describes the use of SC water to treat aqueous solutions containing organic compounds (Haas et al. 1989). The operation of a process based on SCF technology was described to treat waste of recombinant fermentation (Krishna et

The rapid expansion of SCF is a promising new technology for particle formation and distribution of biodegradable polymeric (Debenedetti et al. 1993). Because of the extreme fragility of organic aerogels attempts are made to develop inorganic aerogels. Such microcellular polymers foams can be obtained directly by polymerization in a near critical diluent and SC drying in the same reactor vessel. In polymer industry, polymerization is stopped by adding a termination agent. The polymer solution was contacted with superheated steam to remove unreacted monomer and polymerization solvent (de solvent process). SC-CO2 extraction can be alternative for the de-solvent process of polymer solutions. In fact SC-CO2 can reduce the drying process due to its capability of complete recovery by depressurizing. In addition, SC-CO2 can dissolve the typical polymerization solvents, *n-*hexane or toluene at higher pressures. The design of the de-solvent process requires quantitative information on the distribution of organic solvent between the

Liposomes are non-toxic (mostly) and effective in encapsulation (Mortazavi et al. 2007) and controlled release in food industry (Mozafari and Khosravi-Darani 2008). Manufacturing of liposome by SCF covers three separate methods including: (i) phospholipids solvation in a near critical fluid, mixture with a protein containing buffered solution (ii) decompression of solvated phospholipids prior to injection to solution, (iii) the critical fluid decompression technique in which phospholipids are first hydrated in an aqueous buffer, mixed with SCF, with the mixture being then submitted to decompression. Several parameters can improve the characteristics of the liposomes prepared with SCF ethane. Optimization studies would be necessary to examine whether liposomes of higher quality can be made using SCF

SC antisolvent method has great potential for processing of pharmaceuticals (Mosqueira et al. 1981; Steckel et al. 1997) and labile compounds such as proteins (Debenedetti et al 1993; Winters et al. 1999; Yeo et al. 1994; Yeo et al. 1993) and to obtain various morphologies of biopolymers (Bleich et al. 1996; Debenedetti et al. 1993; Dixon and Johnstone 1993;

polymer solution and the SC-CO2 phase (Inomata et al. 1999).

technology. Also, other SCF should be tested (Frederiksen et al. 1997).

**9.5 Production of different morphologies of biocompatible polymers** 

**9.3 Waste treatment** 

al.1986).

**9.4 Particle formation** 

**9.4.1 Preparation of liposome** 

appropriate by hydrolases (Giebauf et al., 1999). In continuous reaction of acidolysis of triolein with stearic acid, the constants of the reaction and mass transfer such as rate constant, solubility, effective diffusivity, mixing diffusivity and mass transfer coefficient depend on temperature, pressure and flow velocity (Nakamura, 1990).

Immobilized *Candida antarctica* lipase B was successfully used as catalyst to synthesize butyl butyrate from butyl vinyl ester and 1-butanol in SC-CO2) with excellent results. A clear enhancement in the synthetic activity and selectivity was observed with the decrease in fluid density for both liquids and SC-CO2 media (Lozano et al., 2004). Also a commercial solution of free *Candida antarctica* lipase B (Novozyme 525L) was immobilized by adsorption onto 12 different silica supports modified with specific side chains (e.g. alkyl, amino, carboxylic, nitrile, etc.). The best results were obtained for the supports modified with non-functionalized alkyl chains and when the in water activity increased from 0.33 to 0.90. Immobilized derivatives coated with ionic liquids clearly improved their synthetic activity in SC-CO2 by up to six times with respect to the hexane medium (Lozano et al., 2007).

*Pseudomonas cepacea* lipase (PCL) was used to catalyze the trans-esterification reaction between 1-phenylethanol and vinyl acetate in SC-CO2. The catalytic efficiency of enzyme enhances by increasing pressure. Moreover SC sulphur hexafluoride (SCSF6) was used as reaction medium. Results showed high stability of the enzyme in this SC medium in comparison to those achieved in SC-CO2 (Celia et al. 2005).

Thermal stability of proteinase of *Carica papaya* was tested at atmospheric pressure, SC-CO2, nearcritical propane and dimethyl-ether. In SC-CO2 at 300 bar thermal activation of the enzyme was improved in the comparison to ambient pressure. Activity of the enzyme decreased in propane and dimethyl-ether (300 bar). Addition of water in the system increased activity, which was incubated in SC-CO2 for 24 h (Habulin et al. 2005).

Isoamyl acetate was synthesized from isoamyl alcohol in SC-CO2 by enzymatic catalysis. Among several reactants, including acetic acid and two different acetates, acetic anhydride gave best yields. An esterification extent of 100% was obtained in continuous operation using acetic anhydride (acyl donor) and Novozyme 435 (enzyme) (Romero et al. 2005). Cocoa beans had been subjected to various pod storage periods prior to fermentation were analysed for pyrazines and SCE (Sanagi et al. 1997).

#### **9.1 SCFs: puissant media for the modification of biopolymers**

The use of SCFs media for polymer modification has been demonstrated (Yalpani 1993). Treatment of chitosan mixtures with glucose or malto-oligosaccharides in SC-CO2 afforded the corresponding water soluble imine-linked, branched chitosan derivatives with high degrees of conversion. Treatment of starch, maltodextrins, cellulose acetate, poly(vinyl alcohol) and paper in SC-CO2 and O2 (19:1 v/v) led to the corresponding oxidized materials.

#### **9.2 Gasification of straw**

Bioconversion of lignocellulosics consists of substrate pretreatment by high pressure steam (for fractionation into cellulose, hemicellulose and lignin components), enzymatic hydrolyze, followed by fermentation of the liberated sugars to ethanol. The various technoeconomic models developed by network members were used to identify probable process schemes and determine technical "bottlenecks" (Saddler 1992).
