| The importance of enzyme catalysis in chemical manufacturing is widely recognized due to its environment-friendly characteristics, such as mild reaction conditions, high rates and less waste. But the insolublity between organic substrate and bio-enzyme greatly confines the widespread application of enzymes. Emulsification using surfactants as emulsifiers has proven to be a versatile and convenient method. Surfactant-stabilized emulsion systems can increase the interfacial area and mass transfer. However, they may also severely affect the enzymatic activity, and product separation and catalyst recovery is usually a problem. Pickering emulsions (PE), which are emulsions stabilized by nano- to micrometer-sized solid particles (SPs), show the potential to replace the traditional surfactant-based emulsions and is considered to be a typical green biphase reaction system.Firstly, an oil-in-water PE system stabilized by amphiphilic porous carbonaceous microspheres (CMs) was successfully prepared and used in enzyme-catalyzed hydrolysis and esterification reactions. The control of the emulsion droplet size and size distribution were investigated by adjusting the particle content and the dispersed phase volume fraction. More than 99% of the lipase was adsorbed on the interface of the emulsion droplets and showed excellent stability and activity. The highest catalytic efficiency (4.65 μmol/mg min) was achieved at 43.9 m2/L (interfacial area) and 0.57x10-2 g/m2 (interfacial enzyme concentration). For the hydrolysis reaction, the catalytic performance of CRL-PE system was also influenced by agitation speed, olive oil concentration and reaction time; for esterification reaction, the catalytic performance of CRL-PE system was mainly affected by CMs properties, organic solvent and substrate. The effects of CMs mainly include PE emulsion droplet size, lipase distribution and liquid surface tension. The non-polarity of organic solvent and longer chain substrates provide better catalytic efficiency of CRL-PE system. A high hydrolysis yield (100%) and ester yield (99%) can be obtained under the optimum reaction conditions. Additionally, in oil-in-water PE system, CRL showed excellent long-term activity and non-inhibited with high concentration substrate. After 12 circles (x12 h), the catalytic activity of CRL-PE system still reached 81% of the original activity, with pure substrates as the dispersed phase.Secondly, as the similar structure of CMs and PE, a simulated water-in-oil PE system was successfully prepared and TLL was successfully encapsulated on the CMs, which used in lipase-catalyzed esterification and transesterification reactions of organic phase. The water retention of CMs is the key factor affecting the catalyzed efficiency of lipase. For the transesterification reaction, the influence of water retention of CMs was obvious that the highest catalyzed efficiency of lipase was expressed in the highest water retention CM-180-I (4.37%), which ester yield was 40% after 30 min reaction. For the esterification reaction, in the range 1.2%-4.4%, the effects of CMs water retention on lipase catalytic activities were not obvious, which due to the dehydration mechanism of esterification reaction.Finally, combined the PE technology and immobilized enzyme technology, a two-step enzymatic method of preparation biodiesel was successfully established and showed an excllent efficiency. The first step was hydrolysis reaction, which using CRL as catalyst, PE system as the reaction medium,6.75 g rapeseed oil completely hydrolyzed into fatty acid in 3 h under 30 ℃ with the dosage of 2.2 mg CRL/g rapeseed oil. The second step is esterification reaction, which using immobilized lipase (TLL-CMs) as catalyst, in non-solvent medium,1 g fatty acid and methanol completely transformed into biodiesel within 45 min under 40 ℃ with the dosage of 250 mg TLL-CMs (about 95.8 μLTLL). |
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