Lipase Catalyzed Transesterification of Model Long-Chain Molecules in Double-Shell Cellulose-Coated Oil-in-Water Emulsion Particles as Microbioreactors

Abstract

Lipase-catalyzed transesterification is prevalent in industrial production and is an effective alternative to chemical catalysis. However, due to lipases' unique structure, the reaction requires a biphasic system, which suffers from a low reaction efficiency caused by a limited interfacial area. The use of emulsion particles was found to be an effective way to increase the surface area and activity. This research focuses on cellulose as a natural surfactant for oil-in-water emulsions and evaluates the ability of lipase, introduced into the emulsion's aqueous phase, to integrate with the emulsion microparticles and catalyze the transesterification reaction of high molecular weight esters dissolved in the particles' cores. Cellulose-coated emulsion particles' morphology was investigated by light, fluorescence and cryogenic scanning electron microscopy, which reveal the complex emulsion structure. Lipase activity was evaluated by measuring the hydrolysis of emulsified p-nitrophenyl dodecanoate and by the transesterification of emulsified methyl laurate and oleyl alcohol dissolved in decane. Both experiments demonstrated that lipase introduced in the aqueous medium can penetrate the emulsion particles, localize at the inner oil core interface and perform effective catalysis. Furthermore, in this system, lipase successfully catalyzed a transesterification reaction rather than hydrolysis, despite the dominant presence of water.

Keywords: cellulose; emulsions; lipase; transesterification.

Figure 1

Light microscopy images (phase contrast) of cellulose-coated n-decane emulsion microparticles at cellulose:decane wt. ratios of: (A) 1:1, (B) 1:3, (C) 1:5 and (D) 1:8.

Figure 2

Fluorescence microscopy images of a 1:8 wt. ratio cellulose:decane emulsion stained with: (A) calcofluor white, preferentially staining cellulose, and (B) Nile red, preferentially staining the decane core.

Figure 3

Cryo-SEM images of 1:8 (wt. ratio) cellulose:decane emulsion after cryo-fracturing. (A) Lower magnification exhibiting a few microparticles: whole (black arrow), partially fractured (white arrow) and fully fractured (white circle). (B) The two-shell encapsulation structure revealed at higher magnification imaging of the fully fractured emulsion particle: a thick porous inner shell (white arrow) with a thin and more compact outer shell (black arrow).

Scheme 1

Experimental scheme designed to evaluate lipase activity in the emulsion’s inner core interface. Lipase-catalyzed hydrolysis: (a) in the emulsion and (b) in the aqueous medium.

Figure 4

Lipase hydrolysis activity in 1:1, 1:3, 1:5 and 1:8 cellulose:decane emulsions. Comparison of lipase addition before and after centrifugation (Scheme 1a,b, respectively) and to lipase activity in an aqueous medium (control).

Figure 5

Illustration of the reaction sequence for lipase transesterification. Dark yellow—oil core. Gray cellulose outer shell. Light blue—water phase.

Figure 6

Concentrations of products from a lipase-catalyzed reaction in emulsion droplets comprising methyl laurate and oleyl alcohol. An analysis was performed using GC-MS. Bars represent the average results of the concentration, whereas the error bars represent the standard deviation.