Immobilization And Catalytic Activity Of Porcine Pancreatic Lipase On SBA-15 Mesoporous Material

Enzymes catalytic reactions have high chemo-, regio-, and stereo-selectivity. Unfortunately, they are not always suited to industrial or medical applications. They are often unstable and show low activity in organic solvents or at high temperatures. Denaturation of the enzyme, which destroys its catalytic activity, can often be induced by pH and mechanical or thermal treatment as well as the composition of the adsorbent. This is uneconomical as active enzyme being lost. These disadvantages of enzymes can be overcome by immobilization of the enzyme onto solid supports. Immobilization of enzymes on inorganic materials ensures reusability of enzymes, minimizes the cost of production, and overcomes the economic drawback associated with their use. Immobilization also provides operational flexibility and improves enzymes thermal and chemical stability under extreme conditions.Rod-like SBA-15 mesoporous material was synthesized using amphiphilic Pluronic polymer surfactant P123 (EO20PO70EO20) as structure-directing agent. The sample was characterized by SEM, TEM, SAXRD, N2 adsorption-desorption isotherms and FT-IR spectra. Static conditions resulted in the formation of the monodispersed rod-like material with a relatively uniform size of ca. 0.3μm in width and 1–2μm in length. It had a high specific surface area of 490 m2g–1, a total pore volume of 1.04 cm3g–1 and a pore diameter of 7.63 nm. The pore diameter was large enough for accommodating Porcine pancreatic lipase (PPL, 4.6×2.6×1.1 nm3) molecules inside the channels. The physical adsorption immobilization of PPL on the rod-like SBA-15 mesoporous material in buffer solutions was studied via a direct one-step immobilization method. The results obtained by SAXRD and N2 adsorption-desorption isotherms clearly showed that PPL was adsorbed into the channels of SBA-15 and the ordered mesoporous structure of SBA-15 after adsorption of PPL was retained. The catalytic activity, thermal stability, and reusability of the immobilized PPL were measured in phosphate buffer solution by hydrolysis of triacetin. The immobilized PPL showed excellent adaptability at higher pH and better thermal stability comparing with free PPL. The immobilized PPL had a good thermal stability when it was incubated at 60 oC. The relative activity of immobilized PPL on rod-like SBA-15 has been found to be 32% of its original activity after the fifth reuse due to leakage of lipases from the channels of the mesoporous material. Fiber-like SBA-15 material with highly ordered mesostructures have been prepared by a one-step hydrothermal synthesis method of tetraethoxysilane (TEOS) in an acidic solution using surfactant P123 as structure-directing reagent. The prepared materials were characterized by TEM, SAXRD, FT-IR and N2 adsorption-desorption experiments. SBA-15 has a fiber-like morphology, by stacking and coupling of rod-like particles of which respective length and width are ca. 1μm and ca. 0.5μm. An immobilized enzyme had been prepared by incorporation of PPL in the channels of fiber-like SBA-15. The physical adsorptions of PPL on the fiber-like SBA-15 mesoporous material in buffer solution with different pH values (pH = 5–10) and times (0–36 h) had been studied. A high lipase loading (926 mg enzyme per gram silica) can be obtained, but it disagreed with the high catalytic activity. The adsorbed maximum activities were observed at pH = 6.0 and 3 h. The optimal pH of the hydrolysis of triacetin for the immobilized and free PPL was at 7.0. The immobilized PPL showed much more excellent adaptability of the hydrolysis of triacetin compared to free PPL during pH = 6.0–9.0. Meanwhile, the thermal stability of the catalyst and its reusability were tested by performing subsequent reaction cycles of hydrolysis of triacetin. The immobilized PPL had a good thermal stability when it was incubated at 60 oC. The relative activity of immobilized PPL hold over 42% of the initial hydrolysis activity after reaction 2 h, while that of the free PPL remained only 28% of the initial activity after reaction 1 h.The activity of the immobilized PPL fell off rapidly to be 40% of its original activity after five successive batch reactions.