Letter Cite This: ACS Sustainable Chem. Eng. 2018, 6, 9572−9578
Porous Starch-Based Material Prepared by Bioextrusion in the Presence of Zinc and Amylase−Magnesium Complex Enbo Xu,†,§ Zhengzong Wu,‡ Jie Long,† Aiquan Jiao,† and Zhengyu Jin*,† †
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The State Key Lab of Food Science and Technology, School of Food Science and Technology, Joint International Research Laboratory on Food Safety, Jiangnan University, 1800 Lihu Road, Wuxi 214122, China ‡ School of Food Sciences and Engineering, Qilu University of Technology, 3501 University Road, Jinan 250353, China § Whistler Carbohydrate Research Center, Purdue University, 745 Agriculture Mall Drive, West Lafayette, Indiana 47907, United States ABSTRACT: A simple and sustainable preparation route for the creation of macropores in extruded starch was developed. Zinc ions were exogenously added into native corn starch, and then the Znmodified starch was mixed with an enzyme (thermostable α-amylase)− magnesium complex as the biocatalyst prior to extrusion. The porous bioextrudates were characterized using SEM, XRD, FT-IR and water/oil absorption analyses. Results showed that the average pore numbers were increased (97 to 201 in the 160× SEM images), whereas the pore sizes (∼10−20 μm) were almost unchanged with the amylase concentration increased from 0.04 to 0.12%. The crystallinities of the porous extrudates were decreased to 7.9−10.2% compared to that of the native counterpart (21.1%), but their structures all exhibited traces of rigidification. Furthermore, the water absorption capacities were greatly increased (∼450−500%) while the oil absorption capacities were slightly increased (∼40%). Overall, it extends the preparation and the potential application of porous starch in the field of materials. KEYWORDS: Porous biomaterial, Starch, Bioextrusion, Zinc, Enzyme−magnesium complex
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induced pores).6,9 However, most of bio-PSs are deficient in structural strength, thermal resistant or special adsorptivity. It was then studied that the remodification of bio-PS such as cross-linking or grafting, where chemical reagents (e.g., hydroxyapatite (HA),10 ZrOCl211 and N,N′-methylene-bisacrylamide (MBAA),12 etc.) were introduced. A total chemical synthesis route can be used to fabricate goal-oriented PS (chemical-PS is used here for the sake of conciseness) avoiding the chemical reprocessing of bio-PS. However, the chemical method using a mass of organic reagents (even more than starch substrate) has a limitation for both environmental and economic concerns. Furthermore, the porous structure in the transformable networks of chemical-PS, mainly caused by special reagents (e.g., mercaptosuccinic acid (MSA), poly(ethylene-alt-maleic anhydride) (PE-alt-MA) and poly(acrylic acid) (PAA)),13−15 is totally different from that of bio-PS. For physical methods, they are unable yet to create deep/hollow holes inside starch.7 Extrusion is a physical (or thermomechanical) process of converting materials through continuous unit operations of mixing, kneading, shearing and heating under high pressure.16 So far, extruder is not suitable for fabricating PS due to its
INTRODUCTION Starch widely present in plants is a natural biopolymer composed of linear and branched polysaccharide chains, and is usable in food and nonfood fields due to the superiorities of its edible, biodegradable, cost competitive and easy available/ modifiable characteristics.1 There are growing interests of the green and sustainable utilizations of starches as a renewable and eco-friendly resource.2,3 In that sense, one of the effective means is to fabricate porous starch (PS) and PS derivatives by the modification of their native counterparts. In general, PS has a specific hollow structure, a large surface area, and homogeneous/heterogeneous pores. Therefore, PSs are functional biomaterials and have been used as adsorbents, carriers or structure-directing agents for food, agriculture, pharmaceuticals, cosmetics, paper and tissue engineering, etc.4−6 To date, the ways to prepare PSs (mainly by biological, chemical, or physical methods) have made them extremely different in pore-size distributions and physicochemical/ thermal/functional properties. The most common method is to biologically hydrolyze native starches by one or more amylases at subgelatinization temperature (∼50 °C) in an abundant-water system.5,7,8 For this bio-PS, α-amylase is the major biocatalyst to create pores, which is often assisted by a small amount of glucoamylase/amyloglucosidase (whereas cyclodextrin-glycosyltransferase, branching enzyme or βamylase showes almost no or less formation of enzyme© 2018 American Chemical Society
Received: May 9, 2018 Revised: July 14, 2018 Published: July 17, 2018 9572
DOI: 10.1021/acssuschemeng.8b02128 ACS Sustainable Chem. Eng. 2018, 6, 9572−9578
Letter
ACS Sustainable Chemistry & Engineering
Figure 1. SEM images of native (a), bioextruded (b), Mg-bioextruded (c) and Zn/Mg-bioextruded (d) corn starches. Magnification 600×. White box for panel c means magnification 2400×. limit the absorption of Mg2+ into starch granule where there is a competition for the binding of Mg2+ onto the active center of TAM.19 Another novel approach is to simply modify native CS as follows: 250 g of native CS was agitated in 3000 cm3 of 1% aqueous solution of zinc chloride with regular shaking at room temperature for about 8 h; then the sample was filtered off, washed one time with water, and dried in an oven at 40 °C; afterward, Zn-modified CS was mixed with the solution of amylase-Mg complex and extruded under the same operating conditions. Three different levels of enzyme, 0.04, 0.08 and 0.12% (db of starch) were used in the presence of magnesium sulfate (2 mmol/100g starch), and the bioextrudates were labeled as BEPS0.04%, BEPS0.08% and BEPS0.12%, respectively. A Quanta 200 field emission scanning electron microscope (SEM, Thermo Fisher scientific, MA, USA) was used to determine the formation of pores. A D2 PHASER diffractometer (BRUKER AXS GMBH, BadenWuerttemberg, Germany) was used to determine X-ray diffraction (XRD) patterns. A Thermo Nicolet iS 10 FT-IR Spectrometer (Thermo Electron Corp., WI, USA) was used to analyze Fourier transform-infrared (FT-IR) spectra. Water and oil absorption capacities were measured according to the method of Qian et al.5
strong shear stress and high pressure producing a compact extrudate. Given a starch material, it was only observed the bubble/foam with a visible appearance caused by steaminduced (over 100 °C as water flashing off) or CO2-induced (60−70 °C) expansion, which favors the production of cellular-structural and crispy food rather than PS.17,18 When amylase is introduced into an extrusion process (i.e., bioextrusion), the reaction route of enzyme (radiative to starch melt around) differs than the traditional one in a batch (concentrated at starch surface), which indicates a potential to create pores in extrudates. However, it is more difficult to construct a supporting structure to ensure the formation of pores during bioextrusion, where a rapid liquefaction of starch occurs. Herein, we attempted to use zinc ions (with less impact on amylase activity among transition metals)19,20 to strengthen the starch backbone, and use magnesium ions to stabilize the enzyme for preparing PS by extrusion method.
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EXPERIMENTAL SECTION
RESULTS AND DISCUSSION It is feasible to create a uniform distribution of macropores inside the restructured and densified starch extrudate by bioextrusion. SEM analysis indicates the porosity of bioextrudate when starch was modified by appropriate zinc ions prior to extrusion and a TAM enzyme was used in the presence of magnesium during extrusion. As shown in Figure 1a, native CS granule had a smooth surface and an irregular polygonal shape, with an average size of approximately 10−15 μm. A compact structure of bioextruded CS (milled through 100-mesh sieve) was observed resulting from the synergetic actions of thermomechanical stress and enzyme (Figure 1b). During bioextruison processing, starch was subjected to a shearing, heating and hydrolyzing
250 g of native corn starch (13.5% water content) was mixed with an enzyme solution containing 82.7 mL of deionized water and 62 μL of thermostable α-amylase (TAM, 120 000 U/g, with a density of 1.2 g/ mL). The mixture was stored at 4 °C overnight for the equilibrium of water and enzyme distribution, and extruded at 2.5 kg/h feed rate, 150 rpm screw speed, and 50−60−65−88 °C temperature profile by a twin-screw extruder (TSE 24 MC, Thermo Fisher scientific, MA, USA). Enzyme hydrolysis prior to extrusion can be considered negligible under low temperature.21 The bioextrudate was freezedried, milled and determined as the control group. For a novel solution of enzyme−metal complex, magnesium sulfate (2 mmol/ 100g starch) was added in the enzyme solution above at room temperature for 2 h. It was blended with corn starch (CS)22 according to the aforementioned proportion, and extruded under the same conditions. The use of sulfate anions with high charge density was to 9573
DOI: 10.1021/acssuschemeng.8b02128 ACS Sustainable Chem. Eng. 2018, 6, 9572−9578
Letter
ACS Sustainable Chemistry & Engineering
Figure 2. Schematic illustration of Zn/Mg ions assisting a bioextrusion process (using thermostable α-amylase) to create macropores in starch.
salts, is compressed, sheared and micromixed to small fragments during extrusion (Label ② in Figure 2). Given the heterogeneous distribution of Zn ions in starch matrix,30 the fragments of starch may be cross-linking and enzyme aggregation may be obtained in local areas where there is more zinc present. In subsequent processing section (Label ③), pores gradually grew up in the molten starch (from the center of “enzyme-aggregating zone” to around) due the enzymatic degradation of starch. Meanwhile, the crystalline region and/or amorphous region of starch bound with zinc resist to enzymatic hydrolysis, and conversely become the supporting structure to avoid the collapse of those pores (Label ④). Figure 3 indicates the porous morphologies of BEPS samples. Deep holes (those blue arrows in Figure 3) were observed, which is similar to that of traditional bio-PS but with larger average sizes. When a lower concentration of enzyme was used in a batch, the pore size of bio-PS was bigger and more like craters instead of deep holes as reported by Benavent-Gil and Rosell.6 However, in the presence of zinc and magnesium, TAM at higher levels had no prominent influence on pore sizes but obviously increased pore numbers in extrudates (the average numbers of 97, 174 and 201 in the 160× SEM images of BEPS0.04%, BEPS0.08% and BEPS0.12%, respectively). It may be considered the different mechanisms of pore formation between bio-PS and BEPS. The pore size of BEPS should be associated with the scale of “enzymeaggregating zone” (maybe controlled by the types of materials/enzymes/salts) and the mixing performance of feedstock during extrusion. The macromolecule structure of PSs (both bio- and chemical-PS) is weak and liable to collapse limiting its extended applications in food and nonfood fields. For instance, the bio-PS loaded by zirconium showed huge cracks though it had a good ability relating to the adsorptive removal of fluoride.11 Similar cracks were also observed in the starch extrudate with foams visible to the naked eye.18 However, the structure of BEPS seemed to be reinforced, and an increase of structural rigidity might be achieved as
history, and then gelatinized, reorganized, compressed and densified. No significant liquefaction occurred in the bioextrusion of CS, possibly due to the relatively low level of TAM enzyme (0.04%).16 Indistinct pits/holes (white arrows in Figure 1c) were found when magnesium salt was involved (Mg-bioextruded CS), and it may be caused by a certain link between starch and enzyme showing point-attack.19,20 This phenomenon is similar to a “corrosion-type” process but it created no deep holes, the structure of CS against mechanical stress was too weak to act as the backbone. Interestingly, with the further introduction of zinc into native starch feedstock, it was clear that some macropores were formed in BESP0.04% (green circles in Figure 1d). The average size of the pores (∼10−20 μm) in BEPS0.04% is larger than that of traditional bio-PS (
