【摘要】：本研究內(nèi)容隸屬于國家“十二五”科技支撐計(jì)劃項(xiàng)目《乳清蛋白應(yīng)用技術(shù)與開發(fā)(2013BAD18B07)》。本研究以乳清蛋白為原料,采用轉(zhuǎn)谷氨酰胺酶酶交聯(lián)作用,在中性條件下與七水硫酸鋅結(jié)合,再通過乙醇去溶劑化法進(jìn)一步優(yōu)化制備乳清蛋白-鋅納米微粒。通過對(duì)制備條件的優(yōu)化,得到裝載率較高、穩(wěn)定性及各方面性質(zhì)良好的乳清蛋白-鋅納米微粒。隨后,本文對(duì)制備的乳清蛋白-鋅納米微粒的二級(jí)結(jié)構(gòu)、熱變性、外觀形態(tài)學(xué)特性、體外消化特性、細(xì)胞毒性以及貯藏穩(wěn)定性等進(jìn)行了研究。具體研究結(jié)論如下:(1)在酶交聯(lián)法制備乳清蛋白-鋅納米分散液的技術(shù)研究中,本研究分別考察了乳清蛋白濃度、交聯(lián)時(shí)間、加酶量、鋅加入量對(duì)制備的乳清蛋白-鋅納米分散液的粒徑、Zeta電位、裝載率、單位蛋白載鋅量及濁度等因素的影響。結(jié)果顯示:當(dāng)乳清蛋白濃度為5.0%、交聯(lián)時(shí)間為4 h,加酶量為5.0 U/g、鋅加入量為5.0 m M時(shí),制備的乳清蛋白-鋅納米微粒的效果較好,其粒徑大小為100.76±2.17nm,Zeta電位為-29.11±1.96 m V,裝載率可達(dá)到54.56±3.19%,單位蛋白載鋅量為8.28±0.48μg/mg。(2)在乙醇去溶劑化法優(yōu)化乳清蛋白-鋅納米微粒的制備技術(shù)研究中,本研究分別考察了蛋白與乙醇加入比、鋅加入量及加酶量對(duì)制備的乳清蛋白-鋅納米微粒的粒徑、裝載率及單位蛋白載鋅量的影響。結(jié)果顯示:當(dāng)?shù)孜锱c乙醇加入體積比為1:4,鋅加入量為5.0 m M,加酶量為5.0 U/g時(shí),制備的乳清蛋白-鋅納米微粒的效果較好,其粒徑大小為150.83±5.15 nm,裝載率可達(dá)到99.22±3.31%,單位蛋白載鋅量為10.43±0.27μg/mg。(3)在對(duì)酶交聯(lián)及乙醇去溶劑化法制備的乳清蛋白-鋅納米微粒的特性進(jìn)行研究中,結(jié)果顯示:酶交聯(lián)及乙醇去溶劑化處理后乳清蛋白的二級(jí)結(jié)構(gòu)發(fā)生了改變。形成的乳清蛋白-鋅納米微粒在掃描電鏡下呈現(xiàn)圓球狀,粒度大小為100 nm左右,分布較為均勻。乳清蛋白經(jīng)上述方法制備形成納米微粒后,其熱變性有所提高。乳清蛋白-鋅納米微粒在模擬胃液中鋅的釋放率較低,而在腸液中較高。細(xì)胞毒性試驗(yàn)結(jié)果表明,當(dāng)乳清蛋白-鋅納米微粒濃度為0.1875 mg/m L時(shí),對(duì)細(xì)胞作用24 h,乳清蛋白-鋅納米微粒對(duì)細(xì)胞無潛在毒性作用。貯藏期試驗(yàn)表明,制備的乳清蛋白-鋅納米微粒在25℃條件下,14 d內(nèi)的穩(wěn)定性良好。本研究表明通過轉(zhuǎn)谷氨酰胺酶的交聯(lián)及乙醇去溶劑化相結(jié)合的方法能夠制備一種鋅負(fù)載率較高、穩(wěn)定性良好的乳清蛋白-鋅納米微粒,這有可能為解決鋅吸收率低的問題提供了一種新的思路。
[Abstract]:The content of this study belongs to the National Science and Technology support Project of the Twelfth Five-Year Plan, whey protein Application Technology and Development (2013BAD18B07). In this study, whey protein was used as raw material, transglutaminase crosslinking was used to combine with zinc sulfate heptahydrate under neutral conditions, and then further optimized the preparation of whey protein-zinc nanoparticles by ethanol desolvation. By optimizing the preparation conditions, high loading rate, good stability and various properties of whey protein-zinc nanoparticles were obtained. Subsequently, the secondary structure, thermal denaturation, appearance morphology, in vitro digestion, cytotoxicity and storage stability of the prepared whey protein-zinc nanoparticles were studied. The specific conclusions are as follows: (1) in the preparation of whey protein-zinc nano-dispersion by enzyme crosslinking, the concentration of whey protein, the crosslinking time and the amount of enzyme were investigated. The effects of zinc addition on the particle size, Zeta potential, loading rate, zinc load per unit protein and turbidity of the prepared whey protein-zinc nanoparticles were investigated. The results showed that when the concentration of whey protein was 5.0%, the crosslinking time was 4 h, the amount of enzyme added was 5.0 U / g, and the amount of zinc was 5.0 mm, the effect of the prepared whey protein-zinc nanoparticles was better. The particle size is 100.76 鹵2.17nmGV, the Zeta potential is -29.11 鹵1.96mV, and the loading rate is 54.56 鹵3.19mV, and the Zeta potential is -29.11 鹵1.96mV. Zinc content per unit protein was 8.28 鹵0.48 渭 g / mg 路(2). (2) in the optimization of preparation of whey protein-zinc nanoparticles by ethanol desolvation, the ratio of protein to ethanol was studied. The effects of zinc addition and enzyme addition on the particle size, loading rate and zinc content per unit protein of the prepared whey protein-zinc nanoparticles. The results show that when the volume ratio of substrate to ethanol is 1: 4, the amount of zinc added is 5.0 mm and the amount of enzyme is 5.0 U / g, the effect of the prepared whey protein-zinc nanoparticles is better, and the particle size is 150.83 鹵5.15 nm,. The loading rate was 99.22 鹵3.31 and the zinc load per unit protein was 10.43 鹵0.27 渭 g / mg 路(3) in the study of the properties of whey protein-zinc nanoparticles prepared by enzymatic crosslinking and ethanol desolvation. The results showed that the secondary structure of whey protein changed after enzymatic crosslinking and ethanol desolvation. The resulting whey protein-zinc nanoparticles were spherical under scanning electron microscope with a particle size of about 100 nm and a uniform distribution. The thermal denaturation of whey protein was improved after the preparation of nano-particles by the above method. The release rate of zinc in simulated gastric juice was lower than that in intestinal fluid. The results of cytotoxicity test showed that when the concentration of whey protein-zinc nanoparticles was 0.1875 mg/m / L, the leukoprotein-zinc nanoparticles had no potential toxicity to cells at 24 h. The storage test showed that the stability of the prepared whey protein-zinc nanoparticles was good at 25 鈩，

本文編號(hào)：2349955