静电纺丝法制备ZnO构建酪氨酸酶生物传感器检测邻苯二酚

电化学生物传感器在环境监测、生物与食品分析等领域应用广泛。本文采用静电纺丝法制备了氧化锌微纳米纤维材料，并负载酪氨酸酶（Tyr）构建了检测邻苯二酚的生物传感电极。利用X射线粉末衍射（XRD）、扫描电子显微镜（SEM）表征了氧化锌微纳米纤维的形貌结构，采用循环伏安法（CV）和计时电流法（IT）优化了检测邻苯二酚的工作条件。结果表明，制备的Tyr/ZnO/CS/GCE生物传感电极具有良好的电化学性能，在反应过程中为扩散控制，对邻苯二酚的检测在5～50μmol/L的浓度范围内具有较好的线性关系，最低检测限为1.9041μmol/L，灵敏度为376.31μA/(mmol·L·cm²)，氧化锌的加入可增强酪氨酸酶的稳定性，在尿素、多巴胺和葡萄糖3种电化学活性相近物质存在的情况下仍对邻苯二酚的检测有较好的选择性，且具有良好的循环稳定性。     

Discovery and characterization of tyrosinases from sea anemone pedal disc

Abstract

The formation of underwater adhesion is a complicated physiological process and many different types of enzymes are found to be essential apart from structural proteins. Previous studies have shown that various tyrosinases were present in marine adhesives, but little information is available about the over-expression and enzymatic characterization of these enzymes. Specifically, this study first identified four significantly up-regulated tyrosinases in the pedal disc of Haliplanella luciae by means of multi-omics technology, and made preliminary bioinformatics predictions. Sequence alignment showed that the Tyr1_Hl contained six conserved His residues that bind to copper ions, of which a tyrosinase with diphenolase activity named as Tyr1_Hl^Δ, was expressed in Escherichia coli BL21 (DE3) cells and purified by affinity chromatography. Enzymatic characterization showed that the activity of Tyr1_Hl^Δ was Cu²⁺ dependent and maximum catalytic activities were in 20?mM Tris–HCl (pH 8.0) at 37?°C. In summary, we identified novel tyrosinases in the pedal disks of sea anemone for the first time and the Tyr1_Hl^Δ was successfully recombinant expressed. Our study will provide basis for future exploration of bio-adhesion mechanism and design of bio-adhesives derived from sea anemones.     

Simultaneous recovery of glycyrrhizic acid and liquiritin from Chinese licorice root (Glycyrrhiza uralensis Fisch) by aqueous two-phase system and evaluation biological activities of extracts

Aqueous two-phase system (ATPS) was used for simultaneous purification of glycyrrhizic acid (GA) and liquiritin (LQ) from crude extract of Chinese licorice root. It was revealed that 87% GA and 94% LQ were retrieved in the ATPS top phase, under the optimum conditions of 25% (w/w) ethanol, 30% (w/w) K₂HPO₄ and 4% (w/w) loading sample at 10–40°C. Compared with crude extract, the ATPS top-phase extract exhibited the highest antioxidative activity, but no tyrosinase inhibitory effect. Whereas, the ATPS bottom-phase extract was proved to be effective ABTS radical scavenger and tyrosinase inhibitor, suggesting the potency of the alcohol-salt ATPS purification for the different medicinal purposes.     

具有酪氨酸酶抑制作用的酵母多肽的分离纯化及分子量分布

为确定具有较好酪氨酸酶抑制作用的酵母多肽的分子量,对酵母多肽粗制品溶液以截留相对分子量为10 KDa、5 KDa、1 KDa的超滤膜进行超滤,采用生化酶法对不同超滤组分进行酪氨酸酶抑制活性检测。结果显示,各多肽组分的活性随相对分子量的减少而增大,活性多肽的相对分子量主要集中在1 KDa以下。对相对分子量1 KDa的多肽组分依次以DEAE-52阴离子交换柱层析法和Sephadex G-15凝胶层柱析法进行分离,最后得到4个洗脱组分,分别富集各洗脱组分并进行酪氨酸酶抑制活性研究。结果表明,组分d-1活性最佳,其IC50值为3.53 mg/mL。经高效液相空间排阻色谱分析,d-1组分多肽的相对分子质量主要集中在200~800 Da。