Self-Assembled Integrated Nanozyme Cascade Biosensor with Dual Catalytic Activity for Portable Urease Analysis.

In this work, a novel nanozyme (Cu@Zr) with all-in-one dual enzyme and fluorescence properties is designed by simple self-assembly. A nanozyme cascade sensor with disodium phenyl phosphate (PPDS) as substrate was first established by exploiting the dual enzymatic activities of phosphatase and laccase. Specifically, phosphatase cleaves the P-O bond of PPDS to produce colorless phenol, which is then oxidized by laccase and complexed with the chromogenic agent 4-aminoantipyrine (4-AP) to produce red quinoneimine (QI). Strikingly, the NH3 produced by the urease hydrolysis of urea can interact with Cu@Zr, accelerating the electron transfer rate and ultimately leading to a significantly improved performance of the cascade reaction. Moreover, the fluorescence at 440 nm of Cu@Zr is further quenched by the inner filter effect (IFE) of QI. Thus, the colorimetric and fluorescence dual-mode strategy for sensitive urease analysis with LODs of 3.56 and 1.83 U/L was established by the proposed cascade sensor. Notably, a portable swab loaded with Cu@Zr was also prepared for in situ urease detection with the aid of a smartphone RGB readout. It also provides a potentially viable analytical avenue for environmental and biological analysis.

Sensitive chemical sensor array based on nanozymes for discrimination of metal ions and teas.

Tea, originating from China, is an important part of Chinese traditional culture. There are different qualities of and producing areas for tea on the market, therefore it is necessary to discriminate between teas in a fast and accurate way. In this study, a chemical sensor array based on nanozymes was developed to discriminate between different metal ions and teas. The indicators for the sensor array are three kinds of nanozymes mimicking laccase (Cu-ATP, Cu-ADP, Cu-AMP). The as-developed sensor array successfully discriminated 12 metal ions and the detection limit was as low as 0.01 µM. The as-developed sensor array was also able to discriminate tea samples. Different kinds of tea samples appeared in different areas in the canonical score plot with different response patterns. Furthermore, in a blind experiment, we successfully discriminated 12 samples with a 100% accuracy. This sensor array integrates chemistry and food science together, realizing the simultaneous detection of several kinds of teas using a sensitive method. The as-developed sensor array would have an application in the tea market and provide a fast and easy method to discriminate between teas.

Fabrication of superior laccase-mimicking enzyme with catalytic oxidative and photothermal properties for anti-bacterial and dual-mode glutathione S-transferase monitoring.

A novel laccase mimic enzyme Cu-Mn with excellent photothermal properties was firstly prepared via a combination of hydrothermal and in situ synthesis. Cu-Mn nanozymes could catalyze the typical laccase substrate 2,4-dichlorophenol (2,4-DP) to generate the red quinone imine. Further, loading the MnO2 nanosheets with photothermal properties, Cu-Mn nanozymes possessed not only excellent laccase catalytic activity, but also high photothermal conversion efficiency. The presence of glutathione S-transferase (GST) recovered the glutathione (GSH)-induced weakness of the laccase activity and photothermal properties of Cu-Mn. Hence, a GST enzyme-regulated dual-mode sensing strategy was established based on Cu-Mn nanozymes. The detection limits of GST monitoring based on colorimetric and photothermal methods were 0.092 and 0.087 U/L with response times of 20 min and 8 min, respectively. Furthermore, the proposed method enabled the measuring of GST levels in human serum and was successfully employed in the primary evaluation of hepatitis patients. Another attraction, the impressive photothermal behavior also endowed the Cu-Mn nanozymes with promising antimicrobial properties, which exhibited significant antimicrobial effects against Escherichia coli (E.coli) and Staphylococcus aureus (S.aureus). Unsurprisingly, multifunctional Cu-Mn nanozymes certainly explore new paths in biochemical analysis and antimicrobial applications.

2-Methylimidazole-doped nanozymes with enhanced laccase activity for the (+)-catechins detection in dairy products.

In this work, 2-methylimidazole (MI) was doped into the Bpy-Cu nanozyme as a second ligand to form a novel laccase-mimicking enzyme (MI-Bpy-Cu). By comparison, MI-Bpy-Cu nanozyme was identified to have excellent laccase-mimicking activity, high stability and catalytic kinetic properties. It may be that the incorporation of 2-methylimidazole helped the nanozymes to build a structure closer to natural laccase and accelerate the electron transfer rate, thereby achieving the purpose of enhancing the activity. Furthermore, we observed that MI-Bpy-Cu nanozymes could oxidize the colorless (+)-Catechin to generate a yellow radical product, and a novel colorimetric sensing strategy for (+)-Catechin was successfully developed. The method had excellent selectivity and anti-interference properties, and also had good application in the analysis of (+)-Catechin in dairy products.

A novel anthocyanin electrospun film by caffeic acid co-pigmentation for real-time fish freshness monitoring.

In recent years, the use of intelligent and efficient food freshness indicators (FFIs) for monitoring food freshness has been studied widely. In this work, we employed polyacrylonitrile as polymer, blueberry anthocyanins as an indicator, and caffeic acid as a co-pigment and fabricated a novel colorimetric sensing film for real-time monitoring the freshness of fish. The total volatile basic nitrogen (TVB-N) level is one of the potential indicators to evaluate meat freshness. Visual observation confirmed that the polyacrylonitrile-anthocyanin-caffeic acid film changed from pink to light purple, and then to dark purple providing a good indication of spoilage, which correlated well with the TVB-N content and pH values in fish. It is because the volatile ammonia combined with water to form NH3·H2O, and then NH3·H2O is hydrolyzed to form OH- and NH4+. The change of the polyacrylonitrile-anthocyanin film was caused by OH-. Compared with the polyacrylonitrile-anthocyanin film without caffeic acid, the addition of the caffeic acid film had enhanced significantly ammonia responsiveness with a total color difference value of 29.897. And it was also observed that caffeic acid obviously improved the storage stability of the film. This study provided a reference for detecting food freshness using co-pigmentation and electrospinning encapsulation technology in combination.

[Identification of Daturae flos and its adulterants based on DNA barcoding technique].

To identify the original plant of Daturae Flos from its adulterants by DNA barcoding, the sequences of ITS2, psbA-trnH, matK, rbcL of four species including Datura metel, Darura innoxia, Darura stramonium and Brugmansia arborea were compared and analyzed. The PCR and sequencing success rate of the four regions (ITS2, psbA-trnH, matK, rbcL) was 100%, 90%, 100% and 85%, respectively. Sequences were assembled with CodonCode Aligner. K2P distances were calculated and NJ tree was performed by MEGA 4.1. Thirty SNPs were found among ITS2 sequences, and 33 insert/deletes were found among psbA-trnH intergenic regions. The interspecific K2P distance of ITS2 and psbA-trnH was obviously higher than that of the intraspecific one. As to matK and rbcL, there was no "Barcoding Gap" existing between inter- and intra-specific distances. The NJ trees of the four regions/combinations were built separately. Samples of Brugmansia arborea were clustered into one clade, and the other species of Datura L. formed another clade. The results showed that either ITS2 or psbA-trnH was useful to identify Daturae Flos from its adulterants.

A novel selective detection method for sulfide in food systems based on the GMP-Cu nanozyme with laccase activity.

A selective and sensitive colorimetric strategy for sulfide analysis was developed using GMP-Cu nanozymes with a laccase-like activity. This research discovered for the first time that sulfide could significantly enhance the catalytic activity of the GMP-Cu nanozymes by about 3.5 folds. The enhanced laccase activities duo to two reasons. First, Cu2+ in GMP-Cu nanozymes was reduced to Cu+. The other reason was the formation of Cu-S bond which was beneficial to accelerate the electron transfer rate to improve catalytic activity. Therefore, this method showed an excellent selectivity for sulfide. And it had a linear relationship in the sulfide concentration range of 0-220 µmol/L with a detection limit of 0.67 µmol/L. Furthermore, the proposed method was successfully applied to examine sulfide in the food systems. This new method may be used in sulfide detection to improve food quality and safety.