[Objectives]This study was conducted to investigate the effects of embedding conditions on activity and catalytic properties of immobilized polyphenol oxidase.[Methods]Polyphenol oxidase was immobilized in a polymer m...[Objectives]This study was conducted to investigate the effects of embedding conditions on activity and catalytic properties of immobilized polyphenol oxidase.[Methods]Polyphenol oxidase was immobilized in a polymer material by the embedding method,and the optimal immobilization conditions were obtained by single factor tests:CaCl_(2) concentration 2.0%,sodium alginate concentration 2.0%,immobilization time 2 h and mass ratio of enzyme to carrier 10 mg/100 g,under which the immobilized enzyme activity was 93.33 U/g.Under the above conditions,the properties of polyphenol oxidase immobilized by sodium alginate(A-PPO)and free polyphenol oxidase were studied.[Results]The thermostability of A-PPO was better than that of the free enzyme,but the pH stability of A-PPO was inhibited.The Michaelis constant K_(m) values of free polyphenol oxidase and A-PPO were 0.37 and 0.48 mmol/L,respectively,and the maximum reaction rate V_(max) values were 0.38 and 0.51 mmol/(L·g),respectively.[Conclusions]This study provides a theoretical basis for the study of the properties of polyphenol oxidase.展开更多
Litchi(Litchi chinensis Sonn.)is a type of commercially prevalent subtropical and tropical fruit.Since litchi has a highly heterozygous genetic background and a long reproductive cycle,conventional breeding methods(su...Litchi(Litchi chinensis Sonn.)is a type of commercially prevalent subtropical and tropical fruit.Since litchi has a highly heterozygous genetic background and a long reproductive cycle,conventional breeding methods(such as hybridization)have limited ability to nurture new litchi cultivars.Here,an efficient and stable Agrobacterium tumefaciens-mediated genetic transformation of embryogenic callus was established in‘Feizixiao’litchi.Transgenic materials were verified using polymerase chain reaction(PCR)analysis,β-glucuronidase(GUS)assay,and green fluorescent protein(GFP)assay.To implement the technology of the Clustered Regularly Interspaced Short Palindromic Repeats(CRISPR)/associated protein 9(CRISPR/Cas9)technology in‘Feizixiao’litchi and verify the validity of these transformation systems,the litchi polyphenol oxidase gene(LcPPO,JF926153)was knocked out.Various categories of mutations,covering base insertions,deletions,and substitutions,were found in transgenic materials via sequence analysis.The transformation system achieved high feasibility and efficiency,and the system of CRISPR/Cas9 was successfully employed to edit genes in‘Feizixiao’litchi.This work provides an essential foundation for investigating the functions of genes and accelerating litchi genetic improvement.展开更多
Potato(Solanum tuberosum L.)is susceptible to enzymatic browning after fresh processing,resulting in color change and potential alteration in the nutritional quality.In this study,a popular potato cultivar,Feiwuruita,...Potato(Solanum tuberosum L.)is susceptible to enzymatic browning after fresh processing,resulting in color change and potential alteration in the nutritional quality.In this study,a popular potato cultivar,Feiwuruita,was used to profile the metabolites involved in color and nutritional quality changes in fresh shreds stored at 0 and 4 h at 25°C(designated CK and CK4H,respectively).The shreds turned brown within 4 h of storage.In all,723 metabolites consisting 12 classes of compounds were detected in the samples,largely lipids,phenolic acids,alkaloids,amino acids and derivatives,flavonoids,organic acids,nucleotides and derivatives.Of these,163 metabolites accumulated differentially between CK and CK4H shreds.Polyphenolic compounds(phenolic acids and flavonoids)mostly increased in the shreds after 4 h storage.Conversely,the short-term storage drastically reduced lipid compounds(25 LysoPC and 19 LysoPE),while essential alkaloids and terpenoid compounds that are beneficial to human health increased in accumulation.The findings present global metabolome and nutritional composition changes in short-term stored shreds of Feiwuruita.This study provides important foundation for future studies on browning prevention/reduction and for better utilization of Feiwuruita.展开更多
Deactivation of polyphenol oxidase(PPO)in natural products is essential for downstream processing of functional molecules used as food or food additives,particularly those served as antioxidants.In the present work,we...Deactivation of polyphenol oxidase(PPO)in natural products is essential for downstream processing of functional molecules used as food or food additives,particularly those served as antioxidants.In the present work,we identified two proteins with PPO activity from lowbush blueberry using ammonium sulphate precipitation and chromatography procedures.Deactivation of these proteins was studied using aqueous solutions of ethanol of different concentrations.The PPO activity was recovered after ethanol removal for the protein samples previously soaked in a lowconcentration ethanol solution.A complete and unrecoverable deactivation of the proteins was achieved using ethanol with concentration over 70%(v/v),as manifested by the significant changes in circular dichroism(CD)and fluorescence spectroscopy measurements.Based on these findings,we propose a new extraction process for blueberry anthocyanin,in which an ethanol shock,i.e.soaking blueberry fruit in a 70%(v/v)ethanol solution for 1 h,is implemented before subsequent procedures.This new process increases the anthocyanin yield by 55%in comparison to that without the ethanol shock.展开更多
The effect of foliar spray of young lemon trees with Chitosan (CH) (200 - 300 - 400 ppm), on antioxidant enzymes activities (peroxidase POD, polyphenol oxidase PPO) and total phenolic content, and their influence on l...The effect of foliar spray of young lemon trees with Chitosan (CH) (200 - 300 - 400 ppm), on antioxidant enzymes activities (peroxidase POD, polyphenol oxidase PPO) and total phenolic content, and their influence on leaf miner (Phyllocnistis citrella) activity (tunnel length and damage density) under field condition was investigated. The results showed that treatment with chitosan 300 ppm enhanced the total phenolic content (5.975 mg/g) and the POD activity (0.533 min<sup>-1</sup> mg<sup>-1</sup> protein), while PPO activity was increased by chitosan 200 ppm (1.394 ΔA/min/g FW). The results demonstrated as well that chitosan treatment has a beneficial effect in reducing leaf miner activity, by decreasing tunnel length and damage density.展开更多
For building a biosensor, at least two enzyme sources can be employed. The pure enzyme has features with better selectivity and low stability. Crude extract presents better stability and wrong selectivity. Thus, one i...For building a biosensor, at least two enzyme sources can be employed. The pure enzyme has features with better selectivity and low stability. Crude extract presents better stability and wrong selectivity. Thus, one intermediate condition can be feasible joining both benefits of crude extract and pure enzymes. For that result, several procedures of extraction and semi purification of polyphenol oxidase (PPO) enzyme from banana (Musa sp.) were studied. The results showed that cleaned enzymatic extracts presented higher specific activities than crude extracts (2.8 up to 5.3 fold), despite the total protein concentration diminishing from 27% up to 72%, indicating that polyphenol oxidase (PPO) enzyme was preserved in them. The biosensors with 125 AU mL-1 containing cleaned enzymatic extracts performed better by grinding or grinding plus sonication for 30 s. They were linear over the ranges of 5.9 × 10-6 mol·L-1 to 1.4 × 10-3 mol·L-1 and 7.9 × 10-8 mol·L-1 to 4.0 × 10-3 mol·L-1, respectively. The limit of Detection (LOD) was 5.9 × 10-6 mol·L-1 and 7.9 × 10-8 mol·L-1, respectively. The LOD obtained is adequate to adrenaline determination on blood and medicinal samples, and were applied in medicinal samples with satisfactory results.展开更多
After harvest,cassava(Manihot esculenta Crantz)storage roots undergo rapid postharvest physiological deterioration,producing blue-brown discoloration in the vasculature due to the production of polyphenolics(mainly qu...After harvest,cassava(Manihot esculenta Crantz)storage roots undergo rapid postharvest physiological deterioration,producing blue-brown discoloration in the vasculature due to the production of polyphenolics(mainly quinonesand coumarins)by enzymes such as polyphenol oxidase(PPO).Here,we report the application of hen egg-white lysozyme(HEWL),a natural PPO inhibitor,in transgenic cassava to repress the symptoms of postharvest physiological deterioration.The HEWL-expressing transgenic plants had lower levels of the two main cassava coumarins tested,scopoletin and scopolin,compared with wild type.HEWL-expressing cassava also showed increased tolerance of oxidative stress.Overall,the lysozyme-PPO system proved to be functional in plants for repressing PPO-mediated commercial product browning.展开更多
基金Supported by Undergraduate Innovation and Enterpreneurship Training Program(202110514002)Huanggang Normal University High-level Cultivation Project(202108504).
文摘[Objectives]This study was conducted to investigate the effects of embedding conditions on activity and catalytic properties of immobilized polyphenol oxidase.[Methods]Polyphenol oxidase was immobilized in a polymer material by the embedding method,and the optimal immobilization conditions were obtained by single factor tests:CaCl_(2) concentration 2.0%,sodium alginate concentration 2.0%,immobilization time 2 h and mass ratio of enzyme to carrier 10 mg/100 g,under which the immobilized enzyme activity was 93.33 U/g.Under the above conditions,the properties of polyphenol oxidase immobilized by sodium alginate(A-PPO)and free polyphenol oxidase were studied.[Results]The thermostability of A-PPO was better than that of the free enzyme,but the pH stability of A-PPO was inhibited.The Michaelis constant K_(m) values of free polyphenol oxidase and A-PPO were 0.37 and 0.48 mmol/L,respectively,and the maximum reaction rate V_(max) values were 0.38 and 0.51 mmol/(L·g),respectively.[Conclusions]This study provides a theoretical basis for the study of the properties of polyphenol oxidase.
基金supported by grants from the National Key R&D Program of China(Grant No.2019YFD1000900)the Hainan Province Science and Technology Special Fund(Grant No.ZDYF2022XDNY253)the earmarked fund for CARS(Grant No.CARS-32-01)。
文摘Litchi(Litchi chinensis Sonn.)is a type of commercially prevalent subtropical and tropical fruit.Since litchi has a highly heterozygous genetic background and a long reproductive cycle,conventional breeding methods(such as hybridization)have limited ability to nurture new litchi cultivars.Here,an efficient and stable Agrobacterium tumefaciens-mediated genetic transformation of embryogenic callus was established in‘Feizixiao’litchi.Transgenic materials were verified using polymerase chain reaction(PCR)analysis,β-glucuronidase(GUS)assay,and green fluorescent protein(GFP)assay.To implement the technology of the Clustered Regularly Interspaced Short Palindromic Repeats(CRISPR)/associated protein 9(CRISPR/Cas9)technology in‘Feizixiao’litchi and verify the validity of these transformation systems,the litchi polyphenol oxidase gene(LcPPO,JF926153)was knocked out.Various categories of mutations,covering base insertions,deletions,and substitutions,were found in transgenic materials via sequence analysis.The transformation system achieved high feasibility and efficiency,and the system of CRISPR/Cas9 was successfully employed to edit genes in‘Feizixiao’litchi.This work provides an essential foundation for investigating the functions of genes and accelerating litchi genetic improvement.
基金research was funded by Major Science and Technology Project of Xiamen,China(3502Z20211004)Xiamen Science and Technology Assistance Project(3502Z20194509,3502Z20204504-2,3502Z20204501-3).
文摘Potato(Solanum tuberosum L.)is susceptible to enzymatic browning after fresh processing,resulting in color change and potential alteration in the nutritional quality.In this study,a popular potato cultivar,Feiwuruita,was used to profile the metabolites involved in color and nutritional quality changes in fresh shreds stored at 0 and 4 h at 25°C(designated CK and CK4H,respectively).The shreds turned brown within 4 h of storage.In all,723 metabolites consisting 12 classes of compounds were detected in the samples,largely lipids,phenolic acids,alkaloids,amino acids and derivatives,flavonoids,organic acids,nucleotides and derivatives.Of these,163 metabolites accumulated differentially between CK and CK4H shreds.Polyphenolic compounds(phenolic acids and flavonoids)mostly increased in the shreds after 4 h storage.Conversely,the short-term storage drastically reduced lipid compounds(25 LysoPC and 19 LysoPE),while essential alkaloids and terpenoid compounds that are beneficial to human health increased in accumulation.The findings present global metabolome and nutritional composition changes in short-term stored shreds of Feiwuruita.This study provides important foundation for future studies on browning prevention/reduction and for better utilization of Feiwuruita.
基金supported by Xinjiang Tianjianhemu Biotech Co.Ltd under project contract No.20172000941.
文摘Deactivation of polyphenol oxidase(PPO)in natural products is essential for downstream processing of functional molecules used as food or food additives,particularly those served as antioxidants.In the present work,we identified two proteins with PPO activity from lowbush blueberry using ammonium sulphate precipitation and chromatography procedures.Deactivation of these proteins was studied using aqueous solutions of ethanol of different concentrations.The PPO activity was recovered after ethanol removal for the protein samples previously soaked in a lowconcentration ethanol solution.A complete and unrecoverable deactivation of the proteins was achieved using ethanol with concentration over 70%(v/v),as manifested by the significant changes in circular dichroism(CD)and fluorescence spectroscopy measurements.Based on these findings,we propose a new extraction process for blueberry anthocyanin,in which an ethanol shock,i.e.soaking blueberry fruit in a 70%(v/v)ethanol solution for 1 h,is implemented before subsequent procedures.This new process increases the anthocyanin yield by 55%in comparison to that without the ethanol shock.
文摘The effect of foliar spray of young lemon trees with Chitosan (CH) (200 - 300 - 400 ppm), on antioxidant enzymes activities (peroxidase POD, polyphenol oxidase PPO) and total phenolic content, and their influence on leaf miner (Phyllocnistis citrella) activity (tunnel length and damage density) under field condition was investigated. The results showed that treatment with chitosan 300 ppm enhanced the total phenolic content (5.975 mg/g) and the POD activity (0.533 min<sup>-1</sup> mg<sup>-1</sup> protein), while PPO activity was increased by chitosan 200 ppm (1.394 ΔA/min/g FW). The results demonstrated as well that chitosan treatment has a beneficial effect in reducing leaf miner activity, by decreasing tunnel length and damage density.
文摘For building a biosensor, at least two enzyme sources can be employed. The pure enzyme has features with better selectivity and low stability. Crude extract presents better stability and wrong selectivity. Thus, one intermediate condition can be feasible joining both benefits of crude extract and pure enzymes. For that result, several procedures of extraction and semi purification of polyphenol oxidase (PPO) enzyme from banana (Musa sp.) were studied. The results showed that cleaned enzymatic extracts presented higher specific activities than crude extracts (2.8 up to 5.3 fold), despite the total protein concentration diminishing from 27% up to 72%, indicating that polyphenol oxidase (PPO) enzyme was preserved in them. The biosensors with 125 AU mL-1 containing cleaned enzymatic extracts performed better by grinding or grinding plus sonication for 30 s. They were linear over the ranges of 5.9 × 10-6 mol·L-1 to 1.4 × 10-3 mol·L-1 and 7.9 × 10-8 mol·L-1 to 4.0 × 10-3 mol·L-1, respectively. The limit of Detection (LOD) was 5.9 × 10-6 mol·L-1 and 7.9 × 10-8 mol·L-1, respectively. The LOD obtained is adequate to adrenaline determination on blood and medicinal samples, and were applied in medicinal samples with satisfactory results.
基金supported by the National Key R&D Program of China(2018YFD1000500)China Agriculture Research System(CARS-11)。
文摘After harvest,cassava(Manihot esculenta Crantz)storage roots undergo rapid postharvest physiological deterioration,producing blue-brown discoloration in the vasculature due to the production of polyphenolics(mainly quinonesand coumarins)by enzymes such as polyphenol oxidase(PPO).Here,we report the application of hen egg-white lysozyme(HEWL),a natural PPO inhibitor,in transgenic cassava to repress the symptoms of postharvest physiological deterioration.The HEWL-expressing transgenic plants had lower levels of the two main cassava coumarins tested,scopoletin and scopolin,compared with wild type.HEWL-expressing cassava also showed increased tolerance of oxidative stress.Overall,the lysozyme-PPO system proved to be functional in plants for repressing PPO-mediated commercial product browning.
