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Effect of Protocatechuic Acid-Grafted-Chitosan Coating on the Postharvest Quality of Pleurotus eryngii Jun Liu, Chen-guang Meng, Xing-chi Wang, Yao Chen, Juan Kan, and Chang-Hai Jin J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/acs.jafc.6b02468 • Publication Date (Web): 05 Sep 2016 Downloaded from http://pubs.acs.org on September 5, 2016
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Journal of Agricultural and Food Chemistry
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Effect of Protocatechuic Acid-Grafted-Chitosan Coating on the Postharvest
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Quality of Pleurotus eryngii
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Jun Liu*, Chen-guang Meng, Xing-chi Wang, Yao Chen, Juan Kan, Chang-hai Jin*
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College of Food Science and Engineering, Yangzhou University, Yangzhou 225127,
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Jiangsu, China
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* Corresponding authors. Tel: 86-514-87978158, E-mail:
[email protected] (Jun Liu),
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[email protected] (Chang-hai Jin)
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Protocatechuic
acid-grafted-chitosan
(PA-g-CS)
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solution
with
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ABSTRACT:
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antioxidant activity was developed as a novel edible coating material for Pleurotus
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eryngii postharvest storage. The effect of PA-g-CS coating on the postharvest quality of
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P. eryngii was investigated by determination of various physico-chemical parameters
12
and enzyme activities. Results showed that the antioxidant capacity and viscosity of
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PA-g-CS solutions were closely related to the grafting degree, and were much higher
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than that of chitosan (CS) solution. At the end of 15 days of storage, serious mushroom
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browning was observed in the control and CS coating groups. By contrast, PA-g-CS
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coating groups with medium and high grafting degrees maintained better physical
17
appearance. Among all the treatment groups, P. eryngii in PA-g-CS III coating group
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exhibited the highest firmness; however, the lowest weight loss, browning degree,
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respiration rate, malondialdehyde content, electrolyte leakage rate, superoxide anion
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production rate and hydrogen peroxide content. Moreover, P. eryngii in PA-g-CS III
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coating group maintained relatively higher antioxidant enzyme activities but lower
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polyphenol oxidase activity than other treatment groups. Therefore, PA-g-CS III is a
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promising preservation agent for P. eryngii.
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KEYWORDS: chitosan;
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protocatechuic acid
grafted;
Pleurotus
eryngii;
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postharvest
quality;
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■ INTRODUCTION
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Mushrooms are generally considered as functional foods due to their valuable
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nutritional and pharmaceutical properties.1–3 Among various cultivated species of
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mushrooms, the king oyster mushroom (Pleurotus eryngii) is rich in minerals, amino
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acids,
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polysaccharides.4 Because of these nutritional and bioactive ingredients, P. eryngii has
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lots of useful biological activities, such as antitumor, antihypercholesterolemic,
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antihypertensive, antimutagenic, antidiabetic and antioxidant activities etc.4–6 However,
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the fresh P. eryngii possesses high respiration and water loss rates, and is easily attacked
35
by microorganisms.7–8 For these reasons, P. eryngii usually exerts a short shelf-life,
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which greatly restricts its distribution and marketing. Therefore, it is essential to
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develop effective preservation techniques for P. eryngii.
essential
fatty
acids,
vitamins,
peptides,
proteins,
polyphenols
and
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Till now, several methods including cold storage, gamma irradiation, modified
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atmosphere packaging as well as edible coating have been used for the postharvest
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storage of P. eryngii.7–11 Among these methods, edible coating has received increasing
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attention because it can effectively prevent P. eryngii from water loss and gas exchange
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with outside environment. Besides, edible coating can also serve as the carrier of
43
functional components with antimicrobial and antioxidant properties.12 In recent years,
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many different kinds of edible coatings have been successfully developed and further
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applied for the postharvest storage of fruits and vegetables.13–14 Chitosan (CS) is such a
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natural biodegradable polymer that can be used as edible coating to suppress mushroom
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quality changes during storage.11, 15, 16 However, the effect of CS coating on P. eryngii
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preservation is not very ideal, which is probably due to the low solubility, antioxidant
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and antimicrobial activities of CS.11
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Phenolic compounds are essential in human diet due to their potent antioxidant,
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antimicrobial, antidiabetic and anticancer activities.17 In recently years, many phenolic
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compounds have been conjugated with CS through graft copolymerization reaction in
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order to enhance the biological activities of CS.18–19 The graft copolymerization
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techniques, such as the carbodiimide cross-linking, enzyme catalyzation and hydroxyl
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radical graft are regularly adopted.20–24 It has been demonstrated that phenolic grafted
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CS can be used as antioxidant, antimicrobial, antitumor, antidiabetic, adsorptive,
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encapsulation and packing materials.25–28 However, the effect of phenolic grafted CS
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coating on mushroom preservation is still unclear till now.
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In our previous work, a potent antioxidant agent (protocatechuic acid, PA) was
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covalently linked with CS chains via the 1-ethyl-3-(3-dimethylaminopropyl)
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carbodiimide hydrochloride (EDC) cross-linking reaction. The structure and
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physicochemical characteristics of PA-grafted-CS (PA-g-CS) were analyzed in details.29
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Results showed that the conjugation between PA and CS were achieved through amide
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and ester linkages. Antioxidant assay further revealed that PA-g-CS possessed stronger
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2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging capacity than CS, indicating
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PA-g-CS could be a promising antioxidant agent. In the present study, we further
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developed PA-g-CS as a novel edible coating material. The effect of PA-g-CS solution
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coating on the postharvest quality of P. eryngii during storage was investigated. Several
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physico-chemical parameters and enzyme activities of P. eryngii were evaluated
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throughout the storage period. The preservation effect of PA-g-CS on P. eryngii was also
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compared with that of CS. Our study provides novel information about the effect of
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phenolic grafted CS coating on mushroom preservation.
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■ MATERIALS AND METHODS
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Reagents. PA, EDC, N-hydroxysuccinimide (NHS), methionine, riboflavin and
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thiobarbituric acid (TBA) were purchased from Sigma-Aldrich Co. (St. Louis, MO,
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USA). CS with average-molecular weight 2.5 × 105 Da and 71% deacetylated degree,
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trichloroacetic acid (TCA), hydroxylamine hydrochloride, polyvinylpolypyrrolidone
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(PVPP), p-aminophenylsulfonic acid, α-naphthylamine, catechol and glutathione
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oxidized (GSSG) were purchased from Sangon Biotechnology Co. Ltd. (Shanghai,
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China). All other reagents used were of analytical grade.
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Synthesis of PA-g-CS with different grafting degrees. Three PA-g-CS products
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with different grafting degrees were synthesized by an EDC/NHS cross-linking method
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as described previously.29–30 The grafting degrees of PA-g-CS products were controlled
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via altering the molar ratios of reactants (e.g. CS, EDC, NHS and PA). As a result,
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PA-g-CS products with low, medium and high grafting degrees were obtained and
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named as PA-g-CS I (61.64 mg/g), PA-g-CS II (190.11 mg/g) and PA-g-CS III (279.69
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mg/g), respectively.
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Preparation of PA-g-CS and CS coating solutions. To prepare PA-g-CS coating
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solution, 5 g of lyophilized PA-g-CS sample was completely dissolved in 500 mL of
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acetic acid solution (0.5%, v/v) by magnetic stirring and then stored at 4 °C. CS coating
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solution of the same concentration (1%, w/v) was prepared in the same way.
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Determination of antioxidant capacity of PA-g-CS coating solutions.
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Superoxide anion (O2• ) radical scavenging capacity of PA-g-CS coating solutions was
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quantified according to the method of Jing and Zhao.31 Firstly, 0.4 mL of pyrogallol
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(0.025 M) was mixed with 1 mL of coating solution (0.2–1 mg/mL) and 4.5 mL of
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Tris–HCl buffer (0.05M, pH 8.2). Subsequently, 1 mL of HCl (8 mM) was added into
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mixture to initiate the formation of O2•-. Finally, O2•- scavenging capacity of the
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coating solutions could be calculated from the absorbance of reaction mixture at 420
99
nm.
-
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Hydrogen peroxide (H2O2) scavenging capacity of PA-g-CS coating solutions was
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measured using the reported method.32 Firstly, 0.6 mL of H2O2 (0.04 M) was mixed with
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1 mL of coating solution (0.2–1 mg/mL) and 2.4 mL of sodium phosphate buffer (0.1 M,
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pH 7.4). Afterwards, the acquired mixture was further reacted at 20 °C for 10 min.
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Finally, H2O2 scavenging capacity of PA-g-CS coating solutions was calculated from
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the absorbance of mixture at 230 nm.
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Lipid peroxidation inhibition effect of PA-g-CS coating solutions was assayed
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following the reported method.33 Firstly, 1.5 mL of ICR mice’s liver homogenate (1%,
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w/v) was mixed with equal volume of coating solution (0.2–1 mg/mL) followed by
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addition of 75 µL of H2O2 (0.5 mM) and 75 µL of FeCl2 (0.5 mM). The resultant
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mixture was allowed to react at 37 °C for 1 h. Afterwards, the reaction was quenched by
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addition of 2.25 mL of TBA (0.8%, w/v) along with 2.25 mL of TCA (20%, w/v),
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followed by incubation for 15 min at 100 °C and centrifugation for 30 min at 5000 g.
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Finally, lipid peroxidation inhibition effect of the coating solutions could be calculated
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from the absorbance of supernatant at 532 nm.
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Determination of rheological behavior of PA-g-CS coating solutions.
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Rheological behavior of PA-g-CS coating solution was measured on a Kinexus Pro
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Rheometer (Malvern Instruments Ltd., UK) at 20 °C. The viscosity was recorded by
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continuously increasing the shear rate ranging from 2 to 256 s−1.
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Mushrooms treatment and storage. P. eryngii were kindly supplied by Maosheng
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Edible Mushroom Co. Ltd. (Xuzhou, China). Fresh harvested P. eryngii were
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immediately transferred to our laboratory in 2 h under refrigerated conditions. P. eryngii
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with uniform size, weight (105–127 g) and color, but without mechanical damage were
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selected out. Subsequently, the mushroom coating procedure was applied according to
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the literature.15 Selected P. eryngii were equally divided into five groups: (1) control
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(distilled water coating) group, (2) CS coating group, (3) PA-g-CS I (low grafting
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degree) coating group, (4) PA-g-CS II (medium grafting degree) coating group, and (5)
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PA-g-CS III (high grafting degree) coating group. For each treatment, P. eryngii were
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soaked into corresponding coating solution for 30 s and dried on a plastic sieve for 30
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min. The excess coating solution on the surface of P. eryngii was adsorbed by filter
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paper. Afterwards, coated P. eryngii were placed in 28 cm × 30 cm polyethylene bags
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(0.04 mm in thickness) and further stored in a cold room (maintained at 4 ± 1 °C with
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95% relative humidity) for 15 days. During the storage period, five replicates of P.
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eryngii were randomly picked out from each coating group every 3 days for analyzing
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their physico-chemical parameters as well as enzyme activities.
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Firmness and weight loss measurement. TMS-PRO texture analyzer (Food
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Technology Corporation, USA) equipped with a cylindrical probe (diameter of 6 mm)
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was used to measure the firmness of P. eryngii mushroom cap. The penetration speed of
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2.0 mm/s along with the penetration depth of 5 mm was applied. Firmness was acquired
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from the curve of force vs time and expressed as Newton (N). Weight loss of P. eryngii
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was determined by measuring the weight of mushroom every 3 days throughout the
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storage period.
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Respiration rate measurement. To determine the respiration rate, P. eryngii were
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picked out of polyethylene bags and allowed to expose in room condition for 1 h.
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Afterwards, P. eryngii were placed together with a Petri dish containing 10 mL of
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NaOH solution (0.4 M) into an air-tight jar. After 30 min of adsorbance, NaOH solution
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was titrated by using diluted oxalic acid aqueous solution (0.2 M).33 The respiration rate
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of P. eryngii was calculated from the change in the amount of adsorbed CO2.
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Browning degree measurement. To measure the browning degree, P. eryngii (4 g)
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was ground in an ice bath by addition of 20 mL sodium phosphate buffer (0.2 M, pH 6.8)
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which contained PVPP (2.5%, w/v) and NaCl (0.15 M). The obtained mixture was then
151
centrifuged at 10000 g and 4 °C for 10 min.34 The absorbance of supernatant was
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measured at 420 nm and represented the browning degree of P. eryngii.
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Malondialdehyde (MDA) content measurement. MDA content in P. eryngii was
154
determined by the method of Liu and Wang.35 P. eryngii (1 g) was ground in an ice bath
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by addition of 5 mL TCA (5%, w/v). The obtained mixture was centrifuged at 10000 g
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and 4 °C for 20 min. Afterwards, 1 mL of TCA (10%, w/v) that contained TBA (0.67%,
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w/v) was added into 1 mL of supernatant, and further incubated in boiling water for 20
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min. After being cooled to room temperature, reaction mixture was finally centrifuged at
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10000 g and 4 °C for 10 min. The MDA content in P. eryngii was calculated by
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measuring the absorbance of supernatant at three different wavelengths (450, 532 and
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600 nm, respectively).
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Electrolyte leakage rate measurement. The membrane permeability of P. eryngii
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was checked by the electrolyte leakage rate.36 P. eryngii stipe (5 mm×5 mm×5 mm) was
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repeatedly washed using distilled water followed by drying with filter paper. The dried
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sample was further suspended in 25 mL of distilled water. A DDSJ-308A electrical
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conductivity meter (Leici Instrument Co., China) was used to measure the electrical
167
conductivity of sample solution (Co) immediately. Afterwards, sample was placed in
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boiling water for 10 min and the electrical conductivity of cooled sample solution was
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measured and recorded as Ct. Following equation was used to calculated electrolyte
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leakage rate of P. eryngii: electrolyte leakage rate (%) = Co/Ct × 100.
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O2• - production rate measurement. O2•- production rate of P. eryngii was
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measured following the reported method.35 P. eryngii (2 g) was ground in an ice bath by
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addition of 5 mL sodium phosphate buffer (50 mM, pH 7.8). The resultant mixture was
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centrifuged at 10000 g and 4 °C for 15 min. Then, 2 mL of hydroxylamine
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hydrochloride (10 mM) along with 1 mL of sodium phosphate buffer (50 mM, pH 7.8)
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was added into 1 mL supernatant, and further reacted for 1 h at room temperature.
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Finally, the obtained reaction mixture was blended with 1 mL of α-naphthylamine (7
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mM) and 1 mL of p-aminophenylsulfonic acid (50 mM) by incubating at room
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temperature for 20 min. O2•- production rate of P. eryngii was obtained by recording
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the absorbance of reaction mixture at 530 nm.
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H2O2 content measurement. H2O2 content in P. eryngii was assessed by the
182
method of Khan et al.37 P. eryngii (1 g) was ground in an ice bath by addition of 5 mL
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TCA (0.1%, w/v). The homogenate was then centrifuged at 10000 g and 4 °C for 15 min.
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Subsequently, 1 mL of potassium iodide (1 M) along with 1 mL of sodium phosphate
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buffer (0.1 M, pH 7.0) was added into 1 mL of supernatant, and further reacted at room
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temperature for 1 h in dark conditions. H2O2 content in P. eryngii was obtained by
187
recording the absorbance of reaction mixture at 390 nm.
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Determination of enzyme activity. In order to prepare crude enzyme extract, P.
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eryngii (2 g) was ground in an ice bath by addition of 5 mL sodium phosphate buffer
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(50 mM, pH 7.8) which contained PVPP (1%, w/v), Tween-20 (0.3%, v/v) and EDTA (1
191
µM). The homogenate was centrifuged at 10000 g and 4 °C for 20 min to obtain
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supernatant that served as crude enzyme extract.
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Superoxide dismutase (SOD) activity of P. eryngii was assayed by the method of Li
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et al.9 Firstly, 0.2 mL of P. eryngii enzyme extract was homogenized with 0.4 mL of
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EDTA (100 µM), 0.4 mL of nitroblue tetrazolium (0.75 mM), 0.4 mL of methionine
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(0.13 M) and 4.8 mL of sodium phosphate buffer (50 mM, pH 7.8). Afterwards, 0.2 mL
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of riboflavin (20 µM) was added into the homogenate to initial the reaction at 25◦C for
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60 min by 4000 lx of irradiance. SOD activity of P. eryngii was calculated by recording
199
the absorbance of reaction mixture at 560 nm.
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Catalase (CAT) activity of P. eryngii was measured following the reported
201
method.38 Firstly, 0.1 mL of P. eryngii enzyme extract was homogenized with 2.5 mL of
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sodium phosphate buffer (50 mM, pH 7.8). Subsequently, 1 mL of H2O2 (0.1 M) was
203
added into the homogenate to initial the reaction. CAT activity of P. eryngii was
204
calculated by recording the absorbance of reaction mixture at 240 nm.
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Ascorbate peroxidase (APX) activity of P. eryngii was assayed as reported
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previously.37 P. eryngii enzyme extract (0.1 mL) was homogenized with 2.9 mL of
207
sodium phosphate buffer (50 mM, pH 7.8) which contained H2O2 (2 mM), ascorbic acid
208
(0.5 mM) and EDTA (0.1 mM). APX activity of P. eryngii was calculated by recording
209
the absorbance of reaction mixture at 290 nm.
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Glutathione reductase (GR) activity of P. eryngii was determined according to the
211
method of Jiang et al.39 P. eryngii enzyme extract (0.2 mL) was homogenized with 2.7
212
mL of sodium phosphate buffer (0.1 M, pH 7.5) along with 40 µL of nicotinamide
213
adenine dinucleotide phosphate (4 mM) and 0.1 mL of GSSG (5 mM). GR activity of P.
214
eryngii was calculated by recording the absorbance of reaction mixture at 340 nm.
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Polyphenol oxidase (PPO) activity of P. eryngii was assayed as reported
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previously.40 P. eryngii enzyme extract (0.15 mL) was homogenized with 1.5 mL of
217
catechol (50 mM) and 6 mL of sodium phosphate buffer (0.1 M, pH 7.0). PPO activity
218
of P. eryngii was obtained by recording the absorbance of reaction mixture at 420 nm.
219
Scanning electron microscopy (SEM) observation. Micro-structure of P. eryngii
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cap tissue was observed by SEM according to the method of Ogawa et al.41 Small
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pieces of P. eryngii cap tissue were prepared by using the surgical blade. Samples were
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chemically fixed by soaking in formaldehyde solution (10%, v/v) overnight. Afterwards,
223
the resultant fixed tissue pieces were then sequentially dehydrated in ethanol aqueous
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solutions with different concentrations (70%, 80%, 90%, 95% and 100%). The obtained
225
dehydrated samples were freeze-dried and sputtered with gold. XL-30 ESEM scanning
226
electron microscope (Philips Electron Optics, Netherlands) was used for observing the
227
micro-structure of tissue piece.
228
Statistical analysis. Data were expressed as mean ± standard deviation (SD), and
229
analyzed by one-way analysis of variance (ANOVA) as well as Duncan’s multiple-range
230
comparisons. Difference was considered to be statistically significant in the case of p
0.05).
326
Effect of PA-g-CS coating on MDA content of P. eryngii. MDA level is a good
327
indicator to reflect the degree of lipid peroxidation.51 As shown in Figure 6A, MDA
328
contents of P. eryngii in different coating groups increased to 0.46−1.11 µmol/g when
329
storage period reached 15 days. The highest MDA content of P. eryngii was observed in
330
distilled water coating group (p < 0.05). P. eryngii in CS coating group showed
331
relatively lower MDA content than the control group, but higher MDA content than
332
PA-g-CS coating groups (p < 0.05). Notably, there was no statistical difference of MDA
333
content among three PA-g-CS coating groups (p > 0.05). The relatively lower MDA
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contents of P. eryngii in PA-g-CS coating groups could be due to the lipid peroxidation
335
inhibition effect of PA-g-CS.
336
Effect of PA-g-CS coating on electrolyte leakage rate of P. eryngii. Membrane
337
permeability of mushroom is often reflected by the electrolyte leakage rate. Figure 6B
338
showed that the electrolyte leakage rate of P. eryngii in the control and CS coating
339
groups rapidly increased during the storage period. Similar phenomenon was also
340
observed by Li et al.7 By contrast, P. eryngii in PA-g-CS coating groups showed
341
relatively lower electrolyte leakage rate than mushroom in distilled water and CS
342
coating groups (p < 0.05). However, PA-g-CS II coating group exhibited no significant
343
difference in electrolyte leakage rate as compared with PA-g-CS III coating group (p >
344
0.05). When storage period reached 15 days, electrolyte leakage rates of P. eryngii in
345
different coating groups increased to 14.2−48.8%. Results suggested that the membrane
346
integrity of P. eryngii could be maintained by PA-g-CS with medium and high grafting
347
degrees.
348
Effect of PA-g-CS coating on O2•- production rate and H2O2 content of P.
349
eryngii. ROS productions, such as O2•-, OH• and H2O2, are mainly produced in the
350
cellular metabolism process. High level of ROS can cause damage in several cellular
351
components (e.g. DNA, proteins and lipids), which results in mushroom senescence.34
352
As shown in Figure 7A, O2•- production rate of P. eryngii significantly increased after 9
353
days of storage in all the treatment groups, indicating the degree of mushroom
354
senescence accelerated. Among all the treatment groups, P. eryngii in PA-g-CS III
355
coating group showed the slowest O2•- production rate (p < 0.05), suggesting PA-g-CS
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III could greatly reduce the increase of O2• production rate in P. eryngii. Figure 7B
357
showed that H2O2 content of P. eryngii in all the treatment groups significantly
358
increased after 6 days of storage. The highest H2O2 content in P. eryngii was observed in
359
distilled water coating group (p < 0.05). By contrast, P. eryngii in PA-g-CS coating
360
groups exhibited relatively lower H2O2 content (p < 0.05). The relatively slower O2•-
361
production rate and lower H2O2 content in PA-g-CS coating groups should be attributed
362
to the potent antioxidant activity of PA-g-CS, which could scavenge free radicals
363
formed in mushroom.
364
Effect of PA-g-CS coating on antioxidant enzyme activity of P. eryngii. In
365
general, antioxidant enzymes are crucial defense systems in mushrooms.15–16 As shown
366
in Figure 8A, SOD activity in all the treatment groups gradually decreased throughout
367
the storage period. This result was in agreement with the finding of many other
368
reportors.15, 51 The lowest SOD activity of P. eryngii was observed in distilled water
369
coating group (p < 0.05). By contrast, CS and PA-g-CS coatings could significantly
370
delay the decrease of SOD activity. It has been reported that SOD could scavenge most
371
of the O2• - at the early storage stage.34 Thus, O2• - production rate increased
372
significantly at the late storage stage (Figure 7A). CAT plays an important role in
373
H2O2 degradation. Significant increase of CAT activity in P. eryngii was observed after
374
6 days of storage (Figure 8B). The highest CAT activity of P. eryngii was shown in
375
PA-g-CS III coating group (p < 0.05). When storage period reached 15 days, CAT
376
activities of P. eryngii in different treatment groups increased to 1.66−2.36 times of the
377
initial value. As shown in Figure 8C and Figure 8D, remarkable decrease of APX and
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GR activity in P. eryngii was observed in distilled water coating group. CS and PA-g-CS
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coatings could significantly prohibit the decrease in APX and GR activity. Notably,
380
PA-g-CS III coating was more efficient than any other treatment groups (p < 0.05).
381
Effect of PA-g-CS coating on PPO activity of P. eryngii. PPO plays a crucial role
382
in mushroom postharvest browning. When storage period reached 9 days, PPO activity
383
of P. eryngii significantly increased in both distilled water and CS coating groups
384
(Figure 9). By contrast, PA-g-CS coatings could remarkably reduce the increase in PPO
385
activity. However, PA-g-CS II coating group exhibited no significant difference in PPO
386
activity as compared with PA-g-CS III coating group (p > 0.05). Notably, a positive
387
correlation was found between browning degree and PPO activity, indicating that P.
388
eryngii browning was probably caused by PPO.
389
Effect of PA-g-CS coating on micro-structure of P. eryngii. Effect of different
390
coating solutions on the micro-structure of P. eryngii cap tissue was observed by SEM.
391
As shown in Figure 10, the fibrous arrangement of fresh P. eryngii cap tissue was
392
compact. By contrast, distilled water and CS coatings caused tangled or random fibrous
393
structure with enlarged microporous in P. eryngii cap tissue. The micro-structure change
394
should be attributed to the physical deformation of structural polysaccharides, such as
395
chitin and glucan, which could cause severe tissue shrinkage and collapse during
396
storage.41 In PA-g-CS coating groups, less tangled fibrous structure and smaller
397
microporous were observed as compared to distilled water and CS coating groups.
398
These results further confirmed the superior preservation effect of PA-g-CS coating.
399
Overall, PA-g-CS has been demonstrated as a good candidate of edible coating for
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its high antioxidant capacity and viscosity. P. eryngii in the control group showed
401
significant higher ROS production rate and MDA level, as well as lower antioxidant
402
enzyme activity than other treatment groups. The overproduction of ROS and reduced
403
enzyme activity are intrinsic features of mushroom senescence. Although CS coating
404
could improve the postharvest quality of P. eryngii, its effect was not that good as
405
PA-g-CS coating. It should be noted that PA-g-CS III has been demonstrated as the best
406
coating material for the postharvest storage of P. eryngii. The superior preservation
407
effect of PA-g-CS III could be mainly attributed to its highest antioxidant activity and
408
proper viscosity. Thus, PA-g-CS III is a very potential coating material for mushroom
409
postharvest storage.
410
■AUTHOR INFORMATION
411
Corresponding Authors
412
*(Jun Liu) Tel: 86-514-87978158. Email:
[email protected].
413
*(Chang-hai Jin) Tel: 86-514-87978158. Email:
[email protected].
414
Funding
415
This work was supported by grants of National Natural Science Foundation of China
416
(31571788 and 31101216), Natural Science Foundation of Jiangsu Province
417
(BK20151310), Jiangsu Provincial Government Scholarship for Overseas Studies, High
418
Level Talent Support Program of Yangzhou University, and Qing Lan Project of Jiangsu
419
Province.
420
Notes
421
The authors declare no competing financial interest. 20
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FIGURE LEGENDS Figure 1. The superoxide radical scavenging activity (A), H2O2 scavenging activity (B) and lipid peroxidation inhibition effect (C) of CS, PA, PA-g-CS I, PA-g-CS II, PA-g-CS III and Vc. Data are presented as means ± SD of triplicates. Figure 2. Viscosity behavior of CS, PA-g-CS I, PA-g-CS II and PA-g-CS III coating solutions. Figure 3. Effect of CS and PA-g-CS coating on the physical appearance of P. eryngii. A: fresh harvested mushroom; B, C, D, E and F: mushroom coated with distilled water, 1% of CS, PA-g-CS I, II and III solutions respectively, and stored at 4 ± 1 °C with 95% relative humidity for 15 days. Figure 4. Effect of CS and PA-g-CS coating on the firmness (A) and weight loss (B) of P. eryngii during storage at 4 °C for 15 days. Data are presented as means ± SD of five replicates. Figure 5. Effect of CS and PA-g-CS coating on the respiration rate (A) and browning degree (B) of P. eryngii during storage at 4 °C for 15 days. Data are presented as means ± SD of five replicates. Figure 6. Effect of CS and PA-g-CS coating on the MDA content (A) and electrolyte leakage rate (B) of P. eryngii during storage at 4 °C for 15 days. Data are presented as means ± SD of five replicates. Figure 7. Effect of CS and PA-g-CS coating on the O2•- production rate (A) and H2O2 content (B) of P. eryngii during storage at 4 °C for 15 days. Data are presented as means ± SD of five replicates.
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Figure 8. Effect of CS and PA-g-CS coating on the SOD (A), CAT (B), APX (C) and GR (D) activities of P. eryngii during storage at 4 °C for 15 days. Data are presented as means ± SD of five replicates. Figure 9. Effect of CS and PA-g-CS coating on the PPO activity of P. eryngii during storage at 4 °C for 15 days. Data are presented as means ± SD of five replicates. Figure 10. Micro-structure of P. eryngii cap tissue observed by SEM. A: fresh harvested mushroom; B, C, D, E and F: mushroom coated with distilled water, 1% of CS, PA-g-CS I, II and III solutions respectively, and stored at 4 ± 1 °C with 95% relative humidity for 15 days.
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(A)
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Scavenging activity (%)
100 80
Vc PA CS PA-g-CS I PA-g-CS II PA-g-CS III
60 40 20 0 0
0.2
0.4
0.6
0.8
1
Concentration (mg/mL)
Scavenging activity (%)
(B)
100 80
Vc PA CS PA-g-CS I PA-g-CS II PA-g-CS III
60 40 20 0 0
0.2
0.4 0.6 0.8 Concentration (mg/mL)
1
(C) 100 Inhibition effect (%)
80
Vc PA CS PA-g-CS I PA-g-CS II PA-g-CS III
60 40 20 0 0
0.2
0.4 0.6 0.8 Concentration (mg/mL)
1
Figure 1 30
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Viscosity (Pa·s)
1.00 CS
PA-g-CS I
PA-g-CS II
PA-g-CS III
0.10
0.01
0.00 1
10
100
Shear rate (1/s) Figure 2
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(A)
(B)
(C)
(D)
(E)
(F)
Figure 3
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(A)
5.5
CS
PA-g-CS I
PA-g-CS II
PA-g-CS III
4.8
Firmness (N)
Control
4.1 3.4 2.7 2.0 0
3
6
9
12
15
12
15
Storage time (days)
Weight loss (%)
(B)
2.5
Control
CS
PA-g-CS I
PA-g-CS II
2 PA-g-CS III
1.5 1 0.5 0 0
3
6
9
Storage time (days)
Figure 4
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Respiration rate (mg CO2/kg/h)
(A)
420 360 300 240 180
Control
CS
PA-g-CS I
PA-g-CS II
PA-g-CS III
120 0
3
6
9
12
15
Storage time (days)
Browning degree (A 450)
(B) 0.8 0.6
Control
CS
PA-g-CS I
PA-g-CS II
PA-g-CS III
0.4 0.2 0.0 0
3
6
9
12
15
Storage time (days)
Figure 5
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MDA content (μmol/g)
(A)
1.2
Control
CS
1.0
PA-g-CS I
PA-g-CS II
PA-g-CS III
0.8 0.6 0.4 0.2 0.0 0
3
6
9
12
15
12
15
Storage time (days) (B) Relative leakage rate (%)
55 45
Control
CS
PA-g-CS I
PA-g-CS II
PA-g-CS III
35 25 15 5 0
3
6 9 Storage time (days)
Figure 6
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(nmol/min/g)
-
O2• production rate
(A)
8
Control
CS
7
PA-g-CS I
PA-g-CS II
PA-g-CS III
6 5 4 3 2 0
3
6
9
12
15
12
15
Storage time (days)
H2O2 content (μmol/g)
(B)
420
Control
CS
PA-g-CS I
PA-g-CS II
PA-g-CS III
340 260 180 100 0
3
6
9
Storage time (days)
Figure 7
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SOD activity (U/g)
(A)
0.15 0.12
Control
CS
PA-g-CS I
PA-g-CS II
PA-g-CS III
0.09 0.06 0.03 0 0
3
6
9
12
15
12
15
Storage time (days)
CAT activity(U/g)
(B) 6.0 5.0
Control
CS
PA-g-CS I
PA-g-CS II
PA-g-CS III
4.0 3.0 2.0 0
3
6
9
Storage time (days)
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APX activity (U/g)
(C)
60 50
Control
CS
PA-g-CS I
PA-g-CS II
PA-g-CS III
40 30 20 10 0 0
3
6
9
12
15
Storage time (days) Control PA-g-CS I PA-g-CS III
(D) 3.0
GR activity(U/g)
2.5
CS PA-g-CS II
2.0 1.5 1.0 0.5 0.0 0
3
6 9 Storage time (days)
12
15
Figure 8
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PPO activity (U/g)
130 110
Control
CS
PA-g-CS I
PA-g-CS II
PA-g-CS III
90 70 50 30 0
3
6
9
12
15
Storage time (days)
Figure 9
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(A)
(B)
(C)
(D)
(E)
(F)
Figure 10
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OH
OH
O
O HO
NH2
O HO
Pleurotus eryngii
O NHCOCH 3
CS O R= C
OH
PA EDC/NHS
OH
ROS production rate (O2 •- and H2O2 )
OR
OR
O
O RO
Physico-chemical parameters (Firmness, weight loss, respiration rate, browning degree, MDA content and electrolyte leakage rate)
NHR
O RO
O NHCOCH3
PA-g-CS (Enhanced antioxidant activity and rheological property)
Enzyme activities CS and PA-g-CS coating (4 °C storage for 15 days)
(SOD, CAT, APX, GR and PPO)
TOC Graphic
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