LÜ Jingyi, QIU Yanxia, BAI Lin, et al. Combined Effects of Melatonin and Citric Acid on Preservation of Fresh-cut 'Early Crisp' Pears during Cold Storage[J]. Science and Technology of Food Industry, 2025, 46(10): 1−10. (in Chinese with English abstract). doi: 10.13386/j.issn1002-0306.2024070015.
Citation:
LÜ Jingyi, QIU Yanxia, BAI Lin, et al. Combined Effects of Melatonin and Citric Acid on Preservation of Fresh-cut 'Early Crisp' Pears during Cold Storage[J]. Science and Technology of Food Industry, 2025, 46(10): 1−10. (in Chinese with English abstract). doi: 10.13386/j.issn1002-0306.2024070015.
LÜ Jingyi, QIU Yanxia, BAI Lin, et al. Combined Effects of Melatonin and Citric Acid on Preservation of Fresh-cut 'Early Crisp' Pears during Cold Storage[J]. Science and Technology of Food Industry, 2025, 46(10): 1−10. (in Chinese with English abstract). doi: 10.13386/j.issn1002-0306.2024070015.
Citation:
LÜ Jingyi, QIU Yanxia, BAI Lin, et al. Combined Effects of Melatonin and Citric Acid on Preservation of Fresh-cut 'Early Crisp' Pears during Cold Storage[J]. Science and Technology of Food Industry, 2025, 46(10): 1−10. (in Chinese with English abstract). doi: 10.13386/j.issn1002-0306.2024070015.
The aim of this paper was to explore the impacts of melatonin (MT) combined with citric acid (CA) on preservation of fresh-cut 'Early Crisp' pears during cold storage. Fresh-cut 'Early Crisp' pears were treated with 100 μmol·L−1 MT and 2% CA, alone or in combination and then stored at 4±0.5 ℃ for 9 d. Quality-related parameters containing total soluble solids (TSS) content and total number of colonies. Browning-related parameters including browning index (BI), total phenolics content, phenylalanine ammonia lyase (PAL), peroxidase (POD) and polyphenol oxidase (PPO) activities. Antioxidant activity-related parameters involving phenolics, flavonoids, ascorbic acid (AsA) and malondialdehyde (MDA) contents. The 2,2-Di-(4-tert-octylphenyl)-1-picrylhydrazyl (DPPH) radical and ABTS radical cation (ABTS+) radical scavenging activities were analyzed during storage. The results indicated that, in all the parameters considered, significant differences were generally found among all treated and non-treated fresh-cut pears, and combined application of MT and CA showed the best results among treatments. For single treatment, CA treatment exhibited better anti-browning effect than MT treatment, while MT treatment had better antioxidant capacity than CA treatment. Synergistic impacts of combined treatment on antioxidant activity and browning inhibition improvement of fresh-cut 'Early Crisp' pears were observed during cold storage. At 7 d, the BI value in combined treatment group was 30.08%, 10.48% and 5.41% lower than that in control, MT and CA treatment groups, respectively. The combined treatment group presented the highest ABTS+ and DPPH radical scavenging activities compared with other groups during storage. These data indicated that MT combined CA treatment could be used as an effective preservative for fresh-cut 'Early Crisp' pears during cold storage. This combined treatment might have the potential to be employed to food preservation.
'Early Crisp' pear （Pyrus bretschneideri Rehd.） is popular among consumers in China due to its crispy texture, sweet taste and high nutritional value. Owing to the accelerated pace of modern life, people are pursuing more fresh, convenient and healthier foods. Therefore, fresh-cut fruits are more popular to consumers[1]. 'Early Crisp' pears have thin skin, small core and thick flesh, thus having high utilization rate during processing. However, tissue browning can easily occur on cut surface of fresh-cut 'Early Crisp' pears, which reduces sensory quality and nutritional value. In addition, mechanical injury caused by cutting can disrupt the balance between generation and elimination of reactive oxygen species （ROS）, resulting in the highest ROS generation and oxidative damage to tissues, thereby accelerating senescence of fresh-cut fruits and vegetables[2−5].
The phytohormone melatonin （MT） plays essential roles in plant growth, development, senescence and stress resistance[3]. It has been reported as an efficient free-radical scavenger and antioxidant for preservation of many vegetables and fruits[6]. Previous studies have demonstrated that MT treatment can decrease ROS production and malondialdehyde （MDA） content, increase free radical scavenging activities of 2,2-Di-（4-tert-octylphenyl）-1-picrylhydrazyl （DPPH·） and ABTS radical cation （ABTS+·）, thus effectively delaying senescence of many postharvest fruits, including litchi, strawberry and pomegranate[6−8]. Until now, few researches have been carried out on the impacts of exogenous MT on preservation of fresh-cut vegetables and fruits. Studies on fresh-cut European pears （Pyrus communis L.） and fresh-cut avocadoes showed that MT treatment can effectively scavenge DPPH· free radicals and maintain storage quality[9−10].
Browning is one of the major factors restricting shelf life of fresh-cut fruits and vegetables. Browning of plant tissues can be either enzymatic or non-enzymatic. Previous studies have demonstrated that enzymatic browning was the major cause of browning on cut surface of pear fruit[1]. The enzymatic browning in planta occurs when phenolic compounds are oxidized by polyphenol oxidase （PPO） and peroxidase （POD） into quinones, which then polymerize to form dark brown or black polymers[11]. In addition, phenylalanine ammonia lyase （PAL）, a key enzyme in phenolic biosynthesis pathway, also plays essential roles in enzymatic browning[3]. Citric acid （CA） is commonly used as an acidity regulator and flavor enhancer in food industry[12]. Previous studies showed that CA could effectively reduce browning of many fruits and vegetables[13]. The role of CA in preventing tissue browning is mainly due to its dual inhibitory effects on PPO[14]. On one hand, the reactions catalyzed by PPO could be almost completely inhibited by CA, because the optimum pH for PPO activity is 4~7. On the other hand, CA can chelate copper ions at the active site of PPO, leading to lower PPO activity[14]. It has been reported that CA treatment can effectively inhibit PPO activity, maintain total phenolics contents, thus delaying browning and maintaining storage quality of many fresh-cut vegetables and fruits, including fresh-cut sweet potatoes and fresh-cut water chestnuts[3,15]. However, CA treatment is not always effective in reducing browning of fresh-cut vegetables and fruits. Study on fresh-cut fennel showed that browning on cut surface of stem and sheath became much more evident after CA treatment during storage[16]. Suttirak and Manurakchinakorn[17] reported that the anti-browning impacts of exogenous CA on fresh-cut fruits and vegetables might vary among types and cultivars. Until now, the impacts of CA treatment on preservation and browning of fresh-cut 'Early Crisp' pears have not been determined.
It has been demonstrated that CA used in combination with other antioxidants or anti=browning agents can maintain better storage quality of some fruits and vegetables than when used alone[17]. For example, study on fresh-cut apples showed that, compared with CA treatment alone, CA combined with ascorbic acid （AsA） treatment obviously reduced PPO activity and exhibited higher anti=browning efficiency[17]. To date, the effects of CA combined with MT on fresh-cut pear preservation are unknown. The purpose of our work was to explore the impacts of MT combined with CA on preservation of fresh-cut 'Early Crisp' pears during cold storage, and its effects on browning and antioxidant capacity were also discussed.
1.
Materials and methods
1.1
Materials and instruments
'Early Crisp' pears （Pyrus bretschneideri Rehd.） were bought at a local market. Fresh pears without pests, diseases, and mechanical damages were selected in this study. All pears were packed in cartons and immediately transferred to laboratory. Preliminary experiment was conducted to identify the optimum concentration of CA （0%, 0.5%, 1%, 1.5%, and 2%） （w/v） and MT （0, 50, 100, 150, and 200 μmol·L−1）, separately. The results showed that 2% CA and 100 μmol·L−1 MT were the most effective in maintaining fresh-cut 'Early Crisp' pears quality during storage at 4 ℃. Thus, 2% CA and 100 μmol·L−1 MT were used in current research.
CR-300 color difference meter Japan Konica Minolta, WYT-32 handheld refractometer Quanzhou Optical Co. Ltd.; HBM-400B homogenizer Tianjin Hengao Technology Co. Ltd.; Legend Micro21R refrigerated centrifuge America Thermo Fisher Scientific; L9-Plus UV spectrophotometer Shanghai Scientific Instrument Co. Ltd.; LDZX-75KBS autoclave Shanghai Shen'an Medical Device Factory; ICP500 biochemical incubator Shanghai Memmert.
1.2
Experimental methods
1.2.1
Material pre-processing
Pears were washed with distilled water to remove surface dirt and subsequently disinfected with 0.1% （w/v） sodium hypochlorite solution for 2 min. All the tools were sterilized with the sodium hypochlorite solution before use. After natural air-drying, the core of each pear was removed and the remaining tissues were vertically cut into eight equal wedges with fruit sectionizer. All pear wedges were randomly divided into four groups. The first group was dipped in distilled water for 2 min. The second group was dipped in 2% CA for 2 min. The third group was immersed in 100 μmol·L−1 MT for 2 min. The fourth group was immersed in solution containing 2% CA and 100 μmol·L−1 MT for 2 min. Each group had three treatment replicates with 112 pear wedges per treatment replicates. All the treatments were performed at 20 ℃. After treatment, the pear wedges were air-dried, packed in low-density polyethylene （PE） storage bags （100 mm×150 mm, 0.01 mm thick）, and stored at 4±0.5 ℃ for 9 d, 90% RH. Samples were collected every day. 12 wedges were randomly chosen from each group for measuring surface color, and another 30 wedges from each group were cut into pieces, grounded to a fine powder using liquid nitrogen, and stored at −80 ℃. Frozen powders from these 30 wedges were used for determination of the other indicators.
1.2.2
Determination of surface color
Surface color was analyzed by the protocol of Zambrano-Zaragoza et al[18] and expressed as browning index （BI）. The values of L*, a* and b* were measured by color difference meter （CR-300 Konica Minolta Japan）. The BI was calculated using the formula: BI = [100（x−0.31）/0.172], where x = （a*+1.75L*） / （5.645L*+a* − 3.012b*）.
1.2.3
Analysis of TSS, AsA, total flavonoids and phenolics contents
TSS, AsA, total flavonoids and phenolics contents were determined as described by Liu et al[6] with slight modification. TSS content was measured with a handheld refractometer and expressed in percentage （%）. For determination of total phenolics and flavonoids contents, 0.1 g of the frozen tissue was mixed with 1 mL of 50% ethanol, extracted in ultrasonic bath for 40 min and centrifuged. The supernatant was used for assay of total flavonoids and phenolics contents. For assay of total phenolics content, 0.7 mL of the supernatant was mixed with 1.5 mL of Folin-Ciocalteau reagent. 1 mL of 7.5% Na2CO3 solution was then added into the mixture. The resulting solution was incubated in the dark. Absorbance was analyzed at 765 nm. For assay of flavonoids content, 0.5 mL of the supernatant was mixed with 0.2 mL of 5% NaNO2 and 0.2 mL of 10% Al（NO3）3. Then, 1 mol·L−1 NaOH （1.0 mL） was added into the mixture. Absorbance was measured at 510 nm. Content of AsA was determined by 2,6-dichlorophenol indophenol method. 0.5 g of frozen sample was mixed with oxalic acid and CH3COONa solution. The sample was titrated with a calibrated 2,6-dichlorophenol indophenol solution.
1.2.4
Analysis of MDA content, DPPH and ABTS+ radical scavenging activities
Content of MDA was measured as described by Zheng et al[10]. 1 g of the frozen sample was ground with 10 mL of 10% trichloroacetic acid （TCA） （w/v）. The homogenates were centrifuged. The supernatant was mixed with 0.67% TBA, incubated for 20 min and then centrifuged. Absorbance was measured at 450 nm, 600 nm and 532 nm, respectively. Content of MDA was calculated and expressed as nmol·g−1.
Free radical-scavenging capacities （ABTS+· and DPPH·） were measured by the method of Zheng et al[10] with little modifications. Frozen tissue （0.5 g） was mixed with 95% ethanol solution and centrifuged. The supernatant was used for the subsequent determination. For DPPH· radical scavenging activity, the supernatant was mixed with ethanol solution （containing 0.1 mol·L−1 DPPH·） and then reacted for 30 min. Absorbance was measured at 517 nm. DPPH· radical scavenging activity （%） = [100 （Ac−As）/Ac], where Ac represents the absorbance of blank group, and As represents the absorbance of each sample. For ABTS+· radical scavenging activity, the supernatant was mixed with ABTS+· radical reserve solution, and then incubated in the dark. Absorbance was measured at 734 nm. The calculation method of the radical scavenging activity of ABTS+· is the same as that of DPPH· radical scavenging activity.
1.2.5
Determination of PPO, PAL and POD activities
Activities of PPO, POD and PAL were analyzed as described by Peng et al[19] for assay of PAL activity, crude enzyme was extracted using 1 g of frozen sample. PAL activity was determined in a reaction mixture containing 3.4 mL of borate buffer solution （pH8.8）, 100 μL of crude enzyme solution and 500 μL of L-phenylalanine solution （20 mmol·L−1）. Absorbance was measured at 290 nm. For assay of PPO activity, crude enzyme was extracted using 0.5 g of frozen sample. PPO activity was determined in a reaction mixture containing 500 μL of 100 mmol·L−1 4-methylcatechol, 1 mL of 0.1 mol·L−1 phosphate buffer （pH6.8） and 500 μL of crude enzyme solution. Absorbance was determined at 410 nm. For assay of POD activity, crude enzyme was extracted using 0.8 g of frozen fruit sample. POD activity was analyzed in a reaction mixture including 2.7 mL of 0.1 mol·L−1 phosphate buffer, 100 μL of 0.5% H2O2, 100 μL of crude enzyme solution and 100 μL of 4% guaiacol. Absorbance was measured at 470 nm.
1.2.6
Determination of total number of colonies
Total number of colonies was measured by the method of Zhang et al[20] with slight modification. Fresh samples （25 g） were placed into 225 mL of 0.85% sterile NaCl solution and shaken with a homogenizer for 1 min to make 1:10 diluted solution. Then, 1 mL of the 1:10 diluted solution was diluted with 0.85% sterile NaCl solution （9 mL） to make a 1:102 diluted solution. Serially diluted solution was performed as described above and then spread in triplicate onto plate count agar （PCA）. Plates were incubated for 24 h at 36±1 ℃. Total number of colonies on the plate were calculated.
1.3
Statistical analysis
All determinations were conducted in three replicates. The data were compared by ANOVA using SPSS software （version 19.0）. Differences were considered significant when P-values were below 0.05. All values were expressed as means±standard deviation.
2.
Results and Analysis
2.1
Changes in BI and TSS content of fresh-cut 'Early Crisp' pears
At 8 d, the quality of the control fruit became significantly worse and could not be sampled anymore. Therefore, only 0~7 d were measured in the control group. The BI of fresh-cut 'Early Crisp' pears gradually increased during storage （Fig.1A）. Control group presented the highest BI value, whereas combined treatment group presented the lowest BI value. At 6 d, the BI in control pears was 1.64 times higher than that in combined treatment group. The BI in MT-treated pears was higher than in CA-treated pears from 2 d to 8 d. At 7 d, the BI value in combined treatment group was 30.08%, 10.48% and 5.41% lower than that in control, MT and CA treatment groups, respectively.
Figure
1.
Effects of CA, MT and combined treatment on BI （A） and TSS （B） content of fresh-cut 'Early Crisp' pears stored at 4±0.5 ℃
Note: Error bars indicate standard deviations. Different letters show significant differences among treatments at each sampling time point （P<0.05）, the Fig.2~Fig.5 below.
Content of TSS in all groups declined steadily during storage （Fig.1B）. The TSS content in combined treatment group was higher than the other groups. These data showed that combined treatment could effectively retard browning and maintain TSS content of fresh-cut 'Early Crisp' pears compared with single treatment.
2.2
Changes in AsA, total phenolics and flavonoids contents of fresh-cut 'Early Crisp' pears
Contents of AsA, total phenolics and flavonoids in all groups generally decreased during storage （Fig.2）. In combined treatment group, AsA, total flavonoids and phenolics contents were generally higher than the other groups during storage. From 3 d to 7 d, MT treatment group generally maintained higher AsA and total flavonoids contents while lower total phenolics content compared to CA treatment group. At 7 d, AsA content in combined-treated fresh-cut pears was 1.60, 1.30 and 1.29 times higher than that in control, CA- and MT-treated fresh-cut pears, respectively. Content of total phenolics in combined treatment group was 1.51-, 1.28- and 1.11-fold higher than in control, MT and CA treatment groups, respectively. Total flavonoids content in combined-treated group was 1.83-, 1.39- and 1.23-fold higher than in control, CA- and MT-treated groups, respectively.
Figure
2.
Effects of CA, MT and combined treatment on AsA （A）, total phenolics （B） and total flavonoids contents （C） of fresh-cut 'Early Crisp' pears stored at 4±0.5 ℃
2.3
Changes in MDA content, ABTS+· and DPPH· radical scavenging activities of fresh-cut 'Early Crisp' pears
The DPPH· and ABTS+· radical scavenging activities of fresh-cut 'Early Crisp' pears decreased in all groups （Fig.3A and Fig.3B）. Combined treatment group presented the highest ABTS+· and DPPH· radical scavenging activities than the other groups. At 7 d, DPPH radical scavenging activity in combined-treated group was 1.22-, 1.09- and 1.06-fold higher than in control, CA and MT treatment groups, respectively. The ABTS+· radical scavenging activity in combined treatment group was 1.06-, 1.03- and 1.02-fold higher than that in control, CA and MT treatment groups, respectively. These data suggested the combined treatment effectively enhanced antioxidant capacity of fresh-cut pears. In MT treatment group, the ABTS+· and DPPH· radical scavenging activities were higher than that in CA treatment group during storage.
Figure
3.
Effects of CA, MT and combined treatment on DPPH· radical scavenging activity （A）, ABTS+· radical scavenging activity （B） and MDA content （C） of fresh-cut 'Early Crisp' pears stored at 4±0.5 ℃
The MDA content of fresh-cut ‘Early Crisp’ pears generally increased during storage （Fig.3C）. Combined treatment group generally showed the lowest MDA content compared with the other groups. The MDA content in combined treatment group was only 55.85% of that in control group at 7 d, and was 64.50% and 81.69% of that in CA and MT treatment group at 8 d, respectively. These data showed that combined treatment retarded membrane lipid peroxidation of fresh-cut 'Early Crisp' pears more effectively than single treatment.
2.4
Changes in activities of PPO, PAL and POD of fresh-cut 'Early Crisp' pears
The PAL activity gradually increased during storage （Fig.4A）. Combined treatment group showed the highest PAL activity compared with other groups. In combined treatment group, the PAL activity was 1.09 times higher than controls at 6 d. Generally, no significant differences in PAL activity were detected between MT treatment group and CA treatment group.
Figure
4.
Effects of CA, MT and combined treatment on activities of PAL （A）, PPO （B） and POD （C） of fresh-cut ‘Early Crisp’ pears stored at 4±0.5 ℃.
Activities of PPO and POD reached the peak value at 2 d （Fig.4B and Fig.4C）. Their activities in control group were higher than that in combined treatment group during storage. PPO activity in combined treatment group was lower than that in CA treatment group from 1 d to 2 d and from 6 d to 8 d, and its activity in CA treatment group was lower than that in MT treatment group from 2 d to 4 d and from 6 d to 8 d. At 2 d, PPO activity in CA, MT and combined treatment groups were respectively 9.71%, 2.91% and 13.40% lower than that in control group, and POD activities were respectively 10.87%, 10.14% and 16.74% lower than that in control group. At 7 d, PPO activity in combined treatment group was 17.54%, 13.45% and 6.67% lower than in control, MT and CA treatment groups, respectively.
2.5
Changes in total number of colonies of fresh-cut 'Early Crisp' pears
Total number of colonies displayed an increasing tendency in all groups during storage （Fig.5）. Control group showed the highest number of colonies, whereas combined treatment group exhibited the least number of colonies during storage. At 7 d, the number of colonies in control group was 1.92-fold higher than that in combined-treated group. Except 3 d, no significant differences in total number of colonies were detected between MT treatment group and CA treatment group.
Figure
5.
Effects of CA, MT and combined treatment on total number of colonies of fresh-cut 'Early Crisp' pears stored at 4±0.5 ℃
Fresh-cut produce is inevitably subjected to mechanical wounding stress during processing, which will induce high level of ROS accumulation and cause membrane lipid peroxidation[4−5]. In plants, ROS can be scavenged by antioxidant defense systems, including antioxidants （AsA, phenolics and flavonoids） and antioxidases （superoxide dismutase, catalases and POD）[21−22]. MT is a free-radical scavenger with direct antioxidant activity[6]. Exogenous MT application can remove ROS by increasing antioxidants contents and antioxidant enzymes activities, thus maintaining storage quality of many postharvest vegetables and fruits[23]. Our results showed that, compared with control, MT treatment can enhance antioxidant capacity of fresh-cut 'Early Crisp' pears by enhancing contents of antioxidant substances （AsA, phenolics and flavonoids） during storage, as previously demonstrated in fresh-cut European pears （Pyrus communis L.） and avocadoes[9−10]. CA acts as an elicitor of compounds derived from phenylpropanoids and activates signaling cascades to enhance antioxidant activity[24]. Zhao et al[25] suggested that endogenous CA mainly severed as an antioxidant intermediate in TCA cycle and accumulation of CA can improve stress tolerance. Several recent studies have reported that exogenous CA application can induce antioxidant defense systems in plants under abiotic stresses[24]. For fruits and vegetables, previous investigations on relationships between exogenous CA and antioxidant capacity were limited, and particularly for fresh-cut vegetables and fruits, relevant studies were extremely rare. Zhang et al[26] found that CA treatment increased antioxidants contents （total phenolics and total flavonoids） and promoted antioxidant enzyme activities, thus maintaining storage quality of postharvest tomato fruit. Similarly to this previous finding, our results showed that CA treatment can enhance antioxidant activity of fresh-cut 'Early Crisp' pears by increasing antioxidant contents and enhancing ROS scavenging capacity compared with control. However, studies on fresh-cut potato and fresh-cut lettuce indicated that application of CA did not affect total antioxidant activity[27−28]. These results indicated that impacts of exogenous CA on total antioxidant capacity of fresh-cut vegetables and fruits might vary among types. In present study, compared with CA treatment group, MT treatment group generally maintained higher contents of AsA and total flavonoids and higher radical scavenging activities of DPPH· and ABTS+·, suggesting a better antioxidant capacity of MT than CA. Notably, compared with CA treatment, although MT treatment increased content of flavonoids, it decreased content of total phenolics from 3 d to 7 d. Based on solubilities, phenolics can be divided into soluble phenolics （flavonoids, phenolic acids and quinones） and nonsoluble compounds （hydroxycinnamic acids, lignins and condensed tannins）[29]. PPO is a key enzyme involved in enzymatic browning in many fruits and vegetables, including pear fruit[10]. Not all phenolics are the substrate of PPO[30]. This enzyme has substrate specificity for monophenols or o-diphenols[31]. In this study, compared with CA treatment, MT treatment increased content of flavonoids but decreased content of total phenolics, which might be due to the increased PPO activity in fresh-cut pears treated with MT, leading to the oxidation of other phenolic substances except flavonoids. We also found that CA combined MT treatment effectively enhanced antioxidant capacity of fresh-cut 'Early Crisp' pears compared with CA or MT treatment alone. Previous study on postharvest tomato fruit showed that CA combined chitosan treatment effectively enhanced antioxidant capacity by enhancing contents of antioxidant substances （phenolics and flavonoids） and ROS scavenging capacity compared with CA or chitosan treatment alone[26]. These studies indicated a synergistic effect of CA combined with other antioxidants on antioxidant capacity of fruits.
Browning of fresh-cut surfaces is mainly induced by enzymatic browning process, in which PAL, PPO and POD play essential roles[1]. Until now, few researches have been reported impacts of MT treatment on browning of fresh-cut products. Our results showed that MT treatment enhanced PAL activity, reduced PPO activity, increased content of total phenolics, thus decelerating browning of fresh-cut fruit 'Early Crisp' pears, which are consistent with previous findings on fresh-cut European pears （Pyrus communis L.） and fresh-cut taros[10,32]. However, studies on fresh-cut sweetpotato and fresh-cut Chinese water chestnuts indicated that MT treatment inhibited PAL and PPO activities, reduced total phenolics accumulation, thus alleviating browning[3,33]. These contradictory results suggest that the mechanisms of MT in regulation of fresh-cut browning might vary among fruits and vegetables types. It was possibly due to varying concentrations and types of phenolics found in diverse vegetables and fruits[33]. CA is a natural antibrowning agent used to prevent fresh-cut vegetables and fruits from browning[17]. Our results showed that BI in CA-treated group was lower than that in MT-treated group from 2 d to 8 d, indicating that the anti-browning effect of CA treatment was better than MT treatment. From 3 d to 7 d, CA-treated pears had lower activities of PPO and POD and higher content of total phenolics than MT-treated pears. Therefore, we speculated that, compared with MT treatment, CA treatment might reduce phenolics oxidation by reducing activities of PPO and POD, thereby effectively delaying browning of fresh-cut pears. Suttirak and Manurakchinakorn[17] indicated that combined treatment of antibrowning agents increased anti-browning efficiency compared with single treatment, which is possibly due to synergistic inhibitory mechanism of these constituents. In fresh-cut apples, CA combined AsA or oxalic acid （OA） treatment showed higher PPO inhibition and anti-browning efficiency than CA treatment alone[17]. In this work, the combined treatment group exhibited the lowest values of BI and peak of PPO activity compared with control, CA or MT treatment alone, which provides a supportive evidence for the previous finding of Suttirak and Manurakchinakorn[17]. POD plays dual roles in browning promotion and antioxidation[3]. Our results showed that both single and combined treatments reduced POD activity and BI value during storage compared with controls, which was consistent with previous findings on fresh-cut Chinese water chestnut, fresh-cut taro and fresh-cut broccoli[32−35]. However, Li et al[3] indicated that MT treatment increased POD activity and alleviated browning of fresh-cut sweetpotato during storage, and higher activity of POD induced by MT might participate in eliminating ROS produced by wounding stress. No unified mechanism appeared to explain function of POD in different fresh-cut vegetables and fruits.
TSS is an important index to evaluate quality of fruits. To date, researches on impacts of MT on TSS content of fresh-cut vegetables and fruits are limited. Magri et al[9] found that application of MT reduced TSS content of fresh-cut avocado during storage. Conversely, a study on fresh-cut Chinese water chestnuts showed that MT treatment increased TSS content during storage[33]. Zheng et al[10] reported that MT treatment had little effect on TSS content of fresh-cut European pears （Pyrus communis L.）. This article data showed that MT treatment increased TSS content of fresh-cut 'Early Crisp' pears compared with controls. These results demonstrated that the impacts of MT treatment on TSS content of fresh-cut fruits and vegetables varied among types or cultivars. Previous investigations on effects of CA on TSS content of both postharvest fruits and fresh-cut fruits are rare. Our results showed that CA treatment increased TSS content of fresh-cut 'Early Crisp' pears compared with controls during storage, as previously demonstrated in fresh-cut Chinese water chestnut and postharvest peach fruit[12,15]. However, study on tomato fruit showed that there was no significant difference in TSS content between CA treatment group and control group during storage[26]. The role of CA in regulating TSS of fresh-cut fruits during storage should be further investigated. Our work showed that combined treatment increased TSS content more effectively than CA or MT treatment alone, suggesting a synergistic impact of combined treatment on TSS content of fresh-cut 'Early Crisp' pears.
Fresh-cut processing destroys natural barrier （peel） and protective membranes, making fresh-cut products more susceptible to microbial infection and decay[36]. Both MT and CA have been reported to control microbial growth and reduce decay of fruits and vegetables[36−38]. In this study, single MT or CA treatment reduced the increase of total number of colonies during storage compared with controls, which was consistent with previous findings on fresh-cut European pears （Pyrus communis L.）, fresh-cut apples and shredded carrots[13,23,36]. The effectiveness of MT and CA treatment in controlling microbial growth might be partially ascribed to their antioxidant capacity[39]. In addition, CA was reportedly to have antimicrobial activity that affects the elasticity of bacterial biofilm[37]. Combined application of several disinfectant agents has been widely documented[36]. For example, combined CA and alkaline electrolyzed water treatment significantly reduced total bacterial counts of shredded carrots compared with each single treatment[36]. This article result showed that CA combined MT treatment had better antibacterial effect compared with CA or MT treatment alone, which was consistent with the previous finding.
Collectively, our study demonstrated that, compared with CA or MT treatment alone, MT combined with CA treatment enhanced antioxidant capacity of fresh-cut 'Early Crisp' pears by increasing contents of AsA, phenolics and flavonoids and radical scavenging activities of DPPH· and ABTS+· during cold storage. The effectiveness of this combined treatment in controlling microbial growth might be partially ascribed to its higher antioxidant capacity. It also effectively decelerated browning of fresh-cut 'Early Crisp' pears by reducing activities of PPO and POD and increasing PAL activity. This combined treatment is probably a useful strategy to preserve quality of fresh-cut products of other pear cultivars or even other fruits with similar physiological characteristics.
4.
Conclusion
MT combined CA treatment was more effective in maintaining storage quality, improving antioxidant capacity and alleviating browning of fresh-cut 'Early Crisp' pears during cold storage compared with MT or CA treatment alone. With regard to single treatment, MT treatment exhibited better effects on promoting antioxidant of fresh-cut 'Early Crisp' pears than CA treatment, whereas CA treatment showed better performance in alleviating browning of fresh-cut 'Early Crisp' pears than MT treatment. MT combined CA treatment is an effective and eco-friendly method for preservation of fresh-cut 'Early Crisp' pears. This combined treatment might potentially be applied to the preservation of other fresh-cut fruits with similar physiological characteristics. It can serve as an effective and safe preservative for fresh-cut fruits.
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