WO2019223409A1 - 一种蓝莓保鲜剂及其制备方法和应用 - Google Patents

一种蓝莓保鲜剂及其制备方法和应用 Download PDF

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WO2019223409A1
WO2019223409A1 PCT/CN2019/078610 CN2019078610W WO2019223409A1 WO 2019223409 A1 WO2019223409 A1 WO 2019223409A1 CN 2019078610 W CN2019078610 W CN 2019078610W WO 2019223409 A1 WO2019223409 A1 WO 2019223409A1
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preservative
blueberry
gallic
extract
blueberries
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PCT/CN2019/078610
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English (en)
French (fr)
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郜海燕
吴伟杰
陈杭君
房祥军
穆宏磊
韩强
韩延超
刘瑞玲
周拥军
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浙江省农业科学院
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Priority to US16/533,700 priority Critical patent/US20190357555A1/en
Publication of WO2019223409A1 publication Critical patent/WO2019223409A1/zh

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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23B—PRESERVING, e.g. BY CANNING, MEAT, FISH, EGGS, FRUIT, VEGETABLES, EDIBLE SEEDS; CHEMICAL RIPENING OF FRUIT OR VEGETABLES; THE PRESERVED, RIPENED, OR CANNED PRODUCTS
    • A23B7/00—Preservation or chemical ripening of fruit or vegetables
    • A23B7/14—Preserving or ripening with chemicals not covered by groups A23B7/08 or A23B7/10
    • A23B7/153—Preserving or ripening with chemicals not covered by groups A23B7/08 or A23B7/10 in the form of liquids or solids
    • A23B7/154—Organic compounds; Microorganisms; Enzymes
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23B—PRESERVING, e.g. BY CANNING, MEAT, FISH, EGGS, FRUIT, VEGETABLES, EDIBLE SEEDS; CHEMICAL RIPENING OF FRUIT OR VEGETABLES; THE PRESERVED, RIPENED, OR CANNED PRODUCTS
    • A23B7/00—Preservation or chemical ripening of fruit or vegetables
    • A23B7/04—Freezing; Subsequent thawing; Cooling
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs

Definitions

  • the invention relates to the technical field of green preservation and freshness preservation of fresh fruits, in particular to a green safe freshness keeping agent for blueberries, and a preparation method and application thereof.
  • Blueberry fruit has a unique flavor, and is rich in anthocyanins, flavonoids, vitamin C, folic acid, ellagic acid and other physiologically active substances. It has anti-aging, blood pressure lowering, antioxidant, anti-cancer and other health care functions.
  • One of the healthy fruits is well received by consumers and has a very broad market prospect.
  • the blueberry harvest is hot and rainy, the fruit has a high water content, a thin peel, and is extremely susceptible to decay after harvest. Its loss rate is as high as 20% to 40%.
  • blueberries are susceptible to invasion by pathogenic fungi during storage, which causes rapid changes in their structure, nutrition, and physiological and biochemical reactions. The quality of fruits is reduced and the shelf life is shortened. This severely restricts the development of the blueberry industry. Circulation brings great difficulties. Therefore, it is of great significance to reduce the loss of blueberries after doing a good job of preventing and keeping freshness of blueberries after harvest.
  • the commonly used fresh-keeping technologies mainly include spraying antibacterial agents before harvest, soaking chlorine dioxide in post-harvest aqueous solution, high-voltage electrostatic field treatment, and CO 2 controlled atmosphere irradiation for fresh-keeping.
  • Spraying with antibacterial agent and chlorine dioxide solution can effectively control the yeast and mold attached to blueberry.
  • blueberry has no obvious outer skin, and fruits are mostly eaten directly after harvest.
  • Chemical antibacterial agent has hidden safety hazards because it is not easy to remove.
  • Fresh-keeping technology has broad market prospects due to its low energy consumption, hygiene, and easy operation. Then, the treatment technology has a high voltage, has certain risks, requires high environmental humidity, and has poor repeatability of the treatment results. 2
  • the modified atmosphere fresh-keeping technology has the disadvantages of expensive facilities, large energy consumption and high cost. Therefore, the development of a low-cost, significant effect, safe and environmentally-friendly preservative is a hot topic of current research, which has important economic significance.
  • Botrytis cinerea is one of the most important pathogenic bacteria in blueberries after harvest.
  • the probability of carrying this bacteria in blueberries is as high as 81.85%.
  • gray mold fungus can easily multiply and spread rapidly, causing blueberry gray mold and causing blueberry spoilage after harvest. Therefore, the purpose of preventing and keeping freshness of blueberries can be achieved by inhibiting the reproduction of Botrytis cinerea.
  • the sulfur dioxide fumigation method has been proved to be effective in killing rot fungi such as Botrytis cinerea and Penicillium, and to reduce the incidence of gray mold during blueberry storage and transportation.
  • Galla chinensis also known as clams, white worms, wood aconites, etc.
  • clams white worms, wood aconites, etc.
  • the maggots formed in different parts are collectively referred to as having antibacterial, antioxidant and astringent effects.
  • gallica in addition to the use of gallica as a traditional Chinese medicine, various chemical products based on it are widely used in medicine, metallurgy, food, aerospace and other fields.
  • the chemical components of Galla Chinensis mainly include tannin, gallic acid, gallic oil, flavonoids, organic acids, resins, proteins, fats, starches, waxes, etc., among which Galla galanannins (CGTs) are the main functional components and Ingredients with the highest content can reach 50% -70%.
  • Gallic tannin also known as tannic acid, is a mixture of various polygallic gluconates. There are many isomers, which can exert antibacterial and antiviral effects by precipitating proteins, especially enzymes.
  • gallic tannins can effectively inhibit pathogenic bacteria such as Staphylococcus aureus, Escherichia coli, and Salmonella typhimurium, but have no inhibitory effect on fungi such as Saccharomyces cerevisiae, Rhizopus oryzae, and Trichoderma (Tian, F. , et.al. Antioxidant and antimicrobial activities from consectcutive extractions from Gallachinensis: The polarity affects the bioactivities.Food Chemistry, 2009 (1) .113 (1): 173-179; Tian, F., et.al.Identification and structure-- activity relationship of gallotannins separated from Gallachinensis.
  • the present invention studies the inhibitory effect of Galla Chinensis extract on Botrytis cinerea, and aims to develop a green preservative with Galla Chinensis extract as the main component, which is used for the prevention and treatment of postharvest diseases of blueberries, which greatly prolongs the postharvest preservation period of blueberries and improves quality.
  • one of the objectives of the present invention is to provide a blueberry preservative.
  • the preservative is composed of gallic extract, ascorbic acid and chitosan, which can effectively inhibit the main pathogenic bacterium Botrytis cinerea that causes post-harvest spoilage of blueberries, thereby exerting the effect of preventing and keeping freshness of blueberries.
  • the components of the preservative include gallic extract 0.03% -0.5%, ascorbic acid 0.5% -1.0%, and chitosan 1.0% -2.0% by weight percentage.
  • the pH of the preservative is 2.0-6.0.
  • the preparation method of the blueberry preservative is carried out according to the following steps: (1) Preparation of Galla Chinensis Extract: Take high-quality dried Galla Chinensis to remove the internal worm eggs and pulverize it completely with a high-speed powder mill; use ultrasonic extraction at 75% ethanol at room temperature; centrifuge; The supernatant was spin-evaporated to dryness. The crude extract obtained was reconstituted with water and extracted with ethyl acetate. The ethyl acetate layer was evaporated to dryness to obtain gallic extracts. (2) Gallic extracts were added to distilled water by weight percentage. 3. Dissolve ascorbic acid and chitosan; (3) After the mixed solution is thoroughly stirred and mixed, adjust the pH to between 2.0 and 6.0 to obtain the blueberry preservative.
  • the gallic extract material-to-liquid ratio is 1:10 to 1:20 (W / V)
  • the ultrasonic treatment time is 0.5 to 2 hours
  • the temperature of the vacuum rotary evaporation is 60 ° C or lower.
  • the preservative can be solubilized with 0.5% -2% Tween80 during preparation.
  • the present invention provides a method for treating blueberries by using the blueberry preservative, which can effectively reduce the blueberry fruit decay rate, extend the storage period to more than 50 days, and reduce the blueberry decay rate to less than 10%.
  • the weight loss rate is controlled within 5%, which significantly improves the quality of blueberries.
  • the method for treating blueberries by using the blueberry preservative includes the following steps: (1) washing the picked fresh blueberries with water and draining them for use; (2) placing the washed blueberries in the preservative Soak in the solution, take it out to dry, and put it into a plastic box with pores. (3) Put the blueberries treated with preservatives in a pre-cooling room to pre-cool and store them at low temperature.
  • the preservative soaking time is 1-2 minutes
  • the pre-cooling room temperature is 1 ° C-5 ° C
  • the pre-cooling time is 2-4 hours
  • the low temperature storage temperature is 1 ° C-5 ° C.
  • the present invention provides the use of a preservative for the preparation of a reagent for inhibiting Botrytis cinerea on blueberries, wherein the preservative comprises 0.03% -0.5% gallic extract, 0.5% -1.0% ascorbic acid and 1.0% -2.0% chitosan composition, the pH of the preservative is 2.0-6.0.
  • the present invention addresses the problems of easy decay, discoloration, juice, etc. of blueberries during postharvest storage.
  • the pH value of the fresh-keeping agent of the present invention is between 2.0 and 6.0, and it should be used immediately for realizing high-efficiency bacteriostatic effect.
  • the preservative of the invention has the characteristics of high efficiency, safety and environmental protection. Its main raw materials are derived from Chinese medicinal materials, and it is safe to use. The processed blueberries are free of any toxic and harmful substances.
  • the present invention provides a blueberry fresh-keeping method.
  • a blueberry fresh-keeping method By soaking blueberries in a fresh-keeping agent for a short time, pre-cooling them in a pre-cooling room, and then refrigerating them, the fresh-keeping of blueberries can be achieved.
  • the fresh-keeping method is simple in operation and short in processing time. It can effectively alleviate the membrane lipid peroxidation of blueberries after harvest, maintain the integrity of blueberry cell membranes, and extend the storage time of blueberries from 20 days of conventional refrigeration to more than 50 days.
  • the rate of blueberry decay is reduced to Within 10%, the weight loss rate is controlled within 5%, which achieves a relatively long time and high-quality fresh-keeping storage of blueberries.
  • gallic extract As the main component.
  • gallic acid one of the main components of galloid, does not play a bacteriostatic effect in the preservation and preservation of blueberries, and may be gallic tannin. Or it may be the result of the combined action of multiple factors such as pentanogalacyl glucose ( ⁇ -PGG).
  • ⁇ -PGG pentanogalacyl glucose
  • the combined use of ascorbic acid and chitosan enhances the bacteriostatic effect of Galla Chinensis extract, so that the preservative can produce a significant blueberry preservative effect.
  • Figure 1 shows the inhibition rate of Botrytis cinerea on botrytis cinerea.
  • Figure 2 is the effect of pH on the bacteriostatic stability of Galla Chinensis extract.
  • Figure 3 is the effect of temperature on the bacteriostatic stability of Galla Chinensis extract.
  • Figure 4 is the effect of irradiation time on the bacteriostatic stability of Galla Chinensis extract.
  • Fig. 5 is the effect of preservatives on the rot rate and weight loss rate of blueberry fruits, wherein (A) graph shows the blueberry fruit decay rate, and (B) graph shows the blueberry fruit weight loss rate.
  • Figure 6 is the effect of preservatives on membrane permeability and membrane lipid peroxidation of blueberry meat, where (A) shows the effect on membrane permeability and (B) shows the effect on membrane lipid peroxidation.
  • FIG. 7 shows the effect of preservative treatment on the disease resistance-related enzyme activity of blueberries, where (A) shows the effect on phenylalanine ammonialyase activity, and (B) shows the effect on polyphenol oxidase activity.
  • Galla gallica selected in the example was collected to Zhangjiajie, Hunan province; the blueberry variety was "Legsey” and was picked from the “Senzhilan” planting base in Anji County, Hangzhou City, Zhejiang province.
  • Test strains and reagents Botrytis cinerea was isolated from the picked blueberries in this laboratory. Chitosan and ascorbic acid were purchased commercially.
  • Bacteriostatic effect experiment After activating Botrytis cinerea with PDA culture medium, pick bacterial moss and make a bacterial suspension with sterile water for later use. A punch paper was used to make the absorbent filter paper with a diameter of 1 cm. After the filter paper was sterilized by dry heat, it was immersed in gallic extract, chitosan solution and ascorbic acid solution, soaked for 4 hours and dried for later use. The solid PDA medium is melted and poured into the medium. After cooling and solidifying, 0.1 mL of the bacteria suspension is added and coated uniformly. Then, the filter paper sheet impregnated with the above solution is affixed to a bacteria-containing culture dish, and Filter paper as a control. The petri dishes treated above were incubated at 28 ° C for 6 days, and the diameter of the inhibition zone was measured.
  • gallic extract has a strong inhibitory effect on Botrytis cinerea, while chitosan and ascorbic acid have no inhibitory effect on Botrytis cinerea.
  • gallic acid commercially purchased, which is one of the main components of Galla Chinensis, on Botrytis cinerea.
  • the results showed that gallic acid had no inhibitory effect on Botrytis cinerea, suggesting that the substance exhibiting antibacterial effect in gallic extract was not gallic acid.
  • Galla Chinensis extract was fully emulsified in 10% Tween80.
  • the PDA medium to be thawed is cooled to about 50 ° C, a certain volume of gallic suspension is added, and the plate is shaken and mixed thoroughly, and the plate is inverted to make the final concentration of the plate 37.5 ⁇ L / mL, 75 ⁇ L / mL, 150 ⁇ L / mL, 300 ⁇ L / mL, and 600 ⁇ L / mL.
  • a culture plate to which no gallic extract was added was used as a control.
  • Inhibition rate (%) (control colony diameter-treated colony diameter) / (control colony diameter-6mm) ⁇ 100.
  • the minimum concentration of bacterial solution with the inhibition rate of 100% is its minimum inhibitory concentration (MIC). It can be seen from FIG. 1 that the MIC of Galla Chinensis extract on Botrytis cinerea is 300 ⁇ L / mL.
  • the pH value of Galla Chinensis extract was adjusted to 2, 4, 6, 8, 10, 12 with acetic acid, and the inhibitory effects of Galla Chinensis extracts with different pH on Botrytis cinerea were compared.
  • the experimental process is as described in Example 1.
  • the pH has a greater effect on the antibacterial activity of the Galla Chinensis extract.
  • the Galla Chinensis extract gradually reduces the diameter of the zone of inhibition, and the antibacterial activity of the Galla Chinensis extract under acidic conditions. Stronger, and under alkaline conditions, gallic extracts basically lost their antibacterial activity. This is due to the fact that some compounds containing carboxyl and phenolic hydroxyl groups in the Galla Chinensis extract will dissociate to some extent with the increase of the external environment pH value, causing the original structure to be destroyed. Therefore, acidic conditions are more conducive to the antibacterial effect of gallic extracts.
  • gallic tannin is an easily hydrolyzed polyphenol compound, which can be hydrolyzed to produce intermediates such as gallic acid and pentagalloyl glucose ( ⁇ -PGG) under acidic and alkaline conditions. These intermediates are converted into each other. It is possible that the contents of gallic tannin, gallic acid and ⁇ -PGG all show corresponding changes under different pH conditions.
  • Example 2 we speculate that the substance that exerts bacteriostatic effects in gallic extract is not gallic acid, may be gallic tannin, or the result of the combined action of multiple components and multiple factors. The antibacterial effect and its antibacterial mechanism need to be further studied.
  • the bacteriostatic activity of Galla Chinensis extract at different temperatures was measured. As shown in Figure 3, the bacteriostatic activity of Galla Chinensis extract decreased as the temperature increased. Trend, but only decreased by 26.32%, Galla Chinensis extract partially degraded with increasing temperature, but due to the short treatment time, the overall decline was not obvious.
  • the high-temperature treatment in a short time has a certain effect on the activity of gallic extracts, while the long-term high-temperature treatment will cause the degradation of the active substance in the gallnuts and the structure change, which will lead to the reduction of its antibacterial activity.
  • its extraction temperature or use temperature should be lower than 60 °C.
  • the gallic extract, ascorbic acid and chitosan were dissolved in distilled water, solubilized with 2% Tween80, and after stirring and mixing, the pH was adjusted to 4.0 with acetic acid.
  • Embodiment 7 A Method for Preserving Blueberries by a Blueberry Preservative
  • a method for applying blueberry fresh-keeping agent to blueberry fresh-keeping is performed according to the following steps:
  • blueberry green preservatives 0.1% gallic extract, 0.5% ascorbic acid, 1.0% chitosan, the balance of distilled water, solubilize with 0.5% Tween80, stir and mix, and adjust the pH of acetic acid to 6.0.
  • Processing method Choose fresh blueberries with relatively consistent maturity (8-9 mature), uniform size, no mechanical injuries, no pests and diseases, rinse under clear water, drain, and soak in the preservative solution for 1min, remove and dry Then, put it in a plastic box with air holes for storage. After pre-cooling in a cold storage at 4 ° C for 3 hours, it was refrigerated at 4 ° C. Every 10 days, the fruit was checked for mold and rot until the end of storage.
  • the processing method is as follows;
  • the blueberry fruits of the treatment group and the control were tested according to the following methods, and the sampling time was 0d, 10d, 20d, 30d, 40d, 50d, 60d, and 70d, of which:
  • Fruit decay rate (%) number of rotten fruits / total number of fruits ⁇ 100%.
  • Rotten fruit means that at least one place on the surface of the fruit leaks juice, softens or rots.
  • Weight loss rate Determined by weighing method. The initial weight of the sample is subtracted from the weight measured each time during storage. The ratio of the difference to the initial weight of the sample is the weight loss rate, and the unit is expressed in%.
  • the pulp membrane permeability was studied by measuring the relative conductivity. Cut 1mm thick blueberry slices, take 20 slices after mixing, take a total of 3.0g, put in a 25mL test tube, add 25.0mL deionized water, shake for 30min on a shaker, and then use a conductivity meter to measure the solution conductivity P 1 ; After the conductivity, the solution was boiled for 10 minutes, cooled to room temperature, water was added to the original scale, and the conductivity of the solution was measured with a conductivity meter P 2 ; the conductivity of deionized water was measured P 0 .
  • the relative conductivity is calculated by the following formula, which indicates the permeability of the pulp membrane.
  • pulp phenylalanine ammonia lyase (PAL) activity assay 3mL reaction solution comprising 1.5mmol / L ascorbic acid 1mL, 0.1875mmol / L EDTA-Na l.6mL, 1mmol / L, H 2 O 2 0.3mL and 0.1 mL of enzyme solution. Starting with the addition of H 2 O 2 , the change in light absorption at 290 nm was recorded within 1 min. A change of 0.01 per minute represents one unit of enzyme activity (U).
  • the reaction system contains: 2.9 mL of 0.1 mol of catechol and 0.1 mL of the supernatant enzyme solution, and mix well. Put it in a UV spectrophotometer within 15s and measure the change in absorbance at 420nm within 3min. Take the reaction solution without adding catechol as a control. A change of 0.01 per minute indicates an enzyme unit (U).
  • the method can well inhibit water loss during the storage of blueberries, and control the weight loss rate of blueberries within 5%; the freshness keeping agent and the freshness keeping method of the invention can effectively alleviate the membrane lipid peroxidation of blueberries after harvest, and maintain the blueberry cell membrane Completeness; in addition, the present invention can significantly (P ⁇ 0.05) increase the activity of phenylalanine ammonia lyase and polyphenol oxidase related to the disease resistance of blueberries after harvest, effectively improve the disease resistance of blueberries, and prolong the storage time.
  • the application of the freshness keeping agent of the present invention can significantly inhibit the dehydration and dehydration of blueberries after harvest, alleviate membrane lipid peroxidation, improve the disease resistance of blueberries, and maintain the storage quality of blueberries.
  • Embodiment 8 A Method for Preserving Blueberries by a Blueberry Preservative
  • a method for applying blueberry fresh-keeping agent to blueberry fresh-keeping is performed according to the following steps:
  • blueberry green preservative gallic extract 0.03%, ascorbic acid 1.0%, chitosan 2.0%, the balance of distilled water, solubilize with 1.0% Tween80, stir and mix, then adjust the pH to 2.0 with acetic acid.
  • Blueberry fresh-keeping treatment method Choose fresh blueberries with relatively consistent maturity (8-9 mature), uniform size, no mechanical injury, no pests and diseases, rinse under clear water, drain, and soak in fresh-keeping agent solution for 2min, then remove After drying, store in a plastic box with air holes. After pre-cooling in a freezer at 1 ° C for 2h, it was refrigerated at 1 ° C. Every 10 days, the fruit was checked for mold and rot until the end of storage.
  • Embodiment 7 can be achieved through experimental testing, which will not be repeated one by one here.
  • Embodiment 9 A Method for Preserving Blueberries by a Blueberry Preservative
  • a method for applying blueberry fresh-keeping agent to blueberry fresh-keeping is performed according to the following steps:
  • blueberry green preservatives 0.5% gallic extract, 1.0% ascorbic acid, 2.0% chitosan, the balance of distilled water, solubilize with 2.0% Tween80, stir and mix, and adjust the pH to 4.0 with acetic acid.
  • Blueberry fresh-keeping treatment method Choose fresh blueberries with relatively consistent maturity (8-9 mature), uniform size, no mechanical injury, no pests and diseases, rinse under clear water, soak in fresh-keeping agent solution for 1.5min, After taking out and drying, put it into a plastic box with air holes for storage. After pre-cooling in a cold storage at 5 ° C for 4h, it was refrigerated at 5 ° C. Every 10 days, the fruit was checked for mold and rot until the end of storage.
  • Embodiment 7 can be achieved through experimental testing, which will not be repeated one by one here.
  • Example 10 A Method for Preserving Grapes and Black Cloth by a Blueberry Preservative
  • a method for applying blueberry freshness-preserving agent to grapes and black cloth plums is as follows:
  • blueberry green preservatives 0.5% gallic extract, 1.0% ascorbic acid, 2.0% chitosan, the balance of distilled water, solubilize with 2.0% Tween80, stir and mix, and adjust the pH to 4.0 with acetic acid.
  • Treatment methods Choose fresh grapes and black cloth plums of relatively uniform maturity (8-9 mature), uniform size, no mechanical injury, no pests and diseases, rinse them under clear water, drain them, and soak them in preservative solution 1.5min, after taking out and drying, put it into a plastic bag with air holes for storage. After pre-cooling in a cold storage at 4 ° C for 3 hours, it was refrigerated at 4 ° C. Every 10 days, the fruit was checked for mold and rot until the end of storage. A control group without preservative treatment was also set.
  • the preservatives of the present invention can significantly inhibit the dehydration and dehydration of blueberries, relieve the blueberry membrane lipid peroxidation, and significantly improve the disease resistance of phenylalanine ammonia lyase and polyphenols. Enzyme activity causes blueberry disease resistance to increase, thereby effectively maintaining the blueberry storage quality.

Abstract

一种蓝莓保鲜剂及其制备方法和应用，该保鲜剂由0.03％-0.5％五倍子提取物、0.5％-1.0％抗坏血酸和1.0％-2.0％壳聚糖组成。其制备方法包括以下步骤：（1）将五倍子以75％乙醇室温超声提取、离心，取上清液旋转蒸干，所得粗提物用水复溶后，用乙酸乙酯萃取，取乙酸乙酯层蒸干而制得五倍子提取物；（2）按重量百分比分别向蒸馏水中加入五倍子提取物、抗坏血酸、壳聚糖，进行溶解；（3）混合溶液充分搅拌混匀后，调节pH至2.0-6.0即得。将蓝莓在所述保鲜剂溶液中浸泡后，可显著抑制蓝莓灰葡萄孢霉，有效缓解蓝莓采后膜脂过氧化，使贮藏时间延长至50天以上，腐烂率降至10％以下，失重率降至5％以下。

Description

一种蓝莓保鲜剂及其制备方法和应用 技术领域
本发明涉及生鲜水果绿色防腐保鲜技术领域，具体涉及一种蓝莓的绿色安全保鲜剂及其制备方法和应用。
背景技术
蓝莓果实风味独特，且含有丰富的花青素、类黄酮、维生素C、叶酸、鞣花酸等多种生理活性物质，具有抗衰老、降血压、抗氧化、抗癌等保健功效，堪称五大健康水果之一，备受消费者青睐，具有十分广阔的市场前景。然而蓝莓采收期高温多雨，果实含水量高，果皮薄，采后极易腐烂，其损失率高达20％～40％。再加上蓝莓在储藏过程中易受到病原真菌的侵袭，致使其结构、营养和生理生化反应发生急剧变化，果实品质下降，货架期缩短，这严重制约了蓝莓产业的发展，给蓝莓的贮藏及流通带来了很大困难。因此，做好蓝莓采后防害保鲜工作对减少蓝莓损失具有重要意义。
为了缓解蓝莓果实采后腐烂变质，目前常用的保鲜技术主要有采前喷施抗菌剂、采后二氧化氯水溶液浸泡、高压静电场处理、CO 2气调辐照保鲜等。喷施抗菌剂和二氧化氯水溶液浸泡能够有效控制蓝莓附带的酵母菌和霉菌，然而蓝莓无明显外果皮，采后果实多直接食用，化学抗菌剂由于不易去除而存在安全隐患；高压静电场处理保鲜技术以其能耗低、卫生、容易操作等特点而具有广阔的市场前景，然后该处理技术电压较高，具有一定的危险性，对环境湿度要求较高，处理结果可重复性差；而CO 2气调辐照保鲜技术具有设施费用昂贵、能耗大、成本高等缺点。因此，开发一种价格低廉，效果显著，安全环保的保鲜剂是目前研究的热点，具有重要的经济意义。
灰葡萄孢霉是蓝莓采后最主要的致病菌之一，蓝莓带该菌概率高达81.85％。尤其是在雨天过后，湿度大的条件下，灰霉病病菌易迅速繁殖、流行，引起蓝莓灰霉病，导致蓝莓采后腐败。因此，通过抑制蓝莓灰葡萄孢霉得繁殖可以实现蓝莓防害保鲜的目的。二氧化硫熏蒸法被证实能有效杀死灰葡萄孢霉、青霉等致腐菌，降低蓝莓采后贮运过程中灰霉病发病率。但残留的二氧化硫具有一定的毒性，过多摄入对人体有害,且国内外对食品中的SO 2残留量都有严格的限制。植物源抑菌物质以其安全、低毒、高效等特点成为化学保鲜剂的良好替代品，已成为当下研究的热点和发展趋势。
五倍子(Galla chinensis)，又名文蛤、白虫仓、木附子等，是绵蚜科昆虫角倍蚜或者蛋倍蚜在其寄主漆树科植物盐肤木、青麸杨或红麸杨等树上不同部位形成的虫瘿统称，具有抗菌、抗氧化和收敛作用。五倍子除作为中药使用外，以其为基础原料的各种化工产品被广泛应用于医药、冶金、食品、航天等领域。五倍子的化学组分主要包括单宁、没食子酸、五倍子油、黄酮、有机酸、树脂、蛋白质、脂肪、淀粉、蜡质等，其中五倍子单宁(Chinese Gallotannins,CGTs)是五倍子的主要功能成分和含量最多的成分，含量可达50％-70％。五倍子单宁又称单宁酸，是多种聚没食子酰葡萄糖酯的混合物，存在多种异构体，可以通过使蛋白质特别是酶类沉淀而发挥抗细菌和抗病毒的作用。Tian等研究表明五倍子单宁能有效抑制金黄色酿脓葡萄球菌、大肠杆菌、鼠伤寒沙门氏菌等致病细菌；但对酿酒酵母、米根霉、木霉菌等真菌却无抑制效果(Tian,F.,et al.Antioxidant and antimicrobial activities of consecutive extracts from Galla chinensis:The polarity affects the bioactivities.Food Chemistry, 2009(1).113(1):173-179；Tian,F.,et al.Identification and structure–activity relationship of gallotannins separated from Galla chinensis.LWT-Food Science and Technology,2009(2).42(7):1289-1295)。江凯研究发现五倍子单宁对李斯特杆菌等细菌以及苹果青霉等真菌有一定抑菌效果(江凯.五倍子单宁的提取纯化及其抗菌、抗突变作用研究.陕西师范大学硕士学位论文,2011)。没食子酸(又称五倍子酸，Gallic acid)是五倍子的另一主要有效成分，临床研究表明其同样具有抗菌抗病毒作用，广泛应用于医药、食品和农业等领域。
目前关于五倍子对蓝莓病原菌灰葡萄孢霉的抑制的研究还未见相关报道。本发明通过研究五倍子提取物对灰葡萄孢霉的抑制作用，旨在开发一种以五倍子提取物为主要成分的绿色保鲜剂，用于蓝莓采后病害防治，大幅延长蓝莓采后保鲜期，提升品质。
发明内容
针对蓝莓采后病害防治现有技术存在的不足，本发明的目的之一是提供一种蓝莓保鲜剂。该保鲜剂由五倍子提取物、抗坏血酸和壳聚糖组成，其能够有效抑制引发蓝莓采后腐败的主要病原菌灰葡萄孢霉，从而发挥蓝莓防害保鲜的作用。
在一些优选方式中，所述保鲜剂的组分中，以重量百分比计，含五倍子提取物0.03％-0.5％、抗坏血酸0.5％-1.0％、壳聚糖1.0％-2.0％。
在一些优选方式中，所述保鲜剂的pH为2.0-6.0。
所述蓝莓保鲜剂的制备方法按照以下步骤进行：(1)五倍子提取物的制备：取优质干燥五倍子去除内部虫卵后经高速打粉机粉碎完全，用75％乙醇室温超声提取，离心，取上清液旋转蒸干，得到的粗提物用水复溶后，用乙酸乙酯进行萃取，取乙酸乙酯层蒸干，得到五倍子提取物；(2)按重量百分比分别向蒸馏水中加入五倍子提取物、抗坏血酸、壳聚糖，进行溶解；(3)混合溶液充分搅拌混匀后，调节pH至2.0-6.0之间，得到所述蓝莓保鲜剂。
在一些优选方式中，上述保鲜剂制备方法中，五倍子提取料液比为1:10-1:20(W/V)，超声波处理时间为0.5-2小时，真空旋转蒸发的温度为60℃以下。
在一些优选方式中，所述保鲜剂制备时可用0.5％-2％的Tween80进行增溶。
另一方面，本方面提供一种所述的蓝莓保鲜剂处理蓝莓的方法，通过该处理方法，可以有效减低蓝莓果实腐烂率，延长贮藏期至50天以上，使蓝莓腐烂率降至10％以内，失重率控制在5％以内，显著提高蓝莓品质。
在一些优选方式中，运用所述蓝莓保鲜剂处理蓝莓的方法包括以下步骤：(1)将采摘的新鲜蓝莓用清水冲洗，沥干待用；(2)将清洗后的蓝莓在所述保鲜剂溶液中浸泡，取出晾干装入带气孔的塑料盒中；(3)将保鲜剂处理过的蓝莓放入预冷库预冷后，低温保存。
在一些优选方式中，保鲜剂浸泡时间为1-2min，预冷库温度为1℃-5℃，预冷时间为2-4小时，低温保存温度为1℃-5℃。
另一方面，本方面提供一种保鲜剂在制备抑制蓝莓上的灰葡萄孢霉的试剂的用途，其中，该保鲜剂由0.03％-0.5％五倍子提取物、0.5％-1.0％抗坏血酸和1.0％-2.0％壳聚糖组成，所述保鲜剂的pH为2.0-6.0。
与现有技术相比，本发明的有益效果是：
(1)本发明针对蓝莓采后贮藏容易腐烂、变色、出汁等问题，通过研究五倍子提取物以及五倍子提取物与抗坏血酸、壳聚糖复配使用对蓝莓采后腐败的主要病原菌 灰葡萄孢霉的抑制作用，开发了一种以五倍子提取物为主成分的蓝莓绿色保鲜剂。本发明所述保鲜剂的pH值为2.0-6.0之间，并且为实现高效抑菌效果应现配现用。本发明的保鲜剂具有高效安全环保的特点，其主要原料源自中药材，使用安全，处理后的蓝莓无任何有毒有害物质。
(2)本发明提供了一种蓝莓保鲜方法，通过将蓝莓在保鲜剂中短时间浸泡，预冷库预冷之后进行冷藏即可实现蓝莓的保鲜保存。该保鲜方法操作简单，处理时间短，能有效地缓解蓝莓采后膜脂过氧化，保持蓝莓细胞膜完整性，使蓝莓的贮藏时间由常规冷藏的20天延长至50天以上，蓝莓腐烂率降至10％以内，失重率控制在5％以内，实现了蓝莓相对较长时间、高品质的保鲜贮藏。
(3)本发明所开发的保鲜剂以五倍子提取物为主要成分，通过本发明的研究表明，五倍子主要成分之一的没食子酸在蓝莓防腐保鲜中没有发挥抑菌作用，可能是五倍子单宁，或者是五没食子酰基葡萄糖(β-PGG)等多组分多种因素共同作用的结果。并且，抗坏血酸和壳聚糖的复配使用增强了五倍子提取物的抑菌效果，使得该保鲜剂能够产生显著的蓝莓防腐保鲜效果。
附图说明
图1是五倍子提取物对灰葡萄孢霉的抑制率。
图2是pH对五倍子提取物抑菌稳定性的影响。
图3是温度对五倍子提取物抑菌稳定性的影响。
图4是辐照时间对五倍子提取物抑菌稳定性的影响。
图5是保鲜剂对蓝莓果实腐烂率和失重率的影响，其中，(A)图表示蓝莓果实腐烂率，(B)图表示蓝莓果实失重率。
图6是保鲜剂对蓝莓果肉膜通透性和膜脂过氧化的影响，其中(A)图表示对膜通透性的影响，(B)图表示对膜脂过氧化的影响。
图7是保鲜剂处理对蓝莓抗病性相关酶活性的影响，其中(A)图表示对苯丙氨酸解氨酶活性的影响，(B)图表示多酚氧化酶活性的影响。
具体实施方式
下面结合实施例，更具体地说明本发明的内容。应当理解，本发明的实施并不局限于下面的实施例，对本发明所做的任何形式上的变通和/或改变都将落入本发明保护范围。
在本发明中，若非特指，所有的份、百分比均为重量单位，所有的设备和原料等均可从市场购得或是本行业常用的。若无特别指明，实施例采用的方法为本领域通用技术。
实施例中选用的五倍子，采至湖南张家界；蓝莓品种为“莱格西”，采摘于采自浙江省杭州市安吉县“森之蓝”种植基地。
实施例1五倍子提取物、壳聚糖、抗坏血酸对灰葡萄孢霉的抑制
1、五倍子提取物的制备
取优质干燥五倍子去除内部虫卵后经高速打粉机粉碎完全，用75％(V/V)乙醇，料液比1:15(W/V)室温超声提取(功率300W)1h后，离心10min，取上清液50℃旋转蒸干，得到的粗提物用水复溶后，用乙酸乙酯进行萃取，取乙酸乙酯层蒸干，得五 倍子提取物。
2、五倍子提取物、壳聚糖、抗坏血酸对灰葡萄孢霉的抑制试验
供试菌种及试剂：灰葡萄孢霉(Botrytis cinerea)于本实验室分离自所采摘的蓝莓，壳聚糖和抗坏血酸为商业途径购买。
培养基及培养条件：采用PDA培养基，28℃恒温培养6d。
抑菌效果实验：将灰葡萄孢霉用PDA培养基活化后挑取菌苔，用无菌水制成菌悬液备用。用打孔器将吸水性强的滤纸制成直径为1cm的滤纸片，滤纸片经干热灭菌后分别浸泡在五倍子提取液、壳聚糖溶液和抗坏血酸溶液中，浸泡4h后晒干备用。将固体PDA培养基熔化后倒入培养基，冷却凝固后加入0.1mL菌悬液并涂布均匀，然后将浸有上述溶液的滤纸片贴在含菌培养皿上，并用浸有无菌水的滤纸片做对照。经以上处理的培养皿在28℃恒温培养6d，测量抑菌圈直径。
表1五倍子提取液、壳聚糖、抗坏血酸、没食子酸的抑菌效果
Figure PCTCN2019078610-appb-000001
上述结果显示，五倍子提取物对灰葡萄孢霉具有较强的抑制效果，而壳聚糖和抗坏血酸对灰葡萄孢霉均无抑制效果。
为了确认五倍子提取物中发挥抑菌效果的活性物质，研究了五倍子主要成分之一的没食子酸(商业购买)对灰葡萄孢霉的抑菌效果。结果显示，没食子酸对灰葡萄孢霉并无抑制效果，说明五倍子提取物中发挥抑菌效果的物质不是没食子酸。
实施例2不同浓度五倍子提取物的抑菌效果
五倍子提取物溶于10％Tween80中充分乳化。待融化的PDA培养基冷却到50℃左右，加入一定体积的五倍子悬浮液，充分振荡混合后倒平板，使平板终浓度为37.5μL/mL、75μL/mL、150μL/mL、300μL/mL、和600μL/mL。以不添加五倍子提取物的培养基平板为对照。用无菌打孔器在培养7d的病原菌平板上打取直径为6mm菌块放在PDA平板(直径90mm)中央。置于28℃生化培养箱中培养6d，观察抑菌效果。采用十字交叉法测量菌落直径并计算抑制率。每个浓度梯度3个平板，试验重复3次。抑制率(％)＝(对照菌落直径-处理菌落直径)/(对照菌落直径-6mm)×100。以抑制率100％的最低菌液浓度为其最低抑菌浓度(MIC)。由图1可知，五倍子提取物对灰葡萄孢霉的MIC为300μL/mL。
实施例3 pH对五倍子提取物抑菌效果的影响
利用乙酸调节五倍子提取物pH值为2、4、6、8、10、12，分别对比了不同pH的五倍子提取物对灰葡萄孢霉的抑制效果，实验过程如实施例1所述。
从图2可以看出，pH对五倍子提取物的抑菌活性影响较大，随着pH值逐渐增大，五倍子提取物对抑菌圈直径逐渐降低，在酸性条件下五倍子提取物的抑菌活性较强，而在碱性条件，五倍子提取物对抑菌活性基本丧失。这是由于五倍子提取物中某些含 有羧基、酚羟基类的化合物随着外界环境pH值的增大，部分羧基、酚羟基均会发生不同程度的解离，使其原来的结构遭到破坏，因此酸性条件更有利于五倍子提取物发挥抑菌效果。
据悉，五倍子单宁是一种易水解的多酚化合物，其能在酸性、碱性条件下水解产生没食子酸、五没食子酰基葡萄糖(β-PGG)等中间产物，这些中间产物之间存在相互转化的可能，导致在不同的pH条件下，五倍子单宁、没食子酸、β-PGG的含量均呈现相应的变化。
结合实施例1的实验结果，我们推测五倍子提取物中发挥抑菌作用的物质不是没食子酸，可能是五倍子单宁，或者是多组分多种因素共同作用的结果，具体是什么组分起到了抑菌作用以及其抑菌机理还有待进一步研究。
实施例4温度对五倍子提取物抑菌效果的影响
以灰葡萄孢霉的抑菌圈直径为检测对象，测定不同温度下五倍子提取物的抑菌活性，通过图3可以看出，随着温度的升高，五倍子提取物的抑菌活性整体呈下降趋势，但仅下降了26.32％，五倍子提取物随着温度的升高部分降解，但由于处理时间短，整体下降趋势不明显。短时间内高温处理对五倍子提取物的活性有一定影响，而长时间的高温处理则会因导致五倍子中的活性物质降解，结构改变而导致其抑菌活性的降低。为保证五倍子提取物的抑菌活性，其提取温度或使用温度应低于60℃。
实施例5辐照对五倍子提取物抑菌效果的影响
辐照时间对五倍子提取物抑菌效果的影响如图4显示，随着辐照时间的增加，抑菌圈直径呈下降趋势，尤其在连续光照3天以后，抑菌圈直径下降更为明显。这是由于五倍子提取物中含有的单宁类、没食子酸等物质，在不间断的光照下的吸光值下降，导致其稳定性下降，而影响其抑菌效果。五倍子提取液在使用时做到现用现配，避免长时间曝光放置。
实施例6以五倍子提取物为主要成分的保鲜剂对灰葡萄孢霉的抑制
1、保鲜剂的制备
按照重量百分比，将五倍子提取物、抗坏血酸、壳聚糖于蒸馏水中溶解，用2％Tween80增溶，搅拌混匀后，用乙酸调节pH至4.0。
2、不同重量百分比的保鲜剂的抑菌实验
将配制而成的不同重量百分比的保鲜剂分别进行灰葡萄孢霉抑菌实验，并以五倍子提取物为对照，结果如下表所示：
表2不同配比保鲜剂的抑菌效果
Figure PCTCN2019078610-appb-000002
Figure PCTCN2019078610-appb-000003
从结果可以看出，抗坏血酸、壳聚糖与五倍子提取物的复配使用显著提高了五倍子提取物的抑菌活性。五倍子单宁具有强还原性，抗坏血酸的加入对其起到了保护作用。壳聚糖是一种良好的成膜剂，可以在水果表面形成一层相对致密的保护膜，隔绝大部分空气，抑制水果的呼吸代谢。五倍子提取物与抗氧化剂以及成膜剂相互协同，对灰葡萄孢霉产生了显著的防腐抗菌能力，从而达到对蓝莓的防腐保鲜。
实施例7一种蓝莓保鲜剂对蓝莓的保鲜方法
一种蓝莓保鲜剂应用于蓝莓保鲜的方法，按以下步骤进行：
1、五倍子提取物的制备：取优质干燥五倍子去除内部虫卵后经高速打粉机粉碎完全，用75％(V/V)乙醇，料液比1:15(W/V)室温超声提取(功率300W)1h后，离心10min，取上清液50℃旋转蒸干，得到的粗提物用水复溶后，用乙酸乙酯进行萃取，取乙酸乙酯层蒸干，得五倍子提取物。
2、蓝莓绿色保鲜剂的制备：五倍子提取物0.1％，抗坏血酸0.5％，壳聚糖1.0％，蒸馏水余量，用0.5％Tween80增溶，搅拌混匀后，调节乙酸pH至6.0。
3、处理方法：选择成熟度相对一致(8～9成熟)，大小均一，无机械伤、无病虫害的新鲜蓝莓,于清水下冲洗，沥干后，在保鲜剂溶液中浸泡1min，取出晾干后，装入带气孔的塑料盒中储藏。在4℃的冷库预冷3h后，置于4℃温度下冷藏。每隔10天检查果实发霉腐烂情况，直至贮藏结束。
进一步地，以不使用保鲜剂的贮藏方法作为对照，其处理方式如下；
选择成熟度相对一致(8～9成熟)，大小均一，无机械伤、无病虫害的新鲜蓝莓果实，于清水下冲洗，沥干后，装入带气孔的塑料盒中储藏。在4℃的冷库预冷3h后，置于4℃温度下冷藏。每隔10d检查果实发霉腐烂情况，直至贮藏结束。
4、按照下述方法对处理组和对照的蓝莓果实分别进行检测，取样时间为第0d、10d、20d、30d、40d、50d、60d和70d，其中：
(1)果实腐烂率测定：果实腐烂率(％)＝腐烂果实数/总果实数×100％，烂果是指果实表面至少有一处发生汁液外漏、软化或腐烂现象。
(2)失重率：采用称重法测定，用样品的初始重量减去贮藏过程中每次称得的重量，该差值与样品的初始重量之比，即为失重率，单位用％表示。
(3)果肉膜透性测定：通过测定相对电导率来研究果肉膜透性。切取1mm厚蓝莓的薄片，混匀后取20片，共计3.0g，置于25mL试管中，加入25.0mL去离子水，在摇床上震荡30min后，用电导率仪测定溶液电导率P 1；测定电导率后，将溶液煮沸10 min，冷却至室温，加水至原刻度，用电导率仪测定溶液电导率P 2；测定去离子水电导率P 0。按下式计算相对电导率，表示果肉膜透性。
P＝(P 1-P 0)/(P 2-P 0)×100％
(4)丙二醛含量(MDA)的测定：通过MDA含量变化来表征保鲜剂对蓝莓膜脂过氧化的影响。称取1g蓝莓冻样于10mL离心管中，加入5mL 100g/L的TCA溶液，于4℃条件下10 000r/min离心20min，收集上清液。取2.0mL上清液(空白对照加入2.0mL 100g/L TCA溶液)，加入2.0mL 0.67％硫代巴比妥酸溶液，混合后沸水浴20min，分别测定450、532、600nm波长处的吸光值。
(5)果肉苯丙氨酸解氨酶(PAL)活性测定：3mL反应液包括1.5mmol/L抗坏血酸1mL，0.1875mmol/L EDTA-Na l.6mL，1mmol/L，H 2O 2 0.3mL和0.1mL的酶液。以加入H 2O 2开始,记录1min内290nm处光吸收值的变化。每分钟变化0.01表示一个酶活单位(U)。
(6)果肉多酚氧化酶(PPO)定活性测定：反应体系包含：2.9mL 0.1mol邻苯二酚、0.1mL上清酶液，混匀。15s内放入紫外分光光度计中测3min内420nm吸光度的变化，以不加底物邻苯二酚的反应液为对照，每分钟变化0.01表示一个酶活单位(U)。
测定结果如图3、4和5所示。可以看出，随着贮藏时间的延长，蓝莓果实腐烂率和失重率逐渐上升，品质逐渐下降，本发明方法能很好的降低果实腐烂率及失重率，贮藏至30d，一般贮藏方法果实腐烂率达到13.26％，而本发明方法果实尚未发生腐烂；延长贮藏期至50d，一般贮藏方法果实腐烂率达到23.00％，而使用本发明可使腐烂率降至10％以内。同时本方法能很好地抑制蓝莓贮藏过程中水分的流失，使蓝莓失重率控制在5％以内；本发明的保鲜剂及保鲜方法，能有效地缓解蓝莓采后膜脂过氧化，保持蓝莓细胞膜完整性；此外，本发明还可显著(P<0.05)提高蓝莓采后自身抗病性相关苯丙氨酸解氨酶和多酚氧化酶的活性,有效提高蓝莓抗病性，延长贮藏时间。综上所述，应用本发明所述的保鲜剂能明显抑制蓝莓采后腐烂失水，缓解膜脂过氧化，提高蓝莓抗病性，保持蓝莓的贮藏品质。
实施例8一种蓝莓保鲜剂对蓝莓的保鲜方法
一种蓝莓保鲜剂应用于蓝莓保鲜的方法，按以下步骤进行：
1、五倍子提取物的制备：取优质干燥五倍子去除内部虫卵后经高速打粉机粉碎完全，用75％(V/V)乙醇，料液比1:20(W/V)室温超声提取(功率300W)2h后，离心10min，取上清液45℃旋转蒸干，得到的粗提物用水复溶后，用乙酸乙酯进行萃取，取乙酸乙酯层蒸干，得五倍子提取物。
2、蓝莓绿色保鲜剂的制备：五倍子提取物0.03％，抗坏血酸1.0％，壳聚糖2.0％，蒸馏水余量，用1.0％Tween80增溶，搅拌混匀后，用乙酸调节pH至2.0。
3、蓝莓保鲜处理方法：选择成熟度相对一致(8～9成熟)，大小均一，无机械伤、无病虫害的新鲜蓝莓,于清水下冲洗，沥干后，在保鲜剂溶液中浸泡2min，取出晾干后，装入带气孔的塑料盒中储藏。在1℃的冷库预冷2h后，置于1℃温度下冷藏。每隔10天检查果实发霉腐烂情况，直至贮藏结束。
经实验检测可达到实施例7的技术效果，此处不再一一赘述。
实施例9一种蓝莓保鲜剂对蓝莓的保鲜方法
一种蓝莓保鲜剂应用于蓝莓保鲜的方法，按以下步骤进行：
1、五倍子提取物的制备：取优质干燥五倍子去除内部虫卵后经高速打粉机粉碎完全，用75％(V/V)乙醇，料液比1:10(W/V)室温超声提取(功率300W)0.5h后，离心10min，取上清液60℃旋转蒸干，得到的粗提物用水复溶后，用乙酸乙酯进行萃取，取乙酸乙酯层蒸干，得五倍子提取物。
2、蓝莓绿色保鲜剂的制备：五倍子提取物0.5％，抗坏血酸1.0％，壳聚糖2.0％，蒸馏水余量，用2.0％Tween80增溶，搅拌混匀后，用乙酸调节pH至4.0。
3、蓝莓保鲜处理方法：选择成熟度相对一致(8～9成熟)，大小均一，无机械伤、无病虫害的新鲜蓝莓,于清水下冲洗，沥干后，在保鲜剂溶液中浸泡1.5min，取出晾干后，装入带气孔的塑料盒中储藏。在5℃的冷库预冷4h后，置于5℃温度下冷藏。每隔10天检查果实发霉腐烂情况，直至贮藏结束。
经实验检测可达到实施例7的技术效果，此处不再一一赘述。
实施例10一种蓝莓保鲜剂对葡萄和黑布李的保鲜方法
一种蓝莓保鲜剂应用于葡萄和黑布李保鲜的方法，按以下步骤进行：
1、五倍子提取物的制备：取优质干燥五倍子去除内部虫卵后经高速打粉机粉碎完全，用75％(V/V)乙醇，料液比1:10(W/V)室温超声提取(功率300W)0.5h后，离心10min，取上清液60℃旋转蒸干，得到的粗提物用水复溶后，用乙酸乙酯进行萃取，取乙酸乙酯层蒸干，得五倍子提取物。
2、蓝莓绿色保鲜剂的制备：五倍子提取物0.5％，抗坏血酸1.0％，壳聚糖2.0％，蒸馏水余量，用2.0％Tween80增溶，搅拌混匀后，用乙酸调节pH至4.0。
3、处理方法：分别选择成熟度相对一致(8～9成熟)，大小均一，无机械伤、无病虫害的新鲜葡萄和和黑布李于清水下冲洗，沥干后，在保鲜剂溶液中浸泡1.5min，取出晾干后，装入带气孔的塑料袋中储藏。在4℃的冷库预冷3h后，置于4℃温度下冷藏。每隔10天检查果实发霉腐烂情况，直至贮藏结束。同时设置不用保鲜剂处理的对照组。
随着贮藏时间的延长，葡萄和黑布李果实腐烂率和失重率逐渐上升，品质逐渐下降。贮藏至30d，处理组葡萄果实腐烂率达到20.36％，黑布李果实腐烂率达到10.57％，果肉变软。两种水果失水明显，并且电导率与对照组相比无显著降低，即不能有效地缓解葡萄和黑布李膜脂过氧化。以上结果说明，该保鲜剂对葡萄和黑布李并没有产生明显的保鲜结果，具体原因有待进一步分析。
综合上述实施例，本发明所述的保鲜剂能明显抑制蓝莓采后腐烂失水，缓解蓝莓膜脂过氧化，显著提高蓝莓采后自身抗病性相关苯丙氨酸解氨酶和多酚氧化酶的活性而引起蓝莓抗病性增强，从而有效保持蓝莓的贮藏品质。

Claims (8)

  1. 一种蓝莓保鲜剂，其特征在于所述的蓝莓保鲜剂由0.03％-0.5％五倍子提取物、0.5％-1.0％抗坏血酸和1.0％-2.0％壳聚糖组成。
  2. 根据权利要求1所述的保鲜剂，其特征在于所述保鲜剂的pH为2.0-6.0。
  3. 一种制备权利要求1所述的保鲜剂的方法，其特征在于按照以下步骤进行：(1)五倍子提取物的制备：取优质干燥五倍子去除内部虫卵后经高速打粉机粉碎完全，用75％乙醇室温超声提取，离心，取上清液旋转蒸干，得到的粗提物用水复溶后，用乙酸乙酯进行萃取，取乙酸乙酯层蒸干，得到五倍子提取物；(2)按重量百分比分别向蒸馏水中加入五倍子提取物、抗坏血酸、壳聚糖，进行溶解；(3)混合溶液充分搅拌混匀后，调节pH至2.0-6.0之间，得到所述蓝莓保鲜剂。
  4. 根据权利要求1所述的保鲜剂，其特征在于，所述保鲜剂制备时五倍子提取料液比为1:10-1:20(W/V)，超声波处理时间为0.5-2小时，真空旋转蒸发的温度为60℃以下。
  5. 根据权利要求1所述的保鲜剂，其特征在于所述保鲜剂制备时可用0.5％-2％的Tween80进行增溶。
  6. 一种应用权利要求1所述的蓝莓保鲜剂处理蓝莓的方法，其特征在于，包括以下步骤：(1)将采摘的新鲜蓝莓用清水冲洗，沥干待用；(2)将清洗后的蓝莓在所述保鲜剂溶液中浸泡，取出晾干装入带气孔的塑料盒中；(3)将保鲜剂处理过的蓝莓放入预冷库预冷后，低温保存。
  7. 根据权利要求6所述的方法，其特征在于，所述保鲜剂浸泡时间为1-2min，预冷库温度为1℃-5℃，预冷时间为2-4小时，低温保存温度为1℃-5℃。
  8. 一种保鲜剂在制备抑制蓝莓上的灰葡萄孢霉的试剂的用途，其中，该保鲜剂由0.03％-0.5％五倍子提取物、0.5％-1.0％抗坏血酸和1.0％-2.0％壳聚糖组成，所述保鲜剂的pH为2.0-6.0。
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Publication number Priority date Publication date Assignee Title
CN104642528A (zh) * 2013-11-21 2015-05-27 中国科学院大连化学物理研究所 一种壳寡糖复配制剂及其在延长水果货架期中的应用
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Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
ZHANG JUHUA ET AL.: "Effect of cloves schisandra chinensis and chitosan compound preservation on storage quality and disease-resistant enzyme activity of blueberries", vol. 17, no. 12, 31 December 2017 (2017-12-31), pages 139 - 148 *

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