LU501468B1 - Preparation method and application of ionized chitosan-sodium alginate composite film - Google Patents

Preparation method and application of ionized chitosan-sodium alginate composite film Download PDF

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LU501468B1
LU501468B1 LU501468A LU501468A LU501468B1 LU 501468 B1 LU501468 B1 LU 501468B1 LU 501468 A LU501468 A LU 501468A LU 501468 A LU501468 A LU 501468A LU 501468 B1 LU501468 B1 LU 501468B1
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chitosan
solution
sodium alginate
ionized
ionized chitosan
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Xiujuan Tang
Xiaoxiang Han
Chen Yang
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Univ Zhejiang Gongshang
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    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
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    • C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
    • C08B37/006—Heteroglycans, i.e. polysaccharides having more than one sugar residue in the main chain in either alternating or less regular sequence; Gellans; Succinoglycans; Arabinogalactans; Tragacanth or gum tragacanth or traganth from Astragalus; Gum Karaya from Sterculia urens; Gum Ghatti from Anogeissus latifolia; Derivatives thereof
    • C08B37/0084—Guluromannuronans, e.g. alginic acid, i.e. D-mannuronic acid and D-guluronic acid units linked with alternating alpha- and beta-1,4-glycosidic bonds; Derivatives thereof, e.g. alginates
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    • A01N65/00—Biocides, pest repellants or attractants, or plant growth regulators containing material from algae, lichens, bryophyta, multi-cellular fungi or plants, or extracts thereof
    • A01N65/03—Algae
    • A—HUMAN NECESSITIES
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    • A23L29/256—Foods or foodstuffs containing additives; Preparation or treatment thereof containing gelling or thickening agents of vegetable origin from seaweeds, e.g. alginates, agar or carrageenan
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    • C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
    • C08B37/0006—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid
    • C08B37/0024—Homoglycans, i.e. polysaccharides having a main chain consisting of one single sugar, e.g. colominic acid beta-D-Glucans; (beta-1,3)-D-Glucans, e.g. paramylon, coriolan, sclerotan, pachyman, callose, scleroglucan, schizophyllan, laminaran, lentinan or curdlan; (beta-1,6)-D-Glucans, e.g. pustulan; (beta-1,4)-D-Glucans; (beta-1,3)(beta-1,4)-D-Glucans, e.g. lichenan; Derivatives thereof
    • C08B37/0027—2-Acetamido-2-deoxy-beta-glucans; Derivatives thereof
    • C08B37/003—Chitin, i.e. 2-acetamido-2-deoxy-(beta-1,4)-D-glucan or N-acetyl-beta-1,4-D-glucosamine; Chitosan, i.e. deacetylated product of chitin or (beta-1,4)-D-glucosamine; Derivatives thereof
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08L—COMPOSITIONS OF MACROMOLECULAR COMPOUNDS
    • C08L5/00—Compositions of polysaccharides or of their derivatives not provided for in groups C08L1/00 or C08L3/00
    • C08L5/04—Alginic acid; Derivatives thereof
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08L—COMPOSITIONS OF MACROMOLECULAR COMPOUNDS
    • C08L5/00—Compositions of polysaccharides or of their derivatives not provided for in groups C08L1/00 or C08L3/00
    • C08L5/08—Chitin; Chondroitin sulfate; Hyaluronic acid; Derivatives thereof

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Abstract

The preparation method of that new composite membrane comprises the following steps: (1) dissolving sodium alginate in distilled water to prepare sodium alginate solution, and swelling at room temperature for 30 min; (2) heating the solution in step (1) in water bath and stirring at a constant speed; (3) reducing the temperature of the solution in step (2) to 40 degree Celsius, and adding glycerol as a plasticizer; (4) adding ionized chitosan, and finally preparing the ionized chitosan-sodium alginate composite membrane solution with the mass concentration of ionized chitosan of 0.5 percent-1.5 percent. If the perilla anthocyanin solution is added in the third step, the perilla anthocyanin-ionized chitosan-sodium alginate composite film can be prepared. The invention has the characteristics of high dipping film-forming speed, good bacteriostasis and so on, and has wide source of raw materials, low price and wide application prospect.

Description

Description LUS01468 Preparation method and application of ionized chitosan-sodium alginate composite film
TECHNICAL FIELD The invention relates to the technical field of new fresh-keeping materials, in particular to a preparation method and application of a new composite fresh-keeping film of ionized chitosan.
BACKGROUND Litopenaeus vannamei, also known as white-footed shrimp, belongs to the genus Penaeus, with fresh meat and rich nutrition. It is one of the three species with the highest output of cultured shrimp in the world today, and also the main target of shrimp culture in China. However, because of its high water content and strong endogenous enzyme activity, Litopenaeus vannamei is easy to rot and turn black during fishing and refrigerated transportation, so it is very important to explore the new insurance technology of Litopenaeus vannamei.
Using active ingredients in natural plants as preservatives is the development trend of fresh- keeping technology in recent years. Perilla leaves contain many active ingredients, and anthocyanin is a component with high content, which has the functions of antioxidation, anti-aging, antiallergic, antibacterial and anti-tumor. Chitosan is a natural cationic polysaccharide, which has the characteristics of non-toxicity, antibacterial, antioxidant, chemical modifiability and high reactivity. It is widely used in many fields such as chemical industry, medicine, food and biotechnology. Chitosan is soluble in acidic environment, but insoluble in water and alkaline environment, which hinders its wide application. Therefore, in view of its solubility and bacteriostasis, it was modified by ionic liquid, so that it could be dissolved in alkaline solution. On the basis of retaining the original properties of chitosan, it endowed chitosan with new functions, thus expanding the application range of chitosan.
Coating fresh-keeping is another fresh-keeping technology that has attracted the attention of researchers at home and abroad in recent years. Biodegradable biomacromolecules are used as the film-forming substrate, and antiseptic and fresh-keeping components are used as the main components to form the film together, and the barrier properties of the film are used for biological fresh-keeping. At present, the film preservation technology of Litopenaeus vannamei is rarely reported.
SUMMARY
The purpose of the present invention is to provide a preparation method of ionic liqut/501468 chitosan as a new fresh-keeping material by using ionized chitosan and natural antibacterial components. The method has the advantages of simple equipment, simple process, mild action conditions, good repeatability, strong bacteriostasis and good fresh-keeping effect, and provides some insights for the subsequent high value.
The purpose of the invention is realized by the following technical scheme: a preparation method of an ionized chitosan-sodium alginate composite film comprises the following steps: (1) dissolving sodium alginate in distilled water to prepare sodium alginate solution, wherein the ratio of sodium alginate to distilled water is 1%(g/m), and swelling at room temperature for 30 min to obtain a swollen solution; (2) heating the swollen solution in step (1) in a 80°C water bath and stirring at a constant speed for 2 hours; (3) after lowering the temperature of the heated solution in step (2) to 40°C, add glycerin as plasticizer, and the ratio of glycerin to solution is 0.5% (v/v); (4) adding ionized chitosan, and finally preparing the ionized chitosan-sodium alginate composite membrane solution with the mass concentration of ionized chitosan of 0.5%-1.5%.
Furthermore, it is characterized in that the ionized chitosan in step (4) is sulfonated chitosan.
Further, it is characterized in that the preparation process of ionized chitosan is as follows: (a) taking chitosan as raw material, add three times (g/mL) formic acid solution and 12 times (g/mL) distilled water into a three-necked bottle with a volume of 1000 mL, stir at 70°C until the raw material chitosan is dissolved, and then add four times (g/mL) formaldehyde solution. After 5 days of reaction, that mix solution in a three-necked flask was filtered to obtain filtrate. If the raw material chitosan is chitosan with a molecular weight of 10,000 or more, first alkalize it with 1 mol/L sodium hydroxide, after being pasty liquid, then wash it with distilled water to remove impurities, and then acidify it with 1 mol/L hydrochloric acid solution to pH 4 to obtain mixed solution I with a concentration of 5 mg/mL; drop the filtrate or mixed solution I into anhydrous ethanol solution to precipitate white flocculent precipitate, centrifuge at 9000 r/min at 4°C for 8 min to obtain white precipitate I, dissolve in distilled water to obtain a concentration (mg/mL) of mg/mL, put it into MD55 dialysis belt for dialysis for three days (change distilled water twice a day), after freezing in the refrigerator at -80°C for 6 hours, freeze-drying at-50 to - 70°C for 24 hours with a freeze dryer, powdered dimethyl chitosan can be obtained, and the temperature 49501468 preferably-60°C.
(b) Dimethyl chitosan, 80 times volume (g/mL) of reaction medium, 25 times volume (g/mL) of distilled water and 2 times volume (g/mL) of sulfonation reagent were put into a 250 mL three- necked flask, stirred and reacted at 60°C for 6 hours to obtain mixture II, then dropped into acetone solution to precipitate white flocculent precipitate, centrifuge at 9000 r/min at 4°C for 8 min to obtain white precipitate II. Dissolve distilled water to a concentration (mg/mL) of 5 mg/mL, put it into MD55 dialysis belt for dialysis for three days, freeze it in a refrigerator at -80°C for 6 hours, and freeze-dry it with a freeze-dryer at-60°C for 24 hours, and then get powdered ionized chitosan.
Further, at the end of step (3), a perilla anthocyanin solution with a concentration of 0.6 mg/mL is added, and the ratio of perilla anthocyanin solution to solution is 7% (v/v); after adding ionized chitosan, it can be finally prepared into perilla anthocyanin-ionized chitosan-sodium alginate composite membrane solution with the mass concentration of ionized chitosan of 0.5%-
1.5%.
Furthermore, perilla anthocyanins are anthocyanins extracted from perilla leaves. The extraction process is as follows: in the nitrogen atmosphere, the ternary deep eutectic solvents ChCl-ethylene glycol-lactic acid (molar ratio 1:3:0.7) prepared from choline chloride, ethylene glycol and lactic acid is used as the extraction solvent, and the anthocyanins from perilla leaves are extracted with the assistance of ultrasonic-microwave-ultraviolet light, wherein the ultrasonic power is 350W, the microwave power is 416W, and the ultraviolet light intensity is 10 uW/cm?. The extraction process greatly shortened the extraction time, the extraction rate of anthocyanin was 613.9 mg/100 g within 25 minutes, and the solvent could be reused for more than 6 times. Compared with the traditional method, the obtained product had smaller MIC and MBC values.
The application of the prepared ionized chitosan-sodium alginate composite film is used for fresh-keeping film, and the fresh-keeping effect of perilla anthocyanin-ionized chitosan-sodium alginate composite film is higher than that of ionized chitosan-sodium alginate composite film.
Compared with the prior art, the invention has the following beneficial effects: with ionized chitosan and perilla anthocyanin as the active components and sodium alginate as the ligand, a new type of composite coating film was prepared. The invention has the characteristics of high dipping film forming speed, good bacteriostasis and so on, and has wide raw material sources and low price. When the fresh-keeping coating is applied to the fresh-keepirt4/©01 468 of Litopenaeus vannamei, the total number of rotten colonies is reduced by more than 20%, which effectively realizes the biological storage and fresh-keeping of Litopenaeus vannamei, can realize industrial application and has a broad application prospect. The film-forming matrix in the new composite film material is not limited to sodium alginate, but other edible film-forming materials that meet the national standard can also be used.
DESCRIPTION OF THE INVENTION In order to better explain the technical scheme of the present invention, the present invention will be further described in detail with examples below.
Fresh Litopenaeus vannamei was transported to the laboratory. After sudden death with ice, the Litopenaeus vannamei were soaked in the prepared membrane solution for 30s. After being taken out, the prawns were put into a clean tray, and the membrane formation was accelerated under the air-conditioning wind. Then, the prawns were put into sterile sealed bags and refrigerated in a 4°C refrigerator. Taking the sample without film as control, it was repeated three times, and the samples were taken at 0, 1, 2, 3, 4, 5, 6 and 7 days after storage, and the related indexes were determined.
(1) Determination of texture Test room temperature is 25°C, constant test speed is 1.0 mm/s, sample deformation rate 1s 50%, measurement cycle time is 3 s, and trigger force is 0.05 N. Each group measured 5 shrimp meat samples each time, and got the average value.
(2) Determination of weight loss rate Measurement of weight loss rate by calculating the quality change of shrimp samples before and after storage, the weight loss rate is calculated as follows: weight loss rate (%) = (Mo-MT)/Mo Mo is the quality of shrimp at 0 days, and my is the quality of shrimp at 1, 2, 3, 4, 5, 6 and 7 days of storage.
(3) Determination of chromatic aberration The values of L*, a*, b* of the second ventral node of Litopenaeus vannamei were measured by colorimeter, and each group of experiments was parallel for 5 times, and the results were averaged. L*: for brightness, a*: for red and green, and b* for yellow and blue. The overall color difference is calculated according to the formula: LUS01468 AE= |(La — Lo)” + (an — ao)” + (bn — bo)” where: AE, the overall color difference; Ly, the L* value of the nth day; Lo, the L* value of day 0; an, the a* value of the nth day; ao, the a* value of day 0; bn, the b* value of the nth day; bo, the b* value of day 0.
(4) Determination of pH value Weigh 5 g of minced shrimp meat, add 45 ml of ultrapure water, homogenize, stand for 30 min, filter, and measure its pH value.
(5) Determination of thiobarbituric acid value (TBA) Take 5 g chopped shrimp meat, add 25 ml of 7.5% trichloroacetic acid solution, homogenize for 4 min, and centrifuge at 4°C and 5000 r/min for 5 min. Take 5 ml filtrate and 0.02 mol/L thiobarbituric acid solution at a volume ratio of 1:1, water bath at 80°C for 40 min, cool, and centrifuge at 5000 r/min for 10 min. Take the supernatant, add 5 ml chloroform, stand for layering, and measure the absorbance value A at 532 nm and 600 nm with ultraviolet spectrophotometer.
TBA value is expressed by the content of malondialdehyde, TBA= a * 100 (6) Determination of Total Viable Count (TVC) According to GB4789.2—2016 "Determination of the total number of colonies in food microbiological examination of national food safety standards", the total number of colonies of Litopenaeus vannamei was determined. Weigh 5 g of shrimp meat, add 45 ml of sterilized normal saline, select three appropriate dilutions, calculate the total number of colonies in shrimp meat with plate counting agar, pour two plates for each dilution, and select plates with 30-300 colonies for counting. The culture temperature is (30+1)°C and the culture time is (7243) h.
(7) Determination of total volatile basic nitrogen (TVB-N) According to GB5009.228—2016 "Determination of volatile basic nitrogen in food safety national standard", the TVB-N value of fresh-keeping treated Litopenaeus vannamei during 4°C cold storage was determined by semi-micro nitrogen determination method. Each group is repeated for 3 times, and the average value is taken.
Example 1 Taking Litopenaeus vannamei as the target, the antibacterial and fresh-keeping properties of the composite membrane of ionized chitosan and sodium alginate were studied. The results are shown in Table 1-7. LUS01468 Table 1 texture properties Ionized chitosan Storage duration/d mass 0 1 2 3 4 5 6 7 concentration (%) blank | elastic 0.85+ 0.76+ 0.70+ 0.65+ 0.58+ 0.51% 0.43+ 0.36+
0.06 0.07 0.08 0.06 0.07 0.08 0.06 0.09 hardness | 417.08+ | 405.264 | 390.27+ | 371.56+ | 348.55= | 326.58+ | 305.724 | 283.57+
11.28 8.13 13.42 11.31 14.72 8.97 9.34 8.22
0.5 clastic 0.85+ 0.77+ 0.74+ 0.68+ 0.63+ 0.57+ 0.51+ 0.46+
0.06 0.08 0.07 0.12 0.09 0.08 0.09 0.07 hardness | 417.08 | 404.364 | 392.52+ | 378.31+ | 363.983 | 346.13+ | 326.994 | 304.12+
11.28 15.36 14.13 11.82 13.94 14.55 16.23 12.41 1 clastic 0.85+ 0.78+ 0.73+ 0.69+ 0.65+ 0.61+ 0.55+ 0.49+
0.06 0.07 0.05 0.08 0.06 0.07 0.09 0.08 hardness | 417.08+ | 405.12+ | 395.51+ | 383.41+ | 368.22+ | 350,29+ | 330.12 | 306.73+
11.28 12.83 13.57 11.67 14.11 15.36 12.56 13.34
1.5 elastic 0.85+ 0.81+ 0.75+ 0.71+ 0.66+ 0.62+ 0.58+ 0.54+
0.06 0.04 0.08 0.03 0.07 0.09 0.06 0.05 hardness | 417.08 | 408.404 | 396.83+ | 383.92+ | 369.78 | 353.13 | 332.764 | 310.52+
11.28 13.53 12.92 14.46 15.53 15.36 13.57 14.99 From Table 1, it can be concluded that there is no significant difference in hardness and elasticity of the experimental group at the early stage of storage. On the 3rd day or so, the elasticity and hardness of the blank control group and the treatment group are quite different, and the longer the storage time, the more obvious the difference. The elasticity and hardness of the treatment group were higher than those of the control group, which indicated that the coating of membrane could effectively slow down the deterioration of texture of Litopenaeus vannamei, and the higher the content of ionized chitosan, the more obvious the effect.
Table 2 weight loss rate (%) LUS01468 Ionized chitosan Storage duration/d mass 0 1 2 3 4 5 6 7 concentration (%) blank 0.00+ | 1.11+ | 1.83+ | 233+ | 297+ | 3.57 | 4.04+ | 4.53+ Weight 0.00 | 007 | 013 | 011 | 021 | 025 | 0.15 | 0.08 loss 0.5 0.00+ | 0.81+ | 1.274 | 1.714 | 2.26+ | 2.78+ | 3.07+ | 3.43+
0.00 0.13 0.17 0.24 0.08 0.25 0.15 0.31 1 0.00+ | 0.68+ | 1.13 | 1.694 | 2.17+ | 2.62+ | 2.98+ | 3.38%
0.00 0.14 0.09 0.15 0.27 0.22 0.19 0.26
1.5 0.00+ | 055+ | 0.964 | 1.57 | 2.08 | 2.57 | 2.89+ | 3.284
0.00 0.21 0.14 0.26 0.13 0.17 0.33 0.23 Fresh shrimps are usually susceptible to microbial, physical and biochemical influences due to their high water content, rich free amino acids and other non-protein nitrogen compounds during storage after death. During the deterioration of shrimp, water loss, oxidative degradation of protein, oil and other substances will inevitably lead to the decline of quality. As can be seen from Table 2, the weight loss rate of the treatment group 1s generally smaller than that of the control group, which indicates that the coating of film can effectively reduce the water loss and spoilage of shrimp, while the higher the content of chitosan, the lower the weight loss rate of the treatment group, because of the film-forming property and bacteriostasis of ionized chitosan itself, the formed film will be tighter and the bacteriostatic and fresh-keeping effect will be better. Table 3 color difference Ionized Storage duration/d chitosan mass 0 1 2 3 4 5 6 7 concentration (%) blank 0.00+ | 2.04+ | 3.05+ | 4.03+ | 5.21+ | 6.52+ | 7.96+ 9.72+
0.00 0.06 0.14 0.13 0.24 0.32 0.14 0.29 AE 0.5 0.00+ | 1.84 | 240+ | 2.88 | 3.49+ | 4.16+ | 494+ 5.69+
0.00 0.13 0.18 0.25 0.29 0.09 0.15 0.27 1 0.00+ | 1.694 | 2244+ | 2.764 | 3.34% | 4.02+ | 4.744 547+
0.00 0.18 0.23 0.14 0.08 0.32 0.24 0.22
1.5 0.00+ | 1.58 | 2.08+ | 2.50+ | 3.08+ | 3.66+ | 451+ 531+
0.00 0.11 0.08 0.18 0.27 0.16 0.37 0.23 The AE value indicates the total color difference of shrimp. The larger the AE value, the greater the color change of shrimp. From Table 3, it can be seen that with the increase of storage time, the AE value also increases, indicating that shrimp is gradually rotting, and the AE value of the treatment group is lower than that of the blank control group. Therefore, the complex fresh}501468 keeping film treatment group can effectively delay the color change of shrimp. Table 4 pH value Ionized chitosan Storage duration/d mass 0 1 2 3 4 5 6 7 concentration(%o) blank 694+ | 7.17 | 724+ | 7.394 | 7.62+ | 7.764 | 7.854 | 7.93%
0.05 0.13 0.07 0.04 0.05 0.07 0.12 0.06
0.5 694+ | 7.144 | 7.21+ | 7.324 | 7.48+ | 7.60+ | 7.694 | 7.774 pH 0.05 0.10 0.06 0.11 0.04 0.05 0.09 0.06 1 694+ | 7.13% | 7.15+ | 7.264 | 7.384 | 7.55 | 7.614 | 7.73%
0.05 0.05 0.03 0.10 0.12 0.06 0.08 0.13
1.5 6.94+ | 7.114 | 7.13+ | 7.23+ | 730+ | 7.464 | 7.594 | 7.69+
0.05 0.03 0.04 0.05 0.02 0.09 0.07 0.11 As we all know, the reason why the pH value of shrimp increases during storage is that protein and other nitrogen-containing substances are decomposed into volatile bases such as amines and trimethylamine under the action of microorganisms and endogenous enzymes. Therefore, measuring the pH value of each experimental group during storage period can effectively reflect the degree of deterioration of shrimp samples. As can be seen from Table 4, the pH value of the experimental group increased continuously during storage. On the whole, the pH value of the treatment group is lower than that of the blank control group, and the higher the content of ionized chitosan, the lower the pH value, which indicates that the film has strong bacteriostasis, thus effectively reducing the decomposition of nutrients such as protein, while the higher the content of ionized chitosan, the better the bacteriostasis. Table 5 TBA (mg MDA/kg) Ionized chitosan Storage duration/d mass 0 1 2 3 4 5 6 7 concentration(%o) blank 031+ | 0.70+ | 1.11% | 1.71+ | 2.53+ | 3.00+ | 3.59+ | 3.94+
0.07 0.08 0.13 0.09 0.05 0.10 0.07 0.12
0.5 031+ | 0.674 | 099+ | 1.54+ | 2.45+ | 2.95+ | 3.274 | 3.644 TBA 0.07 | 003 | 006 | 007 | 008 | 003 | 005 | 002 1 031+ | 0.60+ | 0.98 | 1.40+ | 2.03+ | 2.60+ | 3.094 | 3.30+
0.07 0.05 0.03 0.08 0.09 0.15 0.03 0.06
1.5 031+ | 0.60+ | 0.94+ | 1.38+ | 1.81+ | 2.364 | 2.774 | 3.104
0.07 0.08 0.04 0.05 0.09 0.06 0.05 0.11 It can be seen from Table 5 that the TBA value of fresh shrimp meat treated with preservative is about 0.31 mg MDA/kg, and the TBA value of each group is on the rise. It is mainly due to tH#/501468 formation of peroxides and final products with the deepening of the oxidation degree of polyunsaturated fatty acids in shrimp meat, which leads to the increasing TBA value. On the whole, the TBA value of the treatment group is lower than that of the blank control group, indicating that the composite membrane can effectively reduce the oxidation degree of Litopenaeus vannamei fat, and because the addition of ionized chitosan makes the composite membrane more compact, the higher the content of ionized chitosan, the better the oxygen isolation effect and the lower the TBA value. Table 6 Total number of colonies (TVC) (Ig (CFU/g)) Ionized chitosan Storage duration/d mass 0 1 2 3 4 5 6 7 concentration(%o) blank 3.68+ | 423+ | 479+ | 331+ | 5.95 | 6.88 | 7.83+ | 8.52
0.10 0.09 0.13 0.17 0.07 0.11 0.25 0.18
0.5 3.68+ | 4.03+ | 437+ | 4.77 | 5.164 | 5.62+ | 627+ | 6.92+ TVC 0.10 0.12 0.08 0.17 0.05 0.15 0.12 0.03 1 3.68+ | 3.97+ | 428+ | 4.65 | 5.08% | 549+ | 6.03+ | 6.80%
0.10 0.07 0.12 0.13 0.14 0.24 0.13 0.13
1.5 3.68+ | 3.88% | 4.12+ | 4.58 | 5.05+ | 546+ | 5.98 | 6.744
0.10 0.16 0.14 0.05 0.17 0.18 0.08 0.10 See Table 6 for the change of the total number of colonies of Litopenaeus vannamei during storage. Generally, when the total number of bacteria reaches 61 g (CFU/g), it is considered to have reached the corruption limit. The initial total number of colonies in Litopenaeus vannamei was
3.681 g (CFU/g), and the total number of colonies in each experimental group increased with the increase of time. The blank control group increased the fastest, and the total number of bacteria on the second day was larger than that on the third day of the treatment group, which exceeded the corruption limit on the fifth day. On the 6th day, most of the experimental groups had exceeded the limit, while the treatment group with high content of ionized chitosan was still within the limit. It shows that the addition of ionized chitosan increases the bacteriostasis of the membrane, and the higher the content of chitosan, the better the bacteriostasis.
Table 7 Total volatile basic nitrogen (TVB-N) (mg/100g) LU501468 Ionized chitosan Storage duration/d mass 0 1 2 3 4 5 6 7 concentration (%) blank 7.674 | 12.67+ | 143+ | 18.17+ | 26.63+ | 35.40+ | 46.174 | 52.40+
0.31 0.62 0.34 0.35 0.35 0.41 0.55 0.46 TVB- 0.5 7.67+ | 11.86+ | 13.22+ | 14.53+ | 18.16% | 20.30+ | 27.61+ | 32.14+ N 0.31 0.46 0.33 0.52 0.38 0.41 0.61 0.34 1 7.674 | 11.164 | 12.60+ | 13.304 | 17.36+ | 18.90% | 27.06 | 31.23+
0.31 0.39 0.42 0.47 0.52 0.36 0.52 0.44
1.5 7.674 | 11.11+ | 11.13+ | 12.524 | 16.04+ | 18.62+ | 26.45+ | 31.074
0.31 0.41 0.37 0.39 0.51 0.46 0.53 0.66 Total volatile basic nitrogen (TVB-N) is dimethylamine, trimethylamine and other ammonia compounds produced by the decomposition of animal food under the action of endogenous enzymes and microorganisms, and it is one of the commonly used indexes to evaluate the spoilage degree of aquatic products. The acceptable range of the content of TVB-N in seawater is not more than 30 mg/100g, among which, when the content of TVB-N is not more than 15 mg/100g, it is the first grade freshness, when it is not more than 20 mg/100g, it is the second grade freshness and when it is not more than 30 mg/100g, it is the third grade freshness.
It can be seen from Table 7 that the TVB-N value of each group showed an upward trend with the extension of storage time. On the 5th day, the TVB-N value of the blank control group has exceeded the specified range, while the low ionized chitosan content treatment group has the third grade freshness, and the high ionized chitosan content treatment group has the second grade freshness. On the 6th day, all the treatment groups were of the third grade freshness; on the 7th day, the TVB-N values of all treatment groups exceeded the specified range. This shows that the complex fresh-keeping film can significantly delay the rise of TVB-N in Litopenaeus vannamei, and the higher the content of ionized chitosan, the more obvious the effect of delaying the rise of TVB-N.
Example 2 The antibacterial and fresh-keeping performance of Perilla anthocyanin-ionized chitosan- sodium alginate composite film on Litopenaeus vannamei was studied. The results are shown in Table 8-14.
Table 8 texture properties
Ionized chitosan Storage duration/d LU501468 mass 0 1 2 3 4 5 6 7 concentration (%) blank | elastic 0.85+ 0.76+ 0.70 0.65+ 0.58+ 0.51% 0.43+ 0.36+
0.06 0.07 0.08 0.06 0.07 0.08 0.06 0.09 hardness | 417.08+ | 405.26+ | 390.27+ | 371.56% | 348.55+ | 326.58+ | 305.72+ | 283.574
11.28 8.13 13.42 11.31 14.72 8.97 9.34 8.22
0.5 clastic 0.85+ 0.81+ 0.78 0.73 0.68+ 0.64+ 0.59+ 0.56+
0.06 0.04 0.08 0.07 0.09 0.06 0.04 0.03 hardness | 417.08+ | 409.17+ | 405.31+ | 401.41% | 388.46+ | 373.17+ | 361.42+ | 344.24+
11.28 15.32 13.56 11.82 14.15 12.97 13.21 14.54 1 clastic 0.85+ 0.82+ 0.79+ 0.75% 0.71+ 0.68+ 0.63+ 0.60+
0.06 0.06 0.07 0.05 0.07 0.09 0.08 0.06 hardness | 417.08+ | 413.56+ | 409.85+ | 402.31+ | 391.44+ | 379.97+ | 364.21+ | 348.344
11.28 16.22 13.97 15.85 14.96 12.84 13.21 11.05
1.5 elastic 0.85+ 0.84+ 0.82+ 0.80+ 0.78+ 0.75+ 0.72+ 0.66+
0.06 0.05 0.06 0.07 0.08 0.05 0.09 0.04 hardness | 417.08+ | 415.34+ | 412.11 | 407.73 | 396.21 | 380.86 | 367.35+ | 355.134
11.28 12.04 13.97 15.23 14.08 15.87 14.23 16.24 Table 8 shows that there is no significant difference in hardness and elasticity of the experimental group at the early stage of storage. On the second day or so, the elasticity and hardness of the blank control group and the treatment group are quite different, and the longer the storage time is, the more obvious the difference 1s. The elasticity and hardness of the treatment group were higher than those of the control group, which indicated that the coating of membrane could effectively slow down the deterioration of texture of Litopenaeus vannamei, and the higher the content of ionized chitosan, the more obvious the effect. Compared with the ionized chitosan- sodium alginate composite film, the elasticity and hardness of the composite film are basically higher than those of the experimental group treated with the ionized chitosan-sodium alginate composite film at different concentrations, which indicates that the addition of perilla anthocyanin can effectively preserve the freshness of Litopenaeus vannamei and better slow down the deterioration of the texture of Litopenaeus vannamei.
Table 9 weight loss rate (%) LUS01468 Ionized chitosan Storage duration/d mass 0 1 2 3 4 5 6 7 concentration (%) blank 0.00+ | 1.11+ | 1.83+ | 2.33+ | 297+ | 3.57 | 4.044 | 4.534 Weight 0.00 | 0.07 | 0.13 | 0.11 | 0.21 | 0.25 | 0.15 | 0.08 loss 0.5 0.00+ | 0.734 | 1.33+ | 1.72+ | 2.08 | 238+ | 2.79+ | 3.194
0.00 0.09 0.11 0.24 0.17 0.15 0.28 0.19 1 0.00+ | 048+ | 1.014 | 148+ | 191+ | 227+ | 2.69+ | 3.164
0.00 0.16 0.13 0.05 0.28 0.21 0.07 0.27
1.5 0.00+ | 040+ | 0.85+ | 127+ | 1.70+ | 2.15+ | 2.594 | 3.054
0.00 0.08 0.22 0.24 0.18 0.15 0.11 0.23 Fresh shrimps are usually susceptible to microbial, physical and biochemical influences due to their high water content, rich free amino acids and other non-protein nitrogen compounds during storage after death. During the deterioration of shrimp, water loss, oxidative degradation of protein, oil and other substances will inevitably lead to the decline of quality. As can be seen from Table 9, the weight loss rate of the treatment group is generally smaller than that of the control group, which indicates that film wrapping can effectively reduce the water loss and spoilage of shrimp, while the higher the content of ionized chitosan, the lower the weight loss rate of the treatment group. Because of the film-forming property and bacteriostasis of the ionized chitosan itself, the formed film will be tighter and the bacteriostatic and fresh-keeping effect will be better. Compared with the ionized chitosan-sodium alginate composite film, the weight loss rate of the composite film is basically lower than that of the experimental group treated with the ionized chitosan-sodium alginate composite film at different concentrations, which indicates that the addition of perilla anthocyanin can effectively preserve the freshness of Litopenaeus vannamei and reduce the change of shrimp weight.
Table 10 color difference LUS01468 Ionized Storage duration/d chitosan mass 0 1 2 3 4 5 6 7 concentration (%) blank 0.00+ | 2.04+ | 3.05+ | 4.03+ | 5.21+ | 6.52+ | 7.96+ 9.72+
0.00 0.06 0.14 0.13 0.24 0.32 0.14 0.29 AE 0.5 0.00+ | 1.43+ | 1.88% | 2.50+ | 3.11 | 3.79+ | 4.434 5.16%
0.00 0.03 0.14 0.16 0.22 0.18 0.27 0.19 1 0.00+ | 1.01+ | 1.54+ | 2.13+ | 2.66+ | 3.32+ | 4.074 4.82+
0.00 0.15 0.19 0.24 0.08 0.11 0.22 0.31
1.5 0.00+ | 0.78+ | 1.21+ | 1.96+ | 2.54+ | 3.16+ | 3.914 4.62+
0.00 0.09 0.21 0.18 0.13 0.33 0.27 0.16 The AE value indicates the total color difference of shrimp. The larger the AE value, the greater the color change of shrimp. From Table 10, it can be seen that with the increase of storage time, the AE value also increases, indicating that shrimp is gradually rotting, and the AE value of the treatment group is lower than that of the blank control group. Therefore, the complex fresh- keeping film treatment group can effectively delay the color change of shrimp. Compared with the ionized chitosan-sodium alginate composite film, the AE value of this composite film is basically lower than that of the experimental group treated with ionized chitosan-sodium alginate composite film at different concentrations, which indicates that the addition of perilla anthocyanin can better delay the color change of shrimp. Table 11 pH value Ionized chitosan Storage duration/d mass 0 1 2 3 4 5 6 7 concentration (%) blank 6.94+ | 717+ | 724+ | 739+ | 7.624 | 7.764 | 7.85+ | 7.93+
0.05 0.13 0.07 0.04 0.05 0.07 0.12 0.06 pH 0.5 6.94+ | 7.11+ | 7.15+ | 7.24+ | 7.40+ | 7.50+ | 7.59+ | 7.70+
0.05 0.04 0.07 0.05 0.02 0.06 0.04 0.03 1 6.94+ | 7.07 | 7.124 | 7.20£ | 7.314 | 742+ | 7.53+ | 7.64+
0.05 0.06 0.07 0.08 0.05 0.04 013 0.05
1.5 6.94+ | 7.01+ | 7.084 | 7.14+ | 7.20£ | 7.35 | 746+ | 7.53+
0.05 0.05 0.08 0.02 0.04 0.06 0.04 0.07 As we all know, the reason why the pH value of shrimp increases during storage is that protein and other nitrogen-containing substances are decomposed into volatile bases such as amines and trimethylamine under the action of microorganisms and endogenous enzymes. Therefore,
i i I i i ffectively reflett/01468 measuring the pH value of each experimental group during storage period can effectively refle the degree of deterioration of shrimp samples. As can be seen from Table 11, the pH value of the experimental group increased continuously during storage. On the whole, the pH value of the treatment group is lower than that of the blank control group, and the higher the content of ionized chitosan, the lower the pH value, which indicates that the film has strong bacteriostasis, thus effectively reducing the decomposition of nutrients such as protein, and the higher the content of ionized chitosan, the better the bacteriostasis. Compared with the ionized chitosan-sodium alginate composite membrane, the pH value of this composite membrane is basically lower than that of the experimental group treated with the ionized chitosan-sodium alginate composite membrane at different concentrations, which indicates that the addition of perilla anthocyanin slows down the decay of Litopenaeus vannamei, reduces the generation of volatile alkali, and better preserves Litopenaeus vannamei. Table 12 TBA (mg MDA/kg) Ionized Storage duration/d chitosan mass 0 1 2 3 4 5 6 7 concentration (7) blank 031+ | 0.70 | 1.11+ | 1.71+ | 2.53 | 3.00+ | 3.594 | 3.944
0.07 0.08 0.13 0.09 0.05 0.10 0.07 0.12 TBA 0.5 031+ | 0.664 | 0.97+ | 1.49+ | 2.22+ | 2.70+ | 3.12+ | 3364
0.07 0.06 0.04 0.09 0.05 0.07 0.06 0.08 1 031+ | 0.64+ | 095+ | 1.291 | 1.76 | 2.37 | 2.754 | 3.164
0.07 0.04 0.02 0.06 0.12 0.03 0.01 0.06
1.5 031+ | 0.524 | 091+ | 1.194 | 1.56 | 2.17 | 2.534 | 2.954
0.07 0.09 0.11 0.08 0.06 0.03 0.04 0.07 It can be seen from Table 12 that the TBA value of fresh shrimp meat treated with preservative is about 0.31 mg MDA/kg, and the TBA value of each group 1s on the rise. It is mainly due to the formation of peroxides and final products with the deepening of the oxidation degree of polyunsaturated fatty acids in shrimp meat, which leads to the increasing TBA value. On the whole, the TBA value of the treatment group is lower than that of the blank control group, indicating that the composite membrane can effectively reduce the oxidation degree of Litopenaeus vannamei fat, and because the addition of ionized chitosan makes the composite membrane more compact, the higher the content of ionized chitosan, the better the oxygen isolation effect and the lower the TBA value. Compared with the ionized chitosan-sodium alginate composite film, the TBA value of this composite film is basically lower than that of the experimental group treated with the ionizé/01468 chitosan-sodium alginate composite film at different concentrations, which indicates that the addition of perilla anthocyanin can better slow down the oxidation of polyunsaturated fatty acids, thus effectively preserving Litopenaeus vannamei. Table 13 Total number of colonies (TVC) (lg (CFU/g)) Ionized Storage duration/d chitosan mass 0 1 2 3 4 5 6 7 concentration (%) blank 3.68+ | 423+ | 479+ | 531+ | 595+ | 6.88+ | 7.83+ | 8.52+
0.10 0.09 0.13 0.17 0.07 0.11 0.25 0.18 TVC 0.5 3.68+ | 3.85+ | 3.96+ | 4.12+ | 441+ | 487+ | 5.474 | 6.244
0.10 0.07 0.14 0.15 0.13 0.16 0.21 0.11 1 3.68+ | 3.78% | 3.864 | 394+ | 424+ | 4.69+ | 531+ | 6.12%
0.10 0.15 0.26 0.07 0.13 0.22 0.13 0.04
1.5 3.68+ | 3.75+ | 3.81+ | 3.89+ | 4.13 | 4.62+ | 527+ | 5.99+
0.10 0.17 0.18 0.09 0.12 0.18 0.11 0.08 See Table 13 for the change of the total number of colonies of Litopenaeus vannamei during storage. Generally, when the total number of bacteria reaches 61 g (CFU/g), it is considered to have reached the limit of corruption. The initial total number of colonies in Litopenaeus vannamei was
3.681 g (CFU/g), and the total number of colonies in each experimental group increased with the increase of time. The blank control group increased the fastest. The total number of bacteria on the second day was larger than that on the fourth day of the treatment group, and it exceeded the corruption limit on the fifth day. On the 7th day, most of the experimental groups had exceeded the limit, while the total number of colonies in the treatment group with 1.5% mass concentration of ionized chitosan was only 5.99, which was still within the spoilage limit, showing strong antibacterial effect, which may be due to the strong antibacterial effect of ionized chitosan on bacteria and strong synergistic antibacterial effect with anthocyanins, which could effectively inhibit the growth of dominant spoilage bacteria in shrimp samples. Compared with the ionized chitosan-sodium alginate composite membrane, the total number of bacteria in this composite membrane is basically lower than that in the experimental group treated with the ionized chitosan- sodium alginate composite membrane at different concentrations, which indicates that the addition of perilla anthocyanin can more effectively inhibit the growth of spoilage bacteria, thus better preserving Litopenaeus vannamei.
Table 14 total volatile basic nitrogen (TVB-N) (mg/100g) LU501468 Ionized chitosan Storage duration/d mass 0 1 2 3 4 5 6 7 concentration(%o) blank 7.671 12.674 | 143+ | 18.174 | 26.63+ | 35.401 | 46.17+ | 52.40+
0.31 0.62 0.34 0.35 0.35 0.41 0.55 0.46
0.5 7.674 | 10.60+ | 12.63+ | 13.774 | 1547+ | 18.801 | 24.63+ | 31.13% TVB- 031 | 035 | 055 | 042 | 040 | 056 | 031 | 035 N 1 7.67Æ | 9.73+ | 11.90+ | 12.53+ | 14.371 | 16.77+ | 21.69+ | 29.43%
0.31 0.62 0.70 0.60 0.50 0.70 0.54 0.31
1.5 7.67Æ | 9.20+ | 10.73+ | 11.26+ | 13.20+ | 16.70+ | 20.90+ | 28.13+
0.31 0.64 0.45 0.25 0.36 0.57 0.67 0.88 It can be seen from Table 14 that the TVB-N value of each group showed an upward trend with the extension of storage time. On the 5th day, the TVB-N value of the blank control group has exceeded the specified range, while the treatment group is the second grade freshness. On the 6th day, all the treatment groups were the third grade freshness; on the 7th day, the treatment group with low ionized chitosan content exceeded the specified range, and the treatment group with high ionized chitosan content had third grade freshness. This shows that the complex fresh-keeping film can significantly delay the rise of TVB-N in Litopenaeus vannamei, and the higher the content of ionized chitosan, the more obvious the effect of delaying the rise of TVB-N. Compared with the ionized chitosan-sodium alginate composite film, the TVB-N value of this composite film is basically lower than that of the experimental group treated with ionized chitosan-sodium alginate composite film at different concentrations, which indicates that the addition of perilla anthocyanin can better preserve the freshness of Litopenaeus vannamei.
The above is only the specific implementation case of the patent of the present invention, but the technical features of the patent of the present invention are not limited to this. Any changes or modifications made by the technicians in relevant fields within the scope of the present invention are covered by the patent scope of the present invention.

Claims (6)

CLAIMS LU501468
1. À preparation method of ionized chitosan-sodium alginate composite film, characterized in that which comprises the following steps: (1) dissolving sodium alginate in distilled water to prepare sodium alginate solution, wherein the ratio of sodium alginate to distilled water is 1%(g/m), and swelling at room temperature for 30 min to obtain a swollen solution; (2) heating the swollen solution in step (1) in a 80°C water bath and stirring at a constant speed for 2 hours; (3) after lowering the temperature of the heated solution in step (2) to 40°C, adding glycerin as plasticizer, and the ratio of glycerin to solution is 0.5% (v/v); (4) adding ionized chitosan, and finally preparing the ionized chitosan-sodium alginate composite membrane solution with the mass concentration of ionized chitosan of 0.5%-1.5%.
2. The preparation method of ionized chitosan-sodium alginate composite film according to claim 1, characterized in that the ionized chitosan in step (4) is sulfonated chitosan.
3. The preparation method of ionized chitosan-sodium alginate composite film according to claim 1, characterized in that the preparation process of ionized chitosan is as follows: (a) taking chitosan as raw material, adding three times (g/mL) formic acid solution and 12 times (g/mL) distilled water into a three-necked bottle with a volume of 1000 mL, stirring at 70°C until the raw material chitosan is dissolved, and then add four times (g/mL) formaldehyde solution; after 5 days of reaction, the mix solution in a three-necked flask was filtered to obtain filtrate; if the raw material chitosan is chitosan with a molecular weight of 10,000 or more, first alkalize it with 1 mol/L. sodium hydroxide, after being pasty liquid, then wash it with distilled water to remove impurities, and then acidify it with 1 mol/L hydrochloric acid solution to pH 4 to obtain mixed solution I with a concentration of 5 mg/mL; drop the filtrate or mixed solution I into anhydrous ethanol solution to precipitate white flocculent precipitate, centrifuge at 9000 r/min at 4°C for 8 min to obtain white precipitate I, dissolve in distilled water to obtain a concentration (mg/mL) of 5 mg/mL, put it into MD55 dialysis belt for dialysis for three days (change distilled water twice a day), after freezing at -80°C for 6 h, and freeze-drying at -50 to- 70 °C for 24 h with a freeze dryer, powdery dimethyl chitosan can be obtained, and the temperature is preferably - 60 °C,
(b) dimethyl chitosan, 80 times volume (g/mL) of reaction medium, 25 times volume (g/mE}501 468 of distilled water and 2 times volume (g/mL) of sulfonation reagent were put into a 250 mL three- necked flask, stirred and reacted at 60°C for 6 hours to obtain mixture II, then dropped into acetone solution to precipitate white flocculent precipitate, and centrifuged at 9000 r/min, 4°C for 8 min to obtain white precipitate II, dissolve distilled water to a concentration (mg/mL) of 5 mg/mL, put it into MD55 dialysis belt for dialysis for three days, freeze it in a refrigerator at -80°C for 6 hours, and freeze-dry it with a freeze-dryer at-60°C for 24 hours, and then get powdered ionized chitosan.
4. The preparation method of ionized chitosan-sodium alginate composite film according to claim 1, characterized in that at the end of step (3), a perilla anthocyanin solution with a concentration of 0.6 mg/mL is added, and the ratio of perilla anthocyanin solution to solution is 7% (v/v); after adding ionized chitosan, it can be finally prepared into perilla anthocyanin-ionized chitosan-sodium alginate composite membrane solution with the mass concentration of ionized chitosan of 0.5%-1.5%.
5. The preparation method of ionized chitosan-sodium alginate composite membrane according to claim 4, characterized in that perilla anthocyanin is anthocyanin extracted from perilla leaves.
6. An application of the ionized chitosan-sodium alginate composite film prepared according to claim 1, which is characterized in that it is used for fresh-keeping film, and the fresh-keeping effect of perilla anthocyanin-ionized chitosan-sodium alginate composite film is higher than that of ionized chitosan-sodium alginate composite film.
LU501468A 2022-02-15 2022-02-15 Preparation method and application of ionized chitosan-sodium alginate composite film LU501468B1 (en)

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