阅读说明：本技术 百里香酚-壳寡糖复合物及其制备方法和应用 (Thymol-chitosan oligosaccharide compound and preparation method and application thereof ) 是由 曹汝鸽 于洋 马乾伟 周中凯 于 2020-03-09 设计创作，主要内容包括：本发明属于食品领域,具体涉及一种百里香酚-壳寡糖复合物及其制备方法和应用。百里香酚-壳寡糖复合物为壳寡糖、百里香酚利用离子交联法交联形成的复合物。本发明以百里香酚和壳寡糖为原料,根据二者活性和结构的特点,通过离子交联法制备了百里香酚-壳寡糖复合物,发现复合物的抑菌性和抗氧化能力均得到显著提高,通过扫描电镜观察发现其呈现球状或块状结构,经过红外与核磁共振分析复合物分子的结构,发现百里香酚分子进入壳寡糖与TPP经电荷作用形成的疏水空腔中,利用分子间的静电作用力及疏水相互作用维持分子结构的稳定。将复合物制成涂膜喷洒于南国梨表面,贮藏实验证明百里香酚-壳寡糖复合物可显著提升南国梨的贮藏保鲜性质。(The invention belongs to the field of food, and particularly relates to a thymol-chitosan oligosaccharide compound and a preparation method and application thereof. The thymol-chitosan oligosaccharide compound is a compound formed by crosslinking chitosan oligosaccharide and thymol by an ion crosslinking method. According to the invention, thymol and chitosan oligosaccharide are used as raw materials, according to the characteristics of activity and structure of thymol and chitosan oligosaccharide, a thymol-chitosan oligosaccharide compound is prepared by an ion crosslinking method, the antibacterial activity and the oxidation resistance of the compound are obviously improved, the compound is observed by a scanning electron microscope to be in a spherical or block structure, the structure of a compound molecule is analyzed by infrared and nuclear magnetic resonance, the thymol molecule enters a hydrophobic cavity formed by charge interaction between chitosan oligosaccharide and TPP, and the stability of the molecular structure is maintained by utilizing the electrostatic acting force and hydrophobic interaction between molecules. The compound is prepared into a coating film and sprayed on the surface of the south China pear, and storage experiments prove that the thymol-chitosan oligosaccharide compound can obviously improve the storage and preservation properties of the south China pear.)

1. A thymol-chitosan oligosaccharide compound is characterized in that the compound is formed by crosslinking chitosan oligosaccharide and thymol by an ion crosslinking method.

2. The thymol-chitosan oligosaccharide compound according to claim 1, wherein the mass ratio of thymol to chitosan oligosaccharide is 1: 2-8.

3. The thymol-chitosan oligosaccharide complex according to claim 1, wherein the mass ratio of thymol to chitosan oligosaccharide is 1: 5.

4. The thymol-chitosan oligosaccharide complex according to claim 1, wherein said chitosan oligosaccharide has a molecular weight of 1500 Da.

5. The thymol-chitosan oligosaccharide complex according to claim 1, wherein said cross-linking agent is sodium Tripolyphosphate (TPP).

6. A method for preparing thymol-chitosan oligosaccharide complex according to any one of claims 1-5, which comprises the following steps:

the preparation method comprises the steps of preparing a thymol solution by using an acetic acid solution as a solvent, adding chitosan oligosaccharide into the thymol solution, and uniformly stirring to obtain a mixed solution of thymol and chitosan oligosaccharide;

preparing a cross-linking agent solution, and dropwise adding the mixed solution of thymol and chitosan oligosaccharide into the cross-linking agent solution;

after the reaction is finished, after the centrifugal collection product is used for removing free polyphenol by a dialysis bag, freeze-drying is carried out for subsequent analysis.

7. The method for preparing a thymol-chitosan oligosaccharide complex according to claim 6, wherein the thymol solution is a 1mg/mL thymol acetic acid solution; the cross-linking agent solution is 1mg/mL sodium tripolyphosphate aqueous solution, and the chitosan oligosaccharide solution is 5mg/mL chitosan oligosaccharide aqueous solution.

8. Use of a thymol-chitosan oligosaccharide complex according to any one of claims 1-5, for storage and preservation of food, fruits and vegetables.

Technical Field

The invention belongs to the field of food, and particularly relates to a thymol-chitosan oligosaccharide compound and a preparation method and application thereof.

Background

Thymol belongs to monophenyl phenol, is widely present in thymus plants, has the effects of bacteriostasis, antioxidation, anti-inflammation, wound healing promotion and the like, and is widely applied in the fields of medicines and foods. The chitosan oligosaccharide is obtained by degrading only positively charged alkaline amino polysaccharide chitosan in natural sugar, has better solubility and absorbability than chitosan, and has multiple biological activity effects of resisting oxidation, enhancing immunity, inhibiting bacteria, resisting tumor and the like. Researches show that the biological activities of the polyphenol, such as oxidation resistance, blood sugar reduction, anticoagulation and the like, can be enhanced through the interaction between the polyphenol and the oligosaccharide, the bioavailability of the polyphenol can be influenced, and the flavor and the nutritional value of the food can be improved.

At present, no report is found in the research of forming a compound by combining chitosan oligosaccharide and thymol by utilizing TPP (thermoplastic vulcanizate). the invention explores the ion crosslinking preparation method of the thymol-chitosan oligosaccharide compound, characterizes the molecular structure of the compound, evaluates the antioxidant and bacteriostatic activity of the compound, applies the compound to the storage experiment of Nanguo pear and provides a new way for the storage and preservation of fruits and vegetables.

Disclosure of Invention

The invention aims to provide a method for preparing a chitosan oligosaccharide-thymol compound by an ion crosslinking method.

In order to achieve the purpose, the invention adopts the following technical scheme:

a thymol-chitosan oligosaccharide compound is prepared by crosslinking chitosan oligosaccharide and thymol by ion crosslinking method.

The mass ratio of the thymol to the chitosan oligosaccharide is 1: 2-8.

Preferably, the mass ratio of the thymol to the chitosan oligosaccharide is 1: 5.

The molecular weight of the chitosan oligosaccharide is 1500 Da.

The cross-linking agent is sodium tripolyphosphate TPP.

The invention also comprises a method for preparing the thymol-chitosan oligosaccharide compound, which comprises the following steps:

the preparation method comprises the steps of preparing a thymol solution by using an acetic acid solution as a solvent, adding chitosan oligosaccharide into the thymol solution, and uniformly stirring to obtain a mixed solution of thymol and chitosan oligosaccharide;

preparing a cross-linking agent solution, and dropwise adding the mixed solution of thymol and chitosan oligosaccharide into the cross-linking agent solution;

after the reaction is finished, centrifugally collecting the product, removing free polyphenol by using a dialysis bag, and freeze-drying for subsequent analysis.

Preferably, the thymol solution is 1mg/mL thymol acetic acid solution; the chitosan oligosaccharide solution is5 mg/mL; the sodium tripolyphosphate solution is 1mg/mL sodium tripolyphosphate aqueous solution;

the invention also comprises the application of the thymol-chitosan oligosaccharide compound as claimed in any one of claims 1 to 4, which is characterized in that the compound is stored and kept fresh in food, fruits and vegetables.

Compared with the prior art, the invention has the beneficial effects that:

according to the invention, thymol and chitosan oligosaccharide are used as raw materials, according to the characteristics of activity and structure of the thymol and the chitosan oligosaccharide, a thymol-chitosan oligosaccharide compound is prepared by an ion crosslinking method and sodium tripolyphosphate is used as a crosslinking agent, the bacteriostatic activity and the antioxidant capacity of the compound are obviously improved, the compound is observed by a scanning electron microscope to be in a spherical or block structure, the structure of the molecule of the compound is analyzed by infrared and nuclear magnetic resonance, the thymol molecule enters a hydrophobic cavity formed by the chitosan oligosaccharide and TPP through charge interaction, and the stability of the molecular structure is maintained by utilizing the electrostatic acting force and the hydrophobic interaction between the molecules. The compound is prepared into a coating film and sprayed on the surface of the south China pear, and storage experiments prove that the thymol-chitosan oligosaccharide compound can obviously improve the storage and preservation properties of the south China pear.

According to the invention, the chitosan oligosaccharide with the molecular weight of 1500Da and the thymol are selected as raw materials, the sodium tripolyphosphate is taken as an ionic crosslinking agent to prepare the compound, the oxidation resistance and the bacterial inhibition of the compound are comprehensively evaluated, and the molecular structure of the compound is systematically analyzed. The antibacterial activity of the compound is evaluated, and the compound is proved to have obvious inhibition effect on escherichia coli, pseudomonas aeruginosa, staphylococcus aureus and bacillus subtilis. The oxidation resistance of the compound is measured, and the ABTS +, DPPH free radical clearance and the total oxidation resistance of the compound are obviously enhanced. Analyzing by scanning electron microscope, ultraviolet spectrophotometer, Fourier transform infrared spectrum, and thermogravimetric analysisPrecipitation,¹H-NMR analysis indicated that the complex was successfully prepared and had a stable conformation.

Drawings

FIG. 1 is a graph of bacterial growth: escherichia coli (a), Pseudomonas aeruginosa (b), Staphylococcus aureus (c), and Bacillus subtilis (d);

FIG. 2 is an electron microscope scan of the bacterial bodies of Escherichia coli (a, a '), Pseudomonas aeruginosa (b, b'), Staphylococcus aureus (c, c '), and Bacillus subtilis (d, d') before and after the thymol-chitosan oligosaccharide complex treatment;

FIG. 3 is a scanning electron micrograph of chitooligosaccharide (a), thymol (b), a mixture of thymol and chitooligosaccharide (c) and thymol-chitooligosaccharide complex (d);

FIG. 4 is a UV spectrum of chitooligosaccharide, thymol and chitooligosaccharide mixture (B-CT), sodium tripolyphosphate and chitooligosaccharide mixture (T-C) and thymol-chitooligosaccharide complex (T-CT);

FIG. 5 is an infrared spectrum of chitosan oligosaccharide, thymol, sodium tripolyphosphate and thymol-chitosan oligosaccharide complex (T-CT);

FIG. 6 of Chitosan oligosaccharide, thymol and thymol-Chitosan oligosaccharide Complex (T-CT)¹H-NMR chart;

FIG. 7 is a thermogravimetric spectrum of chitosan oligosaccharide, thymol, sodium tripolyphosphate and thymol-chitosan oligosaccharide complex (T-CT);

FIG. 8 is a two-dimensional simulated structural diagram of a thymol-chitosan oligosaccharide complex;

FIG. 9 is a graph of weight loss, hardness, soluble solids content, and VC content of Nanguo pears at different storage times.

Detailed Description

In order to make the technical solutions of the present invention better understood by those skilled in the art, the present invention will be further described in detail with reference to the accompanying drawings and preferred embodiments.

Materials and reagents thymol (MW150.22Da) Aladdin reagent, Inc.; chitosan oligosaccharide (MW1500Da) Zhejiang gold Chitosan pharmaceutical Co., Ltd; 2, 2-diaminobis-diammonium salt (ABTS), 6-hydroxy-2, 5, 7, 8-tetramethylchromane-2-carboxylic acid (Trolox), 1-diphenyl-2-trinitrophenylhydrazine (DPPH), and Fulinol are all available from Beijing Soilebao Tech & ltd; 2, 4, 6-Tripyridyltriazine (TPTZ) was purchased from Sigma, USA; escherichia coli (ATCC10305), Staphylococcus aureus (ATCC25923), Pseudomonas aeruginosa (ATCC15442) and Bacillus subtilis (ATCC6633) were obtained from the China center for the Collection of general-purpose microorganisms. Yeast extract and tryptone were purchased from OXOID, United kingdom. Methanol (analytically pure), absolute ethanol (analytically pure), and glacial acetic acid (analytically pure) were purchased from science and technology ltd of kang of Tianjin; hydrochloric acid (analytically pure), sodium hydroxide (analytically pure), beta-mercaptoethanol, SDS solution and borax are purchased from Tianjin north Tianyi chemical reagent factory; sodium chloride (analytically pure) and ascorbic acid (analytically pure) were purchased from Tianjin chemical reagent one factory; agar was purchased from Haimang, Khaki Biotech Co., Ltd; 2, 6-dichloroindophenol (analytically pure) and oxalic acid (analytically pure) were purchased from Shanghai Ika reagent.

The main apparatus and equipment is ALPHA 1-2LD PLUS freeze dryer, Marin Christ, Germany; SU-1510 Scanning Electron Microscope (SEM), hitachi, japan; 89090A ultraviolet spectrophotometer, Agilent technologies, Inc. USA; IS50 fourier infrared spectrometer, nigeroli, usa; TGA-Q50 thermogravimetric analyzer, us TA instruments; FG2pH instrument, mettler-toledo; KQ3200B ultrasonic cleaning instrument, kunshan ultrasonic instruments ltd; an Epoch2 multifunctional microplate reader, Thermo corporation, USA; a 400MHZ nuclear magnetic resonance spectrometer, brueck technologies ltd, germany;