Method for preparing acylated plant pigments and plant pigments

Abstract

The present application discloses a method and a phytochrome for preparing acylated phytochrome, comprising the following steps: subjecting an anthocyanin phytochrome dissolved in an organic solvent and an acylating agent containing a carboxyl group to an acylation reaction under the action of an acylating enzyme to obtain a reaction mixture; and separating the acylated phytochrome from the reaction mixture. By acylating some or all of the -OH on the phytochrome glycoside with an organic acid, the molecular polarity is reduced, and the steric hindrance effect of the acyl group protects the anthocyanin phytochrome; the acylating group associates with the chromophore molecule to form an acyl glycoside, reducing the possibility of bond breakage in the molecular structure and improving its own stability. In addition, the free carboxyl group in the phytochrome has an acidifying ability. In an aqueous solution environment, the proton dissociation of the free carboxyl group -COOH generates H+, thereby reducing the acidity and alkalinity of the solution and reducing the discoloration in an alkaline solution.

Description

Technical Field

[0001] This application belongs to the technical field of chemical material modification, and particularly relates to a method for preparing acylated plant pigments and plant pigments. Background Art

[0002] Plant pigments include fat-soluble chloroplast pigments and water-soluble cell sap pigments. The former exists in chloroplasts and is related to photosynthesis, such as chlorophyll; the latter exists in vacuoles and is particularly related to the color of flowers, such as anthocyanins belonging to flavonoids.

[0003] Red plant pigments include mulberry red, elderberry red, radish red, capsanthin, sorghum red, etc. Among them, anthocyanins are a group of compounds that can present red in flavonoids - a class of substances based on the flavone nucleus. Anthocyanins are compounds formed by the glycosidic bond combination of anthocyanidins and sugars, and are widely present in the cell sap of plant flowers, fruits, stems, leaves and root organs, making them present different colors from red, purple-red to blue. Red plant pigments, such as anthocyanins, are unstable and prone to color change due to the influence of pH, temperature, light, and oxidants. Summary of the Invention

[0004] In view of this, this application provides a method for preparing acylated plant pigments and plant pigments, aiming to solve the problem that red plant pigments are prone to color change or fading.

[0005] In a first aspect, an embodiment of this application provides a method for preparing acylated plant pigments, including the following steps:

[0006] Perform an acylation reaction on the anthocyanin plant pigment dissolved in an organic solvent and an acylating agent containing a carboxyl group under the action of an acylating enzyme to obtain a reaction mixture;

[0007] Separate the acylated plant pigment from the reaction mixture.

[0008] According to an embodiment of one aspect of this application, the acylating agent is selected from aliphatic carboxylic acids, aromatic carboxylic acids or combinations thereof.

[0009] According to an embodiment of one aspect of this application, the aliphatic carboxylic acid is selected from oxalic acid, malonic acid, succinic acid, malic acid, acetic acid, tannic acid, citric acid, lactic acid or combinations thereof;

[0010] The aromatic carboxylic acid is selected from coumaric acid, caffeic acid, ferulic acid, sinapic acid, gallic acid, salicylic acid, syringic acid, ellagic acid or combinations thereof.

[0011] According to an embodiment of one aspect of this application, the anthocyanin plant pigment is selected from mulberry pigment, radish root pigment, elderberry pigment or combinations thereof.

[0012] According to an embodiment of one aspect of the present application, the acylating enzyme is selected from any one of Novozym 435 lipase, Lipozyme TLIM lipase, Lipase PS “Amano” SD lipase, and Lipase PS “Amano” IM lipase.

[0013] According to an embodiment of one aspect of the present application, the molar ratio of the anthocyanin plant pigment, acylating agent, and acylating enzyme is 1-2:50:5-10.

[0014] According to an embodiment of one aspect of the present application, acylating the anthocyanin plant pigment dissolved in an organic solvent with an acylating agent under the action of an acylating enzyme to obtain a reaction mixture includes:

[0015] Under the conditions of heating and vacuum reduction, acylating the anthocyanin plant pigment dissolved in an organic solvent with an acylating agent under the action of an acylating enzyme to obtain a reaction mixture.

[0016] According to an embodiment of one aspect of the present application, the temperature of the heating is 30-60°C, preferably 40-45°C; and / or,

[0017] The degree of vacuum after vacuum reduction is -100 to -80 KPa, preferably -95 to -90 KPa.

[0018] According to an embodiment of one aspect of the present application, stirring is also performed during the acylating reaction, and the rotation speed of the stirring is 10-30 rpm, preferably 15-20 rpm; and / or,

[0019] The time of the acylating reaction is 12-48 h, preferably 24-36 h.

[0020] In a second aspect, an embodiment of the present application provides a plant pigment, and the plant pigment is obtained by the method described in the first aspect.

[0021] Compared with the prior art, the present application has at least the following beneficial effects:

[0022] In the embodiments of the present application, the plant pigment glycoside is acylated by esterifying some or all of the -OH groups with organic acids, which reduces the molecular polarity; at the same time, it also changes the spatial structure of the anthocyanin, and the steric hindrance effect of the acyl group after changing the spatial structure protects the anthocyanin plant pigment and hinders the formation of colorless pseudobase and chalcone; the acylated group associates with the pigment ligand molecule to form an acyl glycoside, reducing the possibility of bond breakage in the molecular structure and improving its own stability; in addition, the free carboxyl group on the plant pigment has an acidifying ability. When acylating with anthocyanin, on the one hand, it acylates the -OH on the sugar group, and on the other hand, the acylated plant pigment still has a free carboxyl group. In an aqueous solution environment, the proton dissociation of the free carboxyl group -COOH generates H + , thus reducing the acidity and alkalinity of the solution and reducing the discoloration in alkaline solutions. Considering the environmental protection and safety of cosmetic materials, acylases are selected for the catalytic reaction, thereby improving the safety of plant pigments. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings required to be used in the embodiments of the present application will be briefly introduced below. For those of ordinary skill in the art, other drawings can also be obtained based on these drawings without creative efforts.

[0024] Figure 1 FIG. is a schematic flow chart of a method for preparing acylated plant pigments provided by an embodiment of the present application. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0025] In order to make the application purpose, technical solutions and beneficial technical effects of the present application clearer, the present application will be further described in detail below with reference to the embodiments. It should be understood that the embodiments described in this specification are only for explaining the present application and not for limiting the present application.

[0026] For simplicity, the present application only explicitly discloses some numerical ranges. However, any lower limit can be combined with any upper limit to form a range not explicitly recorded; and any lower limit can be combined with other lower limits to form a range not explicitly recorded, and similarly any upper limit can be combined with any other upper limit to form a range not explicitly recorded. In addition, although not explicitly recorded, each point or single value between the range endpoints is included in this range. Thus, each point or single value can be used as its own lower or upper limit and combined with any other point or single value or combined with other lower or upper limits to form a range not explicitly recorded.

[0027] In the description of the present application, it should be noted that unless otherwise specified, "above" and "below" include the present number, and "multiple" in "one or more" means two or more.

[0028] The above application content of the present application does not intend to describe every disclosed embodiment or every implementation in the present application. The following description more specifically exemplifies exemplary embodiments. Throughout the application, guidance is provided through a series of embodiments, which can be used in various combinations. In each instance, the enumeration is only a representative group and should not be construed as exhaustive.

[0029] With the trends of "clean beauty" and global sustainable development, products that protect the environment and the ocean from pollution are the trends of modern and future cosmetics. Existing cosmetics tend to be prepared with raw materials of natural origin, preferably plant origin.

[0030] Existing plant pigments in the red color system, such as mulberry red, elderberry red, radish red pigment, capsanthin, and sorghum red pigment, become unstable and prone to color change under the influence of different pH, temperature, light, and oxidants. Considering the structural properties of the pigment, esterification methods can be considered to enhance the stability of the pigment, such as transesterification, esterification of amide and alcohol, addition esterification reaction of alkenes and alkynes, etc.

[0031] The prior art can improve the stability of plant pigments through conventional esterification methods, but the stabilized plant pigments cannot be used in existing cosmetics, their properties have changed, and they contain some substances harmful to the skin, which do not meet the requirements of "clean beauty", and the stabilized plant pigments as raw materials do not belong to natural origin.

[0032] Based on this, the inventors have conducted a large number of studies aiming to improve the stability of plant pigments through a preparation method. At the same time, the stabilized plant pigments can be used in new-generation cosmetics, and then, under the synergistic effect of each natural component, the cosmetics meet people's requirements for naturalness and stability.

[0033] The first aspect of the embodiments of the present application provides a method for preparing acylated plant pigments, as Figure 1 shown, including the following steps:

[0034] S100. React anthocyanin plant pigments dissolved in an organic solvent with a carboxyl-containing acylating agent under the action of an acylating enzyme to obtain a reaction mixture;

[0035] S200. Separate the acylated plant pigments from the reaction mixture.

[0036] According to an embodiment of the present application, on the one hand, anthocyanin plant pigments generally have -OH groups and can undergo an acylation reaction with the carboxyl groups of acylating agents under the action of acylating enzymes; on the other hand, the anthocyanin plant pigments obtained in the natural state are generally combined with sugars to form glycosyl groups, and the acylating agents can bond with the glycosyl groups to achieve acylation; the molecular polarity of the acylated anthocyanin plant pigments is reduced, and their spatial structures are also changed. Due to the steric hindrance effect, the acylated anthocyanin plant pigments protect their own structures, making their chemical properties more stable and also hindering the formation of colorless pseudobases and chalcones. Due to the enhanced bonding effect, the acylated plant pigments achieve intramolecular copigmentation, making the color of the acylated plant pigments more stable or deeper; in particular, the acylated plant pigments may also have free carboxyl groups, and the free carboxyl groups have acidifying ability and can dissociate to produce H + , thereby reducing the acidity and alkalinity of the anthocyanin plant pigments in the solution and further making the color of the plant pigments more stable or deeper.

[0037] According to an embodiment of the present application, the steric hindrance effect mainly refers to the spatial hindrance effect caused by the proximity of certain atoms or groups in a molecule. In the acylated anthocyanin plant pigments of the present application, the steric hindrance caused by the proximity of groups, molecules or atoms in the acyl group.

[0038] According to an embodiment of the present application, the acylating agent may also contain groups such as free carboxylic acids, acid anhydrides, and acyl chlorides. When these groups associate with the pigment ligand molecules to form acyl glycosides, it can further reduce the possibility of bond breakage in the plant pigments, thereby improving the stability of the plant pigments.

[0039] According to an embodiment of the present application, acylating enzymes have the advantages of high regioselectivity, few reaction steps, mild reaction conditions, simple controllability, etc. for acylation reactions. They are more suitable than chemical catalysis and do not contain chemical catalysts in the products, which is more conducive to the purification of acylated plant pigments. In addition, acylating enzymes improve the efficiency and yield of the acylation reaction, improve the stability of plant pigments, do not cause pollution to the environment, have low costs, and are suitable for large-scale production.

[0040] In some embodiments, the organic solvent is selected from isopropanol, methanol, ethyl acetate, ethanol or a combination thereof, which can effectively dissolve anthocyanin plant pigments and can be effectively separated from the acylated plant pigments without residue; the organic solvent can dissolve the anthocyanin plant pigments into a state without particles or turbidity.

[0041] In some embodiments, S200. Separating the acylated plant pigments from the reaction mixture can be carried out under heating and vacuum conditions; the heating and vacuum conditions are conducive to completely separating the acylated plant pigments from other substances. For example, it can completely volatilize the organic solvent ethanol.

[0042] In some embodiments, the acylating agent is selected from aliphatic carboxylic acids, aromatic carboxylic acids, or combinations thereof.

[0043] According to embodiments of the present application, the aliphatic carboxylic acids and aromatic carboxylic acids can be derived from natural substances, preferably plant-derived substances. The molecular structure of aliphatic carboxylic acids does not contain an aromatic ring and contains a straight-chain or branched carbon chain and a carboxyl group. In the molecular structure of aromatic carboxylic acids, the carboxyl group is directly connected to the aromatic ring or connected to the side chain of the aromatic ring. The two major categories of aromatic carboxylic acids are hydroxycinnamic acids (such as coumaric acid, caffeic acid, ferulic acid, and sinapic acid) and hydroxybenzoic acids (such as p-hydroxybenzoic acid and gallic acid). Since the aromatic ring in aromatic carboxylic acids can promote the formation of a copigmentation complex within the anthocyanin molecule, the stability of the plant pigment after acylation with aromatic carboxylic acids is stronger than that with aliphatic carboxylic acids.

[0044] In some embodiments, the aliphatic carboxylic acid is selected from oxalic acid, malonic acid, succinic acid, malic acid, acetic acid, tannic acid, citric acid, lactic acid, or combinations thereof.

[0045] In some embodiments, the aromatic carboxylic acid is selected from coumaric acid, caffeic acid, ferulic acid, sinapic acid, gallic acid, salicylic acid, syringic acid, ellagic acid, or combinations thereof.

[0046] In some embodiments, the anthocyanin plant pigment is selected from mulberry red pigment, radish root red pigment, elderberry pigment, or combinations thereof.

[0047] According to embodiments of the present application, the above pigments are not only of natural origin but also edible. When added to cosmetics, they are safe and environmentally friendly; in high temperature, outdoor light, and alkaline solutions, the anthocyanin plant pigment will change color. In order to fundamentally improve the stability of the phytoerythrin, the anthocyanin plant pigment can be acylated to reduce the possibility of bond breakage in the molecular structure, thereby improving the stability of the plant pigment and extending its storage time under high temperature and light.

[0048] In some embodiments, after the above anthocyanin plant pigment reacts with the acylating agent, the resulting acylated plant pigment can have the structures shown in the following formulas (1), (2), and (3):

[0049]

[0050]

[0051] In formulas (1), (2), and (3), R is selected from a C1-C20 alkyl group substituted or unsubstituted by a substituent, a C3-C20 cycloalkyl group substituted or unsubstituted by a substituent, a C6-C30 aryl group substituted or unsubstituted by a substituent, or a 5-16 membered heteroaryl group substituted or unsubstituted by a substituent;

[0052] The substituents used to replace each group are independently selected from C1-C10 alkyl, C1-C10 alkoxy, C3-C20 cycloalkyl, C6-C20 aryl, 5-20 membered heteroaryl, carboxyl, acid anhydride group, acyl chloride group, etc. each time they appear. Preferably, the acylating agent is an aliphatic carboxylic acid or an aromatic carboxylic acid containing a carboxylic acid.

[0053] Preferably, the number of methoxy groups and hydroxyl groups in the aliphatic carboxylic acid or aromatic carboxylic acid of the carboxylic acid can make the acylated plant pigment weakly acidic or have antioxidant properties, make the color of the plant pigment more stable or darker when used in cosmetics, thereby improving the stability of its color.

[0054] In some embodiments, the acylating enzyme is selected from any one of Novozym435 lipase, Lipozyme TLIM lipase, LipasePS “Amano” SD lipase, and Lipase PS “Amano” IM lipase.

[0055] According to the embodiments of the present application, there is no clear requirement for the type of acylating enzyme, and any enzyme that can achieve acylation can be used. Preferably, the acylating enzyme has little or no residue in the acylated plant pigment.

[0056] In some embodiments, the molar ratio of anthocyanin plant pigment, acylating agent, and acylating enzyme is 1-2:50:5-10.

[0057] According to the embodiments of the present application, within the above ratio range, the acylation of anthocyanin plant pigment can be effectively achieved, the acylating enzyme can be effectively saved, and at the same time, the acylating agent can be reasonably used for acylation, so that the acylated plant pigment is acidic in solution.

[0058] In some embodiments, the anthocyanin plant pigment dissolved in an organic solvent is acylated with an acylating agent under the action of an acylating enzyme to obtain a reaction mixture including:

[0059] Under the conditions of heating and vacuum reduction, the anthocyanin plant pigment dissolved in an organic solvent is acylated with an acylating agent under the action of an acylating enzyme to obtain a reaction mixture.

[0060] According to the embodiments of the present application, under the conditions of heating and vacuum reduction, the maximum catalytic effect of the acylating enzyme can be achieved at a suitable temperature. At the same time, vacuum reduction effectively protects the substances in the reaction system from being oxidized or doped with other substances.

[0061] In some embodiments, the heating temperature is 30-60 °C, preferably 40-45 °C.

[0062] According to the embodiments of the present application, the above temperature can enable the acylating enzyme to exert an excellent catalytic effect, promote the progress of the reaction, and make the acylation reaction fully react.

[0063] In some embodiments, the degree of vacuum after vacuum decompression is -100 to -80 KPa, preferably -95 to -90 KPa.

[0064] According to the embodiments of the present application, at the above degree of vacuum, the substances participating in the reaction can be protected while accelerating the separation of acylated plant pigments.

[0065] In some embodiments, stirring is also carried out during the acylation reaction, and the rotation speed of stirring is 10 to 30 rpm, preferably 15 to 20 rpm.

[0066] According to the embodiments of the present application, stirring at the above rotation speed can promote the full contact of the substances participating in the reaction, make the reaction complete, ensure that the reaction proceeds at a suitable speed, and prevent liquid splashing.

[0067] In some embodiments, the time of the acylation reaction is 12 to 48 h, preferably 24 to 36 h.

[0068] According to the embodiments of the present application, at the above reaction time, the reaction can be made complete, the reaction efficiency can be effectively improved, and the production time can be saved.

[0069] The second aspect of the embodiments of the present application provides a plant pigment, which is obtained by the method of the first aspect.

[0070] According to the embodiments of the present application, the application scenarios of the plant pigment can be the cosmetics and food industries, and the raw materials and reaction conditions of the reaction are environmentally friendly and safe; the generated product is easy to purify, so that the plant pigment does not contain impurities harmful to the application scenario.

[0071] Embodiment

[0072] The following embodiments more specifically describe the content disclosed in the present application. These embodiments are only for illustrative purposes, because various modifications and changes within the scope of the present application disclosure are obvious to those skilled in the art. Unless otherwise stated, all parts, percentages, and ratios reported in the following embodiments are based on weight, and all reagents used in the embodiments are commercially available or synthesized according to conventional methods and can be used directly without further treatment, and the instruments used in the embodiments are all commercially available.

[0073] Example 1

[0074] A method for preparing acylated plant pigment, comprising the following steps:

[0075] Add 50 mg of mulberry red pigment into a 100 mL reactor, dissolve it with 10 mL of ethanol respectively, then add 2 g of oxalic acid and 200 mg of lipase Novozym 435, and finally add ethanol as the solvent to make the total volume of the reaction solution 5 mL; Heat the reaction system on a rotary evaporator at 40 °C under -90 KPa vacuum reaction, stir at a rotation speed of about 20 rpm for 24 h, and after the ethanol has completely evaporated, obtain the reaction product.

[0076] Example 2: A method for preparing acylated plant pigment, comprising the following steps: Add 50 mg of radish root red pigment into a 100 mL reactor, dissolve it with 10 mL of ethanol respectively, then add 2 g of succinic acid and 200 mg of lipase Lipozyme TLIM, and finally add ethanol as the solvent to make the total volume of the reaction solution 5 mL; Heat the reaction system on a rotary evaporator at 40 °C under -90 KPa vacuum reaction, stir at a rotation speed of about 20 rpm for 24 h, and after the ethanol has completely evaporated, obtain the reaction product.

[0077] Example 3

[0078] A method for preparing acylated plant pigment, comprising the following steps:

[0079] Add 50 mg of elderberry pigment into a 100 mL reactor, dissolve it with 10 mL of ethanol respectively, then add 2 g of malic acid and 200 mg of lipase Lipase PS “Amano” SD, and finally add ethanol as the solvent to make the total volume of the reaction solution 5 mL; Heat the reaction system on a rotary evaporator at 40 °C under -90 KPa vacuum reaction, stir at a rotation speed of about 20 rpm for 24 h, and after the ethanol has completely evaporated, obtain the reaction product.

[0080] Example 4

[0081] A method for preparing acylated plant pigment, comprising the following steps:

[0082] Add 50 mg of elderberry pigment into a 100 mL reactor, dissolve it with 10 mL of ethanol respectively, then add 2 g of caffeic acid and 200 mg of lipase Lipase PS “Amano” IM, and finally add ethanol as the solvent to make the total volume of the reaction solution 5 mL; Heat the reaction system on a rotary evaporator at 40 °C under -90 KPa vacuum reaction, stir at a rotation speed of about 20 rpm for 24 h, and after the ethanol has completely evaporated, obtain the reaction product.

[0083] Example 5: A method for preparing acylated plant pigments, comprising the following steps: Add 50 mg of elderberry pigment to a 100 mL reactor, dissolve it with 10 mL of ethanol respectively, then add 2 g of ferulic acid and 200 mg of lipase Novozym 435, and finally add solvent ethanol to make the total volume of the reaction solution 5 mL; Heat the reaction system on a rotary evaporator at 40 °C under a vacuum of -90 KPa, stir at a rotation speed of about 20 rpm for 36 h, and after the ethanol has completely evaporated, obtain the reaction product.

[0084] Example 6

[0085] A method for preparing acylated plant pigments, comprising the following steps:

[0086] Add 50 mg of elderberry pigment to a 100 mL reactor, dissolve it with 10 mL of ethanol respectively, then add 2 g of p-coumaric acid and 200 mg of lipase Novozym 435, and finally add solvent ethanol to make the total volume of the reaction solution 5 mL; Heat the reaction system on a rotary evaporator at 40 °C under a vacuum of -90 KPa, stir at a rotation speed of about 20 rpm for 36 h, and after the ethanol has completely evaporated, obtain the reaction product.

[0087] Example 7

[0088] A method for preparing acylated plant pigments, comprising the following steps:

[0089] Add 50 mg of elderberry pigment to a 100 mL reactor, dissolve it with 10 mL of ethanol respectively, then add 2 g of gallic acid and 200 mg of lipase Novozym 435, and finally add solvent ethanol to make the total volume of the reaction solution 5 mL; Heat the reaction system on a rotary evaporator at 40 °C under a vacuum of -90 KPa, stir at a rotation speed of about 20 rpm for 36 h, and after the ethanol has completely evaporated, obtain the reaction product.

[0090] Example 8

[0091] A method for preparing acylated plant pigments, comprising the following steps:

[0092] Add 50 mg of elderberry pigment to a 100 mL reactor, dissolve it with 10 mL of ethanol respectively, then add 2 g of salicylic acid and 200 mg of lipase Novozym 435, and finally add solvent ethanol to make the total volume of the reaction solution 5 mL; Heat the reaction system on a rotary evaporator at 40 °C under a vacuum of -90 KPa, stir at a rotation speed of about 20 rpm for 24 h, and after the ethanol has completely evaporated, obtain the reaction product.

[0093] Comparative Example 1

[0094] A method for preparing acylated plant pigments. The difference between this comparative example and Example 1 is that the acylating agent is vinyl cinnamate. Since vinyl cinnamate is selected, the acylated plant pigment cannot be dissolved in aqueous solution, is acidic, and has poor solubility.

[0095] Comparative Example 2

[0096] A method for preparing acylated plant pigments. The difference between this comparative example and Example 1 is that the acylating agent is vinyl acetate. Since vinyl acetate is selected, the acylated plant pigment cannot be dissolved in aqueous solution, is acidic, and has poor solubility.

[0097] Test section

[0098] 1. Stability test:

[0099] Take out 5 mg of each sample and add it to 5 mL of a citric acid - sodium dihydrogen phosphate buffer solution with pH = 4 - 5. Place them in a water bath and heat to 65 °C, 80 °C, and 95 °C respectively. Take them out after 1 h and observe.

[0100] For the same sample preparation, place the samples in 10 mL glass test tubes, seal the tube mouths with plastic film, irradiate them under a fluorescent lamp respectively, with the distance between the light source and the samples about 20 cm. Use the dark condition without light as the control, and keep the environmental temperature constant at 20 °C. Compare the color changes after 2 h.

[0101] Table 1 Stability of samples in each example and comparative example.

[0102]

[0103] The untreated anthocyanins will fade significantly with time above 40 °C. The test results of the acylated examples show that the stability of aliphatic carboxylic acid acylation has no significant difference from the plant pigments obtained with other acylating agents, but the improvement of the stability of anthocyanins by aliphatic carboxylic acid acylation is not obvious, and the acylation of low - pressure aromatic carboxylic acids; under sunlight irradiation, some of the acylated pigments do not fade, while some still face the phenomenon of fading; and in the comparative examples, due to vinyl acetate - type acylating agents, the stability of plant pigments may be improved, but under the experimental conditions of this application, due to the pH of the solution and the intensity of irradiation, the color of the plant pigments fades significantly and cannot meet the non - fading requirements for cosmetics.

[0104] 2. Conversion rate detection:

[0105] The conversion rate (%) of the acylated plant pigment is obtained by high - performance liquid chromatography (HPLC). The technical formula is T = A1 / (A1 + A0) × 100%

[0106] Wherein, A0 is the peak area of ​​anthocyanin in the liquid chromatography after the reaction, and A1 is the peak area of ​​the newly generated anthocyanin acylation product.

[0107] Table 2 Conversion rates of various examples and comparative examples.

[0108]

[0109] The data showed that Novozym435 lipase had the highest conversion rate. Although the enzyme activity of Lipozyme TLIM was close to that of Novozym435, its reaction effect was far inferior to that of Novozym435. It was determined that Novozym435 was suitable for catalyzing the acylation reaction of plant pigments. The reaction time had basically no significant effect after 24 hours, and the reaction efficiency of 24 hours and 36 hours was not much different. The conversion rates of different acylating agents (acyl donors) were different, and the conversion rate of aromatic acids was higher than that of fatty acids. Among aromatic acids, the more phenolic hydroxyl groups on the benzene ring, the lower the conversion efficiency, and the farther the phenolic hydroxyl groups on the benzene ring are from the carboxylic acid, the lower the conversion efficiency.

[0110] 3. Antioxidant test

[0111] The DPPH method is used to determine the antioxidant activity of samples. Its mechanism of action can directly act on DPPH free radicals or indirectly consume substances that are easy to generate free radicals to prevent further reactions. DPPH, also known as 1,1-diphenyl-2-trinitrophenylhydrazine, is a very stable nitrogen-centered free radical. Its stability mainly comes from the spatial barriers of the three benzene rings of resonance stabilization, which prevents the unpaired electrons on the nitrogen atom in the middle from playing their due electron pair role. Its anhydrous ethanol emulsion is purple, has maximum absorption at a wavelength of 517nm, and the absorbance is linearly related to the concentration. When a free radical scavenger is added to it, it can be combined with or replaced by the DPPH free radical, so that the number of free radicals is reduced, the absorbance becomes less, and the color of the solution becomes lighter. This can be used to evaluate the ability to scavenge free radicals, that is, by testing the effect of the sample in scavenging DPPH free radicals at a wavelength of 517nm, the antioxidant capacity can be calculated.

[0112] The required materials are DPPH and anhydrous ethanol, and the required instrument is a spectrophotometer.

[0113] 1. Preparation of DPPH stock solution

[0114] Accurately weigh 3.5 mg of DPPH reagent, dissolve it with anhydrous ethanol, transfer it quantitatively into a 10 mL volumetric flask, make up to the mark with anhydrous ethanol, take 2 mL into a 100 mL volumetric flask, shake well to obtain a DPPH stock solution with a concentration of 0.0178 mmol / L, and refrigerate it for later use.

[0115] 2. Determination of DPPH free radical scavenging rate

[0116] In a 10 mL colorimetric tube, successively add 4.0 mL of DPPH solution and 1 mL of the sample in the example (ethanol solution with a sample mass fraction of 1%). Mix evenly and measure the absorbance value (A) at a wavelength of 517 nm using a colorimetric cell. Record the absorbance value as Ai. Then, measure its absorbance value again after storing it in the dark at room temperature for 30 min and record it as Aj. In the control experiment, only add DPPH ethanol solution, and its absorbance value is Ac. The formula for calculating the free radical scavenging rate K is: K(%) = [1 - (Ai - Aj) / Ac] × 100%; Repeat the experiment three times and take the average value of K.

[0117] Table 3 Free radical scavenging rates of each example and comparative example.

[0118]

[0119] As can be seen from the above results, the free radical scavenging rate in the examples reaches more than 40%. The free radical scavenging rate of Example 4 reaches 89.3% and that of Example 5 reaches 90.7%. While the free radical scavenging rate of the comparative example is below 1%. Generally speaking, the free radical scavenging rate of the examples is much higher than that of the comparative example, that is, the antioxidant property is higher than that of the comparative example.

[0120] As described above, it is only the specific implementation manner of the present application, but the protection scope of the present application is not limited thereto. Any person skilled in the art within the technical scope disclosed in the present application can easily think of various equivalent modifications or substitutions, and these modifications or substitutions should all be covered within the protection scope of the present application. Therefore, the protection scope of the present application shall be subject to the protection scope of the claims.

Claims

1. A method for preparing acylated plant pigments to improve the stability of plant pigments, characterized in that, Comprising the following steps: Under the conditions of heating and vacuum decompression, the anthocyanin plant pigment dissolved in an organic solvent is subjected to an acylation reaction with an acylating agent under the action of an acylating enzyme to obtain a reaction mixture; the vacuum degree after vacuum decompression is -100~-80 KPa; The acylated plant pigment is separated from the reaction mixture; The anthocyanin plant pigment is elderberry pigment; The acylating agent is salicylic acid; The acylating enzyme is selected from any one of Novozym 435 lipase, Lipozyme TLIM lipase, Lipase PS "Amano" SD lipase, and Lipase PS "Amano" IM lipase.

2. The method according to claim 1, characterized in that The molar ratio of the anthocyanin plant pigment, the acylating agent, and the acylating enzyme is 1~2:50:5~10.

3. The method according to claim 1, wherein The temperature of the heating is 30~60 °C.

4. The method according to claim 3, characterized in that The temperature of the heating is 40~45 °C.

5. The method according to claim 1, characterized in that The vacuum degree after vacuum decompression is -95~-90 KPa.

6. The method according to claim 1, wherein Stirring is also carried out during the acylation reaction, and the rotation speed of the stirring is 10~30 rpm; and / or, The time of the acylation reaction is 12~48 h.

7. The method according to claim 6, wherein The rotation speed of the stirring is 15~20 rpm.

8. The method according to claim 6, characterized in that, The time of the acylation reaction is 24~36 h.

9. A plant pigment, characterized in that, The plant pigment is obtained by the method according to any one of claims 1~8.

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