阅读说明：本技术 一种提高红曲红色素热稳定性的方法和应用 (Method for improving thermal stability of monascus red pigment and application ) 是由 李利 陈莎 高梦祥 刘彩 于 2021-06-18 设计创作，主要内容包括：本发明公开了一种提高红曲红色素热稳定性的方法和应用,包括：将纯化的红曲橙色素溶解于甲醇中制得红曲橙色素溶液,同时将氨基酸溶解于缓冲液中制得氨基酸溶液；将所述红曲橙色素溶液与所述氨基酸溶液按比例混合,于4～40℃条件下摇床孵育5min～3h,利用红曲橙色素与氨基酸之间的亲氨基反应制备得到红曲红色素,即为热稳定性提高的红曲红色素；其中所述氨基酸选自：组氨酸、苯丙氨酸、酪氨酸或色氨酸。本发明通过在红曲橙色素提取液中加入氨基酸溶液,并对氨基酸种类进行优选,制备得到了水溶性好、且在酸性条件下热稳定显著提高的红曲红色素,可将其应用于酸性食品,如酸性饮料的加工生产中,避免高温杀菌及灌装过程中红曲红色素不稳定导致的失色等问题,具有广泛的应用前景和市场应用价值。(The invention discloses a method for improving thermal stability of monascus red pigment and application thereof, wherein the method comprises the following steps: dissolving the purified monascus orange pigment in methanol to prepare monascus orange pigment solution, and dissolving amino acid in buffer solution to prepare amino acid solution; mixing the monascus orange pigment solution and the amino acid solution in proportion, incubating for 5 min-3 h at 4-40 ℃ in a shaking table, and preparing a monascus red pigment by utilizing an amino-philic reaction between the monascus orange pigment and the amino acid, wherein the monascus red pigment with improved thermal stability is obtained; wherein the amino acid is selected from: histidine, phenylalanine, tyrosine or tryptophan. According to the invention, the amino acid solution is added into the monascus orange pigment extracting solution, and the amino acid type is optimized, so that the monascus red pigment with good water solubility and remarkably improved thermal stability under an acidic condition is prepared, and the monascus red pigment can be applied to the processing production of acidic foods such as acidic beverages, so that the problems of color loss and the like caused by instability of the monascus red pigment in the processes of high-temperature sterilization and filling are avoided, and the monascus red pigment has wide application prospect and market application value.)

1. A method for improving the thermal stability of monascus red pigment is characterized by comprising the following steps:

step 1, dissolving purified monascus orange pigment in methanol to prepare monascus orange pigment solution, and dissolving amino acid in buffer solution to prepare amino acid solution;

step 2, mixing the monascus orange pigment solution with the amino acid solution to ensure that the ratio of monascus orange pigment to amino acid is 1 mg: 0.002-0.1 mmol, adding methanol to a constant volume, and incubating for 5 min-3 h at 4-40 ℃ in a shaking table to obtain the monascus red pigment derived from amino acid, namely the monascus red pigment with improved thermal stability;

wherein the amino acid is selected from: histidine, phenylalanine, tyrosine or tryptophan.

2. The method of claim 1, wherein the method is used to increase the thermal stability of red pigment under acidic conditions.

3. The method according to claim 2, wherein the acidic condition is greater than pH3 and less than pH7, and the temperature of the thermal stability is 60 to 95 ℃.

4. The method according to claim 1, wherein said monascus orange pigment is dissolved in methanol solution containing 0.1% by volume formic acid in step 1.

5. The method of claim 1, wherein the buffer in step 1 is selected from the group consisting of: phosphate buffer, Tris-HCl buffer, HEPES buffer, MOPS buffer, PIPES buffer.

6. The method according to claim 1, wherein the ratio of monascus orange pigment to amino acids in step 2 is 1 mg: 0.08 mmol.

7. The method according to claim 6, wherein 1mL of monascus orange pigment solution with a concentration of 1mg/mL and 5mL of amino acid solution with a concentration of 16mmol/L are mixed in step 2, methanol is added to make the volume to 10mL, and the mixture is subjected to shake culture at 30 ℃ for 2 h.

8. The method of claim 1, wherein step 2 further comprises: and after the incubation is finished, adding deionized water into the reaction system, adjusting the content of methanol, passing through an extraction column, washing with water to remove salt ions in the reaction system, and eluting with methanol after the salt is removed to obtain the monascus red pigment derived from the amino acid.

9. An acid-and heat-resistant monascus red pigment, characterized in that it is prepared by the method according to any one of claims 1 to 8.

10. The use of the acid resistant monascus red pigment according to claim 9 in the production of acidic food products.

Technical Field

The invention belongs to the technical field of monascus pigment production and application, and particularly relates to a method for improving thermal stability of monascus red pigment and application of the monascus red pigment.

Background

The monascus red pigment is a safe and nontoxic natural pigment generated by secondary metabolism of monascus, is also one of three monascus-derived food pigment additives (red yeast rice, monascus red and monascus yellow pigments) approved in the current national standards of China, and is widely applied to daily life of people. Generally, the monascus red pigment is easily soluble in organic solvents such as ethanol and acetone, but is hardly soluble in water, and is relatively stable only in neutral and alkaline environments (pH 6-11), and when the pH is acidic, the color value is reduced. Research shows that the color value of the monascus red pigment is reduced by 20% when the pH value is 2.5-4.5 (the far east, the stability of 2 natural red pigments and the application effect of the natural red pigments in fruit juice beverages are analyzed, 2019). These characteristics of monascus red pigment limit its application in acidic foods such as beverages.

At present, most of the beverages belong to low pH products, such as: the pH value of the carbonated beverage is 2.2-4.9, the pH value of the fruit juice beverage is about 3.0, and the pH value of the yoghourt is less than 4.6. Generally, the heat treatment process of the beverage comprises pasteurization and hot filling, wherein the pasteurization temperature commonly used for beverage production is 63-95 ℃, and the filling process requires 85 ℃ (hot filling) or 68 ℃ (medium filling). Therefore, the color additives used in beverages must have good thermal stability in a low pH environment. Researches have been made to directly apply the monascus red pigment to the production of carbonated beverages, and the result shows that the preservation rate of the pigment is only 46% after the monascus red pigment is subjected to high-temperature treatment and placed in a dark place for 10 days, which indicates that the monascus red pigment has the problem of poor thermal stability in the carbonated beverages, and thus the application of the monascus red pigment is greatly limited. Patent CN201811011840.2 discloses a water-soluble monascus red pigment and a preparation method thereof, wherein the monascus red pigment is modified from the water-solubility angle, so that the monascus red pigment with good water solubility is prepared, but how to improve the thermal stability of the monascus red pigment is not mentioned.

Disclosure of Invention

The invention provides a method and application for improving the thermal stability of monascus red pigment from the aspects of improving the water solubility of the monascus red pigment and the thermal stability in a low pH environment, aiming at the blank of the prior art, the amino acid solution is added into the monascus orange pigment extracting solution, the type of amino acid is optimized, and the monascus red pigment is prepared by utilizing the amino-philic reaction between the monascus orange pigment and the amino acid, so that the water solubility and the thermal stability under an acidic condition are obviously improved, the monascus red pigment can be applied to the processing production of acidic foods such as acidic beverages, the problems of color loss and the like caused by instability of the monascus red pigment in the processes of high-temperature sterilization and filling are avoided, and the monascus red pigment has wide application prospect and market application value.

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

the invention provides a method for improving the thermal stability of monascus red pigment, which comprises the following steps:

step 1, dissolving purified monascus orange pigment in methanol to prepare monascus orange pigment solution, and dissolving amino acid in buffer solution to prepare amino acid solution;

step 2, mixing the monascus orange pigment solution with the amino acid solution to ensure that the ratio of monascus orange pigment to amino acid is 1 mg: 0.002-0.1 mmol, adding methanol to a constant volume, and incubating for 5 min-3 h at 4-40 ℃ in a shaking table to obtain the monascus red pigment derived from amino acid, namely the monascus red pigment with improved thermal stability;

wherein the amino acid is selected from: histidine, phenylalanine, tyrosine or tryptophan.

Further, the method is used for improving the thermal stability of the monascus red pigment under acidic conditions.

Further, the acidic condition is more than pH3 and less than pH7, and the temperature of the thermal stability is 60-95 ℃.

Further, in step 1, the monascus orange pigment is dissolved in a methanol solution containing formic acid with the volume fraction of 0.1%.

Further, in step 1, the buffer is selected from: phosphate buffer, Tris-HCl buffer, HEPES buffer, MOPS buffer, PIPES buffer.

Further, the buffer in step 1 is a phosphate buffer at pH 7.0.

Further, in the step 2, the ratio of monascus orange pigment to amino acid is 1 mg: 0.08 mmol.

Further, in step 2, 1mL of monascus orange pigment solution with the concentration of 1mg/mL and 5mL of amino acid solution with the concentration of 16mmol/L are mixed, methanol is added to the mixture to reach the constant volume of 10mL, and the mixture is incubated for 2 hours in a shaking table at the temperature of 30 ℃.

Further, step 2 further comprises: and after the incubation is finished, adding deionized water into the reaction system, adjusting the content of methanol, passing through an extraction column, washing with water to remove salt ions in the reaction system, and eluting with methanol after the salt is removed to obtain the monascus red pigment derived from the amino acid.

And further adding deionized water to adjust the concentration of methanol to be 20%, passing through an extraction column, washing salt ions in a reaction system with water, desalting, and eluting with a methanol solution with the concentration of 90% to obtain the monascus red pigment derived from amino acid.

The invention also provides the acid-resistant and heat-resistant monascus red pigment prepared by the method.

The invention also provides the application of the acid-resistant and heat-resistant monascus red pigment in the production of acidic foods.

Further, the acidic food is an acidic beverage, and the pH value of the acidic food is more than 3 and less than 7.

Compared with the prior art, the invention has the beneficial effects that: the invention starts from the aspects of improving the water solubility of the monascus red pigment and the thermal stability in the low pH environment, adding amino acid solution into extractive solution of monascus orange pigment (containing monascus purpureus and monascus rubigins) to prepare monascus red pigment, and preferably an amino acid, preferably any one of histidine, phenylalanine, tyrosine or tryptophan, thereby obviously improving the water solubility of the monascus red pigment and the thermal stability in the low pH value environment, namely, the invention provides a method for effectively improving the thermal stability of the monascus red pigment, the monascus red pigment prepared by the method has good water solubility and stable color, can be applied to the processing production of acidic foods such as acidic beverages, thereby widening the application field of the additive of the monascus red pigment, promoting the development of related industries and having extremely high market application prospect and value.

Drawings

FIG. 1 is a graph showing the sensory colors of beverages prepared from monascus red pigment and grape skin red pigment derived from four amino acids in example 2 of the present invention;

FIG. 2 shows the residual pigment ratios of the beverages prepared from the monascus red pigment and the grape skin red pigment derived from the four amino acids in example 2 of the present invention after 65 ℃ treatment;

FIG. 3 shows the residual pigment ratios of the beverages prepared from monascus red pigment and grape skin red pigment derived from four amino acids in example 2 of the present invention after treatment at 75 ℃;

FIG. 4 shows the residual pigment ratios of the beverages prepared from the monascus red pigment and the grape skin red pigment derived from the four amino acids in example 2 of the present invention after being treated at 85 ℃;

FIG. 5 shows the residual pigment ratios of the beverages prepared from the monascus red pigment and the grape skin red pigment derived from the four amino acids in example 2 of the present invention after 95 ℃ treatment;

FIG. 6 is a chromatogram and structural characterization of the histidine-derivatized monascus red pigment in example 2 of the present invention.

Detailed Description

The technical solutions of the present invention will be described clearly and completely with reference to the following embodiments of the present invention, and it should be understood that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

EXAMPLE 1 preparation of Monascus red pigment

1. Preparation of monascus orange pigment

The fermentation and extraction of monascus orange pigment are carried out by adopting the conventional method in the field, and the method comprises the following steps:

(1) preparation of spores: inoculating Monascus ruber M-7 (from CCTCC HF 2008638, China center for type culture Collection) to agar medium of fermentation broth of Saccharomyces boulardii at 28 deg.CCulturing for 5-15 days. Washing spore with appropriate amount of sterile water, filtering with two layers of sterile lens-wiping paper to remove mycelium, counting the obtained spore suspension with blood count plate, and adjusting concentration to 10⁵one/mL. Wherein the agar culture medium of the Chao's yeast extract comprises the following components: NaNO₃ 3.0g/L，K₂HPO₄ 1.0g/L，KCl 0.5g/L，MgSO₄·7H₂O 0.5g/L，FeSO₄·7H₂0.01g/L of O, 30.0g/L of cane sugar and 5.0g/L of yeast extract.

(2) Liquid state fermentation of monascus orange pigment: a two-step fermentation process at low pH (pH 3) was used. The spore suspension (10) was inoculated at 1%⁵one/mL) was inoculated into 50mL of Potato Dextrose Broth (PDB), incubated at 28 ℃ for 40h with shaking at 200rpm, 50mL of a sterilized citric acid-disodium hydrogenphosphate buffer (0.2mol/L, pH 3) was added, and incubation was continued for 5d with shaking at 28 ℃ and 200 rpm.

(3) Extracting monascus orange pigment: after fermentation, the fermentation liquor is vacuumized and filtered by filter cloth, and mycelia are collected. The mycelia were washed with 2 times the volume of the fermentation broth with an acidic solution having a pH of 2, and the water-soluble pigment and the medium remained on the surface were washed off. Drying the water on the surface of the hyphae by using absorbent paper, transferring the hyphae into 70% ethanol solution with the volume 2 times that of the fermentation liquor, wherein the ethanol solution is adjusted to pH2 by using formic acid in advance, so that the orange pigment can be prevented from being converted into red pigment. Homogenizing at 10000rpm for 1min, and filtering with 300 mesh filter cloth to remove mycelium to obtain orange pigment stock solution.

(4) And (3) purifying monascus orange pigment: adding 0.5 times volume of formic acid solution (pH 2) into the above orange pigment stock solution, vortexing at 2800rpm for 5min, mixing thoroughly, standing at-20 deg.C for 2h, taking out, centrifuging at 10000rpm for 20min, and discarding supernatant to obtain orange pigment crystal, i.e. purified monascus orange pigment (mainly containing monascus ruber and monascus rubigin).

2. Preparation of amino acid derivative monascus red pigment

Dissolving the purified monascus orange pigment in methanol (containing 0.1% formic acid) to prepare a monascus orange pigment solution of 1 mg/mL. Respectively dissolving 14 natural amino acids in 200mmol/L phosphate buffer solution (pH 7.0) to obtain a mixture16mmol/L amino acid solution. Then mixing the monascus orange pigment solution with an amino acid solution, wherein each 10mL of reaction system contains 5mL of the amino acid solution, 1mL of the monascus orange pigment solution and 4mL of methanol. Incubate at 30 ℃ for 2h in a shaker at 250 rpm/min. Adding deionized water into the reaction system after the reaction is finished, regulating the content of methanol in the system to 20 percent, and then slowly passing the reaction solution through C₁₈And (3) adding water into the solid phase extraction column to elute and remove the buffer salt, eluting with 90% methanol solution, and performing rotary evaporation and drying on the collected monascus red pigment solution to obtain the desalinized amino acid-derivatized monascus red pigment. Meanwhile, the traditional monascus red pigment (main components of erythrorubimine and monascus purpureus) is prepared by amino-philic reaction between ammonia water and monascus orange pigment and serves as a control group, wherein the concentration of the ammonia water is 1mol/L, and the rest reaction conditions are the same as those of amino acid and orange pigment.

The water solubility of monascus red pigment derived from 14 amino acids and traditional monascus red pigment is respectively determined, including the solubility (g/L) and the color value (AU)_500nm) The data were determined as follows: 25.0mg of monascus red pigment is weighed and added into 1mL of deionized water for full dissolution. If the monascus red pigment is completely dissolved, its solubility in water is recorded>25g/L, measuring the light absorption value at the wavelength of 500nm after diluting by proper times, and according to the formula: color value (AU)_500nm) Calculating the color value (AU) of the monascus red pigment solution at the moment_500nm) The color value of the saturated solution is recorded as being greater than the color value; if the monascus red pigment is not completely dissolved, centrifuging at 10000r/min for 5min, taking out the supernatant, drying and weighing the precipitate, calculating the solubility (g/L) of the precipitate in water according to the reduced mass, and measuring the light absorption value at a wavelength of 500nm after the centrifugation supernatant is diluted by a proper multiple according to the formula: color value (AU)_500nm) Calculating the color value (AU) of saturated aqueous solution of monascus red pigment_500nm). The measurement results are shown in table 1.

TABLE 1 solubility and color number of amino acid derivatized Monascus red pigment and traditional Monascus red pigment (control)

The results show that the solubility of the conventional monascus red pigment (control) is only 3.9g/L, and the color value of the saturated solution is 52.0. Except arginine, the water solubility of the monascus red pigment derived from other amino acids is improved to different degrees compared with the control. Besides the arginine and lysine derived monascus red pigment, the solubility of the rest 12 amino acid derived monascus red pigments is more than 25g/L, the solubility is at least 6.4 times of that of the traditional monascus red pigment (contrast), and the color value is more than 600, so that the color value is obviously improved compared with the contrast.

3. Preliminary screening of acid-resistant and heat-resistant monascus red pigment

The thermal stability of the 14 amino acid-derivatized monascus red pigment and the traditional monascus red pigment under the acidic condition is evaluated and screened, and the method specifically comprises the following steps: adding buffers with different pH values (pH 3,5 and 7) into the 14 amino acid-derivatized monascus red pigment and the traditional monascus red pigment respectively, then placing the mixture at 80 ℃ for heating for 6 hours, and measuring the residual rate (%) of the pigment:

the residual dye ratio (%) × (color value after treatment/color value before treatment) × 100%.

The determination method comprises the following specific steps: 0.2mol/L of a citric acid-phosphate buffer (pH 3, pH5, pH 7) was prepared. Dissolving amino acid-derived monascus red pigment and traditional monascus red pigment with 40% methanol aqueous solution to make the light absorption value of the traditional monascus red pigment at 500nm (2.0 +/-0.2), and mixing the traditional monascus red pigment with citric acid-phosphate buffer solutions with different pH values (pH 3, pH5 and pH 7) at a ratio of 1:1 to obtain monascus red pigment solutions with different pH values. Placing 10mL of red yeast rice red pigment solution with different pH values in a sealed glass bottle, heating at 80 deg.C in the dark for 6h, and measuring color value (AU) of the sample with an ultraviolet-visible spectrophotometer_500nm) The residual ratio of the dye was calculated, and the measurement results are shown in Table 2.

TABLE 2 thermal stability of amino acid derivatized Monascus red pigment and traditional Monascus red pigment (control) at different pH' s

The results show that under acidic conditions, the thermal stability of different amino acid-derivatized monascus red pigments is different, but is obviously higher than that of the traditional monascus red pigment (contrast), wherein the thermal stability of 4 amino acid-derivatized monascus red pigments such as histidine, tyrosine, phenylalanine, tryptophan and the like is superior to that of other monascus red pigments. After heating at 80 ℃ for 6h, the pigment residue rates of the monascus red pigment derived from histidine, tyrosine, phenylalanine and tryptophan at the pH value of 3 are 54.95%, 45.07%, 47.23% and 45.42% respectively; the residual rates of the pigments at pH5 were 49.30%, 45.08%, 43.20% and 37.82%, respectively, which were significantly higher than those of the conventional monascus red pigment (control) and the monascus red pigments derivatized with other 10 amino acids. Namely, the screening provided by the implementation, 4 types of acid-resistant and heat-stable monascus red pigments are obtained in total, and are respectively: monascus red pigment derivatized with tyrosine, tryptophan, phenylalanine and histidine.

Example 2 application and Performance determination of Monascus red pigment

On the basis that 4 monascus red pigments with good water solubility, acid resistance and heat resistance are obtained by screening in example 1, the monascus red pigments are applied to food, specifically acidic beverages, and the performance of the foods is measured.

1. Preparation of acidic beverages

Preparation of the acidic beverage the formulation is prepared according to the formula disclosed in Amruta et al, evaluation of the effect of additives on stability of beta yanin pigments from Basella rubra in a model conversion system for storage, Journal of Food Science and Technology. Adding 13% of white granulated sugar, 0.05% of sodium benzoate and corresponding pigment into 0.1mol/L citric acid-sodium citrate buffer solution with the pH value of 5.0, and uniformly mixing to obtain the acid beverage.

The monascus red pigment derived from tyrosine, tryptophan, phenylalanine and histidine are respectively selected and applied to the acid beverage. Since the conventional monascus red pigment has poor solubility in the beverage system and is liable to generate precipitates, the example adopts the conventional pigment in the field, grape skin red pigment, as a control. The addition amount of the grape skin red is shown in the following literature: the addition of the dosage disclosed in the 'application stability analysis of 4 water-soluble natural red pigments in carbonated beverage'. The other 4 kinds of amino acid derived haematochrome are added according to the sensory color of the prepared grape skin red beverage, and are blended to be consistent with the sensory color of the grape skin red beverage, and the blended color is shown in figure 1, wherein a is grape skin haematochrome (contrast), and b, c, d and e are respectively phenylalanine, histidine, tryptophan and tyrosine derived monascus haematochrome.

2. Characterization of thermal stability

The thermal stability of the 5 beverages containing different red pigments was characterized, specifically:

(1) placing 5 beverages containing different red pigments at 65 deg.C, sampling every 10min, and detecting pigment residue rate, with the detection results shown in FIG. 2;

(2) placing 5 beverages containing different red pigments at 75 deg.C, sampling every 5min, and detecting pigment residue rate, with the detection result shown in FIG. 3;

(3) placing 5 beverages containing different red pigments at heat treatment temperature of 85 deg.C, sampling every 5min to detect pigment residue rate, with the detection result shown in FIG. 4;

(4) the 5 beverages containing different red pigments were subjected to a heat treatment at 95 ℃ and the residual rate of the pigments was measured every 30 seconds, and the results are shown in FIG. 5.

The result shows that after the acid beverage is treated at 65 ℃ for 50min, the residue rate of the grape skin red pigment (contrast) is only 68.37%, the pigment residue rates of the monascus red pigments derived from the rest 4 amino acids are higher than that of the contrast, the residue rates of the monascus red pigments derived from phenylalanine, tyrosine and tryptophan are respectively 79.88%, 82.53% and 85.45%, and the residue rate of the monascus red pigment derived from histidine is the highest and is as high as 96.56%.

After the acid beverage is treated at 75 ℃ for 40min, the residual rate of the grape skin red pigment (contrast) is 55.78%, the residual rates of the monascus red pigments derived from the rest 4 amino acids are higher than that of the contrast, the residual rates of the monascus red pigments derived from phenylalanine, tyrosine and tryptophan are 73.64%, 81.59% and 74.05%, respectively, and the residual rate of the monascus red pigment derived from histidine is 87.55%.

After the acid beverage is treated at 85 ℃ for 40min, the residue rate of the grape skin red pigment (contrast) is only 32.87%, the pigment residue rates of the monascus red pigments derived from the rest 4 amino acids are higher than that of the contrast, the residue rates of the monascus red pigments derived from phenylalanine, tyrosine and tryptophan are respectively 56.19%, 61.53% and 58.10%, and the residue rate of the monascus red pigment derived from histidine is the highest and still reaches 87.53%.

After the acid beverage is treated at 95 ℃ for 660s, the residue rate of the grape skin red pigment (contrast) is 38.73%, the pigment residue rates of the monascus red pigments derived from the rest 4 amino acids are higher than that of the contrast, the residue rates of the monascus red pigments derived from phenylalanine, tyrosine and tryptophan are 74.50%, 82.18% and 77.80%, respectively, and the residue rate of the monascus red pigment derived from histidine is still maintained to be higher and is as high as 94.36%.

The results show that the pigment residue rate of the 4 amino acid-derivatized monascus red pigments is remarkably higher than that of the control grape skin red pigment no matter pasteurization is carried out (63-95 ℃) or filling is carried out (68 ℃ and 85 ℃), further, the pigment residue rate of the histidine-derivatized monascus red pigment is the highest and is between 87% and 98%, and the pigment residue rate is kept stable and does not obviously decrease along with the prolonging of the treatment time, namely, the thermal stability of the histidine-derivatized monascus red pigment is the best. The verification proves that the influence of high-temperature short-time pasteurization (95 ℃) on the pigment is minimum, and the residual rate of the pigment is more than 92% after the pigment is heated for 30-660 s. After sterilization at 95 ℃ for 60s, the microbial indicators in the beverage are detected by a detection method of GB 4789 + 2015 "food hygiene microbiology inspection", and the total number of bacterial colonies, mould and yeast are not detected, so that the beverage meets the national food safety standard (GB 7101-2015).

3. Structural characterization of histidine-derivatized monascus red pigment

And (3) respectively carrying out High Performance Liquid Chromatography (HPLC), ultraviolet-visible spectrum (UV-vis) and high performance liquid chromatography-mass spectrometry (LC-MS) analysis on the screened histidine-derivatized monascus red pigment, determining the molecular weight of each derivatized monascus red pigment according to the excimer ion [ M + H ] + peak obtained by mass spectrometry detection, and comparing the molecular weight with the molecular weight of more than 100 monascus pigments reported at present to carry out qualitative analysis. The chemical structure of histidine-derivatized monascus red pigment is speculated according to the amino group-philic reaction principle of monascus orange pigment (monascus red pigment and monascus rubigin) and amino acid, and is shown in fig. 6.

As can be seen from FIG. 6a, a chromatographic peak appeared in the liquid chromatogram at retention times of 5.728 and 6.293min, indicating the presence of two pigment components in the histidine-derivatized monascus red pigment. As can be seen from FIG. 6b, the maximum absorption peaks of the pigment components corresponding to 5.728min and 6.293min in the ultraviolet-visible spectrum are 526nm and 528nm, respectively, which are consistent with the ultraviolet-visible absorption spectrum characteristics of monascus red pigment, indicating that the colors of the two pigment components are red. As can be seen from FIG. 6C, in the positive ion mode of the mass spectrum, the mass-to-charge ratios of the fragments of the primary parent ion corresponding to 5.728min and 6.293min are 492.2130 and 520.2435, respectively, and the deduced molecular formulas are C₂₇H₂₉N₃O₆(molecular weight 491.54) and C₂₉H₃₃N₃O₆(molecular weight of 519.60) respectively corresponding to two monascus orange pigments (monascus red pigment and monascus rubigins) and products generated by amino-philic reaction of histidine.

In conclusion, the invention provides a method for remarkably improving the stability of monascus red pigment, namely, adding histidine, phenylalanine, tyrosine or tryptophan solution into monascus orange pigment solution to prepare the amino acid-derivatized monascus red pigment, and the result proves that the four amino acid-derivatized monascus red pigments are still stable in a low pH value environment, wherein the pigment residue rate of the histidine-derivatized monascus red pigment is kept at 87% -96% after the heat treatment at 65-95 ℃, and the method can be applied to the processing production of acidic foods such as acidic beverages, maintains the stability of the pigment while carrying out high-temperature sterilization and filling, and has extremely high application value.

The above description is only for the preferred embodiment of the present invention, but the scope of the present invention is not limited thereto, and any changes or substitutions that can be easily conceived by those skilled in the art within the technical scope of the present invention are included in the scope of the present invention.