A method for improving the color of plant-based meat products using recombinant soybean hemoglobin

Abstract

The application discloses a method for improving color of plant-based meat products by using recombinant soybean hemoglobin, and belongs to the field of food biotechnology and plant-based artificial meat processing. In view of the problem of pale color of plant-based artificial meat, the application adopts Bacillus subtilis to ferment and produce recombinant soybean hemoglobin (S-Hb), and adds the S-Hb in the form of purified protein or cell lysate into soybean protein isolate at a proportion of 1% (w / w), and then mixes and heat-treats to prepare plant-based meat products. The plant-based meat products prepared by the method have significantly improved redness value and brightness value, and the color changes from light yellow to meat-like red, effectively solving the problem of poor color of plant-based meat products, and the use of cell lysate containing recombinant soybean hemoglobin can reduce the application cost.

Description

Technical Field

[0001] This invention relates to a method for improving the color of plant-based meat products using recombinant soybean leghemoglobin, belonging to the fields of food engineering and enzyme engineering technology. Background Technology

[0002] In recent years, with consumers' increasing focus on health and environmental protection, the plant-based meat product market has developed rapidly. Plant-based meat products, as a typical example, typically use plant proteins such as soy protein isolate as the main raw material, forming a meat-like texture through methods such as heat-induced gelation. However, compared with real pork, plant-based meat products have significant shortcomings in sensory quality, mainly in color and flavor. In terms of color, soy protein isolate itself is relatively light in color, often turning pale yellow or grayish-white after heat treatment, lacking the bright red color characteristic of pork, leading to lower consumer acceptance.

[0003] To improve the color of plant-based meat products, existing technologies typically add natural pigments such as beetroot red and red yeast rice, or color-developing agents such as nitrites. However, natural pigments generally suffer from poor heat stability and are prone to fading, while nitrites pose potential food safety risks. Adding animal-derived hemoglobin or ferrous heme can impart a red color to the product, but animal-derived ingredients do not align with the positioning of purely plant-based products and are prone to oxidation and browning.

[0004] Soy leghemoglobin has been reported to be used to improve the color of plant-based meat products, but natural soybean leghemoglobin is derived from soybean root nodules, resulting in low yields and high costs, making industrial application difficult. Therefore, there is an urgent need for a technical solution that can stably and effectively improve the color of plant-based meat products at a lower cost. Summary of the Invention

[0005] To address the issue of poor color in plant-based meat products, this invention adds recombinant soy leghemoglobin to give the meat products the color of real meat, thus optimizing the sensory quality of plant-based meat products in terms of both sight and smell.

[0006] The first objective of this invention is to provide a method for improving the color of plant-based meat products by adding recombinant soy leghemoglobin to plant protein raw materials, mixing them, and then subjecting them to heat treatment to obtain plant-based meat products.

[0007] In one embodiment of the present invention, the recombinant soybean hemoglobin is prepared by microbial fermentation.

[0008] In one embodiment, the recombinant soybean hemoglobin may be in the form of, but is not limited to, cell lysate after microbial fermentation or purified protein after separation and purification.

[0009] In one embodiment of the present invention, the microorganism is Bacillus subtilis.

[0010] In one embodiment of the present invention, the plant protein raw material includes, but is not limited to, soy protein isolate.

[0011] In one embodiment of the present invention, the amount of recombinant soybean hemoglobin added is 0.5 to 2% of the total mass of the plant protein raw materials.

[0012] In one embodiment of the present invention, the heat treatment temperature is 90~95°C and the treatment time is 15~25 min.

[0013] This invention also provides a method for preparing plant-based meat products, comprising the following steps: (1) Preparation of an aqueous suspension of soybean protein isolate; (2) Add recombinant soybean hemoglobin S-Hb or cell lysate containing recombinant soybean hemoglobin to the soy protein isolate aqueous suspension prepared in step (1), mix evenly, heat treat, and cool to obtain plant-based meat products.

[0014] In one embodiment, the concentration of the soy protein isolate aqueous suspension in step (1) is 5-15%.

[0015] In one embodiment, the amount of recombinant soybean hemoglobin S-Hb protein added in step (2) is 0.5% to 5% based on S-Hb protein.

[0016] In one embodiment, the amount of cell lysate containing recombinant soybean hemoglobin added in step (2) is 0.5% to 1% based on S-Hb protein.

[0017] In one embodiment, the heat treatment is performed at 95°C for 10 to 30 minutes.

[0018] The present invention also provides plant-based meat products prepared using the method described herein.

[0019] The present invention also provides the application of the method in the food industry.

[0020] The beneficial effects of this invention are: (1) The present invention utilizes recombinant soybean hemoglobin S-Hb or its cell lysate prepared by Bacillus subtilis as a coloring agent for plant-based meat products, which can significantly improve the color of plant-based meat products, significantly increase the redness value a and brightness value L of the product, and change the color from light yellow to red close to real meat, thereby improving the sensory quality of the product and consumer acceptance.

[0021] (2) This invention verifies the feasibility of cell lysate containing recombinant soybean hemoglobin S-Hb in improving plant-based meat products. It can provide good coloring effect for plant-based meat products and will not introduce unpleasant odors. It greatly simplifies the production process, reduces production costs, and is more suitable for industrial-scale production. Detailed Implementation

[0022] Soy protein isolate Suyun 626 (SPI) was purchased from Lianyungang Fengyuan Seed Industry Technology Co., Ltd. After neutralization to pH 7.0, the protein content of SPI was determined by the Kjeldahl method, and the result was 92% on a dry weight basis. SPI was prepared into a 10% (w / v) deionized water suspension and centrifuged at 10,000 g for 20 min to remove particulate matter.

[0023] Example 1: Purification of S-Hb protein Refer to " Metabolic engineering of Bacillus subtilis for the production of active hemoglobins and myoglobins by improving heme supply The method described in the article was used to construct recombinant Bacillus subtilis Heme-K2 expressing S-Hb protein and to prepare S-Hb protein by fermentation. The specific steps are as follows: (1) Construction of pMINI-gntR-S-Hb plasmid and Bacillus subtilis Heme-K2 engineered strain. The constructed engineered strain was inoculated into the culture medium and fermented at 30℃. (2) After the bacterial cells were crushed by a high-pressure homogenizer, they were purified by BeaverBeads™ nickel column affinity chromatography using a C-terminal 6×His tag.

[0024] (3) The purified S-Hb protein was obtained by desalting and concentrating the protein using an Amicon Ultra 10 K ultrafiltration centrifuge tube. The heme binding rate of the recombinant soybean hemoglobin S-Hb was 54.58±2.41%.

[0025] Example 2: Preparation of S-Hb protein lysis buffer The strain was cultured according to the method in Example 1. The bacterial cells in the fermentation broth were collected, and the cells were broken up by high-pressure homogenizer. The supernatant was collected by centrifugation and was used as the S-Hb protein lysis buffer.

[0026] Example 3: Method for preparing plant-based meat protein from S-Hb protein (1) Raw material pretreatment Soy protein isolate (SPI) is pulverized and sieved for later use. Recombinant soy hemoglobin (S-Hb) freeze-dried powder or cell lysate containing S-Hb is refrigerated at 4°C for later use.

[0027] (2) Preparation of plant-based meat product samples Soy protein isolate was prepared into a 10% (w / v) deionized water suspension and centrifuged at 10,000 g for 20 min to remove particulate matter.

[0028] The purified recombinant soybean leghemoglobin prepared in Example 1 and the cell lysate containing recombinant soybean leghemoglobin prepared in Example 2 were added to the soybean protein isolate suspension, respectively. The purified soybean leghemoglobin was added at a ratio of 1% (w / w), and the amount of S-Hb cell lysate added was calculated based on the S-Hb protein content, which was then converted to 1% (w / w). The mixture was stirred for 20 min to achieve homogeneity, allowed to stand at room temperature for 20 min, and then heat-treated at 95°C for 20 min. Subsequently, it was cooled to room temperature in an ice-water bath to obtain the plant-based meat product sample.

[0029] (3) Evaluation of coloring effect After heat treatment, the samples were cooled to room temperature, and the surface color parameters (L, a, b) were measured using a colorimeter. Table 1 shows that the redness value (a) of the sample with 1% purified S-Hb increased from 0.99 to 5.69, and the lightness value (L*) increased from 40.55 to 53.61, with the color changing from pale yellow to a flesh-colored hue. The sample with added S-Hb cell lysate also showed good staining effects without introducing any unpleasant odor, verifying the feasibility of directly using S-Hb-containing cell lysate for addition.

[0030] Table 1. Color parameters of plant-based meat products with different S-Hb addition levels.

[0031] Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Anyone skilled in the art can make various modifications and alterations without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention should be determined by the claims.

Claims

1. A method for improving the color of plant-based meat products, characterized in that, Recombinant soybean leghemoglobin is added to plant protein raw materials, mixed, and then heat-treated to produce plant-based meat products.

2. The method according to claim 1, characterized in that, The recombinant soybean hemoglobin was prepared by microbial fermentation.

3. The method according to claim 2, characterized in that, The recombinant soybean hemoglobin is available in forms including, but not limited to, cell lysates from microbial fermentation or purified proteins after separation and purification.

4. The method according to claim 3, characterized in that, The microorganism in question is Bacillus subtilis.

5. A method for preparing plant-based meat products, characterized in that, Includes the following steps: (1) Preparation of an aqueous suspension of soybean protein isolate; (2) Add recombinant soybean hemoglobin S-Hb or cell lysate containing recombinant soybean hemoglobin to the soy protein isolate aqueous suspension prepared in step (1), mix evenly, heat treat, and cool to obtain plant-based meat products.

6. The method according to claim 5, characterized in that, The concentration of the soy protein isolate aqueous suspension in step (1) is 5-15%.

7. The method according to claim 5, characterized in that, In step (2), the amount of recombinant soybean hemoglobin S-Hb protein added is 0.5% to 5% based on S-Hb protein.

8. The method according to any one of claims 5 to 7, characterized in that, The heat treatment temperature is 90~95℃, and the treatment time is 15~25 min.

9. Plant-based meat products prepared by the method described in any one of claims 5 to 8.

10. The application of the method according to any one of claims 1 to 8 in the food industry.

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