Fading detection method for fruiting bodies of tremella aurantialba in mature period

Abstract

The invention relates to a fading detection method for fruiting bodies of tremella aurantialba in a mature period, and belongs to the technical field of edible mushroom color detection and evaluation. The method comprises the following steps: drying and crushing the tremella aurantialba sporocarp into powder; adding the crushed tremella aurantialba powder into an organic solvent of acetone and ethanol according to a material-liquid ratio of 1: 5 (w / v, g / mL), carrying out ultrasonic extraction in a dark place, and centrifuging under the condition of 8000 g to obtain a beta-carotene extracting solution; and irradiating the beta-carotene extracting solution under ultraviolet light for different times, substituting each measured absorbance into the beta-carotene standard curve to obtain the content of the beta-carotene, calculating the degradation amount of the beta-carotene, and measuring the fading condition of the tremella aurantialba. According to the method, the factors causing the color fading of the tremella aurantialba sporocarp and the color fading detection method are put forward for the first time, the detection method is simple, convenient, safe and efficient, guarantee is provided for rapid evaluation of the commodity value of the tremella aurantialba, and meanwhile an important foundation is laid for further development of a tremella aurantialba sporocarp color protection technology.

Description

Technical Field

[0001] The invention relates to the technical field of edible fungus color detection and evaluation, and in particular to a method for detecting the discoloration of mature golden fungus fruiting bodies. Background Art

[0002] Golden ear (Naematelia aurantialba) belongs to the fungi, phylum Basidiomycetes, class Tremellales, order Tremellales, and genus Auricularia, etc. It is a edible and medicinal fungus mainly distributed in China. It has a unique fruiting body flavor and high nutritional value, and contains many bioactive ingredients, including polysaccharides, active proteins, β-carotene, and flavonoids. It has high edible and medicinal value. Therefore, golden ear is becoming more and more popular among consumers, and its proportion in the diet of Chinese residents is increasing year by year. The surface of the mature golden ear fruiting body is bright orange-yellow, with excellent appearance. However, due to the instability of pigment substances, the color of golden ear is prone to fade during storage and processing, which not only affects its appearance and commodity value, but also affects its flavor and nutritional value. Therefore, it is very important to explore the main factors causing the fading of golden ear fruiting bodies, seek a safe and efficient fading detection method for mature golden ear fruiting bodies, evaluate the color of golden ear, so as to scientifically evaluate the commodity value of golden ear, and carry out research on the color preservation of golden ear on the basis of clarifying the fading mechanism of golden ear. Up to now, the industry's color judgment of golden ear fruiting bodies is limited to naked eye observation. The judgment results are subject to large human differences, inconsistent standards, and only the surface color of the golden ear can be observed. There has been no available scientific and standardized method for detecting the fading of golden ear fruiting bodies. Summary of the invention

[0003] The purpose of the invention is to fill the gap in the prior art and provide a simple, safe and efficient method for detecting the discoloration of mature Tremella fuciformis fruiting bodies.

[0004] The technical solution adopted by the present invention is as follows:

[0005] A method for detecting discoloration of mature Tremella fuciformis fruiting bodies, the method being as follows:

[0006] 1) Dry the fruiting body of Tremella truncatula and crush it into powder;

[0007] 2) The crushed Tremella fuciformis powder was dissolved in an organic solvent of acetone:ethanol 1:1 (v:v) at a solid-liquid ratio of 1:5 (w / v, g / mL), extracted by ultrasound in the dark for 20-40 min, and centrifuged at 8000 g to obtain a β-carotene extract;

[0008] 3) The β-carotene extract was irradiated under ultraviolet light for different time periods, and the absorbance was measured and then inserted into the β-carotene standard curve to obtain the β-carotene content. The β-carotene degradation amount was calculated to measure the fading of golden ear.

[0009] Furthermore, in the above step 1), the Tremella fuciformis fruiting bodies are dried and then crushed into 100-200 meshes.

[0010] Furthermore, the β-carotene standard curve in step 3) is obtained by using β-carotene as a standard substance and preparing solutions at concentrations of 10.0, 7.5, 5.0, 2.5, 1.0, and 0.0 μg / mL to draw a β-carotene standard curve.

[0011] The technical solution of the present invention can reasonably process the golden ear fruiting body, and calculate the degradation amount of beta-carotene based on the absorbance measured after irradiation with ultraviolet light, so as to quickly measure the fading of the golden ear.

[0012] The present invention proposes for the first time the factors causing the fading of golden ear fruiting bodies and a fading detection method. The detection method is simple, efficient and safe, which provides a guarantee for the rapid evaluation of the commercial value of golden ear, and at the same time lays an important foundation for further developing the color protection technology of golden ear fruiting bodies. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Figure 1 The types and contents of carotenoids contained in the fruiting bodies of Tremella truncatula at different growth stages;

[0014] Figure 2 is the standard curve of β-carotene;

[0015] Figure 3 This is the degradation rate change diagram of β-carotene under ultraviolet light;

[0016] Figure 4 This is a graph showing the degradation rate of β-carotene under white light. DETAILED DESCRIPTION

[0017] The content of the present invention will be further described below in conjunction with specific embodiments.

[0018] Main materials and reagents:

[0019] Acetone, acetonitrile, formic acid, methanol, methyl tert-butyl ether, 2,6-di-tert-butyl-p-cresol (BHT) and n-hexane were all chromatographically pure and purchased from Merck Co., Ltd.; anhydrous ethanol was analytically pure and purchased from Shanghai Sinopharm Chemical Reagent Co., Ltd.; β-carotene was purchased from Sigma-Aldrich (Shanghai) Trading Co., Ltd.

[0020] Three different growth periods of golden ear fruiting bodies (from Yunnan Junshijie Biotechnology Co., Ltd.) were selected, including 25 days, 35 days and 45 days, and their apparent chromaticity values ​​were detected and analyzed using a colorimeter [Bio-Gene (Beijing) Biotechnology Co., Ltd.]. The average value of the three measurements was taken as the chromaticity value of the sample. At the same time, sensory judgment was used to observe the color changes of the samples with the naked eye to eliminate the possible errors caused by instrumental measurement.

[0021] Comparison of chromaticity values ​​at different growth stages

[0022] The chromaticity values ​​of the fruiting bodies of three different growing stages of the golden ear were measured, and the results are shown in Table 1

[0023] Table 1 Comparison of chromaticity values ​​of golden ear fruiting bodies at three different growth stages

[0024]

[0025] Among them, the L* value indicates the lightness of the surface color, which mainly depends on the reflectivity of the product surface. The larger the L* value, the whiter and brighter the sample color is. The a* value indicates the redness and greenness of the surface. By measuring the a* value of the sample surface, we can understand the degree to which the sample is biased towards red or green. The larger the a* value, the more red the sample color is. The b* value indicates the yellow-blueness of the surface color. By measuring the b* value of the sample surface, we can understand the degree to which the sample is biased towards yellow or blue. The larger the b* value, the more yellow the sample is. The larger the value, the greater the color difference, and the easier it is to distinguish the difference between the color and the standard color. The formula for the total color difference value is as follows:

[0026]

[0027] In the above formula, Refers to the color difference between each group of samples. It refers to the difference in brightness between samples in each group. It refers to the difference in redness and greenness between samples in each group. b refers to the difference in yellow-blue tint between samples in each group.

[0028] As can be seen from Table 1, the apparent color of the golden ear fruiting bodies at different growth stages was measured by a colorimeter. It was found that with the extension of the growth cycle, the red-green color and yellow-blue color of the golden ear increased significantly (a* and b*), reaching the highest at 45 days, which was consistent with the trend observed with the naked eye.

[0029] Calculate the color difference between each group of samples Compared with the golden ear fruiting bodies with a growth period of 25 days, the color difference value of the golden ear fruiting bodies of 35 days is lower, which is 2.96; compared with the golden ear fruiting bodies with a growth period of 25 days, the color difference value of the golden ear fruiting bodies of 45 days is 45.98, which is much larger than the color difference value of 35 days; combined with sensory evaluation, the golden ear fruiting bodies with a growth period of 25 days and 35 days are lighter beige, and only the golden ear fruiting bodies with a growth period of 45 days (maturity) are obviously orange-yellow. Therefore, the apparent orange-yellow color of the golden ear fruiting bodies mainly accumulates between 35 and 45 days (late fruiting body growth).

[0030] Sample preparation:

[0031] (1) The fruiting bodies of Tremella truncatula at different growth stages were dried and ground into powder using a ball mill (MM400, Retsch, Germany) (30 Hz, 1 min);

[0032] (2) Weigh 50 mg of the ground sample using an electronic balance (AS 60 / 220.R2, RADWAG, Poland, Europe) and extract it with 0.5 mL of a mixed solution of n-hexane / acetone / ethanol (1:1:1, v / v / v) containing 0.01% BHT (g / mL);

[0033] (3) After vortexing (MIX-200, Shanghai Jingxin Industrial Development Co., Ltd.) at room temperature for 20 min, centrifuge at 12,000 g for 5 min at 4°C, take the supernatant, repeat the extraction once, and centrifuge (5424R, Eppendorf, Germany) to combine the supernatants to obtain the extract;

[0034] (4) The extract was concentrated (CentriVap, LABCONCO, USA), reconstituted with 100 μL of a methanol / methyl tert-butyl ether mixture (1:1, v / v), filtered through a 0.22 μm filter, and stored in a brown injection bottle for LC-MS / MS (SCIEX, USA) analysis.

[0035] Chromatography mass spectrometry acquisition conditions:

[0036] The data acquisition instrument system mainly includes ultra-high performance liquid chromatography (UPLC) and tandem mass spectrometry (MS / MS).

[0037] The liquid phase conditions mainly include:

[0038] (1) Chromatographic column: YMC C30 (3 μm, 100 mm × 2.0 mm);

[0039] (2) Mobile phase: Phase A, methanol / acetonitrile (1:3, v / v) with 0.01% BHT and 0.1% formic acid; Phase B, methyl tert-butyl ether with 0.01% BHT;

[0040] (3) Gradient elution program: A / B was 100:0 (V / V) at 0 min, 100:0 (V / V) at 3 min, 30:70 (V / V) at 5 min, 5:95 (V / V) at 9 min, 100:0 (V / V) at 10 min, and 100:0 (V / V) at 11 min;

[0041] (4) Flow rate: 0.8 mL / min; column temperature: 28°C; injection volume: 2 μL.

[0042] Mass spectrometry conditions mainly include:

[0043] The atmospheric pressure chemical ionization source (APCI) temperature was 350°C and the curtain gas (CUR) was 25 psi. In the Q-Trap 6500+, each ion pair was scanned and detected according to the optimized declustering potential (DP) and collision energy (CE).

[0044] Data processing: Substitute the integrated peak area of ​​all detected samples into the linear equation of the standard curve for calculation, and then further substitute it into the calculation formula to finally obtain the content data of the substance in the actual sample.

[0045] Carotenoid content in the sample (μg / g) = c*V / 1000 / m;

[0046] c: Concentration value (μg / mL) obtained by substituting the integrated peak area in the sample into the standard curve;

[0047] V: volume of solution used for reconstitution (μL);

[0048] m: weighed sample mass (g).

[0049] Carotenoids are a general term for a class of natural pigments that are widely present in nature, including β-carotene, lycopene, α-carotene and lutein. They are natural powerful antioxidants and free radical scavengers, and are also involved in plant metabolic processes such as photosynthesis, light protection, and growth and development.

[0050] The results of LC-MS / MS analysis are shown in the attached Figure 1 As shown:

[0051] A total of 16 carotenoids were identified in the fruiting bodies of golden ear, mainly including carotene and lutein. During the growth period of 25-35 days, the contents of β-carotene, α-carotene, γ-carotene, phytoene, β-cryptoxanthin and other substances in golden ear fruiting bodies showed a certain increasing trend; during the growth period of 35-45 days, the above substances increased significantly in multiples, which was consistent with the trend of golden ear chromaticity and color difference value. Therefore, it is believed that the apparent orange-yellow color of golden ear is formed by the gradual and balanced accumulation of orange-red carotene and yellow lutein during the growth process, and the enrichment of this type of substance is consistent with the trend of the apparent color, mainly concentrated between 35-45 days. The golden ear fruiting bodies of 45 days (maturity period) are obviously orange-yellow and have the highest content of carotenoids, and are also most prone to fading.

[0052] The content of β-carotene in the fruiting body of golden ear is the highest, accounting for more than 70% of the total carotene content. Therefore, the content of β-carotene can be used as a characteristic indicator to explore the main factors affecting the apparent color stability of golden ear.

[0053] The orange-yellow color of golden ear may be positively correlated with its carotenoid content, and the β-carotene content is the highest (>70%) in golden ear at 45 days (maturity), so the mature golden ear fruiting bodies were selected, and β-carotene was used as a characteristic indicator to study the effect of different light on its stability. β-carotene is a conjugated double bond composed of isoprene residues. It has very good light absorption performance in the band range of 450-550 nm (blue-green in the visible light spectrum) and can play an important role in photosynthesis as a natural pigment. β-carotene is also the main component of yellow and orange fruits. Because it contains a large number of C=C conjugated systems between its molecules, it is very easy to undergo cis-trans isomerization and oxidative degradation under light conditions, making the pigment unstable.

[0054] Therefore, the β-carotene in the fruiting bodies of golden ear was extracted and prepared to explore the effect of light (white light and ultraviolet light) on the β-carotene in the fruiting bodies of golden ear. The specific steps are as follows:

[0055] (1) The 45-day-old Tremella fuciformis fruiting bodies were dried and crushed (150 mesh) and placed in acetone:ethanol (1:1, v:v) with a solid-liquid ratio of 1:5 (w / v, g / mL). After mixing, the mixture was wrapped in tin foil and sonicated in the dark (KQ5200E, Kunshan Shumei Ultrasonic Instrument Co., Ltd.) for 30 min. The mixture was centrifuged at 8000 g for 20 min and the supernatant was placed in a microplate reader (Synergy HT, BIO-TEK, USA) to measure the absorbance at 450 nm.

[0056] (2) Using β-carotene as the standard, the concentrations were set at 10.0, 7.5, 5.0, 2.5, 1.0, and 0.0 μg / mL, and the following was obtained: Figure 2 β-carotene standard curve shown;

[0057] (3) Divide the supernatant in step 1 into two groups, 8 mL each (three parallels), a UV irradiation group and a white light irradiation group;

[0058] (4) The absorbance of the UV irradiation group was measured every 2 hours for a total of 24 hours; the absorbance of the white light irradiation group was measured every week for a total of 3 weeks. The absorbance was substituted into the standard curve to obtain the β-carotene content, and a curve was drawn based on the degradation rate of β-carotene under UV and white light irradiation.

[0059] from Figure 3 It can be seen that with the extension of UV irradiation time, β-carotene is extremely unstable. From 0 to 10 h, its degradation rate increases linearly. At about 10 h, the degradation rate of β-carotene has reached 100%, which is basically completely degraded. Figure 4 It can be seen that the degradation rate of β-carotene can reach about 90% after 3 weeks of white light irradiation. Compared with ultraviolet light irradiation, the degradation rate of β-carotene under white light irradiation is slower. Therefore, ultraviolet light is the main factor causing the degradation of β-carotene in golden ear.

[0060] Based on the above research analysis and experimental verification, the following unified and standardized fading detection method for mature golden ear fruiting bodies was determined, as follows:

[0061] 1) The Tremella fuciformis fruiting bodies are dried and crushed into powders of 100-200 meshes, and in this embodiment, the powders are crushed into 150 meshes;

[0062] 2) The crushed Tremella fuciformis powder was dissolved in an organic solvent of acetone:ethanol 1:1 (v:v) at a solid-liquid ratio of 1:5 (w / v, g / mL), and subjected to light-protected ultrasonic extraction for 20 to 40 minutes, 30 minutes in this embodiment, and centrifuged at 8000 g for 20 minutes to obtain a β-carotene extract;

[0063] 3) The β-carotene extract was irradiated under ultraviolet light for different time periods, and the absorbance was measured and then inserted into the β-carotene standard curve to obtain the β-carotene content. The β-carotene degradation amount was calculated to measure the fading of golden ear.

[0064] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for detecting the discoloration of mature Tremella fuciformis fruiting bodies, characterized in that: The method is as follows: 1) Dry the fruiting body of Tremella truncatula and crush it into powder; 2) The crushed Tremella fuciformis powder was dissolved in an organic solvent of acetone:ethanol (1:1 (v:v) at a solid-liquid ratio of 1:5 (w / v, g / mL) and subjected to ultrasonic extraction in the dark for 20-40 min at 8000 °C. g Centrifuge under the conditions to obtain β-carotene extract; 3) The β-carotene extract was irradiated under ultraviolet light for different time periods, and the absorbance was measured and then inserted into the β-carotene standard curve to obtain the β-carotene content. The β-carotene degradation amount was calculated to measure the fading of golden ear.

2. The method for detecting discoloration of a mature Tremella fuciformis fruiting body according to claim 1, characterized in that: In the above step 1), the Tremella fuciformis fruiting bodies are dried and then crushed into 100-200 meshes.

3. The method for detecting discoloration of a mature Tremella fuciformis fruiting body according to claim 1 or 2, characterized in that: The β-carotene standard curve is obtained by using β-carotene as a standard product and preparing solutions at concentrations of 10.0, 7.5, 5.0, 2.5, 1.0, and 0.0 μg / mL to draw the β-carotene standard curve.

Images