Ultrasonic Microwave-Assisted Micelle Combined with Fungal Pretreatment of Eucommia ulmoides Leaves Significantly Improved the Extraction Efficiency of Total Flavonoids and Gutta-Percha
Abstract
:1. Introduction
2. Materials and Methods
2.1. Materials
2.2. Dilute Alkali Pretreatment of Biomass
2.3. Process Optimization
2.4. Chromogenic Experiment of Horny Layer
2.5. Removal of Holocellulose by Trichoderma viride Fermentation
2.5.1. Optimization of Fermentation Conditions of Trichoderma viride
2.5.2. Determination of Enzyme Activity
2.5.3. Method for Determination of Lignin, Cellulose, and Hemicellulose
2.6. Extraction and Purification of Gutta-Percha
2.7. Effect of Trichoderma viride Pretreatment on the Yield of Gutta-Percha
2.8. Scanning Electron Microscope (SEM)
2.9. X-ray Diffraction (XRD)
2.10. Fourier Transform Infrared (FTIR)
2.11. 1H NMR Analysis
3. Results and Discussion
3.1. Optimization of Biomass Pretreatment Process for Eucommia ulmoides Leaves
3.1.1. Effects of the Surfactant
3.1.2. Effect of the Liquid–Solid Ratio
3.1.3. Effect of the Microwave Power and Extraction Time
3.2. Optimization of Extraction Technology of Total Flavonoids by CCD Software
3.3. Characterization of Removal Effect of Stratum Corneum
3.4. Optimization of Fermentation Conditions of Trichoderma viride
3.5. Content of Lignin, Cellulose, and Hemicellulose
3.6. Optimization of Extraction Conditions of Gutta-Percha
3.6.1. Effect of the Solid–Liquid Ratio
3.6.2. Effect of the Extraction Temperature
3.6.3. Effect of the Extraction Time
3.6.4. Effect of the Extraction Times
3.6.5. Optimization of Extraction Technology of Gutta-Percha by CCD Software
3.6.6. Comparison of Yield of Gutta-Percha before and after Pretreatment
3.7. SEM Analysis
3.8. XRD Analysis
3.9. FTIR Analysis
3.10. 1H NMR Spectra of Gutta-Percha
4. Conclusions
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Extraction Methods | Extraction Solvent | Extraction Time (min) | Total Flavonoid Extraction Rate (%) | Reference |
---|---|---|---|---|
Ultrasound/microwave-assisted extraction | 41% ethanol | 26 | 2.454 ± 0.230 | Xiang Wang et al., 2020 |
Refluxed | 95% ethanol | 360 | 1.39 | Weixing Huang et al., 2020 |
Ultrasound extraction | 65% ethanol | 30 | - | Daixiu Yuan et al., 2017 |
Supercritical CO2 extraction | 80% ethanol | 150 | 2.032 | Jiaxing Li et al., 2013 |
No. | Central Composite Designs | No. | Central Composite Designs | ||||||||
---|---|---|---|---|---|---|---|---|---|---|---|
X1 | X2 | X3 | X4 | Y | X1 | X2 | X3 | X4 | Y | ||
1 | 1.25 | 600 | 25 | 45 | 0.81 | 16 | 1.75 | 800 | 35 | 55 | 1.67 |
2 | 1.75 | 600 | 25 | 45 | 1.14 | 17 | 1.0 | 700 | 30 | 50 | 0.87 |
3 | 1.25 | 800 | 25 | 45 | 0.69 | 18 | 2.0 | 700 | 30 | 50 | 1.52 |
4 | 1.75 | 800 | 25 | 45 | 1.15 | 19 | 1.5 | 500 | 30 | 50 | 1.30 |
5 | 1.25 | 600 | 35 | 45 | 1.14 | 20 | 1.5 | 900 | 30 | 50 | 0.84 |
6 | 1.75 | 600 | 35 | 45 | 1.37 | 21 | 1.5 | 700 | 20 | 50 | 1.45 |
7 | 1.25 | 800 | 35 | 45 | 0.99 | 22 | 1.5 | 700 | 40 | 50 | 1.68 |
8 | 1.75 | 800 | 35 | 45 | 1.34 | 23 | 1.5 | 700 | 30 | 40 | 1.10 |
9 | 1.25 | 600 | 25 | 55 | 1.39 | 24 | 1.5 | 700 | 30 | 60 | 1.96 |
10 | 1.75 | 600 | 25 | 55 | 1.78 | 25 | 1.5 | 700 | 30 | 50 | 1.61 |
11 | 1.25 | 800 | 25 | 55 | 1.14 | 26 | 1.5 | 700 | 30 | 50 | 1.67 |
12 | 1.75 | 800 | 25 | 55 | 1.64 | 27 | 1.5 | 700 | 30 | 50 | 1.62 |
13 | 1.25 | 600 | 35 | 55 | 1.56 | 28 | 1.5 | 700 | 30 | 50 | 1.58 |
14 | 1.75 | 600 | 35 | 55 | 1.86 | 29 | 1.5 | 700 | 30 | 50 | 1.68 |
15 | 1.25 | 800 | 35 | 55 | 1.20 | 30 | 1.5 | 700 | 30 | 50 | 1.65 |
Analysis of Variance | |||||
---|---|---|---|---|---|
Source | Sum of Square | Degree of Freedom | Mean Square | F-Value | p-Value |
Model b | 3.24 | 14 | 0.23 | 105.75 | <0.0001 c |
X1 | 0.78 | 1 | 0.78 | 355.96 | <0.0001 c |
X2 | 0.19 | 1 | 0.19 | 88.09 | <0.0001 c |
X3 | 0.14 | 1 | 0.14 | 64.82 | <0.0001 c |
X4 | 1.18 | 11 | 1.18 | 540.76 | <0.0001 c |
X1X2 | 0.017 | 1 | 0.017 | 7.95 | 0.0129 |
X2X4 | 0.027 | 1 | 0.027 | 12.35 | 0.0031 |
X3X4 | 0.031 | 1 | 0.031 | 14.22 | 0.0018 |
X12 | 0.37 | 1 | 0.37 | 168.06 | <0.0001 c |
X22 | 0.60 | 1 | 0.50 | 273.26 | <0.0001 c |
X32 | 0.015 | 1 | 0.015 | 6.83 | 0.0196 |
X42 | 0.028 | 1 | 0.028 | 12.91 | 0.0027 |
Residual | 0.033 | 15 | 0.00219 | - | - |
Lack of fit | 0.026 | 10 | 0.002551 | 1.74 | 0.2039 |
Pure error | 0.00735 | 5 | 0.00147 | - | - |
Corrected total | 3.28 | 29 | - | - | - |
Credibility analysis of the regression equations | |||||
Standard deviation | Mean | Coefficient of variation | R2 | ||
0.047 | 1.38 | 3.39 | 0.9852 | ||
Adjust R2 | Predicted R2 | Adequacy precision | |||
0.9761 | 0.9519 | 37.1 |
Variable | pH Value A | Solid–Liquid Ratio B | Inoculum Amount (%) C | Culture Time (days) D | Enzyme Activity |
---|---|---|---|---|---|
1 | 4 | 1:10 | 25 | 3 | 9.21 |
2 | 4 | 1:20 | 30 | 5 | 5.598 |
3 | 4 | 1:50 | 20 | 7 | 3.61 |
4 | 5 | 1:10 | 30 | 7 | 2.348 |
5 | 5 | 1:20 | 20 | 3 | 3.25 |
6 | 5 | 1:50 | 25 | 5 | 2.538 |
7 | 6 | 1:10 | 20 | 5 | 19.882 |
8 | 6 | 1:20 | 25 | 7 | 18.42 |
9 | 6 | 1:50 | 30 | 3 | 13.91 |
K1 | 6.1392 | 10.48 | 10.0525 | 8.7883 | |
K2 | 2.7083 | 9.0892 | 7.2833 | 9.3358 | |
K3 | 17.4025 | 6.6808 | 8.9142 | 8.1258 | |
Range | 14.6942 | 3.7992 | 2.76992 | 1.2100 | |
Optimal level | A3 | B1 | C1 | D2 | |
Optimum condition | 6 | 1:10 | 20 | 5 |
No. | Central Composite Designs | No. | Central Composite Designs | ||||||||
---|---|---|---|---|---|---|---|---|---|---|---|
X1 | X2 | X3 | X4 | Y | X1 | X2 | X3 | X4 | Y | ||
1 | 0.05 | 2 | 60 | 65 | 2.956 | 16 | 0.08 | 4 | 120 | 85 | 2.918 |
2 | 0.08 | 2 | 60 | 65 | 1.38 | 17 | 0.03 | 3 | 90 | 75 | 3.1 |
3 | 0.05 | 4 | 60 | 65 | 2.766 | 18 | 0.1 | 3 | 90 | 75 | 1.21 |
4 | 0.08 | 4 | 60 | 65 | 1.569 | 19 | 0.07 | 1 | 90 | 75 | 2.336 |
5 | 0.05 | 2 | 120 | 65 | 2.876 | 20 | 0.07 | 5 | 90 | 75 | 3.094 |
6 | 0.08 | 2 | 120 | 65 | 2.125 | 21 | 0.07 | 3 | 30 | 75 | 2.892 |
7 | 0.05 | 4 | 120 | 65 | 2.945 | 22 | 0.07 | 3 | 150 | 75 | 2.661 |
8 | 0.08 | 4 | 120 | 65 | 1.706 | 23 | 0.07 | 3 | 90 | 55 | 3.999 |
9 | 0.05 | 2 | 60 | 85 | 2.15 | 24 | 0.07 | 3 | 90 | 95 | 3.986 |
10 | 0.08 | 2 | 60 | 85 | 1.986 | 25 | 0.07 | 3 | 90 | 75 | 3.873 |
11 | 0.05 | 4 | 60 | 85 | 3.653 | 26 | 0.07 | 3 | 90 | 75 | 4.359 |
12 | 0.08 | 4 | 60 | 85 | 3.636 | 27 | 0.07 | 3 | 90 | 75 | 3.714 |
13 | 0.05 | 2 | 120 | 85 | 2.163 | 28 | 0.07 | 3 | 90 | 75 | 3.896 |
14 | 0.08 | 2 | 120 | 85 | 2.236 | 29 | 0.07 | 3 | 90 | 75 | 4.011 |
15 | 0.05 | 4 | 120 | 85 | 3.447 | 30 | 0.07 | 3 | 90 | 75 | 2.932 |
Analysis of Variance | |||||
---|---|---|---|---|---|
Source | Sum of Square | Degree of Freedom | Mean Square | F-Value | p-Value |
Model b | 18.89 | 14 | 1.35 | 7.52 | 0.0002 |
X1 | 3.51 | 1 | 3.51 | 19.57 | 0.0005 |
X2 | 1.65 | 1 | 1.65 | 9.17 | 0.0085 |
X3 | 0.00084 | 1 | 0.00084 | 0.004684 | 0.9463 |
X4 | 0.61 | 1 | 0.61 | 3.42 | 0.084 |
X1X4 | 1.06 | 1 | 1.06 | 5.93 | 0.0278 |
X2X4 | 1.87 | 1 | 1.87 | 10.42 | 0.0056 |
X12 | 6.15 | 1 | 6.15 | 34.29 | <0.0001 c |
X22 | 3.05 | 1 | 3.05 | 17.01 | 0.0009 |
X32 | 2.78 | 1 | 2.78 | 15.48 | 0.0013 |
X42 | 0.0055 | 1 | 0.0055 | 0.031 | 0.8629 |
Residual | 2.69 | 15 | 0.18 | - | - |
Lack of fit | 1.56 | 10 | 0.16 | 0.69 | 0.7129 |
Pure error | 1.13 | 5 | 0.23 | - | - |
Corrected total | 21.58 | 29 | - | - | - |
Credibility analysis of the regression equations | |||||
Standard deviation | Mean | Coefficient of variation | R2 | ||
0.42 | 2.89 | 12.15 | 0.8897 | ||
Adjust R2 | Predicted R2 | Adequacy precision | |||
0.7589 | 0.5084 | 9.758 |
Sample | Sample Quality (g) | Extraction Time (min) | Reaction Temperature (°C) | Extraction Times | Turpentine Volume (mL) | Yield of Gutta-Percha (%) | Amount of Solvent Required (mL/mg) |
---|---|---|---|---|---|---|---|
Eucommia ulmoides leaves (untreated) | 5 | 60 | 85 | 1 | 100 | 0.66 | 2.91 |
Eucommia ulmoides leaves treated with Trichoderma viride | 5 | 60 | 85 | 1 | 100 | 2.77 | 0.96 |
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Wu, M.; Liu, P.; Wang, S.; Zhong, C.; Zhao, X. Ultrasonic Microwave-Assisted Micelle Combined with Fungal Pretreatment of Eucommia ulmoides Leaves Significantly Improved the Extraction Efficiency of Total Flavonoids and Gutta-Percha. Foods 2021, 10, 2399. https://0-doi-org.brum.beds.ac.uk/10.3390/foods10102399
Wu M, Liu P, Wang S, Zhong C, Zhao X. Ultrasonic Microwave-Assisted Micelle Combined with Fungal Pretreatment of Eucommia ulmoides Leaves Significantly Improved the Extraction Efficiency of Total Flavonoids and Gutta-Percha. Foods. 2021; 10(10):2399. https://0-doi-org.brum.beds.ac.uk/10.3390/foods10102399
Chicago/Turabian StyleWu, Mingfang, Peiyan Liu, Siying Wang, Chen Zhong, and Xiuhua Zhao. 2021. "Ultrasonic Microwave-Assisted Micelle Combined with Fungal Pretreatment of Eucommia ulmoides Leaves Significantly Improved the Extraction Efficiency of Total Flavonoids and Gutta-Percha" Foods 10, no. 10: 2399. https://0-doi-org.brum.beds.ac.uk/10.3390/foods10102399