Optimization of the Extraction of Bioactive Compounds from Walnut (Juglans major 209 x Juglans regia) Leaves: Antioxidant Capacity and Phenolic Profile
Abstract
:1. Introduction
2. Materials and Methods
2.1. Reagents and Standards
2.2. Materials
2.3. Extraction
2.4. Box-Behnken Experimental Design
2.5. Analytical Techniques
3. Results and Discussion
3.1. Effect of the Solid/Liquid Ratio on Extraction Yield and Extract Antioxidant Activity
3.2. Relationship between Extract Antioxidant Properties
3.3. Optimization of the Extraction Conditions by a Box-Behnken Design
3.4. Characterization of the Optimum Walnut Leaf Extract
4. Conclusions
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Exp. | S/L Ratio (g/mL) | Extraction Yield (%) | FRAP (nmol AAE/mg Extract) |
---|---|---|---|
A | 1/5 | 25.82 ± 0.25 a | 1246 ± 48 a |
B | 1/7.5 | 27.04 ± 0.42 b | 1350 ± 53 b |
C | 1/10 | 27.86 ± 0.04 b | 1512 ± 61 c |
Exp | Y1 exp | Y1 pred | Y2 exp | Y2 pred | Y3 exp | Y3 pred | Y4 exp | Y4 pred | |||
---|---|---|---|---|---|---|---|---|---|---|---|
1 | −1 | −1 | 0 | 21.03 ± 0.89 | 22.35 | 1181 ± 39 | 1262 | 1.114 ± 0.026 | 1.050 | 0.951 ± 0.026 | 0.964 |
2 | −1 | 0 | −1 | 20.35 ± 0.18 | 19.86 | 842 ± 46 | 908 | 0.663 ± 0.044 | 0.666 | 0.671 ± 0.027 | 0.774 |
3 | −1 | 0 | 1 | 19.15 ± 2.13 | 18.71 | 930 ± 47 | 908 | 0.873 ± 0.013 | 0.852 | 0.807 ± 0.017 | 0.774 |
4 | −1 | 1 | 0 | 23.89 ± 0.28 | 24.16 | 1250 ± 58 | 1262 | 1.008 ± 0.159 | 1.050 | 1.053 ± 0.049 | 0.964 |
5 | 0 | −1 | −1 | 23.13 ± 0.27 | 22.85 | 1160 ± 37 | 1041 | 0.913 ± 0.024 | 0.932 | 0.893 ± 0.041 | 0.875 |
6 | 0 | −1 | 1 | 20.03 ± 1.16 | 19.96 | 1138 ± 17 | 1041 | 0.919 ± 0.039 | 0.932 | 0.866 ± 0.050 | 0.875 |
7 | 0 | 1 | −1 | 24.52 ± 0.11 | 24.66 | 943 ± 65 | 1041 | 0.873 ± 0.037 | 0.932 | 0.869 ± 0.030 | 0.875 |
8 | 0 | 1 | 1 | 21.93 ± 1.76 | 21.77 | 876 ± 18 | 1041 | 1.003 ± 0.036 | 0.932 | 0.875 ± 0.028 | 0.875 |
9 | 1 | −1 | 0 | 29.08 ± 1.43 | 28.40 | 1364 ± 42 | 1529 | 1.493 ± 0.106 | 1.396 | 1.166 ± 0.047 | 1.250 |
10 | 1 | 0 | −1 | 27.01 ± 0.30 | 27.64 | 1199 ± 50 | 1174 | 1.193 ± 0.021 | 1.198 | 1.089 ± 0.035 | 1.060 |
11 | 1 | 0 | 1 | 22.35 ± 0.88 | 23.02 | 1238 ± 32 | 1174 | 1.031 ± 0.023 | 1.012 | 1.095 ± 0.032 | 1.060 |
12 | 1 | 1 | 0 | 30.17 ± 0.09 | 30.21 | 1468 ± 20 | 1529 | 1.318 ± 0.049 | 1.396 | 1.256 ± 0.017 | 1.250 |
13 | 0 | 0 | 0 | 25.85 ± 0.05 | 26.28 | 1389 ± 78 | 1395 | 1.239 ± 0.043 | 1.223 | 1.299 ± 0.049 | 1.315 |
14 | 0 | 0 | 0 | 27.19 ± 0.51 | 26.28 | 1560 ± 49 | 1395 | 1.190 ± 0.031 | 1.223 | 1.330 ± 0.032 | 1.315 |
Independent Variables | Levels | ||||||||||
−1 | 0 | +1 | |||||||||
x1, Temperature (°C) | 25 | 50 | 75 | ||||||||
x2, Time (min) | 30 | 75 | 120 | ||||||||
x3, % EtOH | 10 | 50 | 90 |
Compound | Linear Range (mg/L) | Calibration Curve | R2 |
---|---|---|---|
(–)-Gallocatechin | 1–200 | y = 8068x + 17,932 | 0.9941 |
Catechin hydrate | 1–200 | y = 15,461x + 52,943 | 0.9958 |
Chlorogenic acid | 1–1000 | y = 8181x + 22,563 | 0.9936 |
Ellagic acid | 1–1000 | y = 9647x + 38,987 | 0.9959 |
Epicatechin | 1–1000 | y = 9780x + 17,102 | 0.9976 |
Ferulic acid | 1–200 | y = 3455x + 22,085 | 0.9936 |
Gallic acid | 1–1000 | y = 3814x + 4250 | 0.9977 |
Isorharmnetin | 1–200 | y = 61,453x + 78,552 | 0.9915 |
Kaempferol | 1–200 | y = 61,712x + 75,935 | 0.9923 |
Neochlorogenic acid | 1–200 | y = 10,675x + 15,125 | 0.9989 |
p-Coumaric acid | 1–200 | y = 4442x + 4535 | 0.9972 |
Procyanidin B2 | 1–200 | y = 4014x + 7252 | 0.9925 |
Quercetin | 1–1000 | y = 45,006x + 111,541 | 0.9922 |
Quercetin 3-β-d-glucoside | 1–1000 | y = 13,239x + 42,498 | 0.9836 |
Taxifolin | 1–200 | y = 21,398x + 28,956 | 0.9903 |
Y1 | Y2 | Y3 | Y4 | |||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|
Coeff. | SE | p | Coeff | SE | p | Coeff. | SE | p | Coeff. | SE | p | |
a0 | 26.281 | 0.274 | 0.000 | 1395.250 | 40.672 | 0.000 | 1.223 | 0.019 | 0.000 | 1.315 | 0.028 | 0.000 |
a1 | 3.024 | 0.274 | 0.000 | 13.25 | 40.672 | 0.006 | 0.173 | 0.019 | 0.000 | 0.143 | 0.019 | 0.000 |
a2 | 0.905 | 0.274 | 0.008 | - | - | NS | - | - | NS | - | - | NS |
a3 | −1.444 | 0.274 | 0.000 | - | - | NS | - | - | NS | - | - | NS |
a12 | - | - | NS | - | - | NS | - | - | NS | - | - | NS |
a13 | −0.865 | 0.387 | 0.049 | - | - | NS | −0.093 | 0.027 | 0.005 | - | - | NS |
a23 | - | - | NS | - | - | NS | - | - | NS | - | - | NS |
a11 | - | - | NS | - | - | NS | - | - | NS | −0.083 | 0.028 | 0.012 |
a22 | - | - | NS | - | - | NS | - | - | NS | −0.125 | 0.028 | 0.001 |
a33 | −3.973 | 0.387 | 0.000 | −354.500 | 57.519 | 0.000 | −0.291 | 0.027 | 0.000 | −0.315 | 0.028 | 0.000 |
R2 | 0.964 | 0.789 | 0.947 | 0.951 | ||||||||
R2corr. | 0.947 | 0.757 | 0.934 | 0.933 | ||||||||
SE | 0.774 | 115.038 | 0.053 | 0.055 | ||||||||
p | 0.000 | 0.000 | 0.000 | 0.000 |
Compound | Retention Time (min) | m/z | ng/mg Extract d.b. | mg/100 g Leaves d.b. |
---|---|---|---|---|
(–)-Gallocatechin | 2.4 | 305.06 | 14.17 | 0.45 |
Catechin hydrate | 2.6 | 289.07 | 55.83 | 1.78 |
Chlorogenic acid | 2.7 | 353.09 | 5737.50 | 183.14 |
Ellagic Acid | 4 | 300.99 | 68.92 | 2.20 |
Epicatechin | 2.8 | 289.07 | 12.08 | 0.39 |
Ferulic acid | 3.6 | 193.04 | 158.58 | 5.06 |
Gallic acid | 2.3 | 169.01 | 432.58 | 13.81 |
Isorharmnetin | 5.4 | 315.05 | 3.17 | 0.10 |
Kaempferol | 5.3 | 285.04 | 14.25 | 0.45 |
Neochlorogenic acid | 2.5 | 353.08 | 9125.00 | 291.27 |
p-Coumaric ccid | 3.6 | 163.04 | 10.83 | 0.35 |
Procyanidin B2 | 2.6 | 577.14 | 45.50 | 1.45 |
Quercetin | 4.8 | 301.04 | 163.50 | 5.22 |
Quercetin 3-β-d-glucoside | 3.8 | 463.09 | 15,441.67 | 492.90 |
Taxifolin | 3.4 | 303.05 | 4.50 | 0.14 |
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Fernández-Agulló, A.; Castro-Iglesias, A.; Freire, M.S.; González-Álvarez, J. Optimization of the Extraction of Bioactive Compounds from Walnut (Juglans major 209 x Juglans regia) Leaves: Antioxidant Capacity and Phenolic Profile. Antioxidants 2020, 9, 18. https://doi.org/10.3390/antiox9010018
Fernández-Agulló A, Castro-Iglesias A, Freire MS, González-Álvarez J. Optimization of the Extraction of Bioactive Compounds from Walnut (Juglans major 209 x Juglans regia) Leaves: Antioxidant Capacity and Phenolic Profile. Antioxidants. 2020; 9(1):18. https://doi.org/10.3390/antiox9010018
Chicago/Turabian StyleFernández-Agulló, Adela, Aída Castro-Iglesias, María Sonia Freire, and Julia González-Álvarez. 2020. "Optimization of the Extraction of Bioactive Compounds from Walnut (Juglans major 209 x Juglans regia) Leaves: Antioxidant Capacity and Phenolic Profile" Antioxidants 9, no. 1: 18. https://doi.org/10.3390/antiox9010018