Use of Yarrowia lipolytica Lipase Immobilized in Cell Debris for the Production of Lipolyzed Milk Fat (LMF)
Abstract
:1. Introduction
2. Results and Discussion
2.1. Production of Lipase from Y. lipolytica Immobilized in Cell Debris Induced by Residual Frying Oil
2.2. Characterization of Lipase Immobilized in Cell Debris
2.3. Anhydrous Milk Fat Production
2.4. Lipolysis of Milk Fat
3. Materials and Methods
3.1. Materials
3.2. Strain, Media, and Inoculum Preparation
3.3. Production of Lipases (Fermentation Process)
3.4. Preparation of Lipase Immobilized in Cell Debris
3.5. Lipase Activity
3.6. Effect of Temperature and pH on Lipase Immobilized in Cell Debris
3.7. Thermal Stability and Stability in Different Solvents of Lipase Immobilized in Cell Debris
3.8. Repeated Use of Lipase Immobilized in Cell Debris
3.9. Preparation of Anhydrous Milk Fat
3.10. Milk Fat Hydrolysis Reactions
3.11. Monitoring Lipid Composition during Milk Fat Hydrolysis Reactions
3.11.1. High Performance Liquid Chromatography (HPLC)
3.11.2. Thin Layer Chromatography (TLC)
3.11.3. Analysis of Fatty Acid Composition by Gas Chromatography (GC)
3.12. Statistical Analysis
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
Abbreviations
HPLC | High-performance liquid chromatography |
GC | Gas chromatography |
TLC | Thin layer chromatography |
LMF | Lipolyzed milk fat |
LipImDebri | Lipase immobilized in cell debris |
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Fatty Acids | Contents (%, mol/mol) | Fatty Acids | Contents (%, mol/mol) |
---|---|---|---|
Saturated | Monounsaturated | ||
4:0 | 0.63 ± 0.45 | 14:1 | 0.66 ± 0.02 |
6:0 | 0.48 ± 0.23 | 15:1 | 0.28 ± 0.01 |
8:0 | 1.35 ± 0.33 | 16:1 | 1.30 ± 0.06 |
10:0 | 0.11 ± 0.03 | 17:1 | 0.84 ± 0.01 |
11:0 | 0.16 ± 0.06 | 18:1n-9 | 28.5 ± 2.03 |
12:0 | 1.88 ± 0.17 | Polyunsaturated | |
13:0 | 0.06 ± 0.00 | 18:2n-6 | 2.17 ± 0.03 |
14:0 | 9.13 ± 0.26 | 18:3n-6 | 0.43 ± 0.02 |
15:0 | 0.94 ± 0.03 | 18:3n-3 | 0.44 ± 0.02 |
16:0 | 31.9 ± 0.79 | 20:1n-9 | 0.75 ± 0.02 |
17:0 | 0.50 ± 0.01 | 20:3n-6 | 0.03 ± 0.00 |
18:0 | 16.0 ± 0.15 | 20:3n-3 | 0.10 ± 0.01 |
20:0 | 0.24 ± 0.01 | Conjugated | |
21:0 | 0.06 ± 0.01 | CLA * | 0.35 ± 0.02 |
22:0 | 0.12 ± 0.02 | ||
Branched-chain | |||
i14:0 | 0.08 ± 0.00 | ||
i15:0 | 0.26 ± 0.01 | ||
i18:0 | 0.29 ± 0.00 |
Time of Lipolysis (h) | Lipid Classes | |
---|---|---|
500 mg LipImDebri | 750 mg LipImDebri | |
Free Fatty Acid (FFA) + Monoacylglycerol (MAG) Fraction (%) | ||
0 | 0.00 a,C | 0,00 a,C |
1.5 | 0.98 ± 0.29 a,C | 5.48 ± 2.53 a,B,C |
3.0 | 11.02 ± 1.00 a,B,C | 19.54 ±0.94 b,A,B,C |
4.5 | 17.86 ± 5.84 a,A,B | 11.79 ± 4.58 a,A,B |
6.0 | 28.23 ± 3.45 a,A | 14.30 ± 5.34 a,A,B |
Diacylglycerol (DAG) Fraction (%) | ||
0 | 1.11 ± 0.02 a,A | 1.11 ± 0.02 a,A |
1.5 | 0.31 ± 0.34 a,A | 0.57 ± 0.15 a,A |
3.0 | 1.57 ± 0.94 a,A | 1.27 ± 0.02 a,A |
4.5 | 1.18 ± 0.52 a,A | 0.62 ± 0.20 a,A |
6.0 | 0.56 ± 0.05 a,A | 1.41 ± 0.48 a,A |
Triacylglycerol (TAG) Fraction (%) | ||
0 | 98.96 ± 0.11 a,A | 98.96 ± 0.11 a,A |
1.5 | 98.71 ± 0.63 a,A | 93.95 ± 2.68 a,A,B |
3.0 | 87.41 ± 1.95 a,B | 79.19 ± 0.96 b,C |
4.5 | 80.96 ± 5.30 a,B | 87.60 ±4.37 a,AB,C |
6.0 | 72.71 ± 5.52 a,C | 84.29 ± 4.86 a,B,C |
Fatty Acid | LipImDebri (mg), Lipolysis Time (h) | ||
---|---|---|---|
500, 4.5 | 500, 6 | 750, 3 | |
Saturated | |||
4:0 | 0.74 ± 0.29 a,D | 0.51 ± 0.13 a,D | 0.76 ± 0.53 a,E |
6:0 | 0.45 ± 0.09 a,D | 0.27 ± 0.02 a,D | 0.65 ± 0.49 a,E |
8:0 | 4.40 ± 1.98 a,C,D | 6.79 ± 5.30 a,D | 3.63 ± 3.46 a,E,F |
10:0 | 0.74 ± 0.02 a,C,D | 0.56 ± 0.15 a,D | 1.44 ± 1.16 a,E |
12:0 | 1.99 ± 0.83 a,C,D | 2.42 ± 1.57 a,D | 2.74 ± 0.16 a,D,E |
14:0 | 8.17 ± 0.03 a,C | 6.03 ± 0.99 a,D | 8.14 ± 2.20 a,D |
15:0 | 0.90 ± 0.05 a,C,D | 0.79 ± 0.12 a,D | 0.81 ± 0.01 a,E |
16:0 | 28.80 ± 3.00 a,A | 24.97 ± 6.94 a,B | 24.67 ± 0.71 a,B |
18:0 | 13.94 ± 1.60 a,B | 15.36 ± 3.25 a,C | 12.79 ± 1.60 a,C |
Branched-Chain | |||
i15:0 | 0.22 ± 0.01 a,D | 0.22 ± 0.01 a,D | 0.21 ± 0.03 a,E |
a15:0 | 0.34 ± 0.03 a,C,D | 0.29 ± 0.05 a,D | 0.31 ± 0.06 a,E |
Monounsaturated | |||
14:1 | 0.56 ± 0.09 a,C,D | 0.40 ± 0.06 a,D | 0.69 ± 0.41 a,E |
16:1 | 1.19 ± 0.27 a,C,D | 1.07 ± 0.08 a,D | 1.32 ± 0.39 a,E |
17:1 | 0.69 ± 0.13 a,C,D | 0.64 ± 0.02 a,D | 0.56 ± 0.08 a,E |
18:1n-9 | 31.27 ± 5.16 a,A | 35.18 ± 1.83 a,A | 35.31 ± 1.05 a,A |
Polyunsaturated | |||
18:2n-6 | 4.29 ± 1.24 a,C,D | 3.84 ± 0.54 a,D | 4.82 ± 0.47 a,D,E |
20:1n-9 | 1.33 ± 0.32 a,C,D | 0.67 ± 0.07 a,D | 0.85 ± 0.18 a,E |
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Fraga, J.L.; Penha, A.C.B.; Da S. Pereira, A.; Silva, K.A.; Akil, E.; Torres, A.G.; Amaral, P.F.F. Use of Yarrowia lipolytica Lipase Immobilized in Cell Debris for the Production of Lipolyzed Milk Fat (LMF). Int. J. Mol. Sci. 2018, 19, 3413. https://doi.org/10.3390/ijms19113413
Fraga JL, Penha ACB, Da S. Pereira A, Silva KA, Akil E, Torres AG, Amaral PFF. Use of Yarrowia lipolytica Lipase Immobilized in Cell Debris for the Production of Lipolyzed Milk Fat (LMF). International Journal of Molecular Sciences. 2018; 19(11):3413. https://doi.org/10.3390/ijms19113413
Chicago/Turabian StyleFraga, Jully L., Adrian C. B. Penha, Adejanildo Da S. Pereira, Kelly A. Silva, Emília Akil, Alexandre G. Torres, and Priscilla F. F. Amaral. 2018. "Use of Yarrowia lipolytica Lipase Immobilized in Cell Debris for the Production of Lipolyzed Milk Fat (LMF)" International Journal of Molecular Sciences 19, no. 11: 3413. https://doi.org/10.3390/ijms19113413