Optimization of Fermentation Conditions for the Production of the M23 Protease Pseudoalterin by Deep-Sea Pseudoalteromonas sp. CF6-2 with Artery Powder as an Inducer
Abstract
:1. Introduction
2. Results and Discussion
2.1. Effect of Different Inducers on Pseudoalterin Production
Inducers | Elastolytic activity after different fermentation time (U/mL) | |||||
---|---|---|---|---|---|---|
24 h | 36 h | 48 h | 60 h | 72 h | 84 h | |
Elastin (0.3%) | 16.5 ± 0.7 | 32.4 ± 4.0 | 35.3 ± 3.5 | 42.9 ± 3.2 | 46.9 ± 3.9 | 41.4 ± 3.6 |
Artery (wet, 2.5%) | 12.6 ± 0.8 | 27.5 ± 1.0 | 42.1 ± 2.0 | 45.8 ± 1.6 | 54.6 ± 2.4 | 42.7 ± 1.7 |
Bovine lung (wet, 2.5%) | 6.2 ± 0.8 | 9.2 ± 1.7 | 7.1 ± 1.2 | 7.2 ± 1.3 | 14.5 ± 1.4 | 11.4 ± 0.6 |
Ligament (wet, 2.5%) | 10.3 ± 0.4 | 18.0 ± 0.9 | 11.9 ± 0.6 | 10.0 ± 0.7 | 18.1 ± 0.7 | 16.2 ± 1.2 |
Chicken intestine (wet, 2.5%) | 6.7 ± 1.2 | 9.7 ± 1.0 | 8.3 ± 0.4 | 7.2 ± 0.4 | 14.9 ± 1.0 | 11.1 ± 1.0 |
Beef extract (1%) | 4.8 ± 0.5 | 11.6 ± 0.4 | 11.2 ± 0.0 | 14.2 ± 0.8 | 13.1 ± 0.9 | 12.9 ± 1.3 |
Gelatin (1%) | ND b | 4.3 ± 0.7 | 1.1 ± 0.0 | 0.9 ± 0.2 | ND b | ND b |
Yeast extract peptone dextrose (1%) | ND b | 3.6 ± 0.2 | 2.2 ± 0.1 | 3.8 ± 0.2 | 2.4 ± 0.2 | 2.2 ± 0.2 |
BSA (1%) | 1.1 ± 0.0 | 1.0 ± 0.0 | 1.3 ± 0.1 | 1.0 ± 0.0 | ND b | ND b |
Peptone (1%) | ND b | 1.6 ± 0.1 | 1.3 ± 0.1 | 3.1 ± 0.2 | 1.1 ± 0.1 | 2.0 ± 0.0 |
Soybean meal (1%) | ND b | ND b | 0.9 ± 0.0 | 1.8 ± 0.4 | 1.4 ± 0.4 | 2.2 ± 0.2 |
Casein (1%) | ND b | 3.6 ± 0.3 | 3.1 ± 0.5 | 5.0 ± 0.1 | 1.4 ± 0.0 | 2.9 ± 0.2 |
2.2. Single Factor Experiments
2.3. Central Composite Design (CCD) and Response Surface Analysis
Run | X1 (artery) | X2 (temperature) | X3 (time) | Activity (U/mL) | |||
---|---|---|---|---|---|---|---|
Coded level | Real level (%) | Coded level | Real level (°C) | Coded level | Real Level (h) | ||
1 | 0 | 0.9 | +1.68 | 24.0 | 0 | 25.0 | 54.38 |
2 | 0 | 0.9 | 0 | 19.0 | +1.68 | 33.4 | 75.99 |
3 | +1 | 1.2 | +1 | 22.0 | −1 | 20.0 | 66.51 |
4 | +1 | 1.2 | +1 | 22.0 | +1 | 30.0 | 99.23 |
5 | −1 | 0.6 | +1 | 22.0 | +1 | 30.0 | 63.37 |
6 | 0 | 0.9 | 0 | 19.0 | 0 | 25.0 | 89.15 |
7 | 0 | 0.9 | 0 | 19.0 | −1.68 | 16.6 | 52.76 |
8 | −1 | 0.6 | −1 | 16.0 | +1 | 30.0 | 26.42 |
9 | 0 | 0.9 | −1.68 | 14.0 | 0 | 25.0 | 7.79 |
10 | 0 | 0.9 | 0 | 19.0 | 0 | 25.0 | 91.40 |
11 | 0 | 0.9 | 0 | 19.0 | 0 | 25.0 | 89.83 |
12 | +1 | 1.2 | −1 | 16.0 | +1 | 30.0 | 56.65 |
13 | +1.68 | 1.4 | 0 | 19.0 | 0 | 25.0 | 78.99 |
14 | −1 | 0.6 | +1 | 22.0 | −1 | 20.0 | 50.66 |
15 | +1 | 1.2 | −1 | 16.0 | −1 | 20.0 | 41.81 |
16 | 0 | 0.9 | 0 | 19.0 | 0 | 25.0 | 93.82 |
17 | −1 | 0.6 | −1 | 16.0 | −1 | 20.0 | 16.37 |
18 | −1.68 | 0.4 | 0 | 19.0 | 0 | 25.0 | 50.78 |
19 | 0 | 0.9 | 0 | 19.0 | 0 | 25.0 | 90.16 |
20 | 0 | 0.9 | 0 | 19.0 | 0 | 25.0 | 106.42 |
Source | df | Elastolytic activity of fermented broth a | |||
---|---|---|---|---|---|
Sum of Squares | Mean Square | F Value | p-value Prob > F | ||
Model | 9 | 14610.93 | 1623.44 | 41.99 | <0.0001 b |
X1 | 1 | 1755.06 | 1755.06 | 45.39 | <0.0001 b |
X2 | 1 | 3455.08 | 3455.08 | 89.36 | <0.0001 b |
X3 | 1 | 876.13 | 876.13 | 22.66 | 0.0008 b |
X1X2 | 1 | 1.97 | 1.97 | 0.051 | 0.8258 |
X1X3 | 1 | 76.86 | 76.86 | 1.99 | 0.1889 |
X2X3 | 1 | 52.68 | 52.68 | 1.36 | 0.2702 |
X12 | 1 | 1371.88 | 1371.88 | 35.48 | 0.0001 b |
X22 | 1 | 6815.04 | 6815.04 | 176.25 | <0.0001 b |
X32 | 1 | 1423.06 | 1423.06 | 36.80 | 0.0001 b |
Residual | 10 | 386.67 | 38.67 | ||
Lack of Fit | 5 | 171.62 | 34.32 | 0.80 | 0.5947 |
Pure Error | 5 | 215.05 | 43.01 | ||
Cor Total | 19 | 14997.60 |
2.4. Verification of Optimal Conditions
3. Experimental Section
3.1. Strain and Media
3.2. Inoculum Preparation and Flask Fermentation
3.3. Determination of Elastase Activity
3.4. Effect of Different Inducers on Pseudoalterin Production
3.5. Optimization by Single Factor Experiments
3.6. Optimization by Response Surface Methodology (RSM)
3.7. Statistical Analysis
4. Conclusions
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Zhao, H.-L.; Yang, J.; Chen, X.-L.; Su, H.-N.; Zhang, X.-Y.; Huang, F.; Zhou, B.-C.; Xie, B.-B. Optimization of Fermentation Conditions for the Production of the M23 Protease Pseudoalterin by Deep-Sea Pseudoalteromonas sp. CF6-2 with Artery Powder as an Inducer. Molecules 2014, 19, 4779-4790. https://doi.org/10.3390/molecules19044779
Zhao H-L, Yang J, Chen X-L, Su H-N, Zhang X-Y, Huang F, Zhou B-C, Xie B-B. Optimization of Fermentation Conditions for the Production of the M23 Protease Pseudoalterin by Deep-Sea Pseudoalteromonas sp. CF6-2 with Artery Powder as an Inducer. Molecules. 2014; 19(4):4779-4790. https://doi.org/10.3390/molecules19044779
Chicago/Turabian StyleZhao, Hui-Lin, Jie Yang, Xiu-Lan Chen, Hai-Nan Su, Xi-Ying Zhang, Feng Huang, Bai-Cheng Zhou, and Bin-Bin Xie. 2014. "Optimization of Fermentation Conditions for the Production of the M23 Protease Pseudoalterin by Deep-Sea Pseudoalteromonas sp. CF6-2 with Artery Powder as an Inducer" Molecules 19, no. 4: 4779-4790. https://doi.org/10.3390/molecules19044779