Exploiting 1,3-Propanediol Production by a Clostridium beijerinckii Strain: The Role of Glycerol and Ammonium Sulfate Concentrations
Abstract
1. Introduction
2. Material and Methods
2.1. Microorganism and Inoculum
2.2. Culture Medium
2.3. Central Composite Rotational Design (CCRD)
2.4. Comparison of Culture Media in Kinetic Assays
2.5. Absolute Gene Expression Level
2.6. Analytical Determination
2.7. Statistical Validation
3. Results and Discussions
3.1. Optimization of Glycerol and (NH4)2SO4 Concentrations
3.2. Comparative Assay to Assess the Impact of Media Optimization on Product Formation, Substrate Consumption, and the Expression of Key Metabolic Enzymes
3.3. Sulfate and Ammonium Consumption
3.4. Gene Expression
4. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Enzyme | Primers (5′ 3′) | Reporter 1 (5′ 3′) |
---|---|---|
DNA gyrase (normalizer) | FW: AGATGTTGTGATAACATTAACACATTCAGGAT R: GCTTGAATTCCTTTTCCACCTCTTC | TCTGCCGAAATTCTT |
Glycerol Uptake | FW: CCAGCATTAACAATTGCGTTAGCA R: TGCACCAATACATTGTCCAGCTAAA | CCCATGCCACTTTACC |
Glycerol dehydratase | FW: TGGATTAAATTAAATGATATTAATAAAGTAAGAGATGAGATTTCAACT R: TCTGAATTTTGCCCTCCAACGATTA | ATAGCCGCCAAAATG |
Phosphoenolpyruvate: Dihydroxyacetone phosphotransferase | FW: CAGAGTAAACATCTTCTATTGCAGCTGAT R: GGCTGGAGGAACGTCAGAT | CAGTGCCAACTCTTCC |
1,3-Propanediol dehydrogenase | FW: GGACGCTTTAACTCATGCAATTGAA R: CTTGTATTGCTGAAGCATCTGTCA | CCGGATTTGCATCTTT |
ATP sulfurylase (Adenosine Triphosphate sulfurylase) | FW: GACCAAATCATACTTTCAGAGGATTCCA R: GGAAGTGTCGTAATTTCATCTCCTACA | TTCCAGCTTCAATTTC |
Conditions | Results | ||||||
---|---|---|---|---|---|---|---|
Conditions | Glycerol (mmol L−1) | (NH4)2SO4 (mmol L−1) | Final pH | OD600 | Consumed Glycerol (mmol L−1) | Butyric Acid (mmol L−1) | 1,3-PDO (mmol L−1) |
1 | 200 (−1) | 7.6 (−1) | 5.06 ± 0.10 | 2.09 ± 0.18 | 109.5 ± 6.38 | 18.29 ± 1.13 | 37.41 ± 3.73 |
2 | 200 (−1) | 22.7 (+1) | 5.46 ± 0.05 | 2.01 ± 0.30 | 152.80 ± 8.97 | 10.22 ± 0.01 | 21.30 ± 1.12 |
3 | 400 (+1) | 7.6 (−1) | 5.29 ± 0.06 | 2.06 ± 0.16 | 181.51 ± 14.58 | 19.57 ± 1.84 | 76.27 ± 7.34 |
4 | 400 (+1) | 22.7 (+1) | 5.16 ± 0.04 | 2.36 ± 0.11 | 185.62 ± 5.62 | 18.96 ± 1.26 | 53.88 ± 5.85 |
5 | 158 (−1.41) | 15.1 (0) | 6.03 ± 0.01 | 1.74 ± 0.00 | 51.36 ± 1.87 | 6.03 ± 0.01 | 3.44 ± 2.31 |
6 | 441 (+1.41) | 15.1 (0) | 5.49 ± 0.08 | 2.11 ± 0.08 | 157.42 ± 12.62 | 15.51 ± 0.66 | 41.89 ± 1.28 |
7 | 300 (0) | 4.4 (−1.41) | 5.15 ± 0.02 | 2.20 ± 0.23 | 164.54 ± 1.83 | 17.27 ± 0.29 | 50.98 ± 0.67 |
8 | 300 (0) | 25.8 (+1.41) | 5.16 ± 0.13 | 2.53 ± 0.11 | 183.32 ± 10.26 | 20.34 ± 2.56 | 69.18 ± 2.91 |
9 | 300 (0) | 15.1 (0) | 5.26 ± 0.02 | 2.04 ± 0.04 | 135.51 ± 13.76 | 15.84 ± 0.89 | 38.33 ± 0.66 |
10 | 300 (0) | 15.1 (0) | 5.39 ± 0.30 | 2.10 ± 0.25 | 123.72 ± 19.97 | 12.68 ± 0.66 | 35.70 ± 6.79 |
Factors | Significance | Sum of Squares | Degrees of Freedom | Mean Square | F | p |
---|---|---|---|---|---|---|
Linear glycerol | Significant | 1909.76 | 1 | 1909.76 | 1699.83 | 0.015 |
Quadratic glycerol | Significant | 235.04 | 1 | 235.04 | 209.20 | 0.044 |
Quadratic (NH4)2SO4 | Significant | 561.29 | 1 | 561.29 | 499.59 | 0.028 |
Lack of fit | Not significant | 372.37 | 5 | 74.47 | 66.29 | 0.093 |
Pure error | 1.12 | 1 | 1.12 | |||
Total | 3634.43 | 9 |
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de Castro, P.F.D.L.; Altafini, R.d.M.; Bortolucci, J.; Florêncio, J.; Campos, M.L.A.M.; Reginatto, V. Exploiting 1,3-Propanediol Production by a Clostridium beijerinckii Strain: The Role of Glycerol and Ammonium Sulfate Concentrations. Fermentation 2025, 11, 187. https://doi.org/10.3390/fermentation11040187
de Castro PFDL, Altafini RdM, Bortolucci J, Florêncio J, Campos MLAM, Reginatto V. Exploiting 1,3-Propanediol Production by a Clostridium beijerinckii Strain: The Role of Glycerol and Ammonium Sulfate Concentrations. Fermentation. 2025; 11(4):187. https://doi.org/10.3390/fermentation11040187
Chicago/Turabian Stylede Castro, Pedro Felipe Dassie Leonel, Rafael de Moraes Altafini, Jonatã Bortolucci, Jaques Florêncio, Maria Lucia Arruda Moura Campos, and Valeria Reginatto. 2025. "Exploiting 1,3-Propanediol Production by a Clostridium beijerinckii Strain: The Role of Glycerol and Ammonium Sulfate Concentrations" Fermentation 11, no. 4: 187. https://doi.org/10.3390/fermentation11040187
APA Stylede Castro, P. F. D. L., Altafini, R. d. M., Bortolucci, J., Florêncio, J., Campos, M. L. A. M., & Reginatto, V. (2025). Exploiting 1,3-Propanediol Production by a Clostridium beijerinckii Strain: The Role of Glycerol and Ammonium Sulfate Concentrations. Fermentation, 11(4), 187. https://doi.org/10.3390/fermentation11040187