The Importance of a Comparative Characterization of Saccharomyces Cerevisiae and Saccharomyces Pastorianus Strains for Brewing
Abstract
:1. Introduction
2. Materials and Methods
2.1. Yeast Isolates and Strains
2.2. Genetic Identification and Strain Determination
2.2.1. Real-Time Polymerase Chain Reaction
2.2.2. DNA Fingerprinting (PCR-Capillary Electrophoresis of the IGS2-314 Fragment)
2.3. Phenolic Off-Flavor Test
2.4. Brewing Trials
2.4.1. Wort
2.4.2. Propagation
2.4.3. Fermentation
2.5. Analytical Methods
2.5.1. Determining the Cell Count (Cells in Suspension and Total Cell Count)
2.6. Sensory Evaluation
3. Results
3.1. Genetic Analysis
3.1.1. Real-Time PCR Assays and IGS2-314 PCR-Capillary Electrophoresis
3.2. Brewing Trials
3.2.1. Sugar Utilization
3.2.2. Amino Acid Utilization
3.2.3. Fermentation Kinetics
3.2.4. Flocculation (Cell Count)
3.2.5. Change in pH Value
3.3. Flavor Characterization
3.3.1. Phenolic Off-Flavor
3.3.2 Fermentation by-Products
3.3.3. Sulfur Dioxide
3.3.4. Sensory Evaluation
4. Conclusions
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
References
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TUM Yeast Strains | |||
---|---|---|---|
TUM Yeast Strain | Yeast Species | Industrial Application | Origin |
Frisinga-TUM 34/70® | Saccharomyces pastorianus | lager beer production | Freising-Weihenstephan, Germany |
Securitas-TUM 193® | Saccharomyces pastorianus | lager beer production | Freising-Weihenstephan, Germany |
LeoBavaricus-TUM 68® | Saccharomyces cerevisiae | wheat beer production | Freising-Weihenstephan, Germany |
LunaBavara-TUM 127® | Saccharomyces cerevisiae | wheat beer production | Freising-Weihenstephan, Germany |
Colonia-TUM 177® | Saccharomyces cerevisiae | kölsch beer production | Krefeld, Germany |
Vetus-TUM 184® | Saccharomyces cerevisiae | alt beer production | Dusseldorf, Germany |
Mel-TUM 211® | Saccharomyces cerevisiae | ale and stout beer production | region unknown, Great Britain |
Monacus-TUM 381® | Saccharomyces cerevisiae | trappist beer production | region unknown, Germany |
Tropicus-TUM 506® | Saccharomyces cerevisiae | ale beer production | region unknown, Great Britain |
Harmonia-TUM 511® | Saccharomyces cerevisiae | ale beer production | region unknown, United States of America |
Real-Time PCR Systems, Primer and Probe Sequences (5´–3´) | System Name | Reference | S. cer. | S. cer. var. dia. | S. past. |
---|---|---|---|---|---|
Sbp-f CTTGCTATTCCAAACAGTGAGACT | Sbp | [32,33] | − | − | + |
Sbp-r1 TTGTTACCTCTGGGCGTCGA | |||||
Sbp-r2 GTTTGTTACCTCTGGGCTCG | |||||
Sbp ACTTTTGCAACTTTTTCTTTGGGTTTCGAGCA | |||||
Sc-f CAAACGGTGAGAGATTTCTGTGC | Sce | [32,33] | + | + | + |
Sc-r GATAAAATTGTTTGTGTTTGTTACCTCTG | |||||
Scer FAM-ACACTGTGGAATTTTCATATCTTTGCAACTT-BHQ1 | |||||
Sc-GRC-f CACATCACTACGAGATGCATATGCA | Sc-GRC3 | [30] | + | + | + |
Sc-GRC-r GCCAGTATTTTGAATGTTCTCAGTTG | |||||
Sc-GRC FAM-TCCAGCCCATAGTCTGAACCACACCTTATCT-BHQ1 | |||||
TF-f TTCGTTGTAACAGCTGCTGATGT | TF-COXII | [30] | + | + | − |
TF-r ACCAGGAGTAGCATCAACTTTAATACC | |||||
TF-MGB FAM-ATGATTTTGCTATCCCAAGTT-BHQ1 (MGB probe) | |||||
BF300E CTCCTTGGCTTGTCGAA | BF-300 | [32] | − | − | + |
BF300M GGTTGTTGCTGAAGTTGAGA | |||||
BF300 FAM-TGCTCCACATTTGATCAGCGCCA-BHQ1 | |||||
BF-LRE-f ACTCGACATTCAACTACAAGAGTAAAATTT | BF-LRE1 | [30] | − | − | + |
BF-LRE-r TCTCCGGCATATCCTTCATCA | |||||
BF-LRE FAM-ATCTCTACCGTTTTCGGTCACCGGC-BHQ1 | |||||
Sd-f TTCCAACTGCACTAGTTCCTAGAGG | Sdia | [32] | − | + | − |
Sd-r GAGCTGAATGGAGTTGAAGATGG | |||||
Sdia FAM-CCTCCTCTAGCAACATCACTTCCTCCG-BHQ1 |
Real-Time PCR Internal Amplification Control (IAC135) | ||
---|---|---|
System Name | Primer | Primer Sequence (5′–3′) |
IAC135 | IAC135-f | TGGATAGATTCGATGACCCTAGAAC |
IAC135-r | TGAGTCCATTTTCGCAGATAACTT | |
Probe | Probe Sequence (5′–3′) | |
IAC135-S | HEX-TGGGAGGATGCATTAGGAGCATTGTAAGAGAG-BHQ1 | |
Target DNA | DNA Sequence (5′–3′) | |
IAC135 | TGCTAGAGAATGGATAGATTCGATGACCCTAGAACTAGTGGGAGGATGCATTAGGAGCATTGTAAGAGAGTCGGAAGTTATCTGCGAAAATGGACTCATTCGAGTGGCCTATTGACGGTCGCCCAAGGTGTCGCA | |
IAC135-rev | TGCGACACCTTGGGCGACCGTCAATAGGCCACTCGAATGAGTCCATTTTCGCAGATAACTTCCGACTCTCTTACAATGCTCCTAATGCATCCTCCCACTAGTTCTAGGGTCATCGAATCTATCCATTCTCTAGCA |
Wort composition | |
---|---|
Parameter | Amount |
Original gravity (°P) | 12.40 |
pH | 5.19 |
Spec. weight SL 20/20 °C | 1.05 |
Zinc (mg/L) | 0.15 |
Free amino nitrogen (FAN) (mg/100 mL) | 25.00 |
Total amino acid content (AS) (mg/100 mL) | 203.22 |
Total sugar (g/L) | 83.78 |
EBC-Bittering units | 20.20 |
Glucose (g/L) | 11.46 |
Fructose (g/L) | 2.57 |
Saccharose (g/L) | 1.12 |
Maltose (g/L) | 53.65 |
Maltotriose (g/L) | 14.98 |
RT-PCR-System | ||||||||
---|---|---|---|---|---|---|---|---|
Species Confirmation | TUM Yeast Strain | Sc-GRC3 | Sce | TF-COXII | Sbp | BF-LRE1 | BF-300 | Sdia |
S. pastorianus (S. cerevisiae hybrid strain) | Frisinga-TUM 34/70® | + | + | − | + | + | + | − |
Securitas-TUM 193® | + | + | − | + | + | + | − | |
S. cerevisiae | LeoBavaricus-TUM 68® | + | + | + | − | − | − | − |
LunaBavaricus-TUM 127® | + | + | + | − | − | − | − | |
Colonia-TUM 177® | + | + | + | − | − | − | − | |
Vetus-TUM 184® | + | + | + | − | − | − | − | |
Mel-TUM 211® | + | + | + | − | − | − | − | |
Monacus-TUM 381® | + | + | + | − | − | − | − | |
Tropicus-TUM 506® | + | + | + | − | − | − | − | |
Harmonia-TUM 511® | + | + | + | − | − | − | − |
TUM Yeast Strain | Glucose | Fructose | Sucrose | Maltose | Maltotriose |
---|---|---|---|---|---|
Frisinga-TUM 34/70® | 98.61 ± 0.00 | 96.15 ± 0.00 | 100.00 ± 0.00 | 99.02 ± 0.12 | 94.06 ± 0.91 |
Securitas-TUM 193® | 98.70 ± 0.14 | 96.15 ± 0.00 | 100.00 ± 0.00 | 96.89 ± 0.12 | 86.60 ± 0.52 |
LeoBavaricus-TUM 68® | 100.00 ± 0.00 | 100.00 ± 0.00 | 100.00 ± 0.00 | 99.92 ± 0.02 | 99.65 ± 0.28 |
LunaBavaria-TUM 127® | 100.00 ± 0.00 | 100.00 ± 0.00 | 100.00 ± 0.00 | 99.07 ± 0.10 | 01.05 ± 2.89 |
Colonia-TUM 177® | 100.00 ± 0.00 | 100.00 ± 0.00 | 100.00 ± 0.00 | 99.85 ± 0.02 | 94.80 ± 0.78 |
Vetus-TUM 184® | 100.00 ± 0.00 | 100.00 ± 0.00 | 100.00 ± 0.00 | 98.00 ± 0.53 | 60.92 ± 5.87 |
Mel-TUM 211® | 98.55 ± 0.47 | 98.57 ± 0.21 | 98.51 ± 0.48 | 87.23 ± 0.82 | 26.66 ± 0.26 |
Monacus-TUM 381® | 100.00 ± 0.00 | 100.00 ± 0.00 | 100.00 ± 0.00 | 99.65 ± 0.14 | 97.75 ± 0.09 |
Tropicus-TUM 506® | 98.23 ± 0.12 | 98.05 ± 0.36 | 99.11 ± 0.00 | 98.48 ± 0.93 | 59.28 ± 0.81 |
Harmonia-TUM 511® | 99.42 ± 0.12 | 98.05 ± 0.00 | 95.54 ± 0.00 | 99.28 ± 0.09 | 83.91 ± 0.71 |
Apparent attenuation (AA %) of the final beer | ||
---|---|---|
TUM Yeast Strain | AA (%) | Fermentation time (hours) |
Frisinga-TUM 34/70® | 81.63 ± 0.51 | 216 |
Securitas-TUM 193® | 79.30 ± 0.51 | 216 |
LeoBavaricus-TUM 68® | 86.17 ± 0.05 | 96 |
LunaBavaria-TUM 127® | 76.20 ± 1.76 | 144 |
Colonia-TUM 177® | 84.93 ± 0.37 | 96 |
Vetus-TUM 184® | 80.97 ± 3.02 | 264 |
Mel-TUM 211® | 66.13 ± 0.51 | 240 |
Monacus-TUM 381® | 86.17 ± 0.11 | 168 |
Tropicus-TUM 506® | 77.37 ± 1.34 | 216 |
Harmonia-TUM 511® | 82.70 ± 0.42 | 144 |
Yeast Cell Sedimentation at the end of Primary Fermentation | ||||
---|---|---|---|---|
TUM Yeast Strain | Max. Cell Conc. | Cell Conc. FG | Difference | Flocculation Behavior |
Frisinga-TUM 34/70® | 35.90 | 1.80 | −34.10 | flocculent |
Securitas-TUM 193® | 48.80 | 1.30 | −47.50 | flocculent |
LeoBavaricus-TUM 68® | 62.81 | 32.29 | −30.52 | powdery |
LunaBavara-TUM 127® | 39.38 | 7.50 | −31.88 | flocculent |
Colonia-TUM 177® | 23.00 | 17.20 | −5.8 | powdery |
Vetus-TUM 184® | 17.83 | 0.50 | −17.33 | flocculent |
Mel-TUM 211® | 34.30 | 17.80 | −16.50 | powdery |
Monacus-TUM 381® | 57.65 | 18.12 | −39.53 | powdery |
Tropicus-TUM 506® | 36.45 | 19.36 | −17.09 | powdery |
Harmonia-TUM 511® | 61.30 | 43.07 | −18.23 | powdery |
pH Value Decrease during Primary Fermentation | |||||||||
---|---|---|---|---|---|---|---|---|---|
TUM Yeast Strain | 0 h | 24 h | 48 h | 72 h | 96 h | After primary fermentation | After maturation | Final beer (after lagering) | ∆pH |
Frisinga-TUM 34/70® | 5.2 | 4.6 | 4.5 | 4.5 | 4.5 | 4.5 | 4.5 | 4.4 ± 0.04 | 0.8 |
Securitas-TUM 193® | 5.2 | 4.6 | 4.5 | 4.5 | 4.5 | 4.5 | 4.5 | 4.5 ± 0.01 | 0.7 |
LeoBavaricus-TUM 68® | 5.2 | 4.8 | 4.3 | 4.3 | 4.2 | 4.2 | 4.2 | 4.4 ± 0.01 | 0.8 |
LunaBavaria-TUM 127® | 5.2 | 4.6 | 4.5 | 4.5 | 4.4 | 4.4 | 4.4 | 4.6 ± 0.01 | 0.6 |
Colonia-TUM 177® | 5.2 | 5 | 4.8 | 4.8 | 4.7 | 4.7 | 4.7 | 4.6 ± 0.02 | 0.6 |
Vetus-TUM 184® | 5.2 | 4.7 | 4.6 | 4.6 | 4.5 | 4.5 | 4.6 | 4.5 ± 0.04 | 0.7 |
Mel-TUM 211® | 5.2 | 4.5 | 4.4 | 4.4 | 4.4 | 4.4 | 4.5 | 4.7 ± 0.01 | 0.5 |
Monacus-TUM 381® | 5.2 | 4.6 | 4.5 | 4.5 | 4.5 | 4.5 | 4.5 | 4.5 ± 0.01 | 0.7 |
Tropicus-TUM 506® | 5.2 | 4.6 | 4.5 | 4.5 | 4.5 | 4.5 | 4.6 | 4.7 ± 0.01 | 0.5 |
Harmonia-TUM 511® | 5.2 | 4.6 | 4.4 | 4.4 | 4.4 | 4.4 | 4.4 | 4.4 ± 0.01 | 0.8 |
POF-Test/Sniffing Perception of: | |||
---|---|---|---|
TUM Yeast Strain | 4-Vinylguajacol/ Ferulic Acid | 4-Vinylphenol/ Coumaric Acid | 4-Vinylstyrene/ Cinnamic Acid |
Frisinga - TUM 3470® | − | − | − |
Securitas-TUM 193® | − | − | − |
LeoBavaricus-TUM 68® | + | + | + |
LunaBavaria-TUM 127® | + | + | + |
Colonia-TUM 177® | − | − | − |
Vetus-TUM 184® | − | − | − |
Mel-TUM 211® | − | − | − |
Monacus-TUM 381® | + | + | + |
Tropicus-TUM 506® | − | − | − |
Harmonia-TUM 511® | + | + | + |
Fermentation by-Products (mg/L) | |||||
---|---|---|---|---|---|
FBP | Frisinga-TUM 3470® | Securitas-TUM 193® | LeoBavaricus-TUM 68® | LunaBavaria-TUM 127® | Colonia-TUM 177® |
Isoamyl acetate | 0.63 ± 0.19 | 1.38 ± 0.05 | 4.07 ± 0.46 | 3.97 ± 2.30 | 2.40 ± 0.16 |
Ethyl acetate | 19.77 ± 2.50 | 26.07 ± 1.89 | 32.50 ± 2.97 | 36.97 ± 2.93 | 32.87 ± 1.68 |
∑ Ester (E) | 20.40 ± 2.69 | 27.90 ± 1.94 | 36.57 ± .3.43 | 40.93 ± 3.16 | 35.27 ± 1.83 |
n-Propanol | 11.23 ± 0.77 | 13.43 ± 0.72 | 22.77 ± 2.37 | 15.93 ± 0.70 | 21.30 ± 1.25 |
i-Butanol | 10.63 ± 0.71 | 14.27 ± 0.47 | 62.30 ± 3.51 | 43.70 ± 3.42 | 10.53 ± 0.11 |
Amyl alcohols | 60.53 ± 3.31 | 82.60 ± 2.68 | 127.00 ± 7.33 | 91.60 ± 3.34 | 80.77 ± 1.06 |
∑ Higher alcohols (HE) | 82.40 ± 4.76 | 110.30 ± 3.86 | 212.07 ± 13.15 | 151.23 ± 6.25 | 112.60 ± 2.32 |
4-Vinylguajacol | 0.10 ± 0.00 | 0.10 ± 0.00 | 3.10 ± 0.40 | 1.23 ± 0.30 | 0.10 ± 0.00 |
Diacetyl | 0.18 ± 0.07 | 0.10 ± 0 01 | 0.11 ± 0.01 | 0.17 ± 0.03 | 0.14 ± 0 03 |
2,3-Pentandione | 0.00 ± 0.01 | 0.03 ± 0.00 | 0.01 ± 0.00 | 0.02 ± 0.00 | 0.01 ± 0.01 |
∑ Vicinal diketones | 0.18 ± 0.07 | 0.12 ± 0.02 | 0.12 ± 0.01 | 0.19 ± 0.03 | 0.15 ± 0.04 |
Acetaldehyde | 1.33 ± 0.47 | 11.17 ± 1.46 | 7.27 ± 1.80 | 2.80 ± 0.94 | 8.23 ± 2.02 |
Ratio (∑E:∑HE) | 4.04 | 3.95 | 5.80 | 3.69 | 3.19 |
Fermentation by-Products (mg/L) | |||||
---|---|---|---|---|---|
FBP | Vetus- TUM 184® | Mel- TUM 211® | Monacus- TUM 381® | Tropicus- TUM 506® | Harmonia- TUM 511® |
Isoamyl acetate | 1.80 ± 0.09 | 2.03 ± 0.05 | 4.33 ± 0.05 | 1.53 ± 0.11 | 2.93 ± 0.05 |
Ethyl acetate | 37.40 ± 0.58 | 37.93 ± 0.11 | 50.60 ± 0.85 | 22.57 ± 1.31 | 54.40 ± 0.61 |
∑ Ester (E) | 39.20 ± 0.61 | 39.97 ± 0.14 | 54.93 ± 0.84 | 24.10 ± 1.36 | 57.33 ± 0.65 |
n-Propanol | 15.40 ± 0.46 | 18.30 ± 0.16 | 18.30 ± 0.16 | 20.67 ± 1.01 | 20.77 ± 0.53 |
i-Butanol | 11.37 ± 0.14 | 16.20 ± 0.37 | 24.17 ± 0.66 | 20.90 ± 1.40 | 13.13 ± 0.28 |
Amyl alcohols | 67.37 ± 0.91 | 59.27 ± 1.32 | 101.00 ± 4.23 | 88.50 ± 5.90 | 74.97 ± 1.24 |
∑ Higher alcohols (HE) | 94.13 ± 1.43 | 93.77 ± 1.83 | 142.23 ± 5.26 | 130.07 ± 8.12 | 108.87 ± 1.23 |
4-Vinylguajacol | 0.10 ± 0.00 | 0.10 ± 0.00 | 3.27 ± 0.05 | 0.10 ± 0.00 | 3.33 ± 0.05 |
Diacetyl | 0.06 ± 0.01 | 0.10 ± 0.01 | 0.07 ± 0.01 | 0.12 ± 0.01 | 0.06 ± 0.00 |
2,3-Pentandione | 0.01 ± 0.00 | 0.02 ± 0.00 | 0.01 ± 0.00 | 0.02 ± 0.00 | 0.01 ± 0.00 |
∑ Vicinal diketones | 0.06 ± 0.01 | 0.12 ± 0.01 | 0.08 ± 0.01 | 0.14 ± 0.01 | 0.07 ± 0.00 |
Acetaldehyde | 6.80 ± 0.80 | 4.60 ± 0.48 | 6.23 ± 0.77 | 5.93 ± 0.93 | 4.03 ± 0.46 |
Ratio (∑E:∑HE) | 2.40 | 2.35 | 2.59 | 5.40 | 1.90 |
SO2 Concentration of the Finished Beers | |
---|---|
TUM Yeast Strain | SO2 (mg/L) |
Frisinga-TUM 34/70® | 6.03 ± 0.30 |
Securitas-TUM 193® | 9.47 ± 0.68 |
LeoBavaricus-TUM 68® | 2.87 ± 0.30 |
LunaBavaria-TUM 127® | 1.63 ± 0.51 |
Colonia-TUM 177® | 3.80 ± 0.79 |
Vetus-TUM 184® | 3.10 ± 0.16 |
Mel-TUM 211® | 2.60 ± 0.98 |
Monacus-TUM 381® | 0.50 ± 0.00 |
Tropicus-TUM 506® | 2.23 ± 1.02 |
Harmonia-TUM 511® | 0.50 ± 0.00 |
TUM Yeast Strains | |||||||||
---|---|---|---|---|---|---|---|---|---|
TUM Yeast Strain | Yeast Species | Recommended Main Beer Style | Phenolic Off-Flavor | Flocculation Behavior | Maltotriose-Utilization (%) | ∆pH | SO2(mg/L) | AA(%) | Fermentation Time (days) |
Frisinga-TUM 34/70® | S. pastorianus | lager beer | − | flocculent | 94.06 ± 0.91 | 0.8 | 6.03 ± 0.30 | 81.63 ± 0.51 | 9 |
Securitas-TUM 193® | S. pastorianus | lager bee | − | flocculent | 86.60 ± 0.52 | 0.7 | 9.47 ± 0.68 | 79.30 ± 0.51 | 9 |
LeoBavaricus-TUM 68® | S. cerevisiae | wheat beer | + | powdery | 99.65 ± 0.28 | 0.8 | 2.87 ± 0.30 | 86.17 ± 0.05 | 4 |
LunaBavaria-TUM 127® | S. cerevisiae | wheat beer | + | flocculent | 01.05 ± 2.89 | 0.6 | 1.63 ± 0.51 | 76.20 ± 1.76 | 6 |
Colonia-TUM 177® | S. cerevisiae | kölsch and alt beer | − | powdery | 94.80 ± 0.78 | 0.6 | 3.80 ± 0.79 | 84.93 ± 0.37 | 4 |
Vetus-TUM 184® | S. cerevisiae | alt beer | − | flocculent | 60.92 ± 5.87 | 0.7 | 3.10 ± 0.16 | 80.97 ± 3.02 | 11 |
Mel-TUM 211® | S. cerevisiae | ale beer | − | powdery | 26.66 ± 0.26 | 0.5 | 2.60 ± 0.98 | 66.13 ± 0.51 | 10 |
Monacus-TUM 381® | S. cerevisiae | wheat beer | + | powdery | 97.75 ± 0.09 | 0.7 | 0.50 ± 0.00 | 86.17 ± 0.11 | 7 |
Tropicus-TUM 506® | S. cerevisiae | ale beer | − | powdery | 59.28 ± 0.81 | 0.5 | 2.23 ± 1.02 | 77.37 ± 1.34 | 9 |
Harmonia-TUM 511® | S. cerevisiae | ale and wheat beer | + | powdery | 83.91 ± 0.71 | 0.8 | 0.50 ± 0.00 | 82.70 ± 0.42 | 6 |
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Meier-Dörnberg, T.; Hutzler, M.; Michel, M.; Methner, F.-J.; Jacob, F. The Importance of a Comparative Characterization of Saccharomyces Cerevisiae and Saccharomyces Pastorianus Strains for Brewing. Fermentation 2017, 3, 41. https://doi.org/10.3390/fermentation3030041
Meier-Dörnberg T, Hutzler M, Michel M, Methner F-J, Jacob F. The Importance of a Comparative Characterization of Saccharomyces Cerevisiae and Saccharomyces Pastorianus Strains for Brewing. Fermentation. 2017; 3(3):41. https://doi.org/10.3390/fermentation3030041
Chicago/Turabian StyleMeier-Dörnberg, Tim, Mathias Hutzler, Maximilian Michel, Frank-Jürgen Methner, and Fritz Jacob. 2017. "The Importance of a Comparative Characterization of Saccharomyces Cerevisiae and Saccharomyces Pastorianus Strains for Brewing" Fermentation 3, no. 3: 41. https://doi.org/10.3390/fermentation3030041
APA StyleMeier-Dörnberg, T., Hutzler, M., Michel, M., Methner, F.-J., & Jacob, F. (2017). The Importance of a Comparative Characterization of Saccharomyces Cerevisiae and Saccharomyces Pastorianus Strains for Brewing. Fermentation, 3(3), 41. https://doi.org/10.3390/fermentation3030041