mead production: selection and characterization assays of
TRANSCRIPT
A.P. Pereira1, T Dias 1, J. Andrade1, E. Ramalhosa 1, A. Mendes-Ferreira 2, A. Mendes-Faia 2 , and M.L. Estevinho 1
Mead production: selection and characterization assays of Saccharomyces cerevisiae strains
1 CIMO- Centro de Investigação de Montanha/Escola Superior Agrária de Bragança, Quinta de Sta. Apolónia, Apartado 1172, 5301-855 Bragança, Portugal2 Institute for Biotechnology and Bioengineering-Centro de Genética e Biotecnologia-IBB/ CGB -Universidade de Trás-os-Montes e Alto Douro, Vila Real, Portugal
3
Mead is a traditional alcoholic drink which results from the fermentation of diluted honey. Yeasts used in mead production are, usually, wine Saccharomyces cerevisiae strains.Most of these yeasts are not adapted to the conditions of mead production namely, high sugar levels, low pH values and reduced nitrogen concentrations. The inability of yeaststrains to respond and adapt to unfavorable stressful growth conditions, leads to several problems, such as lack of uniformity of the final product, delays and “pouts”fermentations, as well as the production of off-flavors by the yeasts. Therefore, it is necessary to find yeast strains more suitable for mead production.
BACKGROUND
AIMS
METHODS
The main objectives of this work were to select, from honey, the most appropriate indigenous yeasts for of mead production and to optimize fermentation conditions.
YEAST ISOLATION YEAST IDENTIFICATION CHARACTERIZATION OF TWO HONEY SAMPLES USED INMEAD PRODUCTION
Stress resistance to ethanol, sulfur dioxide and osmoticstress were evaluated in seven Saccharomyces cerevisiaestrains (five isolated from honey, a commercial wine strain and
MEAD PRODUCTION a laboratory strain), using YPD medium.
For mead production, two S. cerevisiae yeasts strains isolatedfrom honey, randomly selected, and the commercial winestrain were used. The fermentation media obtained from twodifferent honeys (a dark and a light honeys) were enrichedwith two different supplements, a commercial one or onedeveloped by the research team.
Honey samples
Rhodotorula mucilaginosa
17%
Saccharomyces cerevisiae
8%
Zygosaccharomyces rouxii
8%
Pichia membranaefaciens
4%
Zygosaccharomyces mellis13% Candida sorbosivorans
8%
RESULTSYEAST SPECIES ISOLATED FROM HONEY SAMPLES YEAST TOLERANCE TO ETHANOL, SO2 AND OSMOTIC STRESS
Strain
μ (h-1)
ControlSO2 10% (v/v)
ethanol20% glucose +20% fructose100 mg/L 250mg/L
1 (honey) 0.18 0.18 0.18 0.10 0.18
2 (honey) 0.16 0.17 0.17 0.11 0.20
3 (reference) 0.18 0.17 0.17 0.10 0.19
4 (honey) 0.18 0.18 0.18 0.10 0.201000
m1
TAB
Z.ro
uxii
ESA5
1
5B3
M1
M70
M12
5
7ER
ESA1
7
ESA1
1
1Lisa
2Lisa
1694
-P01
1
1694
-P01
5
m2
10001000
m1
TAB
Z.ro
uxii
ESA5
1
5B3
M1
M70
M12
5
7ER
ESA1
7
ESA1
1
1Lisa
2Lisa
1694
-P01
1
1694
-P01
5
m2
Trichosporon mucoides4%
Candida parapsilosis13%
Candida magnoliae25%
Size of the PCR-amplified rDNA region of some yeast strains
5 (honey) 0.17 0.18 0.18 0.09 0.19
6 (honey) 0.17 0.17 0.17 0.10 0.19
7 (commercial) 0.17 0.18 0.17 0.10 0.19
μ -Specific growth rates of the strains in absence and presence of SO2 (100 and 250 mg/L), ethanol (10%) and sugars (20% glucose + 20% fructose
CHARACTERIZATION OF HONEYS FOR MEAD PRODUCTIONGROWTH AND FERMENTATION KINETICS
1000
750
500
250
500
400
300
200
100
1000
750
500
250
500
400
300
200
100
1000
750
500
250
500
400
300
200
100
4
5
6
80
100
120
Ln (g/L)
Dark honey enriched with supplement 1
4
5
6
80
100
120
Ln (g/L)
Dark honey enriched with supplement 2
Pollen
Erica (61.91%)Castanea (14.28%)
Lavandula (14.28%)Rubus (9.53%)
Lavandula (52.0%) Trifolium (24.0%)Rubus (16.0%)Others (8.0%)
Moisture (%) 16.80 16.20
pH 4.90 3.84
Acidity (meq.Ac/Kg) 30.00 23.00
Diastase Index 14.60 8.65
HMF (mg/Kg) 3.59 16.02
Electrical Conductance (mS/cm) 0.77 0.32
Total Ashes (%) 0.55 0.17
Reducing Sugars (%) 71 43 68 03
0
1
2
3
0
20
40
60
0 24 48 72 96 120 144 168 192 216
(OD x 10)
[ ] (g
Time (hours)
0
1
2
3
0
20
40
60
0 24 48 72 96 120 144 168 192 216
(OD x 10)
[ ] (g
Time (hours)
2
3
4
5
6
40
60
80
100
120
Ln (OD x 10)
[ ] (g
/L)
Light honey enriched with supplement 1
2
3
4
5
6
40
60
80
100
120
Ln (OD x 10)
[ ] (g
/L)
Light honey enriched with supplement 2
GlucoseFructoseEthanolGlicerolAcetic acidBiomass
This work was supported by FCT project PTDC/AGR-ALI/68284/2006
Reducing Sugars (%) 71.43 68.03
CONCLUSIONSThe results obtained with the three of S. cerevisiae indicate that the success of mead production greatly depends on the fermentation medium composition, namely on the type ofhoney, as well as on the formulation of the supplements used.
0
1
2
0
20
40
0 24 48 72 96 120 144 168 192 216
)
Time (hours)
0
1
2
0
20
40
0 24 48 72 96 120 144 168 192 216
)
Time (hours)