The Effects of Vitamins B6 and C on Yeast Growth By: Gianmarco Frezza Pittsburgh Central Catholic HS Grade 9
Jan 03, 2016
The Effects of Vitamins B6 and C on Yeast Growth
By: Gianmarco FrezzaPittsburgh Central Catholic HS
Grade 9
Rationale
• Humans need vitamins B6 & C from their diets or as supplements
• Vitamin B6 RDA: 1.3 mg/day
• Vitamin C RDA males: 90 mg/day, females: 75 mg/day
• Does vitamin B6 or C affect the microbial flora of the body?
Microbial Flora
• Refers to microorganisms living in or on the body (gastrointestinal tract)
• Little is known about the association between humans and their flora
• Effects are mutualistic, parasitic, pathogenic, and commensal
• Perform functions beneficial to the host, including the manufacture of essential vitamins, and the prevention of colonization by undesirable microbes
• Human foods and supplements may have unintended effects on the flora populations and their functions
Saccharomyces cerevisiae
• A fungi• One of the most
intensively studied eukaryotic model organisms
• Commonly known as yeast
• Can be used as a model for human microflora
Vitamin B6 (Pyridoxine)
• Important for more than 100 enzymatic reactions involved in metabolism and brain development during pregnancy and infancy
• Helps the immune system function properly
Vitamin C (ascorbic acid)
• An antioxidant, protects cells from damage from free radicals
• Helps the immune system function properly• Growth and repair of tissues• Protects against cardiovascular disease• Prevents scurvy
Purpose
• To test if Vitamin B6 or C have an effect on S. cerevisiae growth individually or synergistically
Hypothesis
• Null: the vitamins WILL NOT have an significant effect on yeast growth individually or synergistically.
• Alternate: the vitamins WILL have a significant effect on yeast growth both individually and synergistically.
Materials• 36 sidearm flasks• Aluminum foil• Lab coat• Disposable rubber gloves• Micropipetters• Sterile micropipette tips• Sterile dilution fluid
(100mMKH2PO4, 100mMK2HPO4, 10mMMgSO4, 1mMNaCl)
• Macropippetter• Sterile macropipette tips• 3 Incubators
•1% stock solution vitamin B6•1% stock solution vitamin C•YEP media (1% yeast extract, 2% peptone, 2% glucose)•S. Cerevisiae•Klett Spectrophotometer
Procedure
1. 7.5 mL of YEP media was pipetted into each sterile sidearm flask.
2. Four replicates of the following combinations of variables were prepared as follows.
3. 0.5 mL of S. cerevisiae (KU=100) was pipetted into each flask.4. The flasks were covered with a sterilized aluminum foil cap.5. The 36 sidearm flasks were placed in incubators (32⁰C), and
readings were taken every half hour for six hours in a Klett spectrophotometer.
Mixtures by GroupsS.Cerevisiae(mL)
YEP media(mL)
Concentrations (B6, C) %
Vitamin C (mL)
Vitamin B6 (mL)
SDF (mL)
Group 1 0.5 7.5 [0%] B6 [0%] C 0 0 2
Group 2 0.5 7.5 [0%] B6 [0.01%] C
0.1 0 1.9
Group 3 0.5 7.5 [0%] B6 [0.01%] C
1 0 1
Group 4 0.5 7.5 [0.01%] B6 [0%] C
0 0.1 1.9
Group 5 0.5 7.5 [0.01%] B6 [0.01%] C
0.1 0.1 1.8
Group 6 0.5 7.5 [0.01] B6 [0.1] C 1 0.1 0.9
Group 7 0.5 7.5 [0.1%] B6 [0%] C 0 1 1
Group 8 0.5 7.5 [0.1] B6 [0.01] C 0.1 1 0.9
Group 9 0.5 7.5 [0.1%] B6 [0.1] C 1 1 0
Concentrations of the Vitamins within Groups
[0%] (0) B6 [0.01%] (L) B6
[0.1%] (H) B6
[0%] (0) C 00 L0 H0
[0.01%] (L) C 0L LL HL
[0.1%] (H) C 0H LH HH
Vitamin Effects on Yeast Growth
60 120 180 240 300 3600
20
40
60
80
100
120
140
Group 1 CGroup 2 0L Group 3 0HGroup 4 L0Group 5 LLGroup 6 LHGroup 7 H0Group 8 HLGroup 9 HH
Group 1 CGroup 2 0L Group 3 0HGroup 4 L0Group 5 LLGroup 6 LHGroup 7 H0Group 8 HLGroup 9 HH
Time (min.)
KU
L= [0.01%] H= [0.1%]B, C
Groups at 180 minutes
0% C 0.01% C 0.1% C0
20
40
60
80
100
120
0% B0.01% B0.1% B
Concentrations of vitamin C
Klett Units
Two factor ANOVA synergy P= 0.1162
Single Factor vitamin B P= 0.958538
Single factor vitamin C P= 0.972004
Groups at 360 minutes
0% C 0.01% C 0.1% C0
20
40
60
80
100
120
140
0% B0.01% B0.1% B
Concentrations of Vitamin C
Klett units
Two factor ANOVA synergy P= 0.1785
Single Factor vitamin B6 P= 0.000429
Single factor vitamin C P= 0.16963
Conclusion• The null hypothesis is accepted for all trials other than vitamin B6 at 360 minutes exposure.
• Vitamin B6 and C did not have a significant effect on yeast growth individually or synergistically.
Limitations and Extensions
Limitations• Only two
concentrations of the variables used
• Only growth was tested
• Six hour duration
Extensions•More concentrations
•Longer Duration
•Study survivorship
•More species
Resources• http://en.wikipedia.org/wiki/VitaminC
• www.ods.od.nih.gov/factsheets/VitaminC-HealthProfessional/
• http://en.wikipedia.org/wiki/VitaminB6
• www.ods.od.nih.gov/factsheets/VitaminB6-HealthProfessional/
• http://en.wikipedia.org/wiki/Saccharomyces_cerevisiae
• Mark Krotec, PTEI
• www.epa.gov/epawaste/nonhoz/municipal/
Raw Data at 180 minutes
Groups (Replicant 1) Klett units
Group 1 71
Group 2 73
Group 3 87
Group 4 73
Group 5 59
Group 6 41
Group 7 62
Group 8 79
Group 9 101
Groups (Replicant 2) Klett units
Group 1 70
Group 2 84
Group 3 80
Group 4 72
Group 5 78
Group 6 35
Group 7 57
Group 8 94
Group 9 100
Raw Data at 360 minutesGroups (Replicant 1)
Klett units
Group 1 128
Group 2 107
Group 3 127
Group 4 92
Group 5 105
Group 6 24
Group 7 118
Group 8 105
Group 9 113
Groups (Replicant 2) Klett units
Group 1 125
Group 2 108
Group 3 120
Group 4 94
Group 5 98
Group 6 25
Group 7 119
Group 8 111
Group 9 117
ANOVA at 180 minutes
ANOVA at 360 minutes