1
Q
What is fermentation?
A
any metabolic process that releases energy from a sugar or other organic moleucle that does NOT require oxygen or the electron transport chain
2
Q
What is the crabtree effect?
A
In relation to fermentation the crabtree effect is when there are so many reactants present a reaction can proceed even under adverse conditions, such as fermentation progressing in the presence of oxygen
3
Q
Saccharomyces cerevisiae
A
Top fermenting for ales
- ferments fast
- produces lots of CO2 (what brings it to the top)
4
Q
Saccharomyces carlsbergenesis
A
Bottom fermenting for lagers
- ferments slower
5
Q
Why are grapes not washed before wine making?
A
Because there are natural yeasts on the skins of the grapes and it does not need to be introduced, unlike with barley/ cereals in beer that must be malted before fermentation
6
Q
Describe carbonic maceration
A
un-mashed grapes are put in a vessel with no air present, the vessel is then filled with CO2 and fermentation begins (intracellular). This produces a sweeter wine with fewer tannins.
6
Q
Wild yeasts
A
for lambics/sours
6
Q
Why cant yeasts utilize lactose in fermentation?
A
They do not contain the correct enzymes to break down lactose into lactic acid like lactic acid fermenting bacteira such as Lactobasillicus does
7
Q
What is the main difference between American and Italian/ european fermentation?
A
With American fermentantion bacteria are added into meat and the pH is lowered to prevent spoilage. In Italian/ european fermentation no bacteria is added and the meat is dried to prevent spoilage
7
Q
What three scientists worked on penicillin during WWII?
A
Florey, Heatley, and Chain
- Heatley has gone largely unrecognized
7
Q
Who discovered the antibiotic properties of penicillin?
A
Alexander Fleming observed that the growth of staphyloccocus bacteria was inhibited by the growth of penicillin
7
Q
Four improvements that were made to increase the production of penicillin
A
- Using a corn steep liquor lactose as a medium
- Identified a new strain
- Deep tank fermentation
- ‘Pod’ contactor for liquid/liquid extraction
7
Q
Summarize Butyric acid fermentation
A
- Fermenting genus: Clostridium
- other products: acetic acid, ethanol, isopropanol, and acetone
7
Q
Summarize shikimic acid and D-allulose fermentation
A
Step one: enzymatic hydrolysis with whey/ lactose rich solution
Step two: fermentation with glucose and galactose
Step three: production of shikimic acid and D-allulose
8
Q
D- allulose
A
a sweetner
8
Q
Shikimic acid
A
can produce Tamiflu, an important influenza drug
8
Q
Macrolides
A
hydrophobic, basic antibiotic compounds that slow the growth of bacteria by reducing necessary protein production
8
Q
Summarize the structure of erythromycin
A
- A highly substitued 14-membered lactone ring with 10 asymmetric centers
- a ketone group
- an amino deoxysugar
9
Q
What type of bacteria is used to produce erythromycin?
A
Actinobacteria, a type of gram positive bacteria
- Streptomcyces erythreus
10
Q
Why are macrolide antibiotics best produced by fermentation and not organic synthesis?
A
They have very complex structures that can require dozens of organic syntehsis steps, that process can be shortened by using fermentation
ex: Taxol (40 step synthesis)
11
Q
In the production of ascorbic acid/ vitamin c what enzyme produces 2-keto-gulonic acid (KLG)?
A
Sorbose dehydrogenase
12
Q
Why is 2-KLG important?
A
2-KLG is a direct precursor to ascorbic acid and increases the yield significantly
12
Q
What microorganism produces 2-KLG?
A
K. vulgare
13
Q
Compare the two major ways cells produce energy and their differences in ATP production
A
- Respiration is done in the presence of O2 and produced 30-32 net ATP
- Fermentation is done in anaerobic environments and produces 2 net ATP
14
Summarize the pyruvate to lactate reaction
pyruvate to lactate is catalyzed by the enzyme lactate dehydrogenase
| Embden Meterhof Parnas pathway
15
Indigenous microflora
spontaneous fermentation from microorganisms that live on the skin, air, or surface of the fermenting environment
16
Exogenous microflora
controlled fermentation, stater bacteria that are added in to intitate fermentation
- defined and undefined starters
17
Defined starters
| exogenous
composed of known amounts of identified strains
18
Undefined starters
| exogenous
unknown compositions and amounts of strains
19
Non starter bacteria
do not ferment, added in for flavors and textures
20
Embden Meyerhof Parnas Pathway
pyruvate to lactate via lactate dehydrogenase pathway
- 2 net ATP
21
Homolactic fermentation
- produces 2 mol lactic acid
- yogurt, buttermilk, sour cream, cottage cheese
| homo, one kind of product
22
Heterofermentative fermentation
- 1 mol lactic acid, 1 mol ethanol or acetic acid
- Leuconostoc species
| hetero, two different products
23
Enterdoudoroff pathway
- pyruvic acid and glyceraldehyde three phosphate production
- uses few bacteria
- makes tequila, and mezcal
24
Summarize the pyruvate to ethanol pathway
1. pyruate to acetaldehyde is catalyzed by pyruvate decarboxylase
2. acteldehyde to ethanol is catalyzed by alcohol dehydrogenase
25
Catabolite repression
| crabtree effect
glucose represses the synthesis of respiratory enzymes
26
Catabolite inactivation
| crabtree effect
glucose inactivates respiratory enzymes
27
Explain exploding beers
exploding beers are due to diastatic strains that produce a glucoamylase that breaks down complex sugars which produces lots of ethanol but also produces lots of O2 which leads to too much carbonation and hence exploding beers
28
Malting
controlled germination of grains
29
Wort
liquid product of mashing
30
Lautering
filtering the solid and liquid (wort) from mashing
31
Grist
Malted and then milled barley
- acts as a filter during lautering
32
Adjunct
components that are added in for different, flavors, aromas, etc.
33
Trub
coagulated proteins that are removed after wort boil (solids)
34
What are the three main reasons grains must be malted for beer brewing?
1. To ensure all enzymes in the grain are available to break down sugars
2. If it's not malted it produces a very viscous extract
3. There is a lack of color and flavor if it is not malted
35
Nine steps to beer brewing
1. Malting
2. Milling
3. Mashing
4. Lautering/ wort filtration
5. Wort boil
6. Hops separation
7. Wort cooling
8. Fermentation
9. Maturation
36
Three steps to malting
1. Steeping
2. Germination
3. Drying
37
Explain the steeping step of malting
Kernels are covered with water to increase moisture and produce enzymes that breakdown carbohydrates and proteins in the germination step of malting
38
Explain the germination step of malting
carbohydrates begin to be broken down by enzymes as well as protiens and starch (endosperm walls)
39
Alpha- amylases
- heat stable, survives drying
- cleaves at 1-4 linkages of amylose and amylopectin
40
Beta-amylase
- not heat stable, does not survive drying
- cuts amylose and amylopectin from non reducing ends
41
limit dextrinases
- heat unstable
- cut amylose and amylopectin at 1-6 linkages
42
Explain the drying step of malting
- the longer you dry the darker the color of your beer (but the longer you dry the more likely to kill fermenting enzymes)
- Maillard reaction
43
Explain the three steps of Maillard reaction
1. Carbonyl group reacts with a protien to produce a glycosylamine
2. The glycosylamine isomerizes to a ketosamine
3. Ketosamine can react to provide different products for flavor and aroma
44
Milling
grinding down malted grains, but not powderized
45
Mashing
heating malted milled grains to produce liquid wort and solids (grist)
46
Four mashing methods
1. Double mashing
2. Temperature programmed
3. Infusion mashing
4. Traditional decoction
47
Double mashing
- used for American beers
- two separate kettles
48
Temperature programmed mashing
temperature of mash is changed with heating coils
- can be very precise
49
Infusion mashing
- british, german and belgian ales
- no stirring, grist floats and wort is extracted from the bottom
50
Traditional decoction
- bavarian and czech beers
- part of mixture is removed, boiled and returned to mixture
- decoction can occur more than once
51
wort will contain 15% fermentable sugars ---, ---, and ---
1. glucose
2. fructose
3. sucrose
51
wort will contain 75-78% fermentable sugars --- and ---
1. maltose
2. maltotriose
52
Wort will contain 20% non fermentable sugars --- and ---
1. dextrins
2. amylopectins
53
What two approaches are used for worth filtering/ separation?
1. Mash filter
2. Lauter tun
54
Wort boiling
unique to beer brewing
- sterilizes the wort
- formation of the trub
- extraction of iso-alpha acids
55
Why does beer get skunky?
When exposed to light the beer forms isopentenyl mercaptan which is acutally a chemical in skunk spray
56
Maturation
1. carbonation
2. clarification
3. stabilization
57
Carbonation step in maturation
trap CO2 with green beer to force bubbles into the liquid
58
Clarification step in maturation
centrifuging, filtering to get rid of cloudy haze caused by tannins/ proteins
59
Stabilization step in maturation
pasteurization to remove bacteria and removal of reductants
60
How do you calculate european brewing convention from SRM?
SRM * 1.97 = EBC
61
What is specific gravity?
a liquids density compared to pure water
| 1.080 pronounced ten eighty
62
Original gravity
the specific gravity before fermentation
63
Final gravity
the specific gravity after fermentation
64
Degree brix and degree plato measure what?
sugar content in an aqueous solution
65
--- is used more in the wine field and --- is used more in the beer field
1. degree brix
2. degree plato
66
What are two ways we can measure specific gravity?
Refractometer and hydrometer
67
How do we calculate ABV% from specific gravities?
ABV= (OG - FG)* 132.715
68
Five ways to determine ABV in beer
1. Specific gravity measurements
2. Gas chromatography
3. HPLC
4. IR/ Raman
5. Hydrometers
69
Plate theory
within a chromatography column there are theoretical plates where there is an equilibrium established between the m.p. and s.p., we want to maximize the number of these plates
- derived from distillation
70
H=
| H: height of theoreticall plate
L/N
- want to minimize this value by having as many plates as possible
71
Formula to calculate resolution of two peaks (simplified formula)
Rs1/Rs2 = (N1^1/2/ N2^1/2)
72
Van Deemter equation
H=
A + B/u + u[Cs+ Cm]
- optimum velocity can obtained for maxiumum column efficency
73
A in the Van Deemter equation
Eddy diffusion, not dependent on velocity, relies on particle size
74
B in the Van Deemter equation
Longitudional diffusion, inversely proportional to velocity, main factor that affects band broadening
75
C in the Van Deemter equation
Proportional relationship with velocity, stationary and mobile phase mass transfer, depends on particle size
76
Why are capillary columns better than packed columns?
They can separate more complex mixtures because they are longer and contain more theoretical plates
77
What is the stationary phase for gas-solid chromatography?
Silicone or PEG with PEG being the more polar of the two
78
# True or false?
HPLC is good for thermally stable compounds?
False, gas-liquid chromatography is best for thermally stable compounds
79
Hydrophobic interaction/ normal phase HPLC
- polar stationary phase, non polar mobile phase
- decreasing polarity increases analyte retention time
80
Reverse phase HPLC
- non polar stationary phase, polar mobile phase
- increasing the polarity of the mobile phase increases analyte retention time
81
There are two main steps in fermentation that are skipped in distilling, what are they?
Lautering/ wort separation and wort boil
82
What are the three major stills used for distilling?
1. Pot still
2. Column still
3. Hybrid still
83
Pot still
- used for whiskey, tequila and rum
- 1 distillation 80-120 proof
- 2nd distillation 140-170 proof
84
Column/Coffey still
- each plate in a column still is like one pot still
- constant refluxing where components are separated based on boiling point
85
Stripping section of a column still
bottom part of the still, where components are removed
- bottom part of a still
86
Rectification section of a column still
top part of a column still where components are concentrated
- coolest part of the still
87
Hybrid still
pot still + column still
88
Azeotrope
when a mixture of two or more susbtances behaves as a pure substance such that the components of the mixture cannot be separated
- found at the top of vapor curve where the two curves come together
| ex: water and ethanol, methanol and water
89
# True or false?
The more distillations you perform the smaller your change in proof?
True! look at the vapor liquid equilibrium curve where the azeotrope starts to form
90
Heads
compounds with a bp lower than ethanol
91
Hearts
the ethanol, what you collect to distill and drink
92
Tails
Compounds with bps higher than ethanol
93
Absinthe
- from switzerland
- thought to be hallucinogenic
- anise and fennel flavors
94
Baijiu
- largest selling alcohol in the world
- from china
95
Gin
- juniper berries are required for it to be a gin by defenition
- botanicals are kept in a gin cage
96
Scotch whisky
- must be aged in an oak cask for three years and one day
- must be from Scotland
97
Bourbon (American whiskey)
- must be produced in US
- At least 51% corn
- new charred oak barrel
- aged for at least 2 years
98
How do the flavors develop in barrel aging?
Primary alcohols react to form esters that give flavors
- the charred oak barrel helps to determine color and flavor
99
Beers law
A= Ebc = - log(T) = log(1/T)
100
On a plot of absorbance versus wavelength how do you choose a section that will yield a linear plot of absorbance versus concentration?
A peak in a absorbance vs wavelength plot will yield a linear absorbance versus concentration plot (where the slope is zero)
101
Three limitations to beers law
1. Need a concentration less than 0.01 M
2. Must ensure a single wavelength, espeically if more than one species can absorb
3. Avoid excess light to falsely increase or decrease absorbance measurements
102
How do you select the color for a colorimetry measurement?
find the color of your sample (ex: beer is a yellowish color) and then find the complementary color on the wheel (violet)
103
Why is IR a bad option for beer analysis?
The matrix is often too complicated and contains water which is IR active and can produce distracting peaks or affect the sample holder
104
Why is NIR a good option for beer analysis?
The NIR range does not experience interference from water like IR, so there is no high water background signal and the distinct methyl group can be detected from ethanol
105
How is the color of beer formed?
by the reaction of a carbohydrate/ sugar and an amine
106
How do we test for IBUs?
By extraction with isooctane BUT other compounds can interefere with absorbance at 275 making the test not very selective
| other compounds: polyphenols, alpha and beta acids
107
What amino acids in proteins can be detected by UV-VIS?
The aromatic amino acids, tryptophan and tyrosine absorb at 280 nm because conjugation increases absorbance
108
What two metals in beer can determined by flame emission?
Sodium and calcium
109
Identify M* and Mo as excited or ground state metals
M* is excited state (Atomic emission)
Mo is ground state (Atomic absorption)
110
For atomic emission we want to --- N*/N
maximize
111
For atomic absorption we want to --- N*/N
minimize
112
Atomic absorption
- element specific source
- can use flame or graphite furnace
- inexpesive
- only one element at a time
113
Considering atomic absorbance does the flame or graphite furnace atomizer have a better LOD?
The graphite furnace has a better/ lower LOD than a flame atomizer
| but both are usually in the ppm range
114
Why is an inductively coupled plasma (ICP) torch better than a flame for atomic emission?
It reaches higher temperatures which decreases interferents and maximizes N*/N
115
Why is the monochromator so important in AAS?
Especially with flame AAS we must restrict all flame light from reaching the detector so absorbance is not falsely decreased
116
Mass spectrometry
- most expensive but also highest quality instrument
- can detect isotopes (other methods cannot do this)
- lowest/ best limits of detection (ppt)
117
Two advantages of a PDA over a colorimeter
1. Selection of a wider range of wavelengths
2. Ensures linearity of beers law
118
Is the solvent used to extract isoalpha acids polar or non polar and why?
Isooctane is nonpolar because the structure of isoalpha acids contain many hydrocarbon (nonpolar) side chains
119
IBU=
50 * abs. at 275 nm
120
What happens to hops as a beer ages over time?
Beer loses its iso-alpha acids to oxidation overtime which is (one of) the reason/s oxygen levels must be kept as low as possible during the beer fermentation process
121
SRM=
12.7* DF* abs. at 430 nm
122
Evaporative light scattering
nebulizing and evaporating the mobile phase so carbohydrates and lipids (non volatile non polar compounds) can be detected by the light they scatter
123
Why cant carbohydrates be detected by UV?
They have no structures to promote absorbance (eg. chromophores)
124
What compounds elute first off of GC?
compounds elute from low to high boiling point
- low boiling point indicates low polarity
125
Area under the curve of a chromatogram represents --------
how much sample was injected
126
Why do hops have to be added at the end of the wort boil to affect the beer?
Because the essential oils have low boiling points and will be boiled off
127
Static headspace sampling method
The hops are heated and the vapor from the heating is used as a sample, if it is not concentrated enough an adsorbent trap can be used to collect the necessary concentration for injection
128
How does calcium affect the beer brewing process?
| one good, one bad
Calcium can protect the yeasts from the harmful effects of the product ethanol. Calcium can also inhibit yeasts from interacting with the necessary element of magnesium
128
How does magnesium affect the beer brewing process?
| one good
Magnesium aids fermentation because it is required for many of the fermenting enzymes to function
129
How does a water softner work and why is it bad for beer brewing?
A water softner removes many of the elements essential to beer brewing such as calcium and magnesium and replaces them with sodium to account for water hardness. Sodium can be harmful to enzymes and the flavor of the beer at too high of concentrations
CHM 436 (fermentation)
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