Biotechnology

Question 1

Describe the 7 steps of beer brewing

Answer 1

Malting & milling, Mashing, Lautering, Boiling (w. hops), Fermentation, Maturation, Filtering.

Question 2

Alpha and beta-amylase breakdown of starch

Answer 2

• Beta-amylase cleaves every second a-1,4 bond in starch chains, starting from its non-reducing end. Creating Maltose.
• Alpha-amylase cleaves mentioned bonds in random order. Creating a variety of sugars.

Question 3

What kind of proteins are present in BSG?

Answer 3

• The most soluble proteins will be in the wort, and the less soluble will be in the BSG, furthering the need for emulsification.

Question 4

What is a glycosidic bond?

Answer 4

A glycosidic bond or glycosidic linkage is a type of covalent bond that joins a carbohydrate (sugar) molecule to another group, which may or may not be another carbohydrate.

Question 5

• Yeast fermentation breaks down what, into what?

Answer 5

Maltose uptake into yeast – breakdown into glucose.

1xGlucose –> 2xEthanol and 2xCO2

Question 6

• What is a maltose molecule?

Answer 6

Two glucose molecules linked by a glycosidic bond.

Question 7

• What is the non-reduced end of a sugar?

Answer 7

• Sugar unit with free OH group is known as reducing end.

- The OH group on the other end forms the glycosidic bond - so it is known as a non-reducing end.

Question 8

• BSG LF composition

Answer 8

Around 2% proteins and 2% carbohydrates, essential AA, antioxidants.

Question 9

• Plant-based drink stability is governed by what three main factors?

Answer 9

1) Size of particles
2) Formation of an emulsion
3) Solubility of proteins.

Reducing the size of the dispersed particles can improve the stability of these drinks, however traditional colloid milling introduces the need for pasteurization introducing a heat treatment that can change the properties of starch, proteins and lipids changing viscosity.

Question 10

• Hydrocolloids (such as pectin) increase stability through what mechanism and how?

Answer 10

Steric stabilization by emulsifying, thickening or gelling capabilities increasing kinetic stability.

Question 11

• 1-4 linked α-D-galacturonic acid units (HG) make up what regions of Pectin?

Answer 11

An oxidized version of D-galactose. Has an added ester.

Question 12

• What is an ester? And their polarity?

Answer 12

COOR’

Esters are polar.

Question 13

• Rhamnogalacturonan (RG-I and RG-II) make up what regions of pectin, are built of and have what effect?

Answer 13

1) “Hairy” regions steric stabilization.
2) Draws in water and creates the gelling effect.

HGA backbone interrupted by side chains of neutral sugars, e.g., arabinose, rhamnose, and galactose.

Question 14

• Extraction effects on Pectin

Answer 14

pH and length of extraction. Low pH = higher methoxyl content.

Question 15

• Moving proteins away from pI creates what effect?

Answer 15

Charges the protein so they repel each other and they don’t aggregate.

Question 16

• Ostwald ripening is?

Answer 16

Dispersion of smaller particles into even smaller molecules and the creation of bigger ones.

Question 17

• Esterification is?

Answer 17

The conversion of a Carboxylic acid (like the one on D-galacturonic acids on Pectin) into an ester.
-COOH –> -COOR
In pectin the Carboxyl group can be methylated into a methyl ester group -COOH –> COOCH3 - probably still polar.

Question 18

• Esterification has what effect on Pectin?

Answer 18

1) The degree of esterification (methyl ester groups) and therefore the charge on a pectin molecule significantly affects their commercial use as gelling and thickening agents.
2) HM pectins form gels at low pH (4.0) or in the presence of a low amount of soluble solids, usually sucrose (55%). HM pectin gels are stabilised by hydrophobic interactions.
3) Conversely, LM pectins form electrostatically stabilized gel networks with divalent metal cations, usually calcium.

Question 19

• An acetyl group is? And what is the polarity?

Answer 19

-COCH3

Less polar than the hydroxyl group it adds on to.

Question 20

• An hydroxyl group is? And polarity?

Answer 20

-OH

Polar group.

Question 21

• Stabilization properties of Pectin

Answer 21

Steric stabilization through the long RG-I side chains causing steric repulsion of oil droplets.

Question 22

• Emulsification properties of Pectin

Answer 22

1) Attributed to the protein moiety, methyl, and acetyl groups.
2) Methyl and acetyl groups are hydrophobic while carboxyl groups are hydrophilic and the ester bond between methyl, acetyl and carboxyl groups thus helps determine the degree of amphipathic behaviour that an emulsifier exhibits.
3) Protein works as surface molecules

Question 23

• Typical additives to plant-based milks are?

Answer 23

Fortifications: Ca2+, vitamins

Hydrocolloids: Stabilizers (guar gum, xanthan gum, carrageenan)

Rapeseed or sunflower oil for fat content

Question 24

• What does the higher methoxylcontent (esterification) make pectin?

Answer 24

Methoxyl groups are less polar = hydrophobic because there is less charge (no longer a carboxylic acid group that can make a COO-). So more methoxylation = less carboxylic acid = less polar.

More methoxylated = less carboxylic acid and less carboxylate = less binding to Ca2+.

Question 25

• How does Ca2+ affect pectin especially Low methoxylated pectin?

Answer 25

The strong binding of cations (Ca2+) to the carboxylate (-COO-) groups of Gal residues (monocomplexation) favors:

1) Formation of cross-links
2) and/or the dimerization (or multimerization) of adjacent chains via bridges between neighbouring carboxylate groups.

Question 26

• Why is WRP good?

Answer 26

Balance between esterification, molecular weight, protein content and long RG-I side chains.

Question 27

• Search profiles where setup how?

Answer 27

1) Look at narrow/specific usages first - #1, #2, #3
2) Spread out a bit more to see if our little circle would be within anybody else’s. #5, #6, #,7 #8, #9
3) Major players and uncovering the entire blop in case we missed something. #10, #11.

Question 28

• CSPR is?

Answer 28

Calcium sensitive pectin ratio (CSPR)

Question 29

• CS is?

Answer 29

Calcium sensitivity (CS)

Question 30

• What is the mPa in patent 2?

Answer 30

The unit of viscocity

Question 31

• NCSP is?

Answer 31

Non-calcium sensitive pectin (NCSP)

Question 32

• Rheology is?

Answer 32

A fancy way of saying fluid consistency (often viscosity and elasticity)

Question 33

• It is necessary to do multiple LC-MS because?

Answer 33

We need it for all Agrains types of BSG/cereal types.

Question 34

• Why is it thought that the proteins of WRP work as emulsifiers on droplet surfaces?

Answer 34

Increasing the oil content of the WRP-stabilized emulsions, led to a further decrease in oil droplet size and to a very compacted microstructure formed by a great number of highly packed oil droplets.

Suggestion proteins work as emulsifiers on the droplet surface.

Question 35

• A viscometer is and measures what?

Answer 35

• Viscometers measure the fluid flow and viscosity of liquids. A often used on is a rotational viscometer that measures the torque to overcome the viscous sheer stress at the surface of the spindle.
• Viscosity = sheer stress/shear rate.
• You express the result of this formula in centipoise (cP). Centipoise is the equivalent of 1 mPa s (millipascal second).

Question 36

What is LC-MS?

Answer 36

Coupling of liquid chromatography (LC) with mass spectrometry (MS)

Question 37

LC is?

Answer 37

• Liquid chromatography (LC)

1) The stationary phase (in the column) is chosen to attract a molecule of your choosing.
2) A mobile sample-solution is flushed trough the column with the stationary phase and sample sticks to the surface coated silica in the stationary phase.
3) Some analytes will interact more strongly with the stationary phase than others, resulting in their separation as they pass through the column.
4) The analytes that have the least interaction with the stationary phase emerge from the column first. Those with the strongest interactions emerging last.

Question 38

Examples of LC?

Answer 38

• Partition chromatography – based on the differing solubility and hydrophobicity of the analytes in the stationary phase as compared to the mobile phase.
• Ion-exchange chromatography – separates the analytes on the basis of their ionic charges.
• Size-exclusion chromatography – exploits the differences in the sizes of the analyte molecules to separate them.
• Affinity chromatography – separates the analytes based on their ability to bond with the stationary phase.

Question 39

MS and LC-MS is?

Answer 39

• Mass spectrometry (MS)

1) The separated fractions from the LC an evaporated and ionized.
2) The analyte ions are drawn into the mass spectrometer where they are subjected to electric fields and/or magnetic fields.
3) The flight paths of the ions are altered by varying the applied fields which ensures their separation from one another on the basis of their mass-to-charge (m/z) values.
4) Post-separation, the ions can be collected and detected by a variety of mass detectors.

Question 40

What have we done wring in our patent three?

Answer 40

1) In the “pectin for stabilizing proteins” patent: They didn’t mention the PCT in the national phases in the European Patent Register, so maybe the applications should have originated directly from the priority application.
1b) Could indicate that they were pursued individually even if highly unlikely.
2) We have used publication of applications in our patent trees.

Question 41

Zeta potential should be above what to create a stable solution?

Answer 41

Over 30 mV

Question 42

Draw a glucose molecule

Answer 42

Good job!

Question 43

How does the carboxylic acid groups have to be spaced to work as an emulsifier?

Answer 43

For a pectin to be useful as a stabiliser of an acidified milk drink at least some of its free
carboxyl groups have to be arranged in blocks (i.e. contiguously) rather than being randomly
distributed discretely along the polymer chain.

Question 44

What does PMEs do?

Answer 44

These esterases, usually called pectin methyl esterases (PMEs), demethylate esterified carboxylic groups. The demethylation proceeds in this way forming blocks of carboxyl groups.

Question 45

Calcium sensitive pectin have their blocks distributed how?

Answer 45

CSP is a high ester pectin having a
degree of esterification of at least 65% whose carboxyl groups are arranged predominantly in
blocks.

Question 46

NCSP have their blocks distributed how?

Answer 46

NCSP is also a high ester pectin having a degree of esterification of at least 70% whose free carboxyl groups are randomly arranged along the polymer chain with no significant degree of contiguity.

Question 47

cP is a unit for what?

Answer 47

Centipoise (poise is the unit)

Unit for viscosity