Lecture 5-processing and physiochemical properties

Question 1

what is the purpose of extraction and isolation of food protiens, like what is it used for?

Answer 1

• to prepare comerical products like lactase
• to make nutritional protiens like soy protien isolate
• for certain biopharmaceuticals like insulin
• research/anylatic purposes like protien

Question 2

What is done to a sample in order to prepare it

Answer 2

1. drying
2. reducition of particle size
3. defatting
4. lipid oxidation protection (if working with fat in sample)
5. enzymatic inactivation
6. microbrial growth and contamination

Question 3

what are the three methods of drying to remove moisture content?

Answer 3

1. heating
2. drying reagents
3. freeze drying: this is used in products that have thermosenstiative compounds. First the product is froze, then ice is removed by sublimnation under low pressure. This plays off the idea that as vacum pressure increases boiling point will decrease

Question 4

What is the purpose of reducing the particle size of sample and what are some important notes

Answer 4

This is done to achieve homgenization. The sample should be dry before as a wet sample can result in lots of moisture loss and chemical changes. It is important that when grinding the sample should not be heated so make sure not overload and to cool with liquid nitrogen and then store at low temperatures

Question 5

Explain the importance of defatting and how it is done

Answer 5

If a protien is surrounded by fat which is hydrophobic, the extraction of a protien by any buffer is prevented. So therefore it is important to defat to allow for a homogenoous sample that can be easily accessed. This is done by a fat soluble solvent (non-polar) like hexane

Question 6

If you want to work with fat in sample it is important to protect from lipid oxidation, how is rhat done?

Answer 6

1. store sample under nitrogen or vaccum (no oxygen avaiable)
2. add antioxdiants if it does not interfere with anaylsis
3. control storage conditions→ dark as to avoid light initiated photooxidation
4. store in low temperatures

Question 7

Since enzymes can degrade food components it is important to inactive/eliminate them, how is this done?

Answer 7

1. heat treatmeant to inactivate enzymes in nonheat senstive foods
2. freezer storage
3. change pH
4. salting out
5. addition of reducing agents for oxidative enzymes

Question 8

how can you decrease/prevent microbrial growth/contamination

Answer 8

1. freezing
2. driving
3. chemical perservatives (benzoates, nitrites, sulphites)
4. a combo of methods above

Question 9

why is protien extraction done and what are the products?

Answer 9

this is done to get a higher protien content than the raw material \

products:

1. flour= less than 30% protien
2. concentraate=30-85%
3. isolate= more than 85%

Question 10

distinguish between wet and dry processing methods of protien extractions

Answer 10

wet processing methods are done to get a higher purity, as dry methods cannot give you an isolate. Dry and wet are used to obtain concentrates. Wet methods extract protiens based on 2 properties→ solubility and size and then have coressponding techinques

Question 11

what are the drawbacks of using wet extraction methods?’

Answer 11

• use of chemicals, water and energy have a high cost and negative impact on enviroment
• loss of functional protien properties because of harsh conditixtions

Question 12

what is the flowchart of wet extraction?

Answer 12

1. raw material goes through milling and defating to result in flour
2. the flour is defatted and then the first seperation step happens which is done by alkaline/acid/salt/alchohol extraction
3. this results in a soluble protien the extract and the residue which is insoluble materials and is discarded
4. the extract goes through precipitation/ultrafiltration to result in percipitate wich is non soluble and then the supernatant which is soluble
5. the supernatant goes through ultrafitltration, diafiltration, spray/freeze dry and results in protien isolate 2
   
   1. the percipitate is wash, neutralized and spray/freeze dried to result in protien isolate/concentrate 1

Question 13

what are the steps of alkaline extraction?

Answer 13

whole pulse→milling→ alkali extraction (protien is very soluble in alkaline conditions)→centrifugation (seperates soluble protiens from insoluble)→isolectric precipitation (pH is adjusted to isolectric point, there is no repulsion because no charge which will cause soluble protien to percipitate)→centrifugation (seperates percipated protiens from other soluble compounds)→drying→milling

Question 14

what is the process of salting in/out (micellization)?

Answer 14

1. Salting In: At low salt concentrations, proteins become more soluble in water. This happens because the salt helps reduce electrostatic interactions between protein molecules, which would normally cause them to stick together. By breaking up these interactions, the proteins remain dissolved in the solution. This is called the “salting-in” effect.
2. Salting Out:As the salt concentration increases beyond a certain point, the opposite happens: proteins start to precipitate (come out of solution). This is because the salt binds to water molecules, leaving less water available to hydrate the proteins. Without enough water, the proteins clump together and precipitate out of the solution. This process is called “salting out.”

• Salting in occurs at low salt concentrations and increases protein solubility.
• Salting out occurs at high salt concentrations and causes proteins to precipitate out.

Question 15

what is a common salt used in micellization and why?

Answer 15

ammonium sulfate (NH4)2SO4 because it has a high water solubilitgy

Question 16

what is the problem with the salting in and out techinque?

Answer 16

large concentrations of salt that are needed contaminate the solution and need to be removed before the protien is resolubilized

Question 17

what is the isolectric point of a protien?

Answer 17

the pH where the net charge on the protien is zero

Question 18

what is the principle of isolectric percipitation?

Answer 18

protiens tend to percipitate/aggregate at their pI since there is no electrostatic repulsion from charges keeping them apart. The pI differs from protien to protien, so when one pI is reached that protien will be percipitated and the protiens with different pIs will remain in solution. The percipitated protien can be resolubilized in another solution at a different pH

Question 19

what is the process of acid wash extraction

Answer 19

this is used for soybean protien which is insoluble in water at a pH of 4.2. The acid will change the pH, reaching the isolectric point of soybean protien. This allows for the dissolving of sugars with no special solvents and is a lot safer because of that

Question 20

what are the common steps of preparing soyprotien isolate?

Answer 20

processes is done to seperate fibers of carbohydrates from soy protien concentrate. 1. use alkaline water at pH 8-9 and then 2. isolectric focussing at pH 4.2-4.5 by hydrochloric/phosphoric acid to adjust pH. 3. Then remove insoluble protiens from soluble carbs by centrifugation

Question 21

what is the idea behind solevent fractionation and how is it done?

Answer 21

The idea here is that the solubility of a protien depends on the dielectric constant of the solution surrounded the protien. If there are charges your protien will be stable and soluble→ but remove those charges and the protien will percipitate as it is less soluble. in other words as the dielectric constant of a solution decreases there is an increase in interactions between protien molecules as there is decreased repulsion→ resulting in percipitation.

This is done by adding water soluble ogranic solvents like acetone or ethanol as they decrease the dielectric constant of aqeuous solutions

Question 22

why is solvent fractionation done at 0 degrees or below??

Answer 22

because organic solvent+ water= heat so need to ensure the solution is cool to prevent denaturation by heat

Question 23

what is the amount of organic solvent required in solvent fractionation dependent on?

Answer 23

depends on the protien, normally varies from 5-60%

Question 24

what is aqueos alcohol extraction used for and what is the process?

Answer 24

it is used for soy protien isolate. defatted soy flakes are used as the raw material→ aqueos alchohol is added and sugars are disolved and protien percipitates→alchohol is recovered by evaporation→ protien flakes are dried with hot air and milled

Question 25

explain how heat is used for soy protien extraction

Answer 25

if protien is heat stable like in the case of soy protien, seperation from protiens and other things can be done by heat. By heating the protien the protien is unfolded and coagulated which causes percipitation. Sugars are removed by using hot water

Question 26

what is dialysis

Answer 26

it is a process used to seperate molecules in solution by the use of semipermeable membranes that allow for smaller molecules to pass but prevent larger molecules from passing

Question 27

what is the process of diaylisis?

Answer 27

protien solution is placed in a diaylsis tubing which sealed and placed in a large volume of water or buffer and is slowly stired. This process can take up to 12 hours to be completed. The solutes with low molecular weights will flow through the bag and the large ones will stay in the bag (the protiens).This is often used after salting out to remove salt

Question 28

what is ultrafiltration

Answer 28

a dyalsis process with the addition of pressure, making this a much faster techinque. It is important to note that the semipermeable membranes have a cutoff point, anything larger than that number cannot get out

Question 29

what is the result of ultrafiltration

Answer 29

Retenate and ultrafiltrate retenate is the portion retained by the cell→ the large molecules ultrafiltrate is the the parts that pass through the membrane→ small molecules

Question 30

what is diafiltration?

Answer 30

a speciliazed ultrafiltration process where the retenate is diluted with water and ultrafiltered again to enhance purity and yield

Question 31

explain what gel permeation is

Answer 31

Molecules pass through a hydrophilic polymer and will be seperated by size. The polymer beads are made of cross linked dextran which is very porous so small protiens will enter these beads as they get stuck in the pores and will get out slower, but larger protiens will come out first as they cant fit in the pores.

Question 32

explain what polyacrylamide gel electrophersis is

Answer 32

the electrophersis seperates protiens based on size using an electrical current, the small ones move faster in porous gel than the large ones

Question 33

what is the process of dry extraction (air classification)?

Answer 33

1. material is milled to get a flour (milling mechanically detaches protien from other cellular compounds)→ samples that contain oil must be defatted 2. air stream is applied to the flour

Question 34

what 2 fractions are obtained from dry extraction

Answer 34

1. light or fine fractions (protiens) 2. heavy/coarse fraction (starch)

Question 35

what can be done to get higher purity of protien concentrates?

Answer 35

milling and air classification can be reapeated

Question 36

what are the advantages of dry extraction?

Answer 36

- no solvent used (prep doesnt count) - reduced water consumption and energy - protiens have better functionality since they keep their native properties since there is no heating because no drying step and no exposure to chemicals

Question 37

what are the disadvantages of dry extraction?

Answer 37

- less efficient in seperating protiens compared to wet methods - higher amounts of ANF, oils, fibers - if starch is present and it has similar properties to protien this method may not be abke to serparate strach - moisture and lipid content in flour can affect fractitionation efficiency