exp 4 post lab

Question 1

Thin Layer Chromatography

• the higher the affinity of the compound with the mobile phase, the ______ it will move
• the lower the affinity of the compound with the mobile phase, the ____ it will move

Answer 1

• faster and higher
• slower and lower

Question 2

TLC

• 3 main steps

Answer 2

1. Preparation of TLC plate
2. Preparation of Plate Spotters and Spotting the Plates
3. Developing a TLC plate

Question 3

TLC - Preparation of TLC plate

• stationary phase used is:
• ^^ preferred because its?
• 2 types of ^^ + definition + when used
• preparation of ^^ involves the use of _____ and _____ to prevent _____

Answer 3

• silica gel (made of silicon dioxide joined together)
• higher sample capacity

1. Hydrophilic silica
   - used when mobile is nonpolar
   - polar surface
2. Hydrophobic silica
   - used when mobile is more polar than stationary phase
   - nonpolar surface
   - chemically modified

• face mask and gloves to prevent silicosis

Question 4

TLC - Preparation of Plate Spotters

4 types of spotters used?

Answer 4

1. Capillary Tubes
2. Stirring Rod (not recommended)
3. Toothpick
4. Inert pointed materials

Question 5

TLC - Spotting the Plates

• the sample is spotted ____ cm to ____ cm from the _____ _____ _____
• done by _____ pressing the _____ or using a ____

Answer 5

• 1 cm to 2 cm
• plate’s bottom edge
• manually
• spotter
• syringe

Question 6

TLC - Developing a TLC Plate

• 3 steps + importance (1; 1; 3)

Answer 6

1. Placing of filter paper inside the developing chamber/beaker and equilibration

• ensures that the container is saturated with solvent vapors evenly (for effective separation)

2. Pouring solvent below the sample spots

• prevents dissolution of sample

3. Addition of 2 drops glacial (pure) acetic acid to the developing chamber containing the mobile phase

• prevents “tailing” (changing directions of spots)
• pampabigat ng solvent
• maintains low pH (silica gel plate is acidic) and prevent reactions that may form new products/compounds = error in separation

Question 7

Pano kung masyadong mababa sa 1 - 2 cm yung pag spot sa TLC silica gel plate?

Answer 7

pwedeng ma dissolve yung pigments by the mobile phase rather than tumaas

Question 8

Sample used in TLC

Answer 8

Malunggay leaves

Question 9

Relevance of POUNDING malunggay leaves using mortar and pestle + what was added

+ why was this added??

Answer 9

ETHYL ACETATE
- extracting solvent
- very volatile

• break cells of the leaves and release the pigments into the ethyl acetate

pigments are soluble in ethyl acetate

Question 10

2 possible mobile phases used in TLC

• what is the basis for choosing the better solvent (3)?
• what was chosen at the end?

Answer 10

• 8:2 petroleum ether: ethyl acetate
• 1:1 petroleum ether: ethyl acetate

v/v

1. separated the highest number of components
2. least degree of overlaps
3. clearest color of the spots

8:2 was the best solvent

Question 11

MOBILE PHASES IN TLC

• usually a combination of ____ and _____ compounds
• if the stationary phase is POLAR, the ______ compound in the mobile phase will have larger proportions
• if the stationary phase is NONPOLAR, the ______ compound in the mobile phase will have larger proportions

in the experiment, which is polar and nonpolar in the mobile phase used?

Answer 11

• polar and nonpolar
• nonpolar
• polar

polar (ethyl acetate)
nonpolar (petroleum ether)

Question 12

TYPE OF CHROMATOGRAPHY USED IN TLC

+ stationary and mobile phase used

Answer 12

NORMAL PHASE

• stationary is polar (hydrophilic silica)
• mobile is nonpolar (8:2 petroleum ether:ethyl acetate)

Question 13

ORDER OF SEPARATION OF PIGMENTS in TLC (2)

in normal & reverse phase?

Answer 13

NORMAL PHASE
- less polar components: carried farther from the origin (at the top)
- more polar components: separate at earlier times (at the bottom)

REVERSE PHASE
- more polar components: carried farther from the origin (at the top)
- less polar components: separate at earlier times (at the bottom)

Question 14

PIGMENTS separated from MALUNGGAY LEAVES (5)

+ order of increasing polarity

+ order of separation (lower to higher spots) - normal

+ order of separation (lower to higher spots) - reverse

Answer 14

1. Chlorophyll a
2. Chlorophyll b
3. Carotenoids
4. Xanthophylls
5. Pheophytins

INCREASING POLARITY:
Carotenoids < Xanthophylls < Pheophytins < Chlorophyll a < Chlorophyll b

NORMAL - ORDER OF SEPARATION L->H:
Chlorophyll b > Chlorophyll a > Pheophytins > Xanthophylls > Carotenoids

REVERSE - ORDER OF SEPARATION L->H:
Carotenoids > Xanthophylls > Pheophytins > Chlorophyll a > Chlorophyll b

Question 15

MALUNGGAY LEAVES - PIGMENT

Chlorophyll

• ____ pigments present in ___
• 2 types + color + polarity

Answer 15

• major
• plants

1. Chlorophyll a
   - green and more intense color
   - less polar
2. Chlorophyll b
   - yellow-green
   - more polar

Question 16

MALUNGGAY LEAVES - PIGMENT

Carotenoids

• polarity
• color

Answer 16

• nonpolar
• yellow-orange

Question 17

MALUNGGAY LEAVES - PIGMENT

Xanthophylls

• _____ (color) pigments produced when _____ are ____

Answer 17

• yellow
• carotenes
• oxidized

Question 18

MALUNGGAY LEAVES - PIGMENT

Pheophytins

• ____ derivatives that lack ____
• polarity
• color

Answer 18

• chlorophyll
• magnesium
• nonpolar than chlorophylls
• gray

Question 19

PC (Paper Chromatography)

6 samples used?

Answer 19

1. Blue
2. Red
3. Green
4. Yellow
5. Violet
6. Orange

Question 20

PC - STATIONARY PHASE

• stationary phase used?
• where is ^^ adsorbed

Answer 20

WATER is stationary phase
- adsorbed on Whatman No.1 Filter Paper

Question 21

PC - WHATMAN NO.1 FILTER PAPER

• a grade ___ quality
• the most _____ ____ paper due to ____ ___ and ____ rate
• composition?
• contains how much water by mass?

Answer 21

• 1
• used filter paper
• medium retention
• flow rate
• specially selected cotton cellulose
• 23% water

Question 22

when water is adsorbed to cellulose (whatman no.1 filter paper), it results in a ___-____ ____ that is bonded with ____ ____

Answer 22

water-cellulose complex
- hydrogen bonding

Question 23

PC - MOBILE PHASE

• what is the mobile phase used in the experiment?
• what other mobile phase can be used? + is this more polar than water like the salt?

Answer 23

• 2% NaCl
• isopropyl alcohol can be used: less polar than water

*lahat ng mobile phase maliban sa SALT ay LESS polar than water

Question 24

which is more polar in PAPER CHROMATOGRAPHY: mobile or stationary?

THUS, what type of chromatography was done in PC?

Answer 24

MOBILE PHASE (2% NaCl) is more polar than STATIONARY (water)

so REVERSE CHROMATOGRAPHY was used

Question 25

NaCl (mobile phase) has permanent ____ charges Water (stationary) has permanent ____ charges this explains that? + what bonds is prese

Answer 25

NaCl - full Water - partial thus, NaCl is more polar than water

Question 26

PC - REVERSE CHROMATOGRAPHY - again, why is paper chromatography reverse chromatography? - order of separation (what is carried farther?)??

Answer 26

- mobile phase is more polar than stationary phase ORDER OF SEPARATION - the more polar components of the food dyes will be carried farther from the origin

Question 27

PC - PIGMENTS SEPARATED - 3 types of components that should emerge from all the food dyes? + color it exhbitis + presence or absence of polar groups + number of charged groups and relationship with mobile phase

Answer 27

1. BRILLIANT BLUE FCF - blue - presence of polar groups - has the most charged (7) groups associates the STRONGEST with NaCl mobile phase 2. TARTRAZINE - yellow - presence of 2 polar groups - second most charged groups (6) : associates the second strongest with NaCl 3. ALLURA RED AC - red - presence of 1 polar group - fewest charged groups (4)

Question 28

PC The negative charges of the food dye components will interact with the ____ of the NaCl The positive charges of the food dye components will interact with the ____ of the NaCl both of which will make _____ bond

Answer 28

- positive Na - negative Cl ion-dipole bonds

Question 29

PAPER CHROMATOGRAPHY - RESULTS 1. order of increasing polarity 2. order of separation (lowest to highest) 3. order of separation in reversed chroma 4. order of separation in normal chroma

Answer 29

1. allura red ac < tartrazine < brilliant blue fcf 2. allura red ac > tartrazine > brilliant blue fcf 3. allura red ac > tartrazine > brilliant blue fcf 4. brilliant blue fcf > tartrazine > allura red ac

Question 30

Paper of Chromatography BASIS OF SEPARATION

Answer 30

ionic character more ionic groups = more polar

Question 31

IONIC CHARACTER - found in what type of chromatography - ____ in a compound as the number of ____ ____ increases

Answer 31

- paper chromatography - increases - charged groups

Question 32

NOTES to remember what groups are more polar and less polar

Answer 32

MOST POLAR 1. charged groups (O- or N+) 2. COOH (carboxylic acid) 3. CONH2 (amide) 4. OH 5. NH2 6. CONHR 7. NHR 8. CONR2 9. COOR 10. CO 11. ROR 12. NR2 LEAST POLAR OH > NH > ON > O > N

Question 33

2 possible ERRORS in chromatography + reason (2 per error) + how to counter and why

Answer 33

1. Use of ballpen for sample spots - ballpen ink will smear with spot - ballpen ink also consist of colored pigments INSTEAD, use pencils since pure graphite lang siya which are made of inert carbon 2. Level of solvent is above the sample spots - no separation will occur - solvent will wash away/dissolve the pigments INSTEAD, make sure the solvent will not exceed sample spot

Question 34

for a successful and effective chromatographic separation, ___ ___ of ____ ___ among the ___ participants in the separation process, which are the??

Answer 34

- proper balance of IMFA - 3 participants 1. solute/compound to be separated 2. mobile phase 3. stationary phase

Question 35

selection of an appropriate solvent system/mobile phase is by ____ and ____

Answer 35

trial and error

Question 36

How many cm / mm?

Answer 36

1 cm = 10 mm

Question 37

If whatman no.1 filter paper ang gamit sa paper chromatography, automatic what is the stationary phase?

Answer 37

water