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5) FDSC 300 > Lecture 12- Vitamins > Flashcards

Lecture 12- Vitamins Flashcards

1
Q

What is a Vitamin?

A

Vitamins are defined as relatively low-molecular weight compounds which are required in small quantities for normal metabolism

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2
Q

Why is a property tht most vitamins have and how can it be useful?

A

Ring structure which is important with UV spectroscopy

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3
Q

What is the relevance of determining vitamins? (4)

A
  • nutrition quality
  • regulatory requirements
  • Determination fo shelf life
  • Indicate the effect of food on packaging, processing, storage
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4
Q

What are vitamins classified on?

A

Solubility: fat or water soluble

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5
Q

Why is milk fortifeied?

A

Considered a complete good

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6
Q

Vitamins are normally expressed as?

A

mg or ug

Also internaltional units, daily value

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7
Q

What is an IU?

A

amount of a substance, based on the measured biological acitivty or effect

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8
Q

IS IU universal?

A
  • Definition of one IU varies from substance to substance:
    ○ 1IU of Vit.A –> 0.3 ug retinal, 0.6 ug Bcarotene, 1.2ug other pro vitmain A active carotenoids
    ○ 1IU of Vit C–> 50 ug L-ascorbic acid
    ○ 1IU of Vit D is the equivalent of 0.025ug cholecaciferl/ergocalciferol
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9
Q

What is the process of sample determination?

A

Sample–> extract–> purified extract –> analysis

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10
Q

How do you determine vitamins?

A

Vitamins belong to different classes of organic substance so analysis of vitmains is performed by different chemical, physical and biological methods

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11
Q

How do you extract vitamins?

A

extraction of a vitamin
from its biological matrix prior to analysis.
generally includes one or several of the following treatments:
heat, acid, alkali, solvents, and enzymes. (for extraction)
- In general, extraction procedures are specific for each vitamin and
designed to stabilize the vitamin.
- In some instances, some procedures are applicable to the combined
extraction of more than one vitamin.

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12
Q

Typical extraction procedure for ascorbic acid?

A

cold extraction with metaphosphoric acid/acetic acid

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13
Q

Typical extraction procedure for vitamin B1/B2

A

boiling or autoclaving in acid plus enzyme treatment

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14
Q

Typical extraction procedure for Niacin?

A

autoclaving in acid ( no cereal products) or alkali ( cereal products)

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15
Q

Typical extraction procedure for Folate?

A

enyme extraction with alpha amylase, protease, and gamma glutamyl hydrolase

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16
Q

Typical extraction procedure for Vit A,E,D

A

organic solvent extraction , saponification and re extraction with organic solvents

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17
Q

What does Saponification do?

A

method of making soap, glycerol is left, gets rid of fat

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18
Q

What are some challenges for milk analysis? (3)

A
  • Relatively low concentration
    • Poor stability in response to heat, light, oxidation
    • Possible interference from milk components
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19
Q

What do you want to determine in milk?

A

Vitamin D

20
Q

What is the simple procedure of vit D in milk determination?

A

solvent extraction, saponification, re-extraction

21
Q

What is HPLC?

A

HPLC is a general method for any minor compounds extraction : mobile and stationary phase that can determine a compound

22
Q

What are the three types of vitamin assays?

A

Bioassays
Microbioloical assays
Physiochemical assays

23
Q

What are bio assays?

A

Involving animals

24
Q

What is an example of a bioassay?

A
  • Vit D using rat models
    Line test for vit D is based upon the cure of experimentally induced rickets in rats during a specific period, as measured by the calcification of bones, the extent of calcification determined
25
Q

What is the line test?

A

Bone section are stained with silver for observing calcification and is compared agaisnt the reference amount (goup that is given defined amount of vit D)

26
Q

What are the disadvantages of Bioassay?

A

Costly

Long

27
Q

What are the microbiologial method?

A
  • Principle: growth of microorganism is proportional to their requirement for a specific vitamin
    ○ Vitamin tested will act as the growth limiting nutrietn
    ○ Growth of a certain microorganism in an extract of a vitamin containing sample is compared against the grwoth of this microoganism in the presence of known quantities of that vitamin
28
Q

What are the limitation of microbiological method?

A

Has to be soluble vitamins

29
Q

What vitamin are used in bioassays? (2)

A

B12 and D

30
Q

What type of growth is measured with microbiological methods?

A

turbidity

31
Q

What are the steps of Niacin Analysis?

A

1-Weigh the sample.
2-Add 1N H2SO4.
3-Autoclave 30min at 121-123 0C.
4-Filter the mixture, discard and residue.
5-To the filtrate add DifcoNiacin Assay Medium (assay medium is devoid of Niacin).–> limiting reagent is missing!
6-Autoclave 10min at 121-123 0C.
7-Add Lactobacillus plantarumATCC 8014.
8-Incubate at 37 0C for 16-24hr.
9-Measure the optical density and compare with that of bacteria grown in medium containing known amount of the vitamin (Niacin

32
Q

Analysis of Folate?

A

1-Weigh the sample (2g).
2-Add 50ml buffer
3-Add α-amylase, protease, and conjugate and allow for the digestion of the matrix components
4-Deactivate enzymes by boiling the mixture for 5min.
5-Filter the mixture and discard the residue
6-Add the filtrate with bacterial growth medium
7-Autoclave for 10min at 121-123 0C.
8-Add Lactobacillus casei(spp. rhamnosus) ATCCTM 7469 .
9-Incubate at 37 0C for 20-24hr.
10-Measure the optical density and compare with that of bacteria grown in medium containing known amount of the vitamin (folate)

33
Q

How do you determine a method for vitamin analysis? (8)

A
  1. Method accuracy and precision (Consider the use of official methods)
  2. The need for bioavailability information
  3. Time and instrumentation requirements
  4. Personnel requirements
  5. The type of biological matrix to be analyzed
  6. The number of samples to be analyzed
  7. Regulatory requirements –Must official AOAC International methods be used?
  8. Suitability of the method (Applicability of microbiological assays is limited to water-soluble vitamins (most commonly niacin, B12, and pantothenic acid)).
34
Q

What is Vitamin A sensitive to?

A

ultraviolet (UV) light, air (and any prooxidants, for that matter), high temperatures, and moisture.

35
Q

What is the method to determine vit A?

A
  • High-performance liquid chromatographic (HPLC) (AOAC Method 992.04) methods are considered the only acceptable methods to provide accurate food measurements of vitamin A activity.
36
Q

What is the principle of vit A analysis using HPLC?

A

test sample is saponified with ethanolic KOH vitamin A is extracted into organic solvent and then concentrated. Vitamin A isomer, all trans retinol and ci retinol levels are determined by HPLC on silica column

37
Q

How do you extract samples with a vitamin A sample?

A

No extraction ebcause HPLC will fractionate the compounds already

38
Q

What are the steps of Vitamin A determination?

A

1-Transfer 40 ml of ready-to-use formula or fluid milk to a 100-ml digestion
flask containing a stirring bar.
2-Add 10 ml of ethanolic pyrogallol(absorb oxygen) solution and 40-ml ethanolic KOH (for saponification of lipids, Potassium salts of fatty acids and glycerol).
3-stir at room temperature for 18 hr.
4-Extract vitamin A with hexane: diethyl ether mixture
5-Analyze the organic layer using HPLC for VitA.

39
Q

What is the principle for vitamin C analysis?

A

L ascorbic acids is oxidized to L dehydro-ascorbic acid by the oxidation reduction indicator dye 2,6 dichloroindophenol (DCP)

		- Presence of ascorbic acids in solution/juice keeps the pH acidic and DCP is red colored dye in acidic
		- DCP specifically oxidize ascorbic acid and DCP get reduced to colorless form
		- Endpoint is a rose-pink color
40
Q

What is vitamin C susceptible to?

A

oxidative deterioration that is accelerated by high pH and the presence of ferric and cupric ions.

41
Q

When should you use a chelating agent with vit C?

A
  • If significant amounts of ferric Fe, cupric Cu ions are present in the sample to be analyzed, it is advisable to include a chelating agent such as ethylenediaminetetraaceticacid (EDTA) with the extraction
42
Q

What is the method of thiamine extraction?

A
  • Extraction of thiamine in 0.1N H2SO4 with dilute acid
    - Enzymatic hydrolysis of thiamines phosphate esters at pH 4-4.5 presence of 1.2M sodium acetate
    - Chromatographic cleanup(purification, oxidation to thiochromate, measurement using fluorescence intensity
43
Q

What are the steps of thaimine analysis?

A
  • Extraction of thiamine by heating food sample (2 g) for 30 min in 0.1N HCl
    - Cool, adjust pH to4.0-4.5
    - Enzymatic hydrolysis of thiamine’s phosphate-esters using takadiastasein presence of 2.5 M sodium acetate solution. Incubate overnight at 370C.
    - Cool and dilute to 100 mL with water
    - Purification by passing over ion exchange Chromatographic cleanup (i.e., purification),
    - Oxidation to thiochrome using alkaline potassium ferricyanide
    - Fluorescence intensity measurement using fluorometer
    - Compare the value against known concentration
44
Q

What is used to oxidize ascorbic acid? What happens to it? & when is the end point?

A

DCP, get reduced from red (in acidic solution) to a colorless form, so the end point is a pink colour

45
Q

What 2 vitamins that we saw have an AOAC official method of analysis?

A

thiamine and niacin

5) FDSC 300 (14 decks)

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