Polyphenols 1 Flashcards
1
Q
What does >12 years education do to your life expectancy?
A
+ 5 years
2
Q
What does being overweight do?
A
- 4 months
3
Q
What does being obese do?
A
- 4 years
4
Q
What does smoking do?
A
- 7 years
5
Q
What does increasing fruit and veg consumption from 1 to 5 portions/day do?
A
+1 year
6
Q
What happens with age?
A
There is an increased risk of chronic disease
7
Q
What is hugely linked to chronic disease?
A
oxidative stress
8
Q
What can cause oxidative stress?
A
Living and breathing
Environment
UV exposure
Diet e.g high sugars, high fat
9
Q
What does oxidative stress lead to in general?
A
Chronic
Inflammation and perturbed metabolism
10
Q
What does Chronic
Inflammation and perturbed metabolism lead to?
A
Cancer
Cardiovascular
Diabetes
Accelerated ageing
11
Q
What is oxidative stress?
A
imbalance between the production of free radicals/reactive oxygen species (ROS) and the neutralization of ROS before damage is caused, or subsequent repair of damage
12
Q
What is a free radical?
A
any chemical species (atom, molecule or ion), capable of independent existence, with 1 or more unpaired electrons
13
Q
What are ROS molecules like?
A
highly unstable molecules created normally in vivo and also due to external risk factors
14
Q
What are important radicals?
A
Quantitatively most important: superoxide radical (O2•-)
Other important radicals: hydroxyl radical (OH• ) and nitric oxide (NO•)
15
Q
How is oxidative stress and chronic disease linked?
A
Progression of several chronic diseases linked to oxidative stress in vivo e.g.
Oxidation of LDL –> plaque formation in blood vessels
16
Q
What is the rate of living hypothesis?
A
the higher the metabolic rate of an organism, the greater the production of reactive oxygen species (ROS) and hence the shorter the life span.
17
Q
What goes against the rate of living hypothesis?
A
Birds live longer than predicted by their metabolic rates.
At a given metabolic rate, mitochondria from birds produce fewer ROS so ROS production rather then metabolic rate provides the strongest correlation with overall longevity.
18
Q
What are endogenous sources of oxidative stress?
A
Mitochondria Peroxisomes Lipoxygenases NADPH oxidase Cytochrome P450
19
Q
What are exogenous sources of oxidative stress?
A
Ultraviolet light
Ionizing radiation Chemotherapeutics Inflammatory cytokines Environmental toxins
20
Q
What are antioxidant defences that counteracts and regulates overall ROS levels to maintain physiological homeostasis?
A
Enzymatic systems (CAT, SOD, GPx)
Non-enzymatic systems (Glutathione,
Vitamins (A,C and E))
21
Q
What does too little ROS in body cause?
A
Impaired physiological function
- Decreased proliferative response
- Defective host defences
22
Q
What does the right level of ROS cause?
A
Homeostasis, normal growth and metabolism
23
Q
What does too much ROS in body cause?
A
Impaired physiological function
- random damage to proteins, lipids and DNA
- may also constitute a stress signal that activates specific redox-sensitive signalling pathways
24
Q
What happens in lipid peroxidation?
A
- free radicals “steal” electrons from the lipids in cell membranes, resulting in cell damage.
- commonly affects polyunsaturated fatty acids, they contain multiple double bonds that possess especially reactive hydrogen atoms.
25
What is step 1 of lipid peroxidation?
Initiation: fatty acid radical is produced. Common initiators in living cells are reactive oxygen species (ROS), such as OH· and HOO·, which combines with a hydrogen atom to make water and a fatty acid radical.
Unsaturated lipid + -OH --> lipid radical (H2O removed)
26
What is step 2 of lipid peroxidation?
Propagation: fatty acid radical is not a very stable molecule, so it reacts readily with molecular oxygen, creating a peroxyl-fatty acid radical. This radical is also an unstable species that reacts with another free fatty acid, producing a different fatty acid radical and a lipid peroxide, cycle continues
27
What can this propagation cause?
chains of destruction damaging DNA, proteins, lipids etc.
28
What is step 3 of lipid peroxidation?
Termination: occurs when a radical meets another radical (needs high conc of radical species) or an antioxidant e.g. vitamin C
29
Why are ROS produced?
On purpose: cell proliferation & signalling (H202) or during inflammation
Accidentally: superoxide radical through electron leakage in mitochondria
30
Who came up with the Free Radical Theory of Ageing?
in 1956 by Denham Harman
31
What does evidence suggest regarding obesity and oxidative stress?
Obesity induces oxidative stress
32
How do oxidative stress levels increase in obesity?
Increasing oxidative stress levels with increasing BMI (linked to TBARS, F2-isoprostanes, oxLDL levels) and lower antioxidant capacity and plasma antioxidant levels
33
What does the excessive accumulation of fat lead to?
enhanced production of ROS in adipocytes and systemic tissues
34
What else is ROS positively correlated with (other than obesity)?
incidence of neuronal degeneration, cardiovascular events, diabetes mellitus and cancer
35
What is the antioxidant hypothesis?
Concept that “antioxidants” could cure cancer, CVD or diabetes mellitus
36
What does epidemiology suggest regarding the antioxidant hypothesis?
associations between high consumption of fruit and vegetables (=high in antioxidants) and lower incidence of certain “oxidative stress” related diseases
37
What is an example of an application of this?
“5 a day” campaign in UK since 2002 – based on WHO advice to consume 400g Fruit & Veg per day
38
Oxidation is essential to get energy from fats, but what is the problem?
The functioning of mitochondrial protein can cause superoxides that cause damage/oxidative stress
BUT can be removed by endogenous defences
39
What is the problem with endogenous defences?
May be insufficient
40
What is a solution to insufficient endogenous defences?
Exogenous antioxidant defences from the diet – e.g. vitamin E, C
Increase endogenous defences (boost our own defences)
Inhibit oxidative processes such as superoxide generating metabolic enzymes
41
How do polyphenols work in the body?
Although polyphenols are chemical antioxidants, they do not act by direct chemical antioxidant mechanisms in the body
42
What does the evidence between diet and CVD disease risk show? (Hu and Willett, 2002)
Fruits and some vegetables and WG high in polyphenols
| Causes reduced risk
43
What is important in the definition of antioxidants?
Compounds (polyphenols, vitamin C, E, carotenoids) and food preservatives such as BHT are antioxidants in vitro
- Inhibition of lipid rancidity
- All “antioxidant” assays not involving cells or organisms
- Chemical property to scavenge radicals etc.
44
What are examples of in vitro antioxidant assays?
```
Superoxide scavenging
Hydrogen peroxide scavenging
Hydroxyl radical scavenging
Metal ion-dependent hydroxyl radical scavenging
Peroxyl radical scavenging
Lipid-soluble peroxyl radicals
ORAC, TEAC, TRAP
```
45
What is not an antioxidant?
Concept of an antioxidant has been refined especially based on in vivo experiments
Chemical antioxidant assays such as TRAP, TEAC, ORAC etc. are NOT indicative of a biological condition
Rather they are content, rather than functional assays
46
What is an example of these content rather than functional assays?
Vitamin C – water soluble essential antioxidant
Vitamin E – fat soluble essential antioxidant
Polyphenols – good antioxidants, generally healthy
BHT – excellent antioxidant, some mild toxicity
a-Amanitin – antioxidant but deadly mushroom toxin
47
What does the current more complex view look at?
- how much is absorbed into the body
- specific molecular actions which are affected in the body, resulting in a change of a biomarker and disease risk
- Chemical antioxidant assays on foods give information on content of polyphenols and other redox active compounds but NOT health benefit
48
How can antioxidant activity of plasma by measured?
by the TEAC assay e.g. vitamin C 3%
49
What did EFSA publish on cocoa flavanols and maintenance of normal endothelium-dependent vasodilation?
200 mg of cocoa flavanols should be consumed daily.
This amount could be provided by 2.5 g of high-flavanol cocoa powder or 10 g of high-flavanol dark chocolate, both of which can be consumed in the context of a balanced diet.
The target population is the general population.
*evidence from 5 published and 1 unpublished randomised controlled trials in healthy subjects
50
How can phytochemical, phytonutrients be categorised?
Phenolics (e.g. ferric acid, caffein acid)
Polyphenols
Glucosinolates (sulforaphane)
51
Where are chlorogenic acids found?
In coffee
| An ester of quinic acid and caffeic acid (linked in 3 different positions via ester bond)
52
What is the major form of chlorogenic acid in coffee?
5-caffeoylquinic acid (5-CQA)
53
What strongly affects amount of chlorogenic acid in the diet?
coffee consumption
None: 10-100mg chologenic acid in diet
Heavy: 500-2000
54
What can polyphenols be separated into?
Phenolics
ellagic acid/ellagitannins
Flavonoids
Isoflavones (genistein, daidzein)
55
What can flavonoids be separated into?
Flavonols (e.g. quercetin)
Flavanols
Anthocyanins
Flavanones (e.g. hesperidin)
56
What is high in quercetin?
onions (present as glycosides)
57
How are many flavonoids found in foods?
As glycosides
58
Describe hesperidin?
Hesperidin is a flavanone rhamnoglucoside that is present in Citrus fruits and juices especially oranges.
(a-L-rhamnose + b-D-glucose
+ hesperetin)
59
What do anthocyanins give in food?
Dramatic colour
60
What has high anthocyanin content?
chokeberry, red grapes
61
What can flavanols be separated into?
Catechins
| Pro(antho)cyanidins
62
What are examples of catechin?
```
Catechin
epicatechin
epigallocatechin
epigallocatechin gallate
epicatechin gallate
```
63
Top three foods based on phenol content?
Dark choc, blackberry, high bush blueberry
64
How can we measure polyphenols?
Measurement of polyphenols is by HPLC linked to UV an/or mass spectrometry detection
65
What is the polyphenol % in tea?
30%
Mostly catechins
In green tea mostly EGCG
66
What is the difference between green and black tea?
Green tea has no colour and catechin
After short fermentation
Oolong tea is orange-yellow and has theaflavins
After full fermentation
Black tea is red-amber and has thearubigins
67
What are catechin quantities in leaves?
Green tea dry leaves 12514mg/100g
Black tea dry
3316mg/100g
68
What happens when 2 to 5 of the catechins join?
they are called dimers; and when two to five are linked they are called proanthocyanidins
69
What is the role of polyphenols in the plant?
```
Protection from UV light
Wound healing
Protection against stress
Anti-fungal
Anti-bacterial
Anti-insect
```
70
What does boiling do to quercetin?
Boiling does not change the level of quercetin glucosides in onion, but some is leached into the cooking water.
71
What does storage do?
On storage, some quercetin-4’-monoglucoside is lost whereas quercetin-3,4’-diglucoside increases. There is almost no free quercetin.
72
What does frying do?
There is a 20-25% loss in quercetin (Q) conjugates after frying for 15 min.
73
What does autolysis (destruction of own cells) result in?
conversion of Q-3,4’-diglucoside to Q-4’-glucoside and to free Q, over a period of 24 h.
74
What does microwaving broccoli do?
15 min hardly affects the amount or nature of flavonol glycosides.
75
What is cocoa high in?
Cocoa is high in epicatechin, but processing affects the content dramatically
76
What are transit times in the body?
```
Stomach: 10 min (saline) to
4 h (lipid portion of large meal)
```
Small intestine: 3-4 h
Large intestine: 48-96 h
77
Where does most absorption occur?
in the small intestine
78
How is the mechanism of absorption examined?
Cultured human intestinal Caco-2 cells as a model to examine the mechanism of absorption
79
How does food absorption work in general?
Only a fraction of a food component is absorbed
Only a certain percentage (0-100%) of a compound is absorbed --> metabolized (0-100%) --> and finally available to certain organs of the human body
Some is released into colon (faeces), some through intestine (some goes back to colon) then some into bloodstream and liver
80
How can bioavailability of flavonoids be determined?
Oral faecal balance
Post prandial test
Chronic consumption
81
Oral faecal balance
BV = intake – faecal content
Gut degradation means that flavonoids appear “100%” absorbed
Ileostomists (no colon) more useful for small intestinal absorption
Most useful for minerals
82
Post prandial test
BV = Area Under Curve after oral
consumption of one
dose
Best for comparison between
treatments
Subject is his own control
83
Chronic consumption
BV = amount in urine (e.g. over 24 h)
Closer to consumer situation
Steady state, and can indicate
absorption over a long period of time
Gives a MINIMUM amount or percentage absorbed
Different flavonoids behave differently – so can only compare for the same flavonoid
84
What does Cmax mean in bioavailability?
max concentration it reaches, indicates the max conc in the plasma
85
What does AUC mean?
Area under curve
| - indicates how much has gone into plasma
86
What must be considered in bioavailability?
Effect of
- dose
- matrix
- chemical attachments
- chemical structure
- hydrophobicity
87
What is cmax for flavonoids in humans normalised to?
50mg dose
| assuming xmax increases linearly with dose e.g. with 50mg dose of isoflavones cmax=2um then 100mg rose would give 4um
88
What is 50mg dose of hesperidin?
0.5um cmax
89
What can be said about quercetin glucoside?
Bioavailability almost as high as isoflavones
90
What can be said about Rutin (other form of quercetin)?
much less bioavailable
- it is very similar to the glucoside
- has an extra rhamnose attached so less easily absorbed
91
What can be said about flavanol bioavailability?
EC + ECGC = medium levels, EGC = much more bioavailable
92
What can be said about • Anthocyanidins + proanthocyanidin dimers?
poorly absorbed
93
How can we derive the Half life of the polyphenols?
from the pharmacokinetic curve
94
What can be said about polyphenols in body?
Unlike vitamins/minerals which are used in the body, they aren’t stored in the body – only spend certain amount of time in blood
95
What does half life indicate?
Time spent in body
96
What can be said about cmax and half life?
There is no correlation
97
What would we predict about effectiveness?
Would predict that most effective ones would be ones with highest cmax (best bioavailability) but half life can also affect effectiveness
98
What are the general Half life?
* Isoflavones: 5-7h
* Flavanones: 2h
* Flavonols: 17-20h
* Flavanols: 2-4h
99
How can we find out tmax?
From the curve
| the time the cmax occurs
100
What does tmax effect?
Where the compound is absorbed (under 4 hours is stomach and SI - mostly SI)
101
Where are isoflavones, flavanones and rutin absorbed?
Colon = higher Tmax
102
Where are Quercetin glucoside + flavanols absorbed?
Small intestine = lower Tmax
103
What is rhamnose and glucose attached to?
o Hesperidin
o Naringin
o Rutin
104
What does rhamnose and glucose attached to Hesperidin, Naringin and Rutin mean?
o All have higher Tmax = abs in colon
105
What occurs with only glucose attached?
Lower Tmax = absorption in SI (e.g. quercetin glucoside)
106
What occurs when no sugar is attached e.g. flavanols?
Also low Tmax = abs in SI
107
What does this imply?
glucose is removed in SI and rhamnose cannot be removed in SI so has to be hydrolysed in the colon
108
What are enterocytes in SI responsible for?
deciding if polyphenols move from gut lumen into blood stream
109
What is the arrangement between gut lumen, enterocyte and blood?
Gut lumen boarders enterocyte with apical membrane and enterocyte boarders blood with basolateral membrane
110
What occurs in first step in metabolism of lactose or polyphenol with glucose attached?
LPH = lactase phlorizin hydrolase
o Hydrolyses lactose
o Remove glucose from polyphenols
o Is on surface of enterocyte, so to be hydrolysed the molecule doesn’t have to enter the cell
111
What is an example of this?
quercetin glycoside (3- or 4’-) --> quercetin aglycone
Can now abs by passive diffusion
Membrane is hydrophobic, quercetin aglycone = sufficiently hydrophobic to pass through
112
What is the alternative pathway to this?
o Quercetin 4’-glycoside slowly diffuses through membrane
o Cytosolic b-glucosidase can de-glycosylate
o Minor pathway compared to the other
113
What does passive diffusion depend on?
flavonoid size, hydrophobicity, cellular gradient
114
What occurs with attached rhamnose?
prevents absorption in the small intestine
115
What is an example of this?
Rutin (quercetin with glucose + rhamnose)
o is not a substrate for LPH
o Too big to diffuse through membrane
o Even if it did, no enzyme to remove rhamnose
o Gut microflora can produce enzyme to remove rhamnose
o So in colon = hydrolysed + absorbed
116
What is the deciding factor if something is absorbed in SI?
brush border enzymes
117
What happens after the initial step of metabolism?
A proportion of absorbed polyphenols are conjugated in the enterocyte
118
What happens once quercetin is in enterocyte?
• Chemically changed by various enzymes e.g. UGT (glucuronosyl transferase)
--> adds glucuronide group to quercetin
• This changes properties of quercetin --> more H2O soluble --> can’t diffuse through membrane anymore so needs specific protein transporter
119
What happens after this step?
Polyphenols are exported back to the lumen or absorbed into the blood depending on the specificity of apical and basolateral transporters
120
What happens if quercetin is changed to 3’-glucuronide?
o ABCG2 transporter on apical membrane = specific to this form
o So it is effluxed back out into gut lumen
121
What happens if quercetin is changed to the 2 other forms (3- and 7-glcA)?
ABCC1/C3 transporter on basolateral membrane = transport them into blood
122
What is an aglycone?
No sugar attached (so can enter enterocyte)
123
What does specificity of enzymes e.g. UGT do?
Decides whether it is absorped as different forms require different transporters
124
Why is colonic bacteria important?
* Important for some polyphenols e.g. rutin
| * Can remove rhamnose
125
What can colonic bacteria produce?
glycosidases, glucuronidases, and sulfatases that can strip flavonoid conjugates of their sugar moieties, glucuronic acids and sulfatases
126
What biological effects we associate with flavonoids are linked to cardio-vascular disease?
```
Inflammation
Hypertension
Improvement of vascular function
Antiatherogenic
Platelet aggregation
Proliferation
Cellular and mitochondrial protection
Modulating blood glucose and blood lipids
Modulating NO metabolism
```
127
What does dark chocolate consumption do?
Ingestion of dark chocolate (40 g) significantly improves flow mediated dilation (FMD) after two hours.
White chocolate has no effect.
N = 10 in each group.
The increase in FMD lasts for eight hours after the ingestion of dark chocolate.
128
Does green tea affect FMD?
consumption of 200mg EGCG as isolated EGCG, green tea extract, or green tea
Only green tea increased FMD
129
What is the relationship between EGCG plasma levels and changes in FMD?
EGCG plasma levels not correlating with changes in FMD
HOWEVER
Caffeine correlated with FMD changes
130
Does Orange juice and hesperidin affect CV risk markers?
- Acute trial
- Baseline and 5h post intake
- Men with higher risk 320mg hesperidin
- tested control, OJ, hesperidin supplementation
- Plasma levels only high after juice intake
- No effect on vascular markers of both juice and supplement
131
Do berries effect FMD?
Double blind randomized controlled crossover trial
N=10
FMD % response up to 8h post consumption
1% increase in FMD - decrease in CVD risk of 10-13%
132
How do polyphenols improve vascular function?
Likely involving a number of mechanisms
increase in NO bioavailability as FMD is at least partially NO mediated
Induction of eNOS enzyme to increase NO
Prevention of oxidative damage in endothelial cells by increased Nrf2 signalling
