Nutrient

Bioavailability
(What we get from what we have taken in)

Dereje Tsegaye(MPH,Assiss.Prof.PhD fellow)

1
 Topics
• Definition

• What makes it up

• Critical phase

• How its measured

2
 First basic law of nutrition:

No nutrient is absorbed and

utilized to the full extent that it
is fed
Steven Blezinger

3
Define Bioavailability
•Bioavailability is the fraction of an ingested nutrient that
is available for utilization in normal physiologic functions
and for storage.
–That which becomes bioavailable

–The fraction (or percentage) of nutrient absorbed that

is useful to the body
–The degree to which an absorbed nutrient is available
to the system 4
 Biological Availability
(Bioavailability)

• Definition can be based on either the percentage of a

nutrient ingested or the percentage of a nutrient
absorbed that becomes useful to the organism

• The percentage ingested is preferred by some because

the percentage absorbed is difficult to determine and
relies on an indirect analysis

• The percentage absorbed is, nonetheless, a more

accurate appraisal of bioavailability
5
 Nutritional Definition
• Bioavailability is a post-absorption assessment of how
much of a nutrient that has been absorbed becomes
functional to the system

Food Science Definition

• Bioavailability is a assessment of how much of a
nutrient is retained in the food product after
processing for the consumer

6
 Why Not Absorption Alone as an index of
Bioavailability?
• Assimilation may be a major part of a mineral’s bioavailability
and needs to be assessed separately e.g.
Absorption % Retention %a
75Se as selenite 92 <50
75Se as selenomethionine 96 >80
a
Arbitrary units
• 2-picolinic acid enhances zinc absorption in rats by
nearly 60%.
• But, also increases zinc excretion so there is no net
effect on retention and hence no increase in
bioavailability
7
 Different terms used in bioavailability

• Bioavailability is the fraction of an ingested nutrient

that is available for utilization in normal physiologic
functions and for storage.

• Bioconversion is the fraction of a bioavailable nutrient

(e.g., absorbed provitamin A carotenoids) that is
converted to the active form of a nutrient (retinol).

8
 Terms…
• Bioefficacy is the fraction of an ingested nutrient (e.g., dietary
provitamin A carotenoids) that is absorbed and converted to
the active form of the nutrient (retinol) in the body.

• Functional bioefficacy is the fraction of an ingested nutrient

that performs a certain metabolic function, such as the ability
of ingested provitamin A carotenoids to reverse or prevent
abnormal dark adaptation.

9
• With respect to provitamin A carotenoids, the term
bioefficacy is defined as the product of the fraction of the
ingested amount that is absorbed (bioavailability) and the
fraction of that which is converted to retinol in the body
(bioconversion).

* Bioefficay = bioavailability X bioconversion

10
Components of Bioavailability
• Digestion
• Absorption Absorptive
Phase
• Liver surveillance
• Transport
• Trans membrane movement
• Intracellular movement Assimilation
Phase
• Target binding

11
Bioavailability in toto
Raw Food Product (100%) Total (proximate analysis)

Processed Food Chemically available

Digestion

Absorption
Biologically available
Cellular uptake

Functional Nutrient
12
The
The fraction
fraction of
of the
the total
total amount
amount absorbed
absorbed that
that
performs
performs aa function
function
Digestion
Digestion

Absorption
Absorption

Which is the Blood

BloodTransport
Transport
most critical
Liver
Liver and
andkidney
kidney excretion
excretion
phase for
Nutrients ? Losses
Membrane
Membrane transport
transport along
the
Intracellular
Intracellular movement
movement way

Functional
FunctionalSite
Site
13
 Note that

A nutrient is considered outside the

body until it passes through the
intestinal barrier

14
The amount that gets absorbed depends on:
• Digestibility of the food source
Extrinsic
ExtrinsicFactors
Factors
• Solubility of the nutrient
• Elements in the food source that hinder or facilitate
absorption (enhancers and inhibitors)

With a focus on the organism, bioavailability

depends on:
• Age
• Health
• Nutritional state
Intrinsic
Intrinsic • Physiological state
Factors
Factors • Genetic predisposition
• Gender
• Developmental stage
• Species 15
Bioavailability of
Macronutrients

16
 Absorption & Transport
End-products of digestion:
1. CHO >>> Monosaccharides

2. Fats >>> Glycerol + fatty acids

3. Proteins >>> Amino acids

4. Vitamins, minerals & water – no

digestion 17
Absorption

18
 Absorption & Transport
• Absorption occurs in the small intestine

– Wall of small intestine covered with 100s of folds

– Each fold covered with 1000s of villi

– Each villi contains 100s of microvilli

19
The Small Intestine Villa

20
 Absorption & Transport
• Absorbed nutrients enter either the:
1. Vascular system – water-soluble nutrients
(monosaccharides, amino acids, water- soluble vitamins,
minerals, water, short chain fatty acids) enter the blood
via the portal vein for transport to the liver

2. Lymph system – fat-soluble nutrients (lipids, fat-soluble

vitamins) enter here, eventually entering the blood near
the heart

21
 Absorption & Transport
3. Transport of lipids and lipid soluble vitamins – since fats
are insoluble in water, they must be packaged for
transport as lipoproteins (triglyceride, phospholipid,
protein, cholesterol)

22
Absorption of Carbohydrates
a)Starch and other discharides

23
a)Starch & other disaccharides …

24
b)Digestion of oligosaccharides, resistant starch and non-
starch polysaccharides (Dietary Fiber)
O ligosaccharide s (e g. Raffinose , Stachyo se)
and non-starch po lysaccharide s resistant
starch

Escape digestion in the

upper gut (small intestine

They get fermented in the

colon by anaerobic bacteria

Production of
gases likes co2, Production of
methane and short chain fatty Increased faecal Biomass
hydrogen acids (SCFA) resulting in increased
sulphide  Acetate peristalsis
 Propionate
 Butyrate
25
 Dietary fiber
• Fiber refers to certain types of carbohydrates that our
body cannot digest (oligosaccharides and non-starch
polysaccharides).

• These carbohydrates pass through the intestinal tract

intact and help to move waste out of the body.

• Diets that are low in fiber have been shown to cause

problems such as constipation and hemorrhoids and to
increase the risk for certain types of cancers such as colon
cancer.
26
 Cont…
• Diets high in fiber; however, have been shown to
decrease risks for heart disease, obesity, and they
help lower cholesterol.

• Foods high in fiber include fruits, vegetables, and

whole grain products.

27
 How does fiber prevent different health
problems?
Cancer (Colonic, breast..)
• Prevents secondary bile acid circulation
• Decrease intestinal transit time
• Decrease contact of carcinogens with intestinal
cells
• Fermentation product butyrate has apoptotic
effect
• Decreases absorption fats and sugars

28
 Cont…
Dietary Fiber prevents Constipation, Hemorrhoids &
Diversticulosis by:

• Increasing perystalsis making stool bulk

• Decreasing straining to pass stool

29
 Carbohydrate malabsorption
Lactose intolerance
• Most mammals normally cease to produce lactase,
becoming lactose intolerant, after weaning
• It is estimated that 75% of adults worldwide show some
decrease in lactase activity during adulthood
• The frequency of decreased lactase activity ranges from 5%
in northern Europe through 71% for Sicily to more than 90%
in some African and Asian countries

• This distribution is now thought to have been caused by

recent natural selection favoring lactase-persistent
individuals in cultures in which dairy products are available
30
as a food source.
 Mechanism of Lactose-Induced Diarrhea
and Flatus
Lactase-sufficient
people absorb Glucose
Lactose Small
>80% of lactose Galactose
bowel
Lactose
Lactase-deficient
people absorb
<50% of lactose SCFA
Colon
lactose
6-20 grams malabsorbed glucose
lactose = flatus CO2+H2 galactose
(1 g = 44 ml H2)

>20 grams malabsorbed

lactose = flatus+diarrhea
FLATUS OSMOTIC DIARRHEA
31
 Glycaemic Index
• The glycemic index or glycaemic index (GI) is a measure
of how quickly blood glucose levels (i.e., blood sugar)
rise after eating a particular type of food.

• Glucose (the defining standard) has a glycemic index of

100

32
Glycaemic index...
Glycaemic
Classification Examples of foods
index range
beans (white, black, pink, kidney, lentil, soy,
almond, peanut, walnut, chickpea); small seeds (
sunflower, flax, pumpkin, poppy, sesame); most
Low GI 55 or less whole intact grains (durum/spelt/kamut wheat,
millet, oat, rye, rice, barley); most vegetables,
most sweet fruits (peaches, strawberries, mangos);
tagatose; fructose
not intact whole wheat or enriched wheat, pita
bread, basmati rice, unpeeled boiled potato, grape
Medium GI 56–69
juice, raisins, prunes, pumpernickel bread,
cranberry juice, regular ice cream, sucrose, banana
white bread (only wheat endosperm), most white
rice (only rice endosperm), corn flakes, extruded
High GI 70 and above
breakfast cereals, glucose, maltose, maltodextrins,
33
potato, pretzels, parsnip, bagels
 Absorption of Lipids
• Abnormal to find more than 6 or 7% of ingested lipids still
intact in the feces( Digestibility = 93-94%).

• Most fat absorption takes place in the duodenum or

jejunum – micelles carry monoglycerides and free fatty
acids to the brush border where they diffuse into
enterocytes.
• Cholesterol and other sterols are poorly absorbed.
• Overall, about 50% of dietary cholesterol is absorbed.

• Fiber (especially soluble fiber) decrease cholesterol

absorption 34
 ABSORPTION…
• Bile salts are absorbed in the ileum (enterohepatic
circulation)

• Short-chain fatty acids(< 12 carbon) and glycerol are

absorbed directly into bloodstream.

• They do not enter the lymph system

• Once in the enterocytes, Larger chain fatty acids(>=12

carbon), monoglycerides and free fatty acids are
reformed into triglycerides

35
 ABSORPTION…
• The triglycerides, and cholesterol will be esterfied in to
cholestrol ester and these will be coated with
phospholipids, and protein to form a Lipoprotein called
Chylomicron

• CHYLOMICRONS enter the lymphatic system through the

lacteals

• ChilomicronsVLDLLDLHDL
Lipoprotein lipase lyses contents!

36
Absorption of Lipids…

triglycerides

37
38
 Absorption & Transport
4 basic types of lipoprotein:
1.Chylomicrons – very, very low density (85%
triglyceride); absorbed from small intestine into
lymph & circulated to cells where some of lipid
material is picked off & remnants return to liver

2. VLDL – very low density lipoprotein (50%

triglyceride); made by liver & travels to cells

39
 Absorption & Transport
3. LDL – low density lipoprotein (50% cholesterol);
remains of VLDL; high levels increase risk of
heart attack

4. HDL – high density lipoprotein (50% protein);

removes cholesterol from blood for return to
liver; high levels decrease risk of heart attack

40
 Malabsorption due to
Luminal Maldigestion of Fat (Steatorrhea)
:Common causes

Pancreatic insufficiency: Chronic pancreatitis

Bile salt deficiency: Loss of terminal ileum:

loss of bile salts in stool
insufficient bile salts

Bacterial overgrowth: Deconjugation and loss

of bile acids

Gastric hypersecretion: Acid inactivation of

pancreatic enzymes
41
 Relationship between Pancreatic
Function and Steatorrhea
100

80
Fecal Fat (g/day)

This leads to deficiency of

60 Lipid soluble vitamins
40

20

0
0 20 40 60 80 100
Pancreatic Function (%)
42
 ABSORPTION OF PROTEINS
• Most protein absorption takes place in the duodenum
and jejunum

• Most amino acids are absorbed into the bloodstream,

but some remain in the enterocytes and are used to
synthesize enzymes and new cells

• >99% of protein enters the bloodstream as amino acids

• Absorption of whole protein can cause a severe allergic
reaction

43
 ABSORPTION OF PROTEINS…
• In the enterocyte, other peptidases immediately digest
everything into single amino acids which are absorbed
into the bloodstream

• Some amino acids share the same transport system, so if

you take in a large amount of one particular amino acid,
you may be inhibiting the absorption of others

44
 Free Amino Acid Absorption

• Free amino acids

– Carrier systems
• Basic AA Na+ Na+
• Neutral AA
• Acidic AA
• Imino acids
– Entrance of some AA is
via active transport
• Requires energy
45
 Peptide Absorption
• Form in which the majority
of protein is absorbed
• More rapid than
absorption of free amino
acids
• Active transport
– Energy required
• Metabolized into free
amino acids in enterocyte
• Only free amino acids
absorbed into blood
46
 Absorption of Intact Proteins
• Newborns
– First 24 hours after birth
– Immuno globulins
• Passive immunity
• Adults
– Paracellular routes
• Tight junctions between cells
– Intracellular routes
• Endocytosis
• Pinocytosis
• Of little nutritional significance...
– Affects health (allergies and passive immunity)

47
 Basolateral Membrane
• Transport of
free amino acids
only*
– Peptides are
hydrolyzed
within the
enterocyte
• Transport mainly by
diffusion and
Na-independent
carriers Groff & Gropper, 2000

48
*Whole proteins are nutritionally insignificant...
 Protein Malabsorption
Celiac Sprue I
• Immune-mediated destruction of enterocytes in response
to ingestion of the protein gluten found in wheat and
certain other grains.
• A fraction termed gliadin contains the immunogenic
material (gluten induced eteropathy)
• Small intestinal villi are damaged or destroyed - "flat gut"
appearance.
• Mature digesting and transporting enterocytes are
virtually absent.

49
 Celiac Sprue - II
• Patchy disease - usually affects proximal intestine more
than distal intestine .

• Mucosal digestion and absorption are both severely

impaired.

• Characteristic antibodies used in diagnosis: IgA

antibodies to tissue transglutaminase or gliadin.

50
 Clinical Manifestations of Sprue
• Weight loss, often with increased appetite
• Bulky, oily stools – steatorrhea - fat malabsorption
• Flatus/frothy stools – carbohydrate malabsorption
• Anemia – deficiencies of iron, folate
• Osteopenic bone disease – Vitamin D and calcium
malabsorption
• Edema/hypoproteinemia – protein deficiency and
malnutrition
• Cheilosis and glossitis – B vitamin deficiencies
• The only known effective treatment is a lifelong Gluten
Free Diet 51
 Absorption of
Minerals
(Unifying principles that apply to all minerals)

52
 Digestion
• Preparing for absorption
• Liberating minerals from a bound state to an aqueous
phase
• Digestive enzymes
• Bile acids and salts that work with digestive enzymes
(e.g., lipases)

53
Purpose of digestion to mineral nutrition
• Minerals in a food source are locked within a matrix
composed primarily of proteins, complex
carbohydrates and fats

• The purpose of digestion is to render large composite

molecules into smaller manageable units…minerals are
liberated during this process

• Digestive processes consists mainly of hydrolytic

enzymes that break chemical bonds between modular
units without total destruction (metabolism) of the
liberated components
54
 Digestive Enzymes (hydrolases)
Enzyme Location Target Action

Pepsin gastric juice proteins breaks peptide bonds

Trypsin and duodenum proteins breaks peptide bonds

chymotrypsin
I
Amylases saliva and starch and breaks glycosidic bonds
duodenum glycogen
Lipases duodenum complex breaks ester bonds
lipids
Glycosidases microvilli di- and tri- breaks glycosidic bonds
saccharides
II
Peptidases microvilli small breaks peptide bonds
peptides

Phase I is primarily salivary and pancreatic secretions

Phase II involves enzymes on the surface of absorbing cells
55
Critical factors in Mineral Absorption
• Absorption tends to be selective for the mineral (makes
finding a unified mechanism more difficult)

• A deficiency increases the fraction of that mineral

absorbed (absorption is tuned to internal bodily needs)

• Certain food chemicals (e.g., phytate, oxalate) lower

absorption by tying up the mineral

• There is competition for absorption machinery

56
 Critical factors…
• Metal ions antagonism (Cu-Zn; Zn-Fe; etc.) occurs at ion
channels during the transmural passage phase of
absorption

• Vitamin dependency is seen with Vitamin D and C that

regulate body load of Ca+2 and Fe2+respectively
• Absorptive cells excrete factors that aid in the solubility
of metal ions

• Some transport proteins are in vesicles that fuse with the

membrane and move vectorially within the cell

57
 Steps in mineral absorption
1. Transport through the luminal (apical) cell membrane,
i.e., start of transcellular
2. Handling within the enterocyte, i.e., mediate
transcellular
3. Transport through the antiluminal basolateral
membrane into the circulation, i.e., end of transcellular.
4. Transport between the cells, i.e., paracellular

*Only metals in an aqueous phase can be transported into

the enterocyte

58
Solubility and Metal Ion Absorption

Two categories of ingested metal Ions

1. Solubility not dependent on pH
Examples: Na+, K+, Mg2+, Ca2+
2. Solubility pH dependent
Examples: Cu2+, Fe2+, Mn2+, Zn2+

Category 1 metal ions are soluble throughout

the gastrointestinal pH range (1-8)
Category 2 metal ions are soluble in acid, but form insoluble
hydroxy-polymers at neutral or alkaline pH. 59
Secretions of Digestion

60
 Mucosal Side A large fraction of the
Fe iron can be trapped
Fe Fe Ca (sequestered) within
Ca Ca
the cytosol of the
Microvilli enterocyte)
Apical
surface

Enterocyte

Basolateral To access the serosal side,

Surface the mineral must pass
(antiluminal either through the
surface)
enterocyte (transcellular
Serosal Side 99%) or the junction
between enterocytes
(paracellular <1%))
61
 Role of Vesicles in the Regulation of
Mineral Absorption

Resting Cell Absorbing Cell

Vesicles are internal membrane compartments that move between the

cytosol and membranes. This movement is regulated by external factors
Vesicles contain the transport proteins that absorb the mineral into the lumen of
the vesicle and bring it into the cell

Vesicles that have fused with the membrane are positioned to absorb minerals.
Absorption thus depends on the number of vesicles that fused with
the membrane. 62
MACROMINERALS
• Monovalent cations, Na+, K+

• Monovalent anions, Cl-

• Divalent cations, Ca2+, Mg2+

• Complexes, HPO4=, HCO3-

63
Rules that apply to the absorption of Macrominerals

Rule 1: Macrominerals in general enter intestinal cells

through transport portals designated for the mineral
(major) or between cells (minor).

Rule 2: The energy for entry is provided by a

concentration gradient across the membrane or by
hydrolysis of ATP (active transport)

Rule 3: Electro-neutrality is sought in the operation of

membrane co-transporters
64
 Macrominerals
Na+, K+, Cl-, HPO4-, Mg2+, Ca2+
• The macro-minerals for the most part rely on diffusion
controlled mechanisms combined with specific channel
proteins to pass into the system.

• Gradients across the membrane can be driven by

unidirectional and bidirectional ATPase enzymes

Example
Na+/K+ ATPase
65
Ca2+/H+ ATPase
Properties of Macro-minerals Relative to
Absorption
1. Monovalent ions exist mostly as free ions

2. Monovalent ions are unable to form stable complexes

3. Divalent ions exist partially as free ions

4. Divalent ions are more apt to form complexes with

proteins and organics

5. Complexes exist mainly as free ions

66
 Absorption of Sodium and Chloride
Blood Apical (lumen) side
Na+
Glucose Glucose cotransporter
Na+/K+ ATPase

Amino acids Amino acid transporter

3Na +
ATP
2K+ ase
Na+ Na+/H+ antitporter
H+ H+
Carbonic anhydrase
H2CO3 H+ + HCO3- HCO3-
H2O Anion antiporter
CO2 CO2 Cl-

Intestinal Enterocyte 67
Calcium and Magnesium

68
Duodenum Vitamin D deficient rats
No Vit D Vit D ( 1,25-(OH)2-D3)
100 100

Calcium
Absorbed Non-Saturable

50 50

Saturable

0 0
0 0
100 200 100 200

Dietary Calcium Dietary Calcium

69
 Duodenum Jejunum Ileum
100

Non-saturable
50

Saturable Saturable

0 0 0
0 0 0
100 200 100 200 100 200

Calcium Instilled, mM
Uptake in ileum is by diffusion only; it is, therefore, not regulated
by vitamin D. Thus, most of the Ca2+ is absorbed in the 70
duodenum.
• Our best understanding is that calcium enters the
duodenal cell through calcium channels which may
contain a vitamin D responsive Ca-binding component.
Entry is down an electrochemical gradient( Bonner, 1999)

• CaT1, a Ca channel protein in the brush border of human

enterocyte, is regulated by 1,25-dihydroxyvitamin D.
• The vitamin appears to mediate changes in CaT1-mRNA
levels. CaT1, therefore, could be the primary gatekeeper
regulating homeostatic modulation of intestinal calcium
absorption efficiency.

71
 Calcium Absorption
Lumen Blood
Vitamin D Calcium ATPase
Enterocyte
responsive
Calbindin ATP
CAT1 ase Ca2+
Calcium ATPase
antiporter
Ca2+ Ca2+ Ca2+ ATP
ase Ca2+
Mg2+
(Na+) Albumin

Paracellular
Ca2+

Ca bound to
fiber, phytate, CAT1 is a Ca2+ channel protein located in the brush border of mucosal cells
oxalate, fatty
acids
Calbindin is a small (9 kD) protein in the cytosol of mucosal cells
72
 Magnesium
1. Absorption depends on concentration

Human Study
Fed Fractional Absorption

7 mg 65-75%

36 mg 11-14%

2. Absorption is saturable and non-saturable (7-10%)

3. Fully saturable in ileum but not jejunum (contrast with calcium)

4. Absorption in the colon significant

73
Cation channel protein Magnesium
(transient receptor
protein TRP)
Distal jejunum and ileum

TRPM6

ATP
Mg2+ Mg2+ ATP Mg2+
ase

ADP

Mg2+ -bound to
phytate, fiber, Since TRPM6 operates by
fatty acids Enterocyte diffusion without co-
transporters, Mg2+
absorption efficiency
depends on the amount
of Mg2+ in the diet and
within the cell

74
 Microminerals
Fe , Cu , Mn ,
2+ 2+ 2+

Zn 2+
Because of their very low cellular concentrations, the
micronutrients rely on specific high affinity transporters
and binding proteins for movement. Some collect in
vesicles and use the vesicle as the transport factor.

Redox-sensitive metals (Fe2+/Fe3+, Cu+/Cu2+) rely on

valence state changes to be sequestered or transported
from the cell.

75
Metals such as Fe3+ and Zn2+ are more soluble in acid
solutions due to a shift in the equilibrium towards the free
ion

Fe(OH)3(s) Fe3+(aq) + 3OH-(aq) Pulls equilibria

Zn(OH)2(s) Zn2+(aq) + 2OH-(aq) H+

Solubility Fe(OH)3 solubility

Zn(OH)2 solubility

1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0

pH 76
 Elements of Micromineral Absorption
• Insolubility or iron and zinc is partially overcome by
mucins secreted from the cells

• Fe3+ and Cu+ can engage their respective transporters

• Cytosolic sequestering and regulatory factors have the

potential to lock the mineral within the cell and block its
release

• Internal movement of Zn2+, Cu+ and Fe3+ is primarily via

vesicles

• Basolateral surface release is redox sensitive for Fe and Cu

77
Iron regulation

78
Iron metabolism

79
 Mucins
• Mucins are complex polysaccharides secreted into
the lumen that assist in stabilizing the solubility of
metal ions

• Mucins prevent alkaline-induced polymerization of

category 2 metal ions and make the metal ion available
to transporters on the enterocyte surface

80
Mineral Bioavailability

81
 Methods of
Assessing
Bioavailability
82
1. Depletion-Repletion Techniques
• In the “Sheffield experiment” conducted in the 1940s, in
which male participants were conscientious objectors to
military service, 16 (2 women and 14 men) subjects
consumed a vitamin A–deficient diet and 7 control
subjects (1 woman and 6 men) consumed the same diet
but with additional supplements of retinol or B-carotene
from various sources for periods ranging from 8.5 to 25
mo.

83
 Depletion-Repletion…
• Dark adaptation was used as a functional indicator of
bioefficacy.

• In the control group, either 750 µg retinol/d (n = 2) or

3000 µg B-carotene/d in oil (n = 4) or in margarine (n =
2) prevented impaired dark adaptation.

• In the 16 subjects fed the deficient diet, only 3 men,

who also had the lowest plasma retinol concentrations,
developed impaired dark adaptation (depletion phase
of the study).

84
 Depletion-repletion…
• Either 390 µg retinol/d (n = 1) or 1500µg B-carotene/d in
oil (n = 2) was required to reverse this impaired dark
adaptation (repletion phase).
• Other depletion-repletion studies conducted by Booher
et al (15; n = 5) and by Wagner (16; n = 10) shortly before
World War II reached similar conclusions.

• Whereas in the 1970s Sauberlich et al (17; n = 8)

concluded that, on a weight basis, only twice as much B-
carotene as retinol was required to reverse impaired dark
adaptation.
85
 2. Balance studies
• In oral-fecal balance techniques, the difference between
the amount of -carotene in food consumed and that
excreted in feces is assumed to represent the amount of
nutrient absorbed(eg.B-cartotene).

• With these techniques, gastric or bacterial degradation

of unabsorbed carotenoids may contribute to
overestimation of absorption.

• Rao and Rao found indications that such bacterial

degradation occurs .
86
 Balance study…
• On the other hand, endogenously secreted betacarotene
might be excreted in feces, thus leading to an
underestimate of bioavailability.

• Thus, data on the bioavailability of carotenoids obtained

with balance techniques should be interpreted with care.

87
 3. Radioisotopes
• Nearly all of the early work characterizing body iron pools
and iron bioavailability was undertaken using iron
radioisotopes specifically 55Fe and 59Fe
• Radioactive isotopes have some advantages in human
studies in that they
– require a very small, essentially ‘mass free’,
– amount of iron label and the instrumentation needed
for measurement of iron radioisotopes in biological
samples is found in many research facilities.
– An additional advantage of the gamma emitting 59Fe
radioactive isotope is that whole body counting can be
used to measure the fraction of iron isotope that is
88
retained in the body after ingestion.
 Radioisotopes…
• Disadvantages to the use of radioisotopes include
the fact that they have finite half-lives.
– This may introduce time limitations for shipping isotopes to
isolated settings, storing isotopes before the study is
implemented
– and collecting and analyzing the biological samples post-dosing
• Typical radiation exposures from radioactive iron
isotopes

89
 4. Stable Isotopes
• The potential risk and increased reluctance to use
radioisotopes has stimulated the use of stable isotopes.
• Thus, in recent years, stable isotopes and compounds
labeled with stable isotopes have become increasingly
available.
• Basically, the methodology to assess iron bioavailability
involves the administration of single or multiple oral
stable iron isotope(s).
• Iron absorption is then estimated using one of three
approaches

90
• A metabolic balance study can be undertaken to recover
the amount of oral stable iron isotope(s) excreted in
faeces (faecal recovery method).

• (b)A plasma sample can be obtained several hours post-

dosing to assess plasma appearance kinetics (plasma
appearance method).

• (c)A blood sample can be collected 2 weeks post-dosing to

assess the amount of stable iron isotope(s) incorporated
into RBCs (erythrocyte iron incorporation method).
91
• Administration of stable iron isotopes does not have any
known adverse risks at the doses typically used in human
studies.

• All three minor abundance stable iron isotopes can be

given to the same subject to compare bioavailability of
various foods or to allow for simultaneous administration
of oral and intravenous isotopes.

• Multiple stable isotopes of different minerals can also be

given to the same individual to examine interactions of
iron with other nutrients such as calcium or zinc
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• Stable-isotope-tracer techniques were successfully
developed for studying the metabolism of minerals, such
as iron, magnesium, and zinc, and of vitamins and their
precursors, such as folate and provitamin A carotenoids.

• Because of the limited availability of organic nutrients

labeled with stable isotopes and the difficulty in their
quantification, few studies have been carried out so far.

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 5. Algorithm Method
• The algorithm may be used to translate data from dietary
surveys into amounts of a nutrient expected to be
absorbed.

• The main requirement for such calculations is that

detailed information is available about the meal
composition and its variation over a representative and
sufficiently long period of time.
• A 7-d record, for example, may not represent the iron
absorption from the habitual diet.

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• An important use of the algorithm would be to translate
physiologic requirements into dietary requirements
under different dietary conditions known to prevail in a
certain population.

• In the FAO recommendations, 3 levels of bioavailability

(5%, 10%, and 15%) were used arbitrarily for this
translation.

• The validity of choices of representative bioavailability

values can be tested by using the algorithm.

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• The algorithm may be useful in the future search for
realistic recommendations to be used in food based
strategies to improve iron nutrition in developing
countries.
• Challenges
– Estimation of Phytate
– Estimation of vitamin C

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Thank you!

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