Digestion and Absorption of Minerals I

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Digestion and Absorption of Minerals I
(Unifying principles that apply to all minerals)
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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)

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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
unitsminerals 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
Products of the digestate aid in the
solubilization and absorption of minerals
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Digestive Enzymes (hydrolases)
Enzyme Location Target Action
Pepsin gastric juice proteins breaks peptide bonds
Trypsin and chymotrypsin duodenum proteins breaks peptide bonds
Amylases saliva and duodenum starch and glycogen breaks glycosidic bonds
Lipases duodenum complex lipids breaks ester bonds
Glycosidases microvilli di- and tri- saccharides breaks glycosidic bonds
Peptidases microvilli small peptides breaks peptide bonds
I
II
Phase I is primarily salivary and pancreatic
secretions
Phase II involves enzymes on the surface of
absorbing cells
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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
• 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 Ca2 and
  Fe2respectively
• 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

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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
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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.
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Mucosal Side
Fe
A large fraction of the iron can be trapped
(sequestered) within the cytosol of the enterocye)
Fe
Fe
Microvilli
Apical surface
Enterocyte
Basolateral Surface (antiluminal surface)
To access the serosal side, the mineral must pass
either through the enterocyte (transcellular 99)
or the junction between enterocytes (paracellular
lt1))
Serosal Side
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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.
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MACROMINERALS
Monovalent cations, Na, K Monovalent anions,
Cl- Divalent cations, Ca2, Mg2 Complexes,
HPO4, HCO3-
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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 Electroneutrality is sought in the
operation of membrane co-transporters
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Macrominerals
Na, K, Cl-, HPO4-, Mg2, Ca2
The macrominerals 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
Ca2/H ATPase
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Properties of Macrominerals 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
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Absorption of Sodium and Chloride
Apical (lumen) side
Blood
Na
Glucose cotransporter
Glucose
Na/K ATPase
Amino acids
Amino acid transporter
3Na
ATP ase
2K
Na
Na/H antitporter
H
H
Carbonic anhydrase
HCO3-
HCO3-
H2CO3
H
H2O
Anion antiporter
Cl-
CO2
CO2
Intestinal Enterocyte
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Calcium and Magnesium
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Three stages in intestinal absorption at the
cellular level
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Calcium absorption is the sum of saturable and
unsaturable processes
1. Solubility depends on dietary source
2. CaHPO4 is 18 time more soluble than CaCO3
3. Solubility also depends on pH
4. Transcellular and paracellular transport
processes
5. Transcellular proximal intestine saturable,
regulated
6. Paracellular throughout intestine
unsaturable, unregulated
7. Vitamin D is the major regulator of
transcellular calcium entry
8. Calcium channels in brush border and apical
membranes appear to have a vitamin D-sensitive
element
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Everted Sac and Intestinal Loop Technique to
measure Ca2 Absorption
In situ Intestinal Loop
Inverted sac
Absorption is the amount of Ca2 effusing with
time as measured at different concentrations of
Ca2
Transcellular movement of Ca2 into the sac is a
metabolically active process requiring oxygen and
occurs against a concentration gradient.
Absorption is the sum of two processes saturable
and non-saturable
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(No Transcript)
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In situ intestinal loop experiment showing Ca2
absorption cannot be due to simple diffusion, but
is the sum of two processes, saturated and
unsaturated
absorbed of total sac 45Ca that effused out
1 mM
100
10 mM
Total

Abs
25 mM
Unsaturable
100 mM
50
Saturable
200 mM
Total sum of saturated and unsaturated at each
time point
Time
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Vitamin D deficient rats
Duodenum
-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
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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 duodenum.
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Ficks Law of diffusion The rate of diffusion of
an ion at steady-state transmembrane flux varies
inversely with path length and directly with area
and concentration gradient
A 80 ?m2
L 10 ?m
Dca 3 x 10-3 cm2/min
Adolph Fick
after Bronner
Based on Ficks law, the expected diffusion rate
of Ca across the intestinal cell is 96 x 10-18
mol/min/cell.
Rate observed in the laboratory is 70 times
greater at Vmax, which means duodenal cells have
factors that enhance self diffusion of Ca
Possible factor is Calbindin, a small (9 kD)
Ca-binding protein
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Search for the Vitamin D sensitive Factor

1. Calbindin (9 kd cytosolic Ca-binding protein)
2. CaT1 (a calcium channel protein in brush border
   of intestinal cells)

1,25 dihydroxy vitamin D3 given at time 0
increases the expression of CaT1
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Changes in CaT1 mRNA levels with different
amounts of D3
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Take Home
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.
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Calcium Absorption
Blood
Lumen
Calcium ATPase
Enterocyte
Calbindin
ATP
Ca2
CAT1
ase
Calcium ATPase antiporter
Ca2
Ca2
Ca2
ATP
Ca2
ase
Mg2 (Na)
Albumin
Paracellular Ca2
Ca bound to fiber, phytate, oxalate, fatty acids
CAT1 is a Ca2 channel protein located in the
brush border of mucosal cells
Calbindin is a small (9 kD) protein in the
cytosol of mucosal cells
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Unanswered Questions
1. Where exactly is CaT1 located and does
raising CaT1 protein require it relocation to the
absorbing membrane?
2. Is there any evidence for CaT1 location in
mobile vesicles?
3. Does 1,25-dihydroxy vitamin D3 affect efflux
of Ca2 at the basolateral surface?
4. Does CaT1 also recognize Mg2?
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Phosphorus (phosphate)
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Phosphorous
Phosphorous absorption utilizes a Na/phosphate
cotransporter (Npt2a)
1. Expressed in the brush border membrane
2. Saturable, carrier mediated and responsive to
Vit D.
3. non-regulated diffusion may be the major
absorption pathway with higher
intake
Duodenum, Jejunum
Saturable, carrier-mediated
Npt2a
PO4
PO4
Na
PO4
(Ca2, Mg2)
Complexed with other minerals or as organic
phosphate
Vitamin D stimulated
Enterocyte
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Magnesium
1. Absorption depends on concentration
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
5. Vitamin D has no influence on magnesium
absorption
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Magnesium
Cation channel protein (transient receptor
protein TRP)
Distal jejunum and ileum
TRPM6
ATP
Mg2
Mg2
Mg2
ADP
Mg2 -bound to phytate, fiber, fatty acids
Since TRPM6 operates by diffusion without
co-transporters, Mg2 absorption efficiency
depends on the amount of Mg2 in the diet and
within the cell
Enterocyte
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Microminerals
3d metals Fe, Zn, Cu
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Microminerals
Fe2, Cu2, Mn2, Zn2
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.
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Metals such as Fe3 and Zn2 are more soluble in
acid solutions due to a shift in the equilibrium
towards the free ion
Fe3(aq) 3OH-(aq)
Fe(OH)3(s)
Pulls equilibria
H
Zn(OH)2(s)
Zn2(aq) 2OH-(aq)
Solubility
Fe(OH)3 solubility
Zn(OH)2 solubility
2.0
3.0
4.0
5.0
6.0
7.0
8.0
9.0
1.0
pH
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Elements of Micromineral Absorption

• Insolubility or iron and zinc is partially
  overcome by mucins secreted from the cells
• Only 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
• See Powell et al. The regulation of mineral
  absorption in the gastrointestinal tract. Proc.
  Nutr. Soc. 58(1), 147-153 (1999)

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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
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Correlation of spectra of Fe with iron absorption
Importance of mucins in making insoluble iron
available to membrane transporters
Rudzki et al, 1973 Conrad et al, 1991 as cited in
Powell et al, 1999
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Stomach (pyloric mucosa)
Intestine (colon)
Laminated mucous layer
Mucous layer
Mucosal goblet cells
Pyloric mucosal cells
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Aluminum localization with the mucous layer at
rat villi surfaces
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Events in the Cellular Absorption of Iron
Heme Iron
Non-heme iron
Ferric (Fe3) Iron Pathway
Ferrous (Fe2) Iron Pathway
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Three Pathways in Iron Absorption
Fe2 Pathway Divalent cation transporter (DCT-1,
DCM-1,Nramp2)
Fe3 Pathway Mobilferrin-integrin
Heme Pathway
Heme carrier protein
Dctyb reductase
Fe3
Fe2
DCT1
gastroferrin
integrin
Mobilferrin-Fe3
Mobilferrin
Fe2
Porphyrin ring
Ferroportin 1
Hephaestin
Cu
Cu
Cu
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Iron Absorption (heme and non-heme)
Duodenal Lumen
Duodenal Mucosa
Plasma
Heme- Protein Heme Polypeptides
Biliverdin Bilirubin Bilirubin
HFE
CO CO
Heme Oxygenase
Heme
Ferroportin
Fe2
Fe3
Ferritin
Dcytb reductase
paraferrin
Hephaestin
Fe3
FR
DCT-1
Fe2
Fe3
Fe3
Fe3
Mobilferrin (vesicles)
Fe3
Fe3
B3 integrin
Mucin (gastroferrin)
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Nramp2 (Natural resistance associated macrophage
protein)
Nramp2
Nramp1
(no iron transport)
Nramp2
(DMT1/DCT1)
Transport Mn2,Fe2, Ni2
DMT1 isoform 1
DMT1 isoform 2
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Soluble mucins (gastroferrin)
Fe3 reductase
Integrin anchor
Secretions into the lumin (soluble mucins) retard
hydrolysis of Cu, Fe and Zn permitting binding to
transporters and more efficient uptake.
Efficiency of transport is related to valance
state with M gt M2 gt M3
Redox-active factors reduce Fe3 to Fe2
Divalent cation transporter (DCT1) transports M2
metals (Fe2, Ca2,Cu2, Zn2), keeping out toxic
metals such as Al3. A former name of DCT1 is
Nramp2.
Mobileferrin on the inner side of the apical
membrane receives metal from DCT1 and transfers
it to cytosol.
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Mobilferrin
HFE (human leukocyte antigen H)
Ferritin (or paraferritin) or Fe
?2-microglobulin (for Zn)
HFE may be involved in stabilizing the above
complexes to mobiltransferrin