Clinical Science (1979). 56,407-412
Absorption of inorganic phosphate in the human small
intestine
J. W A L T O N A N D T. K . G R A Y
Departments of Medicine and Pharmacology, University of North Carolina School of Medicine, Chapel Hill,NC, U S A .
(Received 28 April 1978; accepted 29 September 1978)
Summary
1. Intestinal phosphate absorption in human
subjects was studied by the technique of triple
lumen intestinal perfusion in vivo.
2. Ileal phosphate absorption increased as the
intraluminal
phosphate concentration was
increased.
3. Ileal rates of phosphate absorption were
lower at any given intraluminal phosphate concentration than previously described jejunal rates.
Acidification of the ileal lumen did not increase
phosphate absorption.
4. Phosphate absorption was shown in the
jejunum to be dependent on the intraluminal
sodium concentration.
5. Phosphate absorption in the human small
intestine consists of at least two components, one
directly proportional to water movement and the
second apparently independent of water movement.
Key words: absorption, phosphate, small intestine.
Introduction
The intestinal absorption of inorganic phosphate
has traditionally received less attention than calcium absorption. The recent reports that the active
metabolites of vitamin D stimulated phosphate
absorption in vitamin D-deficient animals have
rekindled interest in phosphate absorption as a
process distinct from calcium absorption (Chen,
Castillo, Korycha-Dahl & DeLuca, 1974; Hurwitz
& Bar, 1972; Kowarski & Schachter, 1969;
Wasserman & Taylor, 1973). Despite this resurgent interest, phosphate absorption in the human
small intestine remains incompletely characterized.
A recent review of this topic pointed out that the
literature contained no studies on the site of
phosphate absorption in man (Wilkinson, 1976).
This deficit is due largely to the fact that the bulk of
existing information is derived from balance studies
which measure net retention rather than absorption
rates at specific sites. We have described the jejunal
absorption of phosphate by using the technique of
triple lumen intestinal perfusion in vivo (Juan,
Liptak & Gray, 1976). This previous report
characterized the rates of jejunal phosphate absorption in normal volunteers during the basal state
and after the intravenous administration of salmon
calcitonin. The studies described in this paper
extend our knowledge of phosphate absorption in
the human small intestine by measuring ileal
phosphate absorption and the effect of changing
the luminal pH or sodium concentration on
intestinal phosphate absorption.
Methods
Net absorption or secretion was studied in 40 cm
segments of the human small intestine by using the
triple lumen perfusion technique which has been
previously described (Cooper, Levitan, Fordtran &
Ingelfinger, 1966; Fordtran, 1966). Volunteers
Correspondence: Dr T. Kenney Gray, Box 501, North
Carolina Memorial Hospital, Chapel Hill, NC 275 14,
U.S.A.
407
408
J. Walton and T.K. Gray
were healthy young adults aged 21-29 years. This
technique involves the pumping of a test solution at
a constant rate into the intestinal lumen at a known
site, the infusion point, and the constant aspiration
of luminal fluid from two sites 10 and 50 cm distal
to the infusion point. These latter sites are called
the proximal and distal aspiration points respectively. Jejunal perfusions began after the distal
aspiration site was 120 cm from the volunteer’s lips
and the infusion point was located by fluoroscopy
at the ligament of Treitz. Ileal perfusions began
when the distal aspiration site was 200 cm from the
volunteer’s lips and located by fluoroscopy in the
right lower abdominal quadrant.
Both jejunal and ileal studies used a standard
test solution which contained NaCl (105 mmol/l),
KCl (4 mmol/l), calcium hemigluconate (1.25
mmol/l), NaHCO, (30 mmol/l) and polyethylene
glycol (5 g/l) unless otherwise noted. The polyethylene glycol served as the non-absorbable
volume marker. The phosphate concentration of
the test solution was adjusted from 0 to 10 mmol/l
by the addition of NaH,PO, and Na,HPO, in a
ratio of 9 : 1. Test solutions were pumped at 10
ml/min and luminal fluid was aspirated from the
proximal and distal sites at 1 ml/min. Proximal
aspiration preceded distal aspiration by 20 rnin for
jejunal perfusions and 10 min for ileal perfusions
(Whalen, Harris, Geenen & Soergel, 1966).
Each perfusion consisted of two study periods,
each period being 2 h in duration and including an
equilibration (the first 40-50 rnin) during which
luminal samples were collected and discarded
followed by a 1 h collection of luminal samples for
later analysis. The second period involved a reequilibration (40-50 min) and another 1 h collection. In the studies of ileal phosphate absorption
this protocol allowed measurement of net ileal
phosphate absorption from two test solutions
differing only in phosphate concentration (0-10
mmol/l). The same protocol involving two test
solutions per perfusion was utilized to study the
effect of independently changing two luminal
factors, pH and sodium concentration.
To assess the effect of ileal pH change, the test
solution described above was acidified by the
substitution of equimolar amounts of NaCl for the
NaHCO,. Four subjects undergoing ileal perfusion
received both solutions, bicarbonate-containing
and bicarbonate-free, in random order. The effect
on jejunal phosphate absorption of altering the
luminal sodium concentration was studied in a
similar manner in 16 subjects undergoing jejunal
perfusion. Luminal sodium concentration was
altered by the substitution of choline chloride (53
mmol/l) for equimolar amounts of NaCl, both
solutions being perfused in a random order. In five
of these 16 subjects the two test solutions were
calcium-free, and in four others the solutions were
without phosphate.
Luminal aspirates were centrifuged and the
supernatants were analysed for phosphate, calcium, sodium, potassium, chloride, bicarbonate and
polyethylene glycol. The standard analytical
methods have been described by Juan et al. (1976).
Osmolality of the luminal aspirates was measured
by Advanced Digimatic Osmometer, model 3DII
(Advanced Instruments Inc., Needham Heights,
MA 02194, U.S.A.) and pH of the luminal
aspirates was measured immediately after the study
by using the Beckman Expandomatic pH meter
(model SS2).
Net absorption rates were calculated from the
pumping rate and the concentration of polyethylene glycol and ions in the proximal and distal
luminal aspirates. The calculated rates from two
consecutive 30 min pools of luminal aspirates were
averaged and expressed as a single datum (mmol,
p o l or m1/30 min per 40 cm length). Comparisons
were made by analysis of variance or paired t-test
of the mean difference (Remington & Schork,
1970). Values are expressed as mean & SEM.
Results
Ileal absorption ofphosphate
The ileal absorption of phosphate increased as
the phosphate of the perfusion solution was raised
from 0 to 10 mmolll (Table 1). Increases in
[phosphate] of the perfusion solution did not
significantly change the net absorption of water,
sodium and calcium. Potassium, chloride and
bicarbonate absorption did not change with
increasing [phosphate]. When the solution was
phosphate-free, a slight net secretion of phosphate
was observed. This secretion diminished when 0.25
rnmol/l [phosphate] was perfused, and with
[phosphate] ranging from 0.5 to 10.0 mmol/l net
absorption of phosphate was seen (Table 1).
The phosphate concentration of the perfusion
solution differs from the intralurninal concentration
because the solution mixes with endogenous fluids
before entering the ileal segment. The intraluminal
[phosphate] determined by averaging the values in
the corresponding proximal and distal samples is
the value actually present within the ileal lumen.
Fig. 1 shows the net phosphate absorption rates in
Zntestinalphosphate absorption
409
TABLE1. Effect of phosphate in the perfusion solution on the net movement
( A ) of water, Nu, P and Ca in the human ileum
Net movement of water is expressed as ml 30 min-l 40 cm-', of Na as
mmol 30 min-I 40 cm-I and of P and Ca as jmol 30 min-a 40 cm.
-, Net absorption; +, net secretion. Values are expressed as means & SEM.
n = number of volunteer subjects.
~
~
~~~
~~
AH,O
ANa
APhosphate
ACa
0
n=5
0.25
n=5
0.50
n=7
-70 f 15
-9.9 f 2.2
+7.0 f 5.0
-58 i 18
-71 f 8.0
-9.6 f 1.4
+2.0 f 5.0
-56 f 8.0
-84 f 8.4
-11.9 f 1.3
-28 f 7.0
-83 f 9.0
1.0
-79 f 8.0
-10.5 f 1.1
-59 f 9.0
-73 f 5.0
-83 f 6.0
-11.4 f 1.0
-84 f 12
-86 f 5.0
-76 f 10
-10.9 f 1.4
-295 f 38
-75 f 13
-78 f 14
-11.6 f 2.2
-485 t 81
-92 f 30
Phosphate in
solution (mmol/l)
n=7
2.0
n=6
5.0
n=7
10.0
n=5
-
h
1500
-
1000
Mean intralurninal [phosphatel (mmol/l)
FIG. 1. Net absorption of inorganic phosphate in the
human small intestine. Each point represents the mean of
studies in at least five subjects. Bars indicate 1 SEM.
Jejunal data previously reported (Juan et al., 1976). 0 ,
Ileum; 0 ,jejunum.
the ileum together with the jejunal rates reported by
Juan et al. (1976), which have been recalculated to
correspond to the current results plotted against the
mean intraluminal [phosphate]. Ileal perfusion with
the phosphate-free solution was associated with a
mean intraluminal [phosphate] of 0.19 mmol/l and
the net secretion rate of 7 ,umol, the latter
representing the endogenous ileal secretion of
phosphate. At the highest intraluminal [phosphate],
9.58 mmol/l, the ileal rate was 485 pmol. A
comparison of the ileal and jejunal phosphate
absorption rates (Fig. 1) shows that the ileal rates
.
are lower at any given intraluminal [phosphate]
than the jejunal rates. The differences between
these anatomical sites were significant (P < 0.01) at
each lphosphatel tested. Below an intraluminal
[phosphate] of 1.0 mmol/l, both ileal and jejunal
phosphate absorption rose sharply with increasing
intraluminal [phosphate]. Above 1.O mmol/l, the
magnitude of increase in absorption with increasing
intraluminal [phosphate] diminished but saturation
was not reached even at intraluminal values of 10.0
mmol/l (Fig. 1).
The marked differences in phosphate absorption
observed between the ileum and jejunum correspond to similar although smaller differences in the
net absorption of water, sodium and calcium
(Table 2). The ileal absorption of water was 77.6
ml 30 min-I 40 cm-1 (f 3.5) compared with the
reported jejunal rate of 103-4 m l 3 0 min-I 40 cm-1
(f5.0).Since the changes in [phosphate] did not
significantly alter water movement at either site, all
the values were pooled for analysis and the
differences between the ileum and jejunum was
highly significant (P < 0.001). Mean sodium
absorption in the ileum was 10.9 mmol 30 min-I
40 cm-I (f0.53) in contrast with the mean jejunal
rate of 13.4 (f0.65) and the mean calcium
absorption rates for the ileum and jejunum were 73
p o l 30 min-I 40 cm-I (54.0) and 113 p o l 30
min-' 40 cm-1 (k5.0) respectively, all these
differences being significant (P< 0.01).
Ileal perfusion with the standard test solution
containing various amounts of phosphate was
associated with a mean intraluminal pH of 7.37 f
J. Walton and T. K . Gray
410
TABLE2. Comparison of the net absorption of water, sodium and calcium
in the jejunum and the ileum
Values are expressed as means f SEM; n = number of subjects. Data were
pooled from studies at all [phosphate]values tested at both sites. Jejunal data
previously reported (Juan et al., 1976).
Calcium
(pmol30 min-I cm-l)
Sodium
(mmol 30 min-l40 cm-l)
Water
(ml30 min-l40 cm-l)
Ileum
Jejunum
P
73 f 4.0
n=37
113 f 5.0
n=42
<0.01
10.9 -I 0.53
n=42
13.4 f 0.65
n=41
<0.01
77.6 & 3 . 5
n=42
103.4+ 5.0
n=41
t0.01
TABLE3. Eflect of intraluminal [Nal on the net movement of water
and ions in the jejunum
The plasma-like solution and choline chloride solution were perfused
in random sequence. -, Net absorption; +, net secretion. Values are
expressed as means ~ S E M , n = number of volunteer subjects.
*P (0.005 for differences by paired t-test; **P(0.001 for differences
by paired t-test.
Intraluminal “a1 (mmoV1). . .
Phosphate
@mo1/40 cm)
n = 12
Calcium
@mol/40 cm)
n = 11
Sodium
(mmol/40 cm)
n = 16
Water
(m1/40 cm)
n = 16
0.04. In contrast, the mean intraluminal pH in the
jejunum was 6-79 i-0.03 during perfusion with the
same solution. When the ileum was perfused with
the test solution lacking bicarbonate, the mean ileal
pH was 6.65 f 0.06. No significant change in the
ileal absorption rates of water, sodium phosphate
and calcium occurred despite this manipulation of
the ileal pH to the jejunal range.
Jejunal phosphate absorption and intraluminal
sodium concentration
Table 3 shows the net jejunal absorption rates
during perfusion with the siandard test solution and
the solution in which 53 mmol of choline chloride
was substituted iso-osmotically for NaCl. The mean
138
-291
+ 40
-101
+ 10
93
-193 f 30.
-40 f 10”
-8.9 f 4.4
-3.7 5 2.0**
-91 + 9
+11 f 14**
intraluminal “a] was 138 & 5 mmol/l when the
perfusate was the standard test solution and 93 & 5
mmol/l when choline chloride was substituted. N o
significant change in intraluminal [phosphate]
occurred. The fall in intraluminal “a] was
associated with lowered jejunal rates of water, Na,
phosphate and calcium absorption compared with
the rates observed when intraluminal “a1 was 138
mmol/l. Net jejunal phosphate absorption was
reduced from 291 pmol 30 min-’ 40 cm-1 to 193
pmol whereas net calcium absorption fell from 101
to 40 p o l 30 min-’ 40 cm-I. Net water
absorption seen with the standard test solution was
converted into net secretion during perfusion with
the choline chloride solution. All these differences
in jejunal absorption were highly significant (P <
0.005).
Intestinal phosphate absorption
Discussion
The intestinal absorption of inorganic phosphate
has been reviewed by Wilkinson (1 976), known
factors influencing absorption including the dietary
amount, the anatomical site, the metabolites of
vitamin D, and the endogenous secretion and other
ions, such as Na+, Ca+ and magnesium, to name
but a few. The role of these factors in humans are
often extrapolated from studies in experimental
animals or derived from balance studies which have
provided the bulk of the information about human
intestinal phosphate absorption. We have focused
on the intestinal absorption of phosphate in
healthy human subjects measured in uiuo by triple
lumen intestinal perfusion, which permits the
anatomical localization of the absorption rates and
control over intraluminal factors such as pH or
ionic composition (Cooper et af., 1966; Fordham,
1966). In previous studies we concentrated on
jejunal phosphate absorption (Juan et al., 1976),
describing the relationship between jejunal phosphate absorption and the intraluminal [phosphate],
the inhibition of phosphate absorption induced by
salmon calcitonin and the positive correlation
between water and phosphate absorption from isoosmotic solutions. We now describe the influence of
intraluminal “a] on jejunal phosphate absorption.
When the intraluminal “a] was 138 mmol/l, water
was absorbed. When the choline chloride solution
decreased the intraluminal “a] to 93 mmol/l,
water was secreted and Na, C a and phosphate
absorption decreased. Under these circumstances
jejunal absorption of water and phosphate were
positively correlated (r = 0.488, P < 0.025). A
positive correlation between jejunal water and
phosphate absorption was observed previously
when jejunal secretion was induced by the administration of salmon calcitonin (Juan et al.,
1976). Linear regression analysis of jejunal water
and phosphate absorption in the previous report
revealed the y-intercept, which represented phosphate absorption at zero water absorption, to be 150
pmol 30 min-I 40 cm-I. When the same analysis
was performed on the jejunal absorption rates from
the standard test solution and the choline chloride
solution, the y-intercept was 180 pmol 30 min-I 40
cm-I. These two comparable values indicate the
rate of jejunal phosphate absorption which was not
due to diffusional movement as a result of solvent
drag from water and sodium absorption and
provide indirect evidence that jejunal phosphate
absorption in the human small intestine is at least a
two component process.
41 1
The extension of our studies in uiuo to the ileum
were performed under the same conditions as the
previously reported jejunal studies so that comparisons between the two anatomical sites could be
made. Ileal phosphate absorption increased as the
intraluminal phosphate was raised by increasing
the phosphate of the perfusion solutions. Nevertheless, at each value the ileal rates were significantly
less than the previously reported jejunal rates
demonstrating clearly the anatomical differences in
phosphate absorption. Ileal absorption of water,
Na and C a was also lower than the previous jejunal
rates. These differences in water and ion absorption
are consistent with other studies of ileal absorption
(Fordtran, Levitan, Bikerman, Burrows &
Ingelfinger, 196 1; Vergne-Marini, Parker, Pak,
Hull, DeLuca & Fordtran, 1976). Although the
ileal lumen was more alkaline than the jejunal
lumen, this pH difference was not an adequate
explanation for the anatomical differences in phosphate absorption since acidification of the ileal
lumen did not increase phosphate absorption.
Intrinsic differences in the jejunal and ileal mucosa
are more likely explanations for these differences.
For example, the pore size of the jejunal mucosa
was estimated from human studies to be larger than
the ileal pore size (Fordtran, Rector, Ewton, Soter
& Kinney, 1965). In rats the absorptive area of
mucosal surface per unit length has been calculated
to be greater in the jejunum than in the ileum
(Fisher & Parsons, 1950).
Rates of phosphate secretion or absorption
during intestinal perfusion with phosphate-free
solutions demonstrate the role of endogenous
phosphate since the intraluminal phosphate in these
instances represented only phosphate secreted into
the lumen proximal to the test segment or in the test
segment. Under such circumstances the mean
jejunal absorption was 20 pmol 30 min-’ 40 cm-’
(+5.0), a net absorption of endogenous phosphate. The ileum secreted phosphate in small
amounts, 7 p n o l 30 min-’ 40 cm-I (+5.0),
another indication of anatomical differences between the two intestinal sites.
Our results support to some degree the concepts
about phosphate absorption derived from studies in
other species. McHardy & Parsons (1956) reported
that jejunal phosphate absorption in the rat was
higher than in the ileum, a linear relationship
between phosphate absorption and intraluminal
phosphate and the dependence of phosphate
absorption on intraluminal “a]. The N a dependency of phosphate absorption was confirmed by
Taylor (1974) who used everted gut sacs prepared
412
J . Walton and T.K.Gray
from rachitic chick ileum, and by Walling (1977)
who used rat jejunal mucosa in the Ussing
chamber. Walling (1977) also noted that active
phosphate transport was maximal at pH 6.0 and
decreased as the pH was raised to 7-0. The linear
relationship between phosphate absorption and
intraluminal phosphate was evident in both the
jejunum and the ileum at values above 1.0 mmol/l.
Below this value the relationship is more complex,
presumably due to the movement of phosphate
down the chemical gradient from blood to the
lumen or the presence of an active transport
component being more obvious at lower intraluminal phosphate. A similar phosphate absorption
curve generated by perfusion studies in a normal
subject has been reported by Wilkinson (1976),
which was interpreted to suggest that phosphate
absorption was probably an active process at low
intraluminal concentrations and passive at higher
concentrations. Our results are entirely consistent
with this hypothesis but not sufficient to prove it.
Acknowledgments
The authors appreciate the typing and clerical
assistance of Robin Mays and Sheila Braxton, and
technical assistance of M. E. Williams and D. Pool.
This investigation was supported by Public Health
Service Research Grant no. RR-46, from the
General Clinical Research Centers Branch of the
Division of Research Resources, and NIH Grant
AM17835 and National Foundation Grant 6-43.
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