SPECIAL COMMUNICATION
Whole body protein kinetics using Phe and Tyr tracers: an evaluation
of the accuracy of approximated flux values
Kevin R.
Short1,
Shon E.
Meek1,
Niels
Moller1,
Karin
Ekberg2, and
K. Sreekumaran
Nair1
1 Endocrinology Research Unit,
Mayo Clinic, Rochester, Minnesota 55905; and
2 Clinical Physiology, Karolinska
Institute, S-17177 Stockholm, Sweden
 |
ABSTRACT |
Phenylalanine
(Phe) kinetics are increasingly used in studies of amino acid kinetics,
because the metabolic fate of Phe is limited to incorporation into
protein (protein synthesis, Sp) and catabolism via hydroxylation
(Qpt) to
tyrosine (Tyr). Besides an infusion of labeled Phe to measure Phe flux
(Qp), a priming dose of Tyr and an independent Tyr tracer are used to measure Tyr flux
(Qt) and
Qpt.
Alternatively,
Qt,
Qpt, and
Sp can be approximated by using
equations, based on Phe and Tyr concentrations in body proteins, that
eliminate the need for a Tyr tracer. To evaluate the accuracy of this
approach, data were obtained from 12 type I diabetic patients and 24 nondiabetic control subjects who were studied with the full complement
of tracers both with and without insulin infusion.
Sp approximations closely matched
measured values in both groups (mean difference <2%, all values
<5%), but the agreement was poor for
Qpt (error range =
8 to +43%) and
Qt (error range
22 to +41%). Insulin status had no effect on these comparisons. The lower approximation error for
Sp vs.
Qpt is due to the
small contribution (~10%) of
Qpt to
Qp. Approximation
error for Qpt (r > 0.99) can be explained by
variability in the ratio of Tyr to Phe coming from protein breakdown,
(Qt
Qpt)/Qp.
Ideally, all fluxes should be directly measured, but these data suggest that whole body Sp can be
approximated with an acceptably small margin of error. However, the
same equations do not yield reliably accurate values for
Qpt or
Qt.
protein metabolism; stable isotopes
 |
INTRODUCTION |
THE USE OF STABLE ISOTOPES has become an invaluable
tool for measuring whole body protein synthesis rates in humans. Among the useful tracers for such studies is isotopically labeled
phenylalanine (Phe), as proposed by Clarke and Bier (3) and further
developed by Thompson et al. (16). Phe is an ideal tracer because it is not synthesized endogenously and has only two fates in the body: incorporation into protein (protein synthesis,
Sp) and conversion to tyrosine
(Tyr) through hydroxylation. As described by Thompson et al.,
Phe-to-Tyr hydroxylation
(Qpt) can be
determined from 1) incremental Tyr
enrichment derived from a Phe tracer during an infusion of labeled Phe
and 2) Tyr flux
(Qt) by use of
an independent Tyr tracer. Both Phe and Tyr enrichment in plasma can be
determined by use of established gas chromatography-mass spectrometry methods.
As an alternative to directly measuring
Qt and
Qpt, Thompson et
al. (16) developed equations to approximate both of these parameters
and, therefore, whole body Sp.
Using this indirect approach does not require the independent Tyr
tracer infusion. It was originally proposed that excluding the Tyr
infusion had both practical and financial benefits (16). From a
practical standpoint, tyrosine's low solubility can necessitate large
infusion volumes, which may be difficult in some patients, i.e.,
children or subjects with renal complications. At least four
investigations involving newborns or young children have used the
approximation equations, presumably for these reasons (4, 7, 15, 17). However, Tyr infusions have been successfully performed in premature infants (5). In many clinical conditions, such as renal and cardiac
failure, however, there is obvious advantage if the volume of the
infusion can be restricted.
Regardless of the reason for using the approximations, the method has
yet to be adequately tested for its validity. The assumptions of the
model are that Phe and Tyr appear only from protein breakdown in the
fasted state and that the ratio of their appearance should resemble the
proportional content of Tyr and Phe in whole body proteins. For their
calculations, Thompson et al. (16) selected a Tyr-to-Phe ratio of 0.73 on the basis of animal studies (10). The validity of their
approximation equations was originally demonstrated in four healthy
adults (16). In this small group approximated and measured averages
differed by only 1% for both
Qpt and
Sp. Subsequent comparisons have
been less consistent, however. In six type I diabetic patients,
approximated Sp was only 5% lower than measured, but
Qpt
approximations averaged 51% higher (14). Likewise,
Qpt
approximations in six premature infants ranged from 11% under to 25%
over the measured values (15). In contrast, less than 2% variation
between the approximated and measured parameters was reported in three
pediatric cancer patients (4).
With the increasing use of labeled Phe in human metabolic studies, it
is important that the models and equations employed are rigorously
tested to determine the validity of the approximation equations.
Previous reports raise the possibility that use of approximations could
produce variable results. It is uncertain whether the variability in
these earlier studies is due to shortcomings in the model and
equations, limited sample sizes, or differences in the
metabolic state of the subjects studied. The current study was
performed with these concerns in mind. We have evaluated the accuracy
of the indirect method for determining whole body Phe kinetics by using
a much larger study population than has previously been reported.
Furthermore, the data were obtained from studies of patients with type
I diabetes and nondiabetic control subjects, each studied with and
without administration of exogenous insulin, so that the influence of
metabolic status could be assessed.
 |
METHODS |
Materials.
L-[15N]Phe,
L-[ring-2H5]Phe,
L-[ring-2H2]Tyr,
and
L-[ring-2H4]Tyr
were purchased from Cambridge Isotope Laboratories (Andover, MA).
L-[15N]Tyr
and additional
L-[ring-2H4]Tyr
were purchased from Isotec (Miamisburg, OH). All isotopes were 99 atom
percent excess. The chemical, isotopic, and optical purities of these
compounds were confirmed before use. Sterile solutions were prepared
and shown to be bacteria and pyrogen free before use in human subjects.
Subjects.
Data from 72 studies were obtained from a total of 36 men and women who
provided informed written consent to participate in three different
institutionally approved protocols (Table
1). Subjects were classified as either
having type I diabetes or as being nondiabetic controls. All diabetic
volunteers were studied in the insulin-treated and insulin-deprived
states on separate occasions at least 3 wk apart. Results from six
members of the diabetic group (referred to here as
subgroup A) have already been published (12). Data from the other six diabetic subjects, referred to
as subgroup B, have not previously
been published. The control group was composed of 24 young healthy men
and women. Control studies were performed in two phases (basal and
insulin infusion) on a single day, as described below. Additional
results from the control group appear elsewhere (9).
Protocol.
For 3 days preceding each study, participants received a
weight-maintaining diet. All subjects were admitted to the hospital 24-48 h before each investigation. No food or caloric beverages were consumed after the evening meal the night before the study. All of
the type I diabetic subjects were switched from long- or intermediate-acting insulin to regular insulin 72 h before the study to
avoid the carry-over effects of the longer-acting insulin (12). Their
subcutaneous regular insulin injections were discontinued at 6:00 PM,
and either saline or insulin was infused overnight and throughout the
duration of the protein turnover measurements on the following morning.
During insulin treatment, insulin dosage was regularly adjusted to
maintain blood glucose within the normal range.
Isotope infusions were started at ~7:00 AM via forearm vein
catheters. Members of the control group and diabetic
subgroup B received primed, continuous
infusions of
L-[15N]Phe
(3.9 µmol · kg
1 · h
1,
3.9 µmol/kg prime) and
L-[ring-2H4]Tyr
(2.9 µmol · kg
1 · h
1,
2.9 µmol/kg prime), and a priming dose of
L-[15N]Tyr
(1.4 µmol/kg). Diabetic subgroup A
received primed, continuous infusions of
L-[ring-2H5]Phe
(4.6 µmol · kg
1 · h
1,
4.6 µmol/kg prime) and
L-[ring-2H2]Tyr
(3.5 µmol · kg
1 · h
1,
3.5 µmol/kg prime), and a priming dose of
L-[ring-2H4]Tyr
(1.5 µmol/kg) (12). Isotope infusions were maintained for up to 5 h.
Arterial blood samples were collected from either brachial or femoral
lines, immediately before the isotope infusion was started and then at
10- to 20-min intervals after isotopic plateau had been reached
(2-3 h after start of the infusion). The protocol for controls
included a second study phase on the same day. Immediately on
conclusion of the basal period, insulin was infused at either 0 (saline), 0.25, 0.50, or 1.00 mU · kg body
weight
1 · h
1
for 2.5 h (9). Six subjects were assigned to each insulin dose. Four
arterial blood samples were collected at 10-min intervals beginning 2 h
after the start of insulin in this group.
Sample analysis and calculations.
Plasma enrichment of Phe and Tyr were determined using gas
chromatography-mass spectrometry, as described (9).
M-4 mass abundance of
[2H2]Tyr
was adjusted for m-2 mass distribution
by use of a matrix correction. The reproducibility and stability of the
measurements were determined for each isotope measured. Interassay
coefficients of variation, determined from replicates of a
representative plasma sample, were between 3.8 and 6.0%. Isotopically
enriched standards were used to calculate the intra-assay coefficients
of variation, which ranged from 2.7 to 6.7%.
Whole body kinetics for Phe and Tyr were calculated using the equations
described by Thompson et al. (16) and outlined below. Infusion and flux
rate units are micromoles per kilogram per hour. The rates of flux
(Q) of Phe and Tyr (measured) were
obtained from isotope dilution
|
(1)
|
where
i is the rate of tracer infusion and
Einfusate and
Eplasma correspond to the
enrichments of infusate and plasma amino acids, respectively.
Conversion rate of Phe to Tyr
(Qpt) in the measured model was calculated as
|
(2)
|
where
Qt and
Qp are the flux
rates for Tyr
([2H4]Tyr
or
[2H2]Tyr)
and labeled Phe, respectively. Et
and Ep are the Tyr
([15N]Tyr or
[2H4]Tyr)
and labeled Phe enrichments in plasma, respectively, and ip is the infusion rate of the Phe tracer.
The approximations of
Qt and
Qpt were obtained
from the following equations
|
(3)
|
|
(4)
|
where
Pt and
Pp refer to the protein contents
of Tyr and Phe, respectively, and the other quantities are as
previously described. Equation 3 is a
mathematical combination of Eqs. 2 and 4. The molar ratio of
Pt to
Pp, according to Thompson et al.
(16), was assumed to be 0.73 on the basis of measurements in animal studies (10). When
Qt was calculated
with Eq. 4, the approximated value for
Qpt
(Eq. 3) was used.
The rate of whole body incorporation of Phe into proteins,
Sp, was calculated as the
difference between
Qp and
Qpt. Measured and
approximated Sp values were
determined by using either the measured or approximated
Qpt, respectively.
To determine the agreement of approximated values with measured values,
comparisons between means of each treatment were first performed, as
shown in Figs. 1-3. The graphic method of Bland and Altman (1) was
also used, with slight modification.
Differences between measured and approximated values were plotted against the
measured values in Figs.
4-6, with the assumption that the measured value was accurately determined.
Approximation error was calculated as the percent deviation of the
approximated value from the measured value. Analysis of variance was
used for comparisons within and between groups. Pearson product
correlations were used to determine the strength of association for
selected variables. Significant effect for all tests was accepted at
P < 0.05.

View larger version (14K):
[in this window]
[in a new window]
|
Fig. 1.
Comparison of protein synthesis
(Sp) derived from measured
(solid bars) and approximated (open bars) equations. Data are means ± SE. Control subjects were studied in basal phase
(n = 24) and then again after infusion
of saline [I(0)] or insulin at 0.25, 0.5, or 1.0 mU · kg 1 · min 1
[I(0.25), I(0.5), or I(1.0)]. Type I diabetics were studied
during well-controlled insulin replacement [I(+)] and
during insulin withdrawal [I( )]. * Significant
difference between basal and insulin in controls or from I( ) in
diabetics.
|
|

View larger version (14K):
[in this window]
[in a new window]
|
Fig. 2.
Comparison of phenylalanine hydroxylation
(Qpt) derived
from measured (solid bars) and approximated (open bars) equations.
Subject groups and treatments are the same as in Fig. 1.
* Significant difference between basal and insulin in controls;
significant difference from measured value.
|
|

View larger version (14K):
[in this window]
[in a new window]
|
Fig. 3.
Comparison of tyrosine flux
(Qt) derived
from measured (solid bars) and approximated (open bars) equations.
Subject groups and treatments are the same as in Fig. 1.
* Significant difference between basal and insulin in controls or
from I( ) in diabetics; significant difference from
measured value.
|
|

View larger version (13K):
[in this window]
[in a new window]
|
Fig. 4.
Difference between measured and approximated
Sp plotted against measured
Sp value. Dotted horizontal lines,
mean difference; solid lines, mean ± 2 SD.
A: control subjects studied during
basal conditions; B: control subjects
studied after saline ( ) or insulin infusion [0.25 ( ), 0.5 ( ), or 1.0 ( )
mU · kg 1 · h 1];
C: type I diabetic patients studied
under insulin-treated (subgroup A,
; subgroup B, ) and
insulin-deprived (subgroup A, ;
subgroup B, ) conditions.
Horizontal lines in B and
C, means of pooled results of
conditions, since there were no differences between them.
|
|

View larger version (13K):
[in this window]
[in a new window]
|
Fig. 5.
Difference between measured and approximated values for Phe-to-Tyr
hydroxylation
(Qpt) plotted
against measured
Qpt value.
Symbols, subjects, and treatments are the same as in Fig. 4.
|
|

View larger version (13K):
[in this window]
[in a new window]
|
Fig. 6.
Difference between measured and approximated Tyr flux
(Qt) plotted
against measured
Qt value.
Symbols, subjects, and treatments are the same as in Fig. 4.
|
|
 |
RESULTS |
Isotopic enrichment values are shown in Table
2. All subjects studied achieved isotopic
plateau for plasma amino acid enrichments during the study period (9,
12). To increase the sample size in the diabetic group, data from
subgroups A and
B were pooled, despite the fact that
they received differentially labeled
(15N or deuterated) Phe and Tyr
tracers as described in METHODS. Data
pooling was performed only after it was confirmed that there were no
systematic differences in any of the outcome variables of interest
between the two data sets. For the sake of clarity, no distinction
between subgroups A and
B is made in the remainder of the
text. However, to demonstrate the overlap of the results, different
symbols are used in Figs. 4-6 to denote the two diabetic subgroups. Agreement between
[15N]Phe and
[2H5]Phe
kinetics has been demonstrated previously by others (8).
As previously reported and shown in Fig. 1, insulin infusion decreased
Sp in both the diabetic (12) and
control groups (9). A slight dose-dependent effect was evident in
controls receiving insulin, but this was not statistically significant.
The agreement between measured and approximated
Sp was not affected by insulin status (see Fig. 4 and Table 3).
Correlational analysis indicated excellent agreement between measured
and approximated Sp values in both
the control (r = 0.99) and diabetic
(r = 0.98) groups. Approximated values
tended to be lower than measured
Sp, especially in the control
subjects, but the average error for the
Sp approximations was <2%, and
the range was 4.8% below to 1.1% above the measured value (Table 3).
However, Fig. 4 reveals a tendency for greater separation between
measured and approximated values as measured values increase.
Agreement between the direct and indirect models was not as strong for
Qpt as it was for
Sp. Exogenous insulin reduced
Qpt in controls
but not diabetics (Fig. 2), although there was no systematic effect of
insulin on the relationship between measured and approximated values
(Fig. 5 and Table 3). The correlation coefficients between the measured
and approximated
Qpt values were
high in each subject group and treatment
(r = 0.89). By comparing Figs. 4 and
5, it is clear that the absolute differences between measured and
approximated Qpt
are the same as for Sp (±2
µmol · kg
1 · h
1).
The percent error of the approximations is an order of magnitude higher
for Qpt than
Sp, however. The range of
Qpt approximation error in all subjects was
8% to +43%, with all of the control data and one-half of the diabetic data higher than measured. Unlike Sp, the differences between
measured and approximated
Qpt were evenly
distributed (Fig. 5).
Diabetics and controls both had reductions in Tyr flux in response to
the insulin administration (Fig. 3). Again, there was no effect of
insulin on the agreement between measured and approximated values (Fig.
6, Table 3). Correlations between approximated and measured values were
lower than for Sp or
Qpt
(r < 0.77, controls; r < 0.45, diabetics). The range of
error was
22% to +41%, and the approximated values were most
likely to be overestimates of the measured values (Table 3). Mean
differences between measured and approximated
Qt were
statistically significant in controls but not diabetics.
Flux ratios are shown in Table 4.
Qpt and
Qt data for these
calculations were obtained from the measured model with all three of
the Phe and Tyr isotopes. The ratio of
Qpt to
Qp indicates the
relative contribution of Phe hydroxylation to total
Qp. It can be
seen that Qpt
accounted for 9-11% of total
Qp in all cases, with no differences between groups or treatments. The ratio
(Qt
Qpt)/Qp
in Table 4 represents the ratio of Tyr and Phe flux coming from whole
body protein breakdown (16). Theoretically, this ratio should
approximate 0.73, which is the value used for Pt/Pp
in Eqs. 3 and 4. The results show that the control
group value was significantly lower than 0.73 in both study phases. Diabetic subjects were evenly distributed above and below the 0.73 value, although the average was <0.73 in both conditions.
When the measured
(Qt
Qpt)/Qp
ratios were plotted against the
Qpt approximation
errors (Fig. 7), a wide range of
(Qt
Qpt)/Qp values (0.52-0.81) was evident, and the relationship between the two variables was very strong (r > 0.99). In Fig. 7 all 72 data points are pooled to illustrate that a
single line can be used to describe the relationship between
(Qt
Qpt)/Qp
and the Qpt approximation error, regardless of metabolic status. The line of best
fit was slightly curvilinear, as denoted by the equation (y = 266x2
527x + 244) in Fig. 7.

View larger version (19K):
[in this window]
[in a new window]
|
Fig. 7.
Distribution of error for approximated Phe hydroxylation
(Qpt) in
relation to Tyr and/or Phe flux values adjusted for
Qpt,
(Qt Qpt)/Qp.
Data have been pooled from all subjects and treatments, and line of
best fit is shown with formula. Dotted lines highlight the ratio of
whole body protein contents of Tyr and Phe
(Pt/Pp;
0.73, x-axis) used in equation to
estimate Qpt and
level of zero estimation error (y-axis).
|
|
 |
DISCUSSION |
The purpose of the current study was to determine the accuracy of the
approximation equations for whole body Phe kinetics by use of a
previously developed model. Several studies in the literature have used
the approximation equations (4, 6, 7, 15, 17), but to date a limited
amount of data has been presented that addresses their accuracy. For
the purpose of analysis, we have evaluated the approximated values by
using the measured values as a benchmark, assuming that the measured
values are accurate.
The results show that whole body
Sp approximations differed from
measured values by <2%, or <0.75
µmol · kg
1 · h
1,
on average. The error was biased toward underapproximation in all of
the control subjects. Values for subjects with diabetes, however, were
equally under- and overapproximated. There was also a slight tendency
for the size of error to increase with the absolute value of
Sp. In practical terms, however,
the discrepancies between measured and approximated values are
negligible. Figure 1 demonstrates that the variability among subjects
within groups is typically equal to or greater than that of the
approximation errors. A natural concern when the indirect model of Phe
kinetics is used is that agreement between the measured and
approximated outcomes could change among treatments or patient groups.
Our data show no effect of insulin on accuracy of approximations in
either the group with type I diabetes or the group without diabetes.
There was a small difference in average approximation error between the
diabetic and nondiabetic groups, but, again, in practical terms this
difference should not prevent the identification of physiologically
important effects in similar patients. Other metabolic states or
treatment conditions need to be examined, but in general these results
suggest that in studies in which whole body
Sp is the variable of interest, the indirect model provides acceptable results.
In contrast, the approximation equations provided unacceptably variable
results for Qpt
and Qt. As shown
in Table 3, the error rates for these two parameters were 10-fold
higher than for Sp. The indirect
model produced
Qpt values that
were significantly higher than the measured approach, which in turn
caused the underapproximation of
Sp.
Sp is calculated as the difference
between Qp and
Qpt, so any error
associated with approximating
Qpt will be
reflected in the Sp value.
However, Qpt was
only 10% of the total
Qp, so it makes
only a minor contribution in the final
Sp calculation. Other authors tend
to report higher hydroxylation rates, i.e., 15-25% of
Qp (2-4,
13-16), so there is potential for
Qpt approximation error to make a greater impact on
Sp, but the contribution should still remain fairly small. Approximations of
Qt, on the other hand, give the same relative error as
Qpt because of
the method of calculation. In absolute terms,
Qt approximations
were as much as 8 µmol · kg
1 · min
1
above to 11 µmol · kg
1 · min
1
below the measured values in these subjects, which is too high to be of
use in nearly any study.
A closer look at the
Qpt approximation
error showed that it was clearly related to the ratio of
Qt and
Qp coming from
protein breakdown,
(Qt
Qpt)/Qp
(Fig. 7). When Thompson et al. (16) developed the equation for
estimating Phe hydroxylation (Eq. 3), it was assumed that Tyr and Phe appeared from
catabolism in the same ratio as their relative distributions in whole
body proteins (Pt/Pp = 0.73). The data in Fig. 7 show that this ratio did not accurately fit
the subjects in this study, particularly the control group, because the
majority of values were lower than 0.73.
To derive the 0.73 Pt/Pp
value, Thompson et al. (16) used the whole body protein amino acid
composition from a hen
(Pt/Pp = 0.68) and extrapolated from multiple tissue analyses in rat (0.75)
and pig (0.75), which were published in the monograph of Munro and
Fleck (10). There is a lack of similar data for humans, so most
subsequent authors have opted to use the 0.73 ratio in their
calculations of Phe hydroxylation. However, for their study on Phe
kinetics in infants, Kilani et al. (7) used a ratio of 0.71 to estimate
Phe hydroxylation, citing previous work on the human fetus (18). On
closer examination, however, the ratio appears to have been
miscalculated by those authors (7). The original report of Widdowson et
al. (18) presented the amino acid content of fetal bodies in relative
mass units (g/g N), which would yield a Tyr-to-Phe mass ratio of 0.71. The number that should be used in the flux equations should be the
Tyr-to-Phe molar ratio, which would be 0.64 when the data of Widdowson
et al. (18) are used.
In light of the different ratios available, two important points about
using them in the approximation equations should be made clear. First,
the ratio of amino acids in whole body proteins (Pt/Pp)
is not likely to match the ratio of those amino acids coming from whole
body protein breakdown
(Qt
Qpt)/Qp.
This discrepancy stems from the fact that there are multiple protein
pools in the body that have different amino acid ratios (10), different
pool sizes, and different turnover rates (11, 12). For example, skin
and muscle have a high
Pt/Pp
and large protein pools but slow turnover, whereas liver and gut have a
low
Pt/Pp,
smaller pool size, and high turnover. The
Pt/Pp
estimate greatly oversimplifies the complexity of factors determining
(Qt
Qpt)/Qp.
To further complicate matters, changing physiological conditions could
affect the (Qt
Qpt)/Qp
ratio if some protein pools respond differently than others.
Second, in view of the current results, selecting a "better"
ratio to use in the approximation equations is not really possible. The
main reason is the high variability of
(Qt
Qpt)/Qp
among subjects (range = 0.52-0.81). The cause of this variability
is not apparent. Given the high degree of precision for measuring isotopic enrichment in plasma, the variability is more likely physiological than methodological. Whatever the explanation, this variability among patient groups and studies is common. Pacy and colleagues (13, 14) reported average values between 0.52 and 0.59 irrespective of nutritional state or diabetic status. In three
pediatric cancer patients (4) and four healthy adults (16), the average
ratio was 0.76, whereas in nine healthy infants, the average was 0.88 (2). These differences help explain why previous attempts to validate
the approximation equations have met with mixed success. In the studies
of young cancer patients and healthy adults,
(Qt
Qpt)/Qp
averaged 0.76, and estimate error for Phe hydroxylation was <2% (4,
16). However, in six type I diabetics, the average ratio was 0.53 under
both insulin-treated and insulin-deprived states, leading to an average
hydroxylation estimation error of 51% (14). These results are
compatible with those shown in Fig. 7.
In conclusion, use of the approximation equations for Phe kinetics can
provide reasonably accurate rates of whole body protein synthesis, but
approximation of Phe hydroxylation and Tyr flux is associated with
unacceptably high and variable levels of error. We have demonstrated
that the approximation errors are closely related to the ratio of Tyr
to Phe coming from protein breakdown and that this parameter varies
widely among subjects. It is unclear why that ratio is so different
among subjects, but it underscores the need to use the full measured
model in all studies, if possible.
 |
ACKNOWLEDGEMENTS |
We gratefully acknowledge the skillful technical assistance of G. C. Ford, M. Persson, M. Bigelow, and the Clinical Research staffs of
the Mayo Clinic and the Karolinska Hospital.
 |
FOOTNOTES |
This work was supported in part by National Institutes of Health Grant
DK-41973 and General Clinical Research Center Grant RR-00585.
The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement"
in accordance with 18 U.S.C. §1734 solely to indicate this fact.
Address for reprint requests and other correspondence: K. S. Nair,
Endocrine Research Unit, 5-194 Joseph, Mayo Clinic and Foundation,
Rochester, MN 55905 (E-mail: nair.sree{at}mayo.edu).
Received 25 August 1998; accepted in final form 19 February 1999.
 |
REFERENCES |
1.
Bland, J. M.,
and
D. G. Altman.
Statistical methods for assessing agreement between two methods of clinical measurement.
Lancet
1:
307-310,
1986[Medline].
2.
Clark, S. E.,
C. A. Karn,
J. A. Ahlrichs,
J. Wang,
C. A. Leitch,
E. A. Liechty,
and
S. C. Denne.
Acute changes in leucine and phenylalanine kinetics produced by parenteral nutrition in premature infants.
Pediatr. Res.
41:
568-574,
1997[Abstract].
3.
Clarke, J. T. R.,
and
D. M. Bier.
The conversion of phenylalanine to tyrosine in man. Direct measurement by continuous intravenous tracer infusions of L-[ring-2H5]phenylalanine and L-[1-13C]tyrosine in the postabsorptive state.
Metabolism
31:
999-1005,
1982[Medline].
4.
Daley, S. E.,
A. D. J. Pearson,
A. W. Craft,
J. Kernahan,
R. A. Wyllie,
L. Price,
C. Brock,
C. Hetherington,
D. Halliday,
and
K. Bartlett.
Whole body protein metabolism in children with cancer.
Arch. Dis. Child
75:
273-281,
1996[Abstract].
5.
Denne, S. C.,
C. A. Karn,
J. A. Ahlrichs,
A. R. Dorotheo,
J. Wang,
and
E. A. Liechty.
Proteolysis and phenylalanine hydroxylation in response to parenteral nutrition in extremely premature and normal newborns.
J. Clin. Invest.
97:
746-754,
1996[Abstract/Free Full Text].
6.
Dworzak, F.,
P. Ferrari,
C. Gavazzi,
C. Maiorana,
and
F. Bozzetti.
Effects of cachexia due to cancer on whole body and skeletal muscle protein turnover.
Cancer
82:
42-48,
1998[Medline].
7.
Kilani, R. A.,
F. S. Cole,
and
D. M. Bier.
Phenylalanine hydroxylase activity in preterm infants: is tyrosine a conditionally essentially amino acid?
Am. J. Clin. Nutr.
61:
1218-1223,
1995[Abstract].
8.
Krempf, M.,
R. A. Hoerr,
L. Marks,
and
V. R. Young.
Phenylalanine flux in adult men: estimates with different tracers and routes of administration.
Metabolism
39:
560-562,
1990[Medline].
9.
Meek, S. E.,
M. Persson,
G. C. Ford,
and
K. S. Nair.
Dose effect of insulin on protein dynamics: differential responses of splanchnic and skeletal muscle beds in healthy human subjects.
Diabetes
47:
1824-1835,
1998[Abstract].
10.
Munro, H. N.,
and
A. Fleck.
Analysis of tissues and body fluids for nitrogenous constituents.
In: Mammalian Protein Metabolism, edited by H. N. Munro. New York: Academic, 1969, p. 423-525.
11.
Nair, K. S.
Assessment of protein metabolism in diabetes.
In: Clinical Research in Diabetes and Obesity, Part I: Methods, Assessment, and Metabolic Regulation, edited by B. Draznin,
and R. Rizza. Totowa, NJ: Humana, 1997, p. 137-170.
12.
Nair, K. S.,
G. C. Ford,
K. Ekberg,
E. Fernqvist-Forbes,
and
J. Wahren.
Protein dynamics in whole body and in splanchnic and leg tissues in type I diabetic patients.
J. Clin. Invest.
95:
2926-2937,
1995[Medline].
13.
Pacy, P. J.,
G. M. Price,
D. Halliday,
M. R. Quevedo,
and
D. J. Millward.
Nitrogen homeostasis in man: the diurnal responses of protein synthesis and degradation and amino acid oxidation to diets with increasing protein intakes.
Clin. Sci. (Colch.)
86:
103-118,
1994[Medline].
14.
Pacy, P. J.,
G. N. Thompson,
and
D. Halliday.
Measurement of whole-body protein turnover in insulin-dependent (type 1) diabetic patients during insulin withdrawal and infusion: comparison of [13C]leucine and [2H5]phenylalanine methodologies.
Clin. Sci. (Colch.)
80:
345-352,
1991[Medline].
15.
Shortland, G. J.,
J. H. Walter,
P. J. Fleming,
and
D. Halliday.
Phenylalanine kinetics in sick preterm neonates with respiratory distress syndrome.
Pediatr. Res.
36:
713-718,
1994[Abstract].
16.
Thompson, G. N.,
P. J. Pacy,
H. Merritt,
G. C. Ford,
M. A. Read,
K. N. Cheng,
and
D. Halliday.
Rapid measurement of whole body and forearm protein turnover using a [2H5]phenylalanine model.
Am. J. Physiol.
256 (Endocrinol. Metab. 19):
E631-E639,
1989[Abstract/Free Full Text].
17.
Treacy, E.,
J. J. Pitt,
K. Seller,
G. N. Thompson,
S. Ramus,
and
R. G. H. Cotton.
In vivo disposal of phenylalanine in phenylketonuria: a study of two siblings.
J. Inherited Metab. Dis.
19:
595-602,
1996[Medline].
18.
Widdowson, E. M.,
D. A. Southgate,
and
E. N. Hey.
Body composition of the fetus and infant.
In: Fifth Nutricia Symposium: Nutrition and Metabolism of the Fetus and Infant, edited by H. K. A. Visser. The Hauge: Martinus Nijhoff, 1979, p. 169-177.
Am J Physiol Endocrinol Metab 276(6):E1194-E1200
0002-9513/99 $5.00
Copyright © 1999 the American Physiological Society