deficiency, the substrate accumulates intracellularly,
undergoes spontaneous hydrolysis, and diffuses out
of the cells, thereby raising the plasma concentration
of methylmalonate. Plasma methylmalonate concen-
tration can therefore be used as a marker of mutase
activity and, in turn, of tissue cobalamin supply
(Nexř
et al.
1994, Chanarin and Metz 1997). As a
rule, enzyme substrates increase in concentration in
plasma and are excreted in the urine at greater than
normal rates as a consequence of micronutrient
deficiency. The
in vivo
activity of micronutrient-
dependent enzymes can also be assessed by measure-
ment of the usual products of the metabolic
transformation that is catalyzed or by measurement
of substances or processes that are metabolically
downstream of the catalytic step. Trace substance
deficiency leads to a reduction in the plasma concen-
tration of the catalysis products and a reduced rate of
urinary excretion of the products. Lastly, direct,
in
vitro
assay of the activity of micronutrient-dependent
enzymes can be undertaken when the enzymes are
present in marrow cells. Enzyme activities
measured
in vitro
decrease in deficiency states.
Nonenzymatic
in vitro
assay of micronutrients in
marrow cells is also possible; the most important
example being the semi-quantitation of erythroblast
iron by light microscopy using iron-specific histo-
logic stains.
Another laboratory approach that can be used to
evaluate trace substance supply is measurement of a
precursor substance in the metabolic pathway of the
trace substance. When a trace substance is not
freely available for incorporation into a product
because of deficiency of the substance, precursors
accumulate intracellularly and, if able to diffuse out
of the cell, they accumulate in the plasma and are
excreted at increased rates in the urine. In the case
of iron, in the later stages of deficiency, when there
is iron-deficient erythropoiesis, protoporphyrin, the
immediate precursor of heme, accumulates in red
cells. Red cell protoporphyrin concentration then
correlates with the severity of the iron deficiency
(Hastka
et al.
1996).
In deficiency states, there is typically a decrease
in the synthesis of the intracellular micronutrient-
binding protein with an associated decline in the
plasma concentration of the protein. In iron
deficiency, decreased synthesis of apoferritin leads
to a fall in the intracellular and plasma concentra-
tions of ferritin. Because it is the protein concentra-
tion of ferritin rather than the ferritin-bound iron
concentration that is measured when ferritin is
assayed in the laboratory, it is the decreased synthe-
sis of apoferritin that underlies the inverse relation-
ship between plasma ferritin concentration and the
status of the iron stores.
Deficiency states can also lead to a number of
physiologic alterations that serve to improve tissue
access to a deficient micronutrient. There may be an
increase in the absorptive capacity of the intestine
due to increased expression of enterocyte receptors
for the micronutrient; there may be an increased rate
of hepatic secretion of the plasma transport protein,
and there may be an increase in the tissue expression
of cellular receptors for the micronutrient or the
transport complex. Many of these alterations occur
in response to cobalamin deficiency (Herbert 1994)
and all of them are seen in iron deficiency (Melefors
and Henze 1993). Changes in enterocyte iron recep-
tors can be assessed by performance of a radio-iron
absorption study but absorption studies are less
reliable than the much more easily performed blood
studies. Changes in tissue transferrin receptor
expression are reflected in the concentration of the
receptor in plasma (Skikne
et al.
1990). Because
transferrin receptors enter the plasma as a result of
cell turnover, the increased turnover of red cells in
the marrow and in the circulation that is present in
iron-deficiency leads to a diagnostically useful
exaggeration of the plasma receptor concentration in
that state. Alterations in the rate of secretion of
apotransferrin are quantified by measurement of the
plasma concentration of the protein, usually as total
iron binding capacity. It is common practice to
measure the concentration of the plasma iron as well
as total iron binding capacity and to calculate the
ratio of the two, which is expressed as percent trans-
ferrin saturation. In iron deficiency, once iron
stores are depleted and plasma iron concentration
begins to fall, transferrin saturation is directly
related to micronutrient stores and in a fashion that
is much more steep, and therefore much more sensi-
tive and precise, than that of either of the two
measures separately.
Deficiency of a trace substance leads to a
decreased cellular concentration of the substance and
thereby directly increases tissue access for the
substance if the substance readily diffuses into cells.
This phenomenon underlies the use of load tests for
the evaluation of deficiency states of the water-
soluble vitamins and some of the trace minerals. In
a load test, the urinary excretion of a test dose of
Nutritional Status
8-6
