Chapter 34. Homocysteine, Cysteine, and Taurine

(predominantly CySSCy and HcySSCy) and with very low concentrations of free thiols. The Cys-containing peptides, cysteinylglycine (CysGly), g-glutamylcysteine

(gGluCys), and glutathione (GSH, or gGluCysGly), are also present in plasma and tissues.



Figure 34.2. Concentrations of various forms of the major aminothiols in human plasma. Cys, cysteine; Hcy, homocysteine; CysGly, cysteinylglycine; PS, sulfhydryl

group of cysteinyl residue in protein; RS, unspecified thiol, usually Cys in plasma. The designation RSH is used to represent the reduced thiol form, RSSR or RSSR'

to represent the disulfide of the thiol with itself or another thiol, and PSSR to represent protein-bound disulfides. Mean fasting values for plasma thiols are based on

the data of Manssoor et al. (16) for 34 male and 31 female control subjects. Other plasma thiols include glutathione (total GSH » 7 mmol/L) and g-glutamylcysteine

(total g-GluCys » 3 mmol/L) (17).



The protein binding and redox status of different plasma aminothiols are interactive because of presumed ongoing redox cycling and disulfide exchange reactions. For

example, Hcy will displace protein-bound Cys or CysGly ( 18). After ingestion of a methionine load or a protein-containing meal, protein-bound Cys tends to decrease,

probably because of displacement of protein-bound Cys by Hcy ( 16, 19).

Measures of tHcy or tCys are useful in clinical studies because they are not affected in vitro by disulfide exchange reactions and redistribution between forms. Mean

fasting plasma tHcy was 11.9 µmol/L (median, 11.6) with a range of 3.5 to 66.8 µmol/L in 1160 subjects aged 67 to 95 years ( 20); mean plasma tHcy is slightly lower

for younger adults than for older ones and for women than for men ( 21, 22 and 23). Mean fasting plasma tCys concentrations in healthy adults range from about 220

to 320 µmol/L (16, 17, 18 and 19).

A wide range of plasma taurine concentrations has been reported for human subjects. Trautwein and Hayes ( 24) reviewed values reported in the literature and found

the reported mean plasma concentration of taurine in human subjects ranged from 39 to 116 µmol/L. Whole blood taurine ranged between 160 and 320 µmol/L with a

mean of 225 µmol/L in a small sample of adults (24). Plasma taurine concentrations change more rapidly in response to changes in taurine intake than do whole blood

concentrations, and whole blood taurine concentrations are not correlated with plasma taurine concentrations except during periods of depletion or excess intake.

Plasma taurine concentrations are somewhat lower in vegans than in omnivores and somewhat lower in females than in males ( 25, 26).

Careful handling of blood samples is essential for measurement of plasma concentrations of aminothiols and taurine. Plasma tHcy concentrations may increase with

storage of blood because of transsulfuration in blood cells unless the blood is rapidly cooled and processed ( 27). Hemolysis or contamination of the plasma fraction

with platelets or white cells interferes with analysis of plasma taurine but not with measures of whole blood taurine ( 24).



DIETARY CONSIDERATIONS AND TYPICAL INTAKES

Methionine and Cyst(e)ine

The sulfur amino acids, methionine and cyst(e)ine, are normally consumed as components of dietary proteins. Normal Western diets provide 15 to 20 mmol (~2.25–3

g) of sulfur amino acids per day. Mixtures of proteins consumed in the United States contain about 35 mg methionine plus cyst(e)ine per gram of protein ( 28). Sulfur

amino acids tend to be more abundant in animal and cereal proteins than in legume proteins, and the ratio of methionine to cysteine tends to be higher in animal

proteins than in plant proteins ( Table 34.1) (29).



Table 34.1 Methionine and Cysteine Content of Selected Foods



The 1989 RDA committee recommended a sulfur amino acid intake of 13 mg/kg/day, a protein intake of 0.75 g/kg/day, and a desirable amino acid pattern for adults

that includes at least 17 mg methionine plus cyst(e)ine per gram of protein ( 28). This suggested intake of sulfur amino acids is easily met by mixtures of protein

commonly consumed in the United States. However, Young et al. (30) and Storch et al. (31) have proposed that the sulfur amino acid allowance set by the FAO/WHO

(32) and the NRC (28) at 13 mg/kg/day (0.087 mmol/kg/day, or 6 mmol/day for a 70-kg adult) represents the average requirement of healthy adults rather than the

upper end of the normal distribution of requirements (mean + 2 SD). Their studies indicated that an intake of 25 mg/kg/day (0.17 mmol/kg/day or 12 mmol/day for a

70-kg adult) was necessary to ensure an adequate intake for the entire population.

Despite the availability of food proteins that provide ample amounts of sulfur amino acids, some individuals probably have inadequate intakes because of either

inadequate intake of total protein or selection of a restricted variety of proteins that provide inadequate sulfur amino acids. Analysis of diets of long-term vegans living

in California indicated an average protein intake of 64 g/day and a sulfur amino acid intake of 1.04 g (7.6 mmol) per day ( 26); this is equivalent to an intake of ~15

mg/kg/day of sulfur amino acids and an amino acid pattern of 16 mg methionine plus cyst(e)ine per gram protein. This level of intake would meet but not exceed the

average requirement as estimated by Young et al. ( 30) and Storch et al. (31) and would be marginal for adults with higher-than-average requirements. Careful

selection of plant proteins to ensure an adequate intake of sulfur amino acids may be very important for strict vegan adults and even more so for children fed a strict

vegan diet.

Taurine

Although taurine is an end product of sulfur amino acid metabolism, it is usually obtained from the diet as well. Food taurine content has not been widely determined,

but data from several reports ( 33, 34, 35 and 36) are summarized in Table 34.2. Taurine is present in most animal foods and is either absent or present in very low

levels in most plant foods. Relatively high concentrations of taurine have been reported for some lower plants such as seaweeds ( 36, 37). Analysis of the diets of strict

vegans living in England yielded no detectable taurine, whereas the diets of omnivores contained 463 ± 156 (SE) µmol/day ( 25). The analyzed taurine intake of adults



fed omnivorous diets in a clinical study center in the U.S. was 1000 to 1200 µmol/day ( 33).



Table 34.2 Taurine Content of Selected Foods



The taurine content of milk from lactating women was 41.3 ± 7.1 (SE) µmol/100 mL for early milk (1–7 days) and 33.7 ± 2.8 µmol/100 mL for later milk (>7 days) ( 38).

Taurine is added to infant formulas at levels comparable to those in human milk or at somewhat higher levels in formulas for premature infants ( 33). The mean taurine

content of milk of lactating vegan women is lower than that of lactating omnivores, but values overlap considerably between the two groups, and the taurine

concentration in milk of vegan mothers is still about 30 times the level in the cow's milk–based infant formulas used prior to the mid-1980s ( 25).



ABSORPTION, TRANSPORT, AND EXCRETION

Absorption of the products of protein digestion across the intestinal epithelium is highly efficient (~95–99%). Dietary methionine, a precursor of Cys, is transported by

neutral amino acid transport systems (B 0,+, ASC, and L), and as methionine-containing peptides by peptide transport systems. Dietary Cys is absorbed as CySH,

CySSCy, and Cys-containing peptides by a variety of L-amino acid and peptide transport systems in the small intestinal mucosa. Cysteine is transported by neutral

amino acid transporters including system B in the apical (brush border) membrane and system ASC in both the apical and basolateral plasma membranes of the

intestinal mucosa cells; cysteine uptake is largely Na + dependent. Cystine is transported by system b 0,+, a Na+-independent system present in the apical membranes

of the intestinal mucosa, which serves cationic amino acids as well as zwitterionic amino acids. Efficient absorption of taurine is facilitated by the b-amino acid or

taurine transport system, a Na+ and Cl–-dependent carrier that serves taurine, b-alanine, and g-aminobutyric acid, which is present in the apical membrane of

intestinal mucosa cells.

Amino acids enter the plasma and circulate as free amino acids until they are removed by tissues. The liver removes a substantial proportion of the sulfur-containing

amino acids from the portal circulation and uses them for synthesis of protein and glutathione or for catabolism to taurine and sulfate. GSH is exported into plasma,

and this cysteine-containing tripeptide as well as its metabolites, CysGly and g-GluCys, can be a source of cysteine to tissues. Hcy is not normally present in the diet,

and only very low amounts are normally released from tissues into the plasma.

The reabsorptive epithelium of the kidney proximal tubule has transport systems similar to those of the absorptive epithelium of the intestine, and the kidney efficiently

reabsorbs amino acids from the filtrate. Renal reabsorption of Cys and methionine is normally very high (³94%), and the loss of amino acids in the urine is normally

negligible (39). Urinary methionine excretion has been reported to be 22 to 41 µmol/day ( 25, 40). Urinary cyst(e)ine excretion by adults has been reported to be 63 to

285 µmol/day (25, 40).

Cystinuria is an inherited disorder of cystine and dibasic amino acid transport. One cause of cystinuria is a defect in the gene that encodes the rBAT (related to b 0,+

amino acid transporter) protein, a subunit of the system b 0,+ transporter that is expressed by the kidney and small intestine ( 41, 42 and 43). Urinary cystine excretion

exceeding 1.2 mmol/day (~2.5 mmol Cys/day) is usually diagnostic of homozygous cystinuria ( 44). Cystine is very insoluble and can cause cystine stones if it is

present above its aqueous solubility limit (250 mg/L, or 1 mmol/L).

Urinary excretion of extracellular Hcy is limited, even in individuals with defective Hcy metabolism, because the extensive binding of plasma Hcy to proteins limits

filtration and because of the normally active renal reabsorption of free Hcy. Of the plasma Hcy filtered by the kidney, only about 1 to 2% is excreted in the urine ( 45).

Normal urinary Hcy excretion ranges from 3.5 to 9.8 µmol/day (45). Higher levels of Hcy in urine indicate very high plasma tHcy concentrations and an inborn error of

metabolism. For example, Hcy excretion in urine of patients with N5,10-methylenetetrahydrofolate reductase deficiency ranged from 15 to 667 µmol/day ( 46).

Unlike most amino acids, taurine is not usually completely reabsorbed, and fractional excretion may vary over a wide range. Normally, the kidney regulates the body

pool size of taurine and adapts to changes in dietary taurine intake by regulation of the proximal tubule brush-border membrane transporter for taurine (b system).

During periods of inadequate dietary intake of taurine or its sulfur amino acid precursors, more taurine is reabsorbed from the filtrate because of enhanced taurine

transporter activity, less taurine is excreted in the urine, and more of the tissue taurine stores are maintained. The renal taurine concentration seems to be the signal

for changes in renal taurine transporter activity ( 47, 48).

Consistent with differences in taurine intake and with adaptive regulation of taurine reabsorption, urinary taurine levels vary widely. Urinary taurine levels of 250

µmol/day have been reported for adult vegans consuming diets with no preformed taurine, whereas excretion of taurine by adult omnivores usually exceeds 600

µmol/day, and values above 1000 µmol/day are not uncommon (25, 26, 40).



METHIONINE/HOMOCYSTEINE METABOLISM AND CYSTEINE FORMATION

Metabolic Pathways

Transmethylation

The essential amino acid methionine is activated by ATP to form S-adenosylmethionine in a reaction catalyzed by S-adenosylmethionine synthetase (EC 2.5.1.6).

S-Adenosylmethionine serves primarily as a methyl donor via reactions catalyzed by a variety of methyl transferases and involving a variety of acceptors. These

S-adenosylmethionine-dependent methylations are essential for the biosynthesis of a variety of cellular components including creatine, epinephrine, carnitine,

phospholipids, proteins, DNA, and RNA ( Fig. 34.3).



Figure 34.3. Methionine metabolism. Numbered reactions are catalyzed by the following enzymes: (1) methionine adenosyltransferase; (2) various

methyltransferases; (3) adenosylhomocysteine hydrolase; (4) N 5-methyltetrahydrofolate-homocysteine methyltransferase; (5) N5,10-methylenetetrahydrofolate



reductase; (6) betaine-homocysteine methyltransferase; (7) cystathionine b-synthase; (8) cystathionine g-lyase; (9) enzymes involved in polyamine synthesis; and

(10) enzymes involved in methylthioadenosine salvage pathway.



S-Adenosylhomocysteine, the byproduct of these methyl transfer reactions, is hydrolyzed by S-adenosylhomocysteine hydrolase (EC 3.3.1.1), thus generating

adenosine and homocysteine. Although the equilibrium of S-adenosylhomocysteine hydrolase actually favors formation of S-adenosylhomocysteine, the reaction is

normally driven forward by rapid removal of the products.

The homocysteine generated by hydrolysis of S-adenosylhomocysteine has two likely metabolic fates, remethylation or transsulfuration. In remethylation,

homocysteine acquires a methyl group from N5-methyltetrahydrofolate or betaine to form methionine. In transsulfuration, the sulfur is transferred to serine to form

cysteine, and the remainder of the homocysteine molecule is catabolized to a-ketobutyrate and ammonium.

Remethylation

The remethylation pathway allows methionine to be regenerated from homocysteine by use of new methyl groups synthesized in the folate coenzyme system or

preformed methyl groups, both of which may subsequently be transferred to acceptors via S-adenosylmethionine-dependent methyltransferase reactions. The

remethylation of homocysteine by transfer of a methyl group from N5-methyltetrahydrofolate is catalyzed by N5-methyltetra hydrofolate-homocysteine

methyltransferase (EC 2.1.1.13), commonly called methionine synthase. Methionine synthase is widely distributed in mammalian tissues and contains

methylcobalamin as an essential cofactor. The methyl group of N5-methyltetrahydrofolate is synthesized de novo in the folate coenzyme system. The final step of

N 5-methyltetrahydrofolate synthesis is the irreversible reduction of N5,10-methylenetetrahydrofolate, which is catalyzed by the flavoenzyme,

N 5,10-methylenetetrahydrofolate reductase (EC 1.1.1.68), using NADH as the electron donor.

The other homocysteine methyltransferase, betaine-homocysteine methyltransferase (EC 2.1.1.5), is present only in liver and kidney of humans and requires betaine

as the methyl donor (49). This reaction uses preformed methyl groups because betaine is derived from choline, which is either obtained from the diet or synthesized

through successive S-adenosylmethionine-dependent methylations of phosphatidylethanolamine.

Transsulfuration

The transsulfuration of homocysteine to cysteine is catalyzed by two pyridoxal 5'-phosphate (PLP)-dependent enzymes, cystathionine b-synthase (EC 4.2.1.22) and

cystathionine g-lyase (EC 4.4.1.1; cystathionase). Cystathionine b-synthase catalyzes the condensation of homocysteine and serine to form cystathionine.

Cystathionine is then hydrolyzed by cystathionine g-lyase to form cysteine and a-ketobutyrate plus ammonium. Thus, the transsulfuration pathway is responsible for

both the catabolism of homocysteine derived from methionine and the transfer of methionine sulfur to serine to synthesize cysteine.

Regardless of the extent of remethylation, in the steady-state metabolic condition, intake of methionine sulfur is balanced by metabolism of an almost equivalent

amount of homocysteine sulfur through the transsulfuration pathway ( 50, 51). Although some methionine (via decarboxylated S-adenosylmethionine acting as a donor

of aminopropyl groups) is used for polyamine synthesis, the methylthioadenosine that is a byproduct of this pathway is effectively recycled to methionine. The sulfur

and methyl carbon of methionine and the carbon chain of ribose are reused in methionine synthesis by the methylthioadenosine salvage pathway ( 51, 52). Hence,

little sulfur is oxidized or lost during methionine metabolism, and essentially all methionine sulfur is transferred to cysteine prior to oxidation/excretion of the sulfur

atom.

Regulation of Remethylation versus Transsulfuration

Response to Changes in Methionine Intake

The remethylation and transsulfuration pathways can be considered to be competing for available homocysteine. Studies of whole body methionine kinetics

demonstrated that in young adult men fed a diet with adequate methionine (~14 mmol/day), about 17 mmol/day of homocysteine was formed by transmethylation;

approximately 38% of this homocysteine was remethylated to methionine, and 62% was catabolized by transsulfuration ( 31). In another study, transmethylation or

homocysteine formation decreased markedly in subjects fed a sulfur amino acid–free diet, from approximately 20 mmol/day in men on an adequate diet to 6 mmol/day

in men on a sulfur amino acid–free diet; the percentage of homocysteine remethylated to methionine increased from 36% in men on the adequate diet to 67% in men

on the sulfur amino acid–free diet ( 53). As a result of decreased transmethylation or homocysteine formation and the greater percentage remethylation of

homocysteine, transsulfuration or oxidation of methionine was reduced from 12 mmol/day in men fed the methionine-adequate diet to 2 mmol/day in subjects fed the

sulfur amino acid–free diet.

Response to S-Adenosylmethionine as an Effector

Whether homocysteine is metabolized by remethylation or transsulfuration seems to be coordinated in response to cellular S-adenosylmethionine concentrations or

the need to generate methionine methyl groups (54). S-Adenosylmethionine is both an allosteric inhibitor of N5,10-methylenetetrahydrofolate reductase and an

activator of cystathionine b-synthase. Hence, when the cellular S-adenosylmethionine concentration is low, synthesis of N 5-methyltetrahydrofolate proceeds

uninhibited and cystathionine synthesis is suppressed, which conserves homocysteine for methionine synthesis. Conversely, when the S-adenosylmethionine

concentration is high, inhibition of N 5-methyltetrahydrofolate synthesis is accompanied by diversion of homocysteine through the transsulfuration pathway because of

stimulated cystathionine synthesis. This coordinate control results in both regulation of cellular S-adenosylmethionine concentration and maintenance of a

homocysteine concentration that is compatible with the need for methyl groups synthesized de novo.

Methionine-Sparing Effect of Cyst(e)ine

Cyst(e)ine is said to have a “methionine-sparing” effect by reducing methionine catabolism via the transsulfuration pathway. The effect of supplemental cyst(e)ine

added to a sulfur amino acid–free diet or a low-methionine diet may be at least partially due to promotion of methionine incorporation into protein so that less

methionine is catabolized (53, 55). The effect of cyst(e)ine used to replace part of the dietary methionine (keeping total sulfur amino acid level the same) may be

explained by a reduction in the hepatic concentrations of methionine and S-adenosylmethionine and, hence, less activation and reduced activity of hepatic

cystathionine b-synthase. Less homocysteine catabolism by transsulfuration would result in increased recycling of homocysteine to methionine, using methyl groups

generated by the folate coenzyme system.

Response to Supplemental Betaine

Normal adult subjects given a control diet with a betaine supplement had increased rates of methionine transmethylation and transsulfuration ( 56), suggesting that an

increased dietary supply of methyl groups may increase methionine catabolism. A high dietary intake of betaine when coupled with a marginal intake of methionine

could interfere with the normal coordinated regulation of remethylation versus transsulfuration by increasing S-adenosylmethionine concentration and stimulating

methionine catabolism (56). Presumably, increased remethylation induced by betaine increases S-adenosylmethionine concentrations, resulting in inhibition of

N 5-methyltetrahydrofolate-dependent remethylation and stimulation of homocysteine catabolism.



HOMOCYSTINURIA AND HYPERHOMOCYSTEINEMIA

Homocystinuria Due to Inborn Errors of Metabolism

Severe forms of hyperhomocysteinemia result in excretion of homocysteine, homocystine, and mixed disulfides of homocysteine in the urine. Homocystinuria (urinary

Hcy > 10 µmol/24 h) is rare and results from severe hyperhomocysteinemia caused by several inborn errors of metabolism. The most common inborn error of sulfur

amino acid metabolism and the most common cause of homocystinuria is a lack of cystathionine b-synthase activity, which is commonly associated with plasma tHcy

concentrations above 200 µmol/L in untreated patients. Two clinical forms have been described: one not responsive to treatment with the coenzyme precursor



pyridoxine and a pyridoxine-responsive form that improves with a high intake of vitamin B 6. The latter is usually associated with residual cystathionine b-synthase

activity and milder disease (57). Based on newborn-screening programs, the estimated incidence of cystathionine b-synthase deficiency is 1:170,000 ( 58). The actual

incidence of cystathionine b-synthase deficiency is thought to be closer to 1:340,000, however, because newborn-screening programs fail to detect most infants with

the pyridoxine-responsive form of the disease. Cystathionine b-synthase deficiency is inherited as an autosomal recessive disorder and results in dislocation of optic

lenses, skeletal abnormalities, mental retardation, neurologic disorders, and widespread thromboembolic phenomena. This disorder has been shown to be due to

heterogeneous mutations, and most patients are compound heterozygotes; more than 18 mutations have been identified in patients with cystathionine b-synthase

deficiency (59, 60).

A second inborn error of metabolism that causes homocystinuria is a lack of N5,10-methylenetetrahydrofolate reductase activity. This is the major known inborn error

affecting folate metabolism and the second leading known cause of homocystinuria. N5,10-Methylenetetrahydrofolate reductase deficiency is inherited as an autosomal

recessive disorder with an incidence about 1/10th that of cystathionine b-synthase deficiency. Individuals with severe deficiencies have residual activity that ranges

from undetectable to 20% of normal activity. At least nine different mutations have been identified in patients with a severe lack of N 5,10-methylenetetrahydrofolate

reductase activity (61, 62). Homocystinuria and hyperhomocysteinemia in these patients are usually less severe than in patients with cystathionine b-synthase

deficiency, and a wide range of neurologic and vascular disturbances have been observed in these patients (see Chapter 26 and Chapter 95).

A third group of inborn errors giving rise to homocystinuria are those affecting various steps in the synthesis of methylcobalamin, an essential cofactor for methionine

synthase (63). Cobalamin metabolism loci C, D, E, F, and G have been shown to result in reduced activity of methionine synthase and homocystinuria (see Chapter

27).

Hyperhomocysteinemia and Its Genetic, Nutritional, and Metabolic Bases

Milder forms of hyperhomocysteinemia have been recognized more recently and are much more prevalent than is homocystinuria. Heterozygosity for one of the inborn

errors of metabolism resulting in homocystinuria may be one cause of milder hyperhomocysteinemia. However, the predicted incidence of heterozygosity for these

inborn errors is too small to account for a large proportion of the observed hyperhomocysteinemia.

A milder form of N 5,10-methylenetetrahydrofolate reductase deficiency that results in approximately 50% residual enzyme activity in homozygotes is caused by a point

mutation (677C®T) in the N5,10-methylenetetrahydrofolate reductase gene (62, 64, 65). This genetic deficiency of N5,10-methylenetetrahydrofolate reductase is also

described as the “thermolabile variant” because of the marked thermolability of the altered enzyme in vitro ( 66). Approximately 5% of subjects studied by Kang et al.

(66) and 12% of those studied by Rozen et al. ( 67) were homozygous for this 677C®T mutation in the N5,10-methylenetetrahydrofolate reductase gene. This mutation

is hypothesized to be an underlying cause of much of the hyperhomocysteinemia and associated increased risk for vascular disease and neural tube defects ( 62, 64,

68, 69).

Nutritional disorders that potentially lead to hyperhomocysteinemia, particularly in individuals with underlying genetic predispositions, are deficiencies of folate,

vitamin B12, and vitamin B6 (20, 70, 71 and 72). Selhub et al. (20) estimated that 67% of cases of hyperhomocysteinemia are at least partially due to inadequate

B-vitamin status. As noted above, de novo synthesis of methionine methyl groups requires both vitamin B 12 and folate coenzymes, whereas transsulfuration requires

PLP. Evidence for an inverse correlation between vitamin intake or status and hyperhomocysteinemia is stronger and more consistent for folate than for vitamin B 12 or

vitamin B6 (20, 21, 71, 72, 73, 74 and 75). The thermolabile variant of N5,10-methylenetetrahydrofolate reductase is very responsive to folate status;

hyperhomocysteinemia is observed most frequently in homozygotes who also have a low folate intake and rarely in homozygotes who have a higher folate intake ( 65,

75).

Mild-to-moderate hyperhomocysteinemia and increased incidence of atherosclerotic disease are also observed in patients with renal disease. Plasma tHcy levels are

significantly increased in patients with moderate renal failure and rise steeply in terminal uremia ( 76, 77). The rise in plasma tHcy levels in patients with renal failure is

attributed to loss of renal parenchymal uptake and metabolism of plasma Hcy rather than to decreased urinary excretion of Hcy.

Clinical Significance of Hyperhomocysteinemia

Apparent associations of mild hyperhomocysteinemia with atherosclerotic disease, venous thromboses, neural tube defects, placental abruption/infarction, and

unexplained pregnancy loss have been reported ( 8, 9, 10, 11, 22, 69, 74, 77, 78, 79, 80, 81 and 82). In populations of individuals with arteriosclerotic disease, mild

hyperhomocysteinemia is observed about as frequently as hypercholesterolemia or hypertension. Hyperhomocysteinemia is now considered an independent risk

factor for vascular disease of the coronary, cerebral, and peripheral arteries, with an increase in plasma tHcy of only 5 µmol/L estimated to result in a 50 to 80%

increase in risk for cerebrovascular or coronary artery disease ( 73).

The basis of the association of hyperhomocysteinemia with vascular disease is not clear ( 9, 83, 84, 85 and 86). Hypotheses related to a direct adverse effect of

homocysteine (endothelial cell damage, oxidative damage, promotion of thrombogenesis via alterations in the coagulation and fibrinolytic systems, impaired regulation

of vasoconstriction, stimulation of vascular smooth muscle cell proliferation) are currently being investigated. It is also possible that a decreased intracellular ratio of

S-adenosylmethionine to S-adenosylhomocysteine or reduced availability of S-adenosylmethionine, either of which may be associated with hyperhomocysteinemia,

interferes with essential methylation reactions and leads to the adverse effects associated with hyperhomocysteinemia. Similarly, impaired methylation of proteins has

been implicated as a possible cause of neural tube defects ( 87, 88).

Detection of Homocystinuria and Hyperhomocysteinemia

Inborn errors giving rise to homocystinuria may be detected by newborn-screening programs (see Chapter 61). Most current screening programs detect individuals

with cystathionine b-synthase deficiency by primary screening for hypermethioninemia and secondary screening for elevated urine and/or plasma tHcy levels, low

plasma tCys concentration, and deficient cystathionine b-synthase activity in fibroblasts, leukocytes, or liver biopsy specimens. These screening programs fail to

detect many pyridoxine-responsive forms of cystathionine b-synthase deficiency, which may not give rise to abnormal plasma Hcy or methionine concentrations in

infants, especially if the protein intake is low or if testing is done within the first few weeks of life ( 58). Screening for hypermethioninemia will also not detect

homocystinuria due to defects in folate or cobalamin metabolism because plasma methionine concentrations in these patients are either normal or low ( 46). Patients

with defects in folate or cobalamin metabolism tend to have lower urinary Hcy levels than do patients with cystathionine b-synthase deficiency, and analysis of urine or

plasma Hcy levels often yields a negative test result in affected newborns. Genotyping will likely be used more widely in the future to identify both homocystinuric

individuals and heterozygous carriers.

Milder forms of hyperhomocysteinemia (heterozygotes for cystathionine b-synthase or N5,10-methylenetetrahydrofolate reductase deficiency; B-vitamin deficiency;

thermolabile N5,10-methylenetetrahydrofolate reductase; renal failure; etc.) may be indicated by elevated fasting plasma tHcy concentrations (~16–30 µmol/L). Normal

reference intervals for plasma tHcy vary, depending on methodology and the selection of reference individuals. Because as much as 40% of the population, including

individuals with relatively low plasma tHcy values, may respond to folate or B-vitamin supplementation with a reduction in plasma tHcy ( 20, 71, 74, 80) and because

even small elevations in plasma tHcy seem to be associated with increased risk of vascular disease, it has been difficult to establish exact desirable or reference

values for the normal population. Use of 90th percentile values as cutoffs has resulted in use of plasma fasting tHcy values above 14 to 16 µmol/L as indicators of

hyperhomocysteinemia (21, 89). A lower cutoff for the normal range may be appropriate, however, because the frequency distribution of plasma tHcy concentrations is

positively skewed (74). Use of reference intervals based on presupplementation measurements of plasma tHcy in subjects who can be classified as weak responders

to folate supplementation has been suggested. Rasmussen et al. ( 23) found a mean ± SD of 7.76 ± 1.54 µmol/L for these weak responders, compared with 12.33 ±

2.04 µmol/L for the high responders; they derived age- and gender-specific 95% confidence intervals including values between 4.5 and 11.9 µmol/L for the weak

responders. Ubbink et al. ( 74) suggested using a mathematical model to predict the plasma tHcy that could be expected for a population with improved vitamin status;

their calculated 95% reference range was 4.9 to 11.7 µmol/L for a population with improved folate, vitamin B 12, and vitamin B6 status. Hence, about 12 µmol/L would

seem to be the upper end of the normal “desirable” range for adult fasting plasma tHcy.

The use of both fasting and post–methionine load measurements may have benefits in screening for mild forms of hyperhomocysteinemia ( 54, 90). In one study,

fasting plasma tHcy determination alone failed to identify more than 40% of persons classified as hyperhomocysteinemic when both fasting and post–methionine load

measurements were used (89). Measurement of plasma tHcy 2 or 4 hours after administration of an oral load dose of methionine (~0.1 g or 670 µmol/kg) yielded

plasma tHcy concentrations (mean ± SD) of 29.2 ± 10.4 µmol/L, compared with 9.6 ± 4.2 µmol/L for fasting concentrations; the 90th percentile cutoff value used to



define hyperhomocysteinemia was 41.4 µmol/L for post–methionine load values, compared with 13.6 µmol/L for fasting tHcy ( 89).

Dietary Treatment of Homocystinuria and Hyperhomocysteinemia

The goal of dietary treatment of homocystinuria is to decrease the biochemical abnormalities (reduce plasma Hcy, maintain normal concentrations of plasma

methionine and Cys) and to minimize development of clinical complications. Treatment of individuals with inborn errors of transsulfuration (cystathionine b-synthase

deficiency) usually involves restriction of dietary methionine intake while providing adequate cyst(e)ine intake (see Chapter 61). Small, frequent feedings are also

recommended to minimize a methionine load effect. Newborns diagnosed with cystathionine b-synthase deficiency are fed low-protein formula in controlled quantities

or formula low in methionine and supplemented with cystine in an effort to maintain normal plasma methionine concentrations (20–40 µmol/L) and very low plasma

concentrations of tHcy. Inclusion of choline or betaine (25–165 mmol/day; 100–600 mg/kg/day) as a methyl donor is also beneficial ( 58, 91); these levels of betaine

supplementation are very high compared with the normal total choline intake in the adult (5.0–8.3 mmol/day), of which only a portion becomes available as betaine

(92). Pyridoxine at levels ranging from 150 to 1200 mg/day has been used successfully in treatment of cystathionine b-synthase-deficient adults; these levels greatly

exceed the normal recommended intake of approximately 2 mg/day for adults.

In homocystinuria due to inborn errors in the methionine remethylation pathway, the goal of dietary treatment is to maintain sufficient levels of methionine methyl

groups without elevating the plasma Hcy concentration; this usually involves relatively high levels of B-vitamin supplementation (folate, vitamin B 12) to stimulate the

N 5-methyltetrahydrofolate-dependent homocysteine re-methylation pathway, betaine or choline to stimulate the betaine-dependent homocysteine remethylation

pathway, and in some cases methionine supplementation to ensure sufficient availability of S-adenosylmethionine (93).

Modest levels of vitamin supplementation (~0.6 mg folic acid, 0.4 mg cyanocobalamin, and 10 mg pyridoxine) reduce plasma tHcy concentrations in a proportion of

individuals with mild hyperhomocysteinemia (74, 77, 94, 95 and 96). Some individuals with plasma tHcy levels in the reference range (fasting values <12 µmol/L) also

respond to folate or B-vitamin supplementation with a reduction in plasma tHcy ( 74, 94, 95). Although Hcy-lowering therapy reduces the vascular risk of patients with

severe hyperhomocysteinemia due to inborn errors of metabolism, the outcome of treatment of mild hyperhomocysteinemia on atherosclerotic and thromboembolic

disease is yet to be reported (97). Periconceptual supplementation with folate reduces the incidence of neural tube birth defects in newborns ( 79). To increase the

folate intake of women of childbearing age, the U.S. Food and Drug Administration ruled in 1996 to require fortification of most breads and grain products as of 1998.

Fortification of foods with folate may have some beneficial effects on additional pregnancy outcome measures and the incidence of vascular and thromboembolic

disease.



METABOLISM OF CYSTEINE

Pathways of Cysteine Metabolism

Cysteine, whether formed from methionine and serine via transsulfuration or supplied preformed in the diet, serves as a precursor for synthesis of proteins and

several other essential molecules as shown in Figure 34.4. These metabolites include GSH, coenzyme A, taurine, and inorganic sulfur.



Figure 34.4. Pathways of cysteine metabolism. Numbered reactions are catalyzed by the following enzymes or pathways: (1) protein synthesis; (2) protein

degradation; (3) GSH-thioltransferase or nonenzymatic thiol-disulfide exchange of cystine with GSH; (4) g-glutamylcysteine synthetase; (5) glutathione synthetase; (6)

glutathione transpeptidase; (7) dipeptidase; (8) pathway of coenzyme A synthesis; (9) cysteine dioxygenase; (10) cysteine sulfinate decarboxylase; (11) enzymatic or

nonenzymatic oxidation of hypotaurine; (12) aspartate (cysteine sulfinate) aminotransferase; (13) cysteine sulfinate-independent or desulfhydration pathways of

cyst(e)ine catabolism; (14) sulfide oxidase; (15) thiosulfate sulfurtransferase or GSH-dependent thiosulfate reductase; and (16) sulfite oxidase.



At intakes near the requirement, a large proportion of available cysteine is used for synthesis of proteins and GSH. In addition to the specific metabolic functions of

GSH, GSH serves as a reservoir of Cys and as a means for transporting Cys to extrahepatic tissues ( 86). A large proportion of the sulfur amino acid intake is

converted to GSH by the liver and released into the circulation. g-Glutamyl transpeptidase, an enzyme located on the outer surface of the plasma membrane of cells

in a number of tissues, hydrolyzes GSH (or its disulfide) to yield CysGly (or its disulfide), which can be further degraded by peptidases to release cysteine (or cystine)

into the plasma. The normal turnover of GSH in humans is estimated to be 40 mmol/day (99, 100), nearly two to three times the typical sulfur amino acid intake of 15

to 20 mmol/day and six to seven times the estimated sulfur amino acid requirement. This estimate suggests that the magnitude of turnover of the Cys pool as a result

of GSH turnover may be slightly greater than that resulting from protein turnover in the body (~30 mmol/day; [ 31]).

Cysteine is also a precursor for synthesis of coenzyme A and for production of taurine and inorganic sulfate. These three fates of cysteine involve loss of the cysteine

moiety as such. Cysteine is substrate for coenzyme A synthesis in that it donates the cysteamine (decarboxylated cysteine) moiety of the coenzyme A molecule and,

hence, contributes the reactive sulfhydryl group. Coenzyme A turnover is thought to be slow, such that coenzyme A synthesis does not consume a large amount of

dietary cysteine. The functions of coenzyme A are discussed in Chapter 25.

Both taurine and inorganic sulfur are products of cysteine catabolism. As shown in Figure 34.4, there are several pathways for cysteine catabolism. Cysteine

catabolism may occur by desulfuration of cysteine to yield pyruvate and reduced sulfur (often in the form of a persulfide such as thiocysteine, mercaptopyruvate, or

thiosulfate). Cysteine desulfuration can be catalyzed by the b-cleavage of cyst(e)ine by cystathionine g-lyase or by transamination of cysteine to b-mercaptopyruvate

followed by de- or transsulfuration by mercaptopyruvate sulfurtransferase ( 51). Individuals with a rare inborn error of metabolism in which b-mercaptopyruvate

sulfurtransferase is deficient excrete the mixed disulfide of cysteine and b-mercaptolactate, suggesting that transamination of cysteine to mercaptopyruvate occurs to

some extent in humans (101). However, these patients excrete normal levels of urinary sulfate, indicating that overall cysteine catabolism is not impaired. The

reduced sulfur may be used in synthesis of molecules requiring a source of reduced sulfur, or it may be oxidized to thiosulfate (inner sulfur), sulfite, and finally sulfate.

Although most of the inorganic sulfur is eventually oxidized to sulfate, mammals depend upon the cysteine sulfinate–independent or desulfhydration pathways of

cysteine metabolism as a source of reduced forms of inorganic sulfur because they cannot reduce sulfate or sulfite to thiosulfate or sulfide.

In animals fed high-protein or high sulfur amino acid–containing diets, the major pathway of cysteine catabolism involves oxidation of cysteine to cysteine sulfinate by

cysteine dioxygenase (EC 1.13.11.20), which is an inducible enzyme in rat liver ( 51, 102). Cysteine sulfinate may be decarboxylated to hypotaurine, which is

subsequently oxidized to taurine, or cysteine sulfinate may be transaminated (with a-ketoglutarate) to the enzyme-bound intermediate, b-sulfinylpyruvate, which gives

rise to pyruvate and sulfite. Sulfite is further oxidized to sulfate by sulfite oxidase (EC 1.8.3.1).

In all catabolic pathways except that resulting in taurine formation, the carbon chain of cysteine is released as pyruvate, the sulfur is released as inorganic sulfur, and

the amino group is released as ammonium or transferred to a keto acid acceptor. When taurine is the end product, only the carboxyl carbon of the cysteine is

released, and the other three carbons as well as the nitrogen and sulfur atoms remain in the end product. Thus, the distribution of Cys among its catabolic pathways

potentially affects the use of amino acid carbon chains for energy, the net production of acid or fixed anions (sulfate), and the synthesis of essential metabolites

(inorganic sulfur and taurine). Although taurine and sulfate can be regarded as end products of cellular cysteine catabolism, both of these compounds participate in

conjugation reactions and have a variety of essential physiologic functions prior to their ultimate excretion.



Adult human subjects remain in sulfur balance, with sulfur excretion essentially equivalent to sulfur intake (15–20 mmol/day). Both sulfate and taurine are excreted in

the urine. In studies of total urinary sulfur excretion by children and adults, inorganic sulfate accounted for about 77 to 82%, ester sulfate about 8 to 9%, taurine about

3%, and cyst(e)ine about 0.6 to 0.7%. Other sulfur-containing compounds found in urine in trace amounts (<0.2% of total sulfur) include methionine, Hcy,

cystathionine, N-acetylcysteine, mercaptolactate, mercaptoacetate, thiosulfate, and thiocyanate ( 40, 103).

Cysteine Oxidation and Taurine Synthesis in Humans

Relatively little is known about the specific pathways of cysteine metabolism in humans. Taurine synthesis requires both cysteine dioxygenase and cysteine sulfinate

decarboxylase (EC 4.1.1.29). Significant expression of the cysteine dioxygenase gene was indicated by the presence of mRNA for cysteine dioxygenase in human

liver, kidney, and lung ( 104). S-Carboxymethyl-L-cysteine (SCMC) has been used as a marker substrate for cysteine metabolism, presumably by cysteine

dioxygenase, in vivo (105). Metabolism of SCMC is polymorphic, and about 20% of healthy individuals are poor S-oxidizers, based upon conversion of SCMC to

urinary metabolites, SCMC sulfoxide, or methylcysteine sulfoxide. A low capacity to oxidize SCMC has been observed in some individuals with liver diseases or

rheumatoid arthritis (105, 106). Individuals who exhibited low capacities for SCMC oxidation also had elevated cysteine:sulfate plasma ratios, excreted a smaller

percentage of a dose of acetaminophen as the sulfate (vs. the glucuronide conjugate), and had a lower sulfate concentration in synovial fluid, all of which are

consistent with impaired cysteine oxidation.

The human liver has been reported to have low cysteine sulfinate decarboxylase activity ( 107). Nevertheless, the adult human seems to have a significant ability to

synthesize taurine. In vivo assessment of the ability of adults to synthesize taurine, based upon incorporation of 18O (from inhaled 18O2) into taurine, resulted in

conservative estimates of synthesis in the range of 200 to 400 µmol/day ( 108). These estimates are equivalent to 1 to 3% of the total sulfur amino acid intake and

compare favorably with the mean taurine excretion observed in strict vegans consuming an essentially taurine-free diet (approximately 250 µmol/day [ 25, 26]). Thus,

the percentage of the sulfur amino acid intake or total urinary sulfur excretion that is represented by urinary taurine in humans fed taurine-free diets is similar to that

observed in rats fed taurine-free diets (2–6%) ( 109), which seems to dispute the often-made statement that the rat has a high capacity for taurine synthesis whereas

humans have a low capacity. Possibly, relatively high hepatic cysteine dioxygenase activity in man permits high rates of cysteine catabolism to cysteine sulfinate, and

relatively high concentrations of cysteine sulfinate allow adequate rates of taurine synthesis despite relatively low cysteine sulfinate decarboxylase activity.



ESSENTIAL FUNCTIONS OF CYSTEINE AND ITS METABOLITES

Synthesis of Protein and Glutathione

Cysteine, either preformed or synthesized from methionine and serine, is required for protein synthesis and, hence, growth or nitrogen balance. Both the reactive

sulfhydryl group of cysteinyl residues in proteins and the ability of these residues to form disulfide linkages play important roles in protein structure and function.

The Cys-containing tripeptide GSH has a number of essential functions in the body in addition to serving as a storage or circulating reservoir of Cys ( 110, 111).

Because GSH has a reactive sulfhydryl group, it can readily form disulfides with itself (oxidized glutathione or GSSG) or with other thiol compounds (GSSR). The ratio

of GSH to GSSG in most cells is greater than 500, so GSH serves as a supply of reducing equivalents or electrons. Glutathione is involved in protection of cells from

oxidative damage because of its role in reduction of hydrogen peroxide and organic peroxides via glutathione peroxidases and because of its ability to inactivate free

radicals by donating hydrogen to the radical; these processes result in oxidation of GSH to GSSG. GSSG and GSH can be interconverted via the glutathione

reductase reaction, which uses NADP +/NADPH as the oxidant/reductant; hence, glutathione plays a role in maintenance of the cellular redox state. GSH is an

important source of reducing equivalents for the intracellular reduction of cystine to cysteine, which can occur by thiol-disulfide exchange or enzymatically via

thioltransferase, with GSH providing the reducing equivalents.

GSH may participate in the transport of amino acids via the membrane-bound enzyme g-glutamyl transpeptidase, the same enzyme responsible for extracellular

hydrolysis of GSH. This enzyme catalyzes the transfer of the g-glutamyl group of GSH to the a-amino group of an acceptor amino acid such as cystine or glutamine.

The g-glutamyl amino acid is transported into the cell, where the amino acid is released and the glutamyl moiety cyclizes to 5-oxoproline, which is then hydrolyzed to

regenerate glutamate. The CysGly dipeptide that is the byproduct of g-glutamyl-transpeptidation can be hydrolyzed to Cys and glycine either extracellularly or

intracellularly by dipeptidases; hence, no net consumption of amino acids results from this transport cycle. The contribution of this proposed transport system to amino

acid transport cannot exceed the rate of GSH turnover, which is ~40 mmol/day in an adult.

GSH also serves as a cosubstrate for several reactions, including certain steps in leukotriene synthesis and melanin polymer synthesis. GSH is the substrate for a

group of enzymes, glutathione S-transferases, that form GSH conjugates from a variety of acceptor compounds including various xenobiotics ( 112). These conjugates

are normally degraded by the enzymes of the g-glutamyl cycle to yield cysteinyl derivatives that may be acetylated using acetyl-CoA to mercapturic acids, which are

excreted in the urine. This is usually a detoxification and excretion process.

Functions of Inorganic Sulfur

Reduced sulfur is required for synthesis of iron-sulfur proteins and other compounds. The activated form of sulfate, 3'-phospho-5'-phosphosulfate (PAPS), serves as

the substrate for a variety of sulfotransferase reactions. Many structural compounds are sulfated; in particular, the oligosaccharide chains of proteoglycans contain

many sulfated sugar residues. In addition, many compounds of both endogenous and exogenous origin are excreted as sulfoesters; sulfoesters of steroid hormones

and the drug acetaminophen are examples. Inorganic sulfur is largely obtained from the metabolism of cysteine in the body, but animal studies have suggested that

dietary inorganic sulfate may improve growth, feed efficiency, and sulfation of cartilage proteoglycans when sulfur amino acid intake is insufficient ( 113).

Functions of Taurine

The only physiologic function of taurine that is well understood is its role in bile acid conjugation ( 37). Taurine conjugates are the major metabolites of taurine formed

in vertebrates. Taurocholate is a very efficient bile salt because of the low pK a of the sulfonic acid group, which faciliates its ionization and hence detergent action,

solubility, slower reabsorption, and higher intraluminal concentration. Taurine is also a conjugation substrate for certain other compounds, such as all-trans-retinoic

acid, increasing polarity, aqueous solubility, and, in most cases, clearance from the body.

Humans can conjugate bile acids with both taurine and glycine. In adults, the taurocholate:glycocholate ratio is about 3:1, but this ratio varies from individual to

individual and with changes in the hepatic concentration of taurine. In contrast, the human fetus and neonate are exclusive taurine conjugators. Glycine conjugation is

not usually observed until about the 3rd week of life, but it appears sooner in infants deprived of dietary taurine ( 114). Taurine supplementation resulted in lower

cholesterol synthesis and higher bile acid excretion and fatty acid absorption in preterm infants with a gestational age less than 33 weeks but not in older preterm or

full-term infants (115).

Taurine is present in high concentrations in many human tissues (~25 µmol/g wet wt in retina and leukocytes), and a number of other physiologic actions of taurine in

various tissues have been hypothesized ( 15, 37). Unfortunately, these actions are not well understood despite several decades of intensive work ( 15, 37, 48). Taurine

is involved in osmoregulation in many marine invertebrates and fish and may also function as an organic osmolyte in mammals. Taurine seems to modulate many

Ca2+-dependent processes and to be involved in phospholipid/Ca +2 interactions. Taurine has been said to have “antioxidant or radioprotective” functions, but these

are probably largely secondary to its membrane-stabilizing actions. The metabolic precursor of taurine, hypotaurine, can function as an antioxidant; and the cysteine

derivative, cysteamine, derived from coenzyme A catabolism, is a good radioprotectant. Additionally, taurine facilitates removal of hypochlorite, a strong oxidant

generated from peroxide and Cl– by myeloperoxidase. Taurine reacts with hypochlorite to form N-chlorotaurine, which can then be reduced to taurine and Cl –.

N-Chlorotaurine may itself have a regulatory role in the inflammatory process.

Taurine is involved in development, with substantial evidence supporting a crucial role during the pre- and postnatal development of the central nervous and visual

systems. The specific manner in which taurine participates in these events is not clear. In primates deprived of taurine, retinal changes, impaired visual acuity, and

degenerative ultrastructural changes in photoreceptor outer segments have been observed, with changes being more severe in younger animals ( 37, 48). Some

human infants and children whose only nutrition was taurine-free parenteral infusion or taurine-devoid formulas have exhibited ophthalmoscopically and

electrophysiologically detectable retinal abnormalities and immature brainstem auditory-evoked responses ( 37, 48).



POSSIBLE CAUSES OF DEFICIENCY OF CYSTEINE OR TAURINE

Inability to Synthesize or Conserve

Immaturity

Immaturity may be associated with a conditional requirement for both cysteine and taurine. Preterm infants (£32 weeks gestation) have a low capacity for

transsulfuration (low cystathionine g-lyase activity), low plasma Cys concentrations, elevated plasma cystathionine concentrations, and a low rate of GSH synthesis

from methionine in erythrocytes (116, 117). These observations all suggest that transsulfuration may be insufficient to meet the cysteine requirements of the very

premature infant. Full-term formula-fed infants have also been observed to have increased cystathionine and decreased taurine levels in urine, suggesting a limited

capacity for transsulfuration even in term infants ( 118).

In addition to a limited capacity to convert methionine to cysteine and hence to taurine (low synthetic rate), several other characteristics of premature infants

contribute to their conditional requirement for taurine and/or cysteine ( 15, 48). First, the premature infant may have a greater requirement for cysteine because of

more rapid growth and for taurine because of a likely role of taurine in development of the nervous and visual systems. The brain and retina of developing animals

have high taurine concentrations, and morphologic and functional impairments have been observed in animals deprived of taurine during development. Second,

premature infants are born with lower stores of taurine than are mature infants. Third, the b-amino acid transport system in the immature kidney does not adapt to

poor taurine status by increasing reabsorption of taurine. The urinary taurine content of premature neonates is markedly elevated, with a fractional excretion ranging

from 38 to 60%, compared with fractional excretion below 10% in term infants. Premature infants who received parenteral nutrition solutions devoid of taurine had high

urinary taurine excretion rates despite very low plasma taurine values ( 47, 119, 120). By contrast, term neonates given a taurine-deficient parenteral nutrition solution

can maintain plasma taurine concentrations by increased renal reabsorption of taurine with as little as 1% of the filtered taurine load being excreted. (See also

Chapter 51 concerning taurine administration parenterally.)

Hepatic Dysfunction

Because liver is the major site for transsulfuration and taurine synthesis, hepatic dysfunction can adversely affect sulfur amino acid status. Patients with advanced

forms of liver dysfunction or cirrhosis had low plasma taurine, Cys, and glutathione concentrations, an elevated plasma cystathionine concentration, decreased

urinary taurine excretion, a decreased ratio of urinary sulfate to total sulfur, and increased urinary excretion of Cys and cystathionine ( 121, 122). These patients

appeared to have a decreased ability to metabolize methionine (to cysteine, with cystathionine accumulation) and cysteine (to taurine and inorganic sulfate, with

thiosulfate, Cys, and N-acetylcysteine accumulation).

Inadequate Supply Relative to Need

Total Parenteral Nutrition

Patients on long-term total parenteral nutrition (TPN) have experienced adverse effects on their sulfur amino acid status, both because of the route of administration

and the composition of the TPN solutions. The amino acid mixtures used for TPN solutions usually contain little if any cysteine, because cysteine is rapidly converted

to its disulfide, cystine, which is very insoluble in aqueous solution. Taurine is not routinely added to adult TPN solutions. Hence, patients on TPN must synthesize

both cysteine and taurine from the methionine provided by TPN (see Chapter 101). However, synthesis of cysteine and taurine from methionine is restricted when

first-pass metabolism by the liver is bypassed with parenteral alimentation. In adult subjects given parenteral alimentation solutions free of Cys via different routes,

plasma Cys concentration dropped markedly when the feeding was via the parenteral route whereas it rose when feeding was switched to the oral route ( 123). The

liver apparently removes much of the methionine on the first pass when solutions are administered by the oral route, thus facilitating cysteine and taurine synthesis

from methionine.

Vegan Diets

Because strict vegan diets tend to be lower in total sulfur amino acid content and virtually free of taurine, adult vegans and particularly children consuming vegan diets

are at somewhat greater risk of inadequate sulfur amino acid status. Adult humans who consume a strict vegetarian diet have been reported to have lower plasma

taurine concentrations and greatly reduced urinary taurine excretion compared with omnivores. However, vegans consuming little or no preformed taurine are healthy,

and the children born to and nursed by vegan mothers have normal growth and development ( 25). (See Chapter 106.)

Drug Metabolism

Various drugs and toxins are partially metabolized and excreted by conjugation with sulfate, glutathione (mercapturic acid synthesis), or even taurine. Rats fed up to 1

g (6.6 mmol) of acetaminophen per 100 g diet experienced dose-dependent inhibition of growth that was independent of hepatotoxicity and that could be overcome by

addition of methionine or cyst(e)ine to the diet ( 99, 124). Lauterburg and Mitchell ( 99) found that therapeutic doses of acetaminophen (600 and 1200 mg, or 4 and 8

mmol) administered to healthy adult subjects markedly stimulated the rate of turnover of the pool of Cys available for synthesis of GSH. Patients and volunteers with

prolonged ingestion of acetaminophen in doses of 2 to 4 g (13–26 mmol) per day had a decreased urinary output of inorganic sulfate but no decrease in plasma

sulfate concentrations (125). Subjects produced a maximum of 0.6 mmol/h of acetaminophen sulfate, whereas total sulfur excretion was 7.5 to 26.7 mmol/24 h

(0.3–1.1 mmol/h). A marginal sulfur amino acid intake accompanied by prolonged ingestion of high doses of drugs or toxins that are metabolized by sulfate and/or

glutathione conjugation could have adverse effects on both sulfur amino acid status and drug metabolism.



MEASURES OF TAURINE STATUS AND OF CYSTEINE AND/OR SULFUR AMINO ACID STATUS

Sulfur amino acid adequacy has generally been assessed by measures of nitrogen balance or growth. Although growth and nitrogen balance have been used to

define the nutritional requirements for amino acids, they are not necessarily good indicators of whether or not sulfur amino acid intake is sufficient for optimal rates of

production of glutathione, inorganic sulfur, or taurine.

Plasma and blood taurine concentrations, plasma tCys concentration, and the plasma glutathione concentration have been used as indicators of sulfur amino acid

status. Normal values for these measures are discussed above in this chapter.

Because most of the sulfur from sulfur amino acids is excreted in the urine, primarily as inorganic sulfate, measures of total urinary sulfur or of urinary sulfate are

useful indicators of sulfur amino acid intake and/or metabolism. The urinary taurine level can also be used as an indicator of adequate supply of sulfur amino acids or

taurine because taurine excretion increases as plasma taurine concentration and/or taurine intake increases.



RECOMMENDED INTAKES FOR HUMANS

The estimated upper-end requirement for sulfur amino acids (methionine plus cyst(e)ine) is 58 mg/kg/day for infants, 27 mg/kg/day for children (~age 2 years), 22

mg/kg/day for older children (~10–12 years), and 13 mg/kg/day for adults ( 28). A conservative estimate of methionine replacement by cyst(e)ine, up to 30% of the

methionine requirement, is suggested ( 28). In cases of limited ability to convert methionine to cysteine (whether due to hepatic dysfunction, inborn errors of

methionine metabolism to cysteine, or prematurity), the total amount of sulfur amino acids in the diet, the balance of cysteine and methionine, and the adequacy of

taurine should all be considered.

By consensus, taurine is considered conditionally essential during infant development and probably for adults in some special circumstances. Because brain and

retina of human infants are not fully developed at birth and may be vulnerable to the effects of taurine deprivation, it has been judged prudent to supplement human

infant formulas and pediatric feeding solutions with taurine ( 15, 37). During the 1980s, manufacturers of infant formulas began adding taurine to their products, and

taurine is presently added to virtually all human infant formulas and pediatric parenteral solutions throughout the world. The taurine content of human milk has been

used as a guideline for supplementation levels.



TOXICITY

Large doses of cysteine or cystine are neuroexcitotoxic in several species. The effect of cysteine seems to involve the N-methyl-D-aspartate subtype of the glutamate

receptor for which cysteine sulfinate and cysteic acid are agonists ( 125, 126, 127, 128 and 129). Single injections of cysteine (0.6–1.5 g/kg) into 4-day-old rat pups

resulted in massive damage to cortical neurons ( 130), permanent retinal dystrophy (131), atrophy of the brain (132), and hyperactivity (126). Cats fed a 5% cystine

diet had no obvious immediate ill effects but exhibited acute neurotoxic symptoms after several months ( 133). The onset of symptoms in the cats was sudden, with

rapid progression to a moribund state or death, usually within 48 hours. The morphologic changes observed in the retinas of cats fed the cystine-rich diets were

comparable to those described in the retinas of young rodents treated with glutamate or certain other acidic or excitotoxic amino acids. It is not clear whether cysteine

or a metabolite (cysteine sulfinate) is responsible for the cytotoxicity. These observations have given rise to concerns about administration of excess cyst(e)ine to

humans, especially to infants.

Studies in rodents also demonstrated influences of dietary sulfur amino acids on lipid metabolism, with 2 to 5% (by wt) L-cystine resulting in elevated plasma

cholesterol concentration, increased hepatic cholesterol biosynthesis, and depressed plasma ceruloplasmin activity ( 134, 135). Excess L-cysteine (0.8 or 2% of diet

by wt) did not result in an elevation in plasma cholesterol whereas addition of 0.8% L-methionine did ( 135, 136).

Sturman and Messing (137) found no evidence of adverse effects of prolonged feeding of high taurine diets (up to 1 g [8 mmol]/100 g diet) on adult female cats or

their offspring. In fact, taurine may protect against toxic effects of some other compounds. Taurine addition to cat diets provided some protection against the adverse

effects of a high level of cystine, supporting a neuroprotective role of taurine against the excitotoxic damages in the mammalian nervous system ( 133). Studies in

rodents have suggested that dietary taurine also has hypolipidemic and antiatherosclerotic effects ( 138, 139).

Abbreviations: Cys—cysteine (any form), with thiol and disulfide forms indicated as CySH, CySSCy, and CySSR; tCys—sum of all forms of Cys including that present as thiol, half-disulfide, mixed

disulfide, and protein-bound disulfide; Cyst(e)ine—cysteine and/or cystine; Hcy—homocysteine (any form) with thiol and disulfide forms indicated as HcySH, HcySSHcy, and HcySSR; tHcy—sum of

all forms of Hcy; Homocyst(e)ine—homocysteine and/or homocystine; Glu—glutamate; Gly—glycine; GSH, glutathione; SCMC—S-carboxymethylcysteine; THF, tetrahydrofolate.



CHAPTER REFERENCES

1.

2.

3.

4.

5.

6.

7.

8.

9.

10.

11.

12.

13.

14.

15.

16.

17.

18.

19.

20.

21.

22.

23.

24.

25.

26.

27.

28.

29.

30.

31.

32.

33.

34.

35.

36.

37.

38.

39.

40.

41.

42.

43.

44.

45.

46.

47.

48.

49.

50.

51.

52.

53.

54.

55.

56.

57.

58.

59.

60.

61.

62.

63.

64.

65.

66.

67.

68.

69.

70.

71.

72.

73.

74.

75.

76.

77.

78.

79.

80.

81.



Osborne TB, Mendel LR. J Biol Chem 1915;20:351–78.

Mueller JH. Proc Soc Exp Biol Med 1921;19:161–3.

Jackson RW, Block RJ. J Biol Chem 1932;98:465–77.

Womack M, Kemmerer KS, Rose WC. J Biol Chem 1937;121:403–10.

Rose WC, Wixom RL. J Biol Chem 1955;216:763–73.

du Vigneaud VE. Trail of research in sulfur chemistry and metabolism and related fields. Ithaca, NY: Cornell University Press, 1952.

Carson NAJ, Neill DW. Arch Dis Child 1962;37:505–13.

Clarke R, Daly L, Robinson K, et al. N Engl J Med 1991;324:1149–55.

Robinson K, Mayer E, Jacobsen DW. Cleve Clin J Med 1994;61:438–50.

Steegers-Theunissen RPM, Boers GHJ, Trijbels FJM, et al. Metabolism 1994;43:1475–80.

Wouters MCAJ, Boers GHJ, Blom HJ, et al. Fertil Steril 1993;60:820–5.

Tiedemann F, Gmelin L. Ann Physik Chem 1827;9:326–37.

Hayes KC, Carey RE, Schmidt SY. Science 1975;188:949–51.

Sturman JA, Rassin DK, Gaull GE. Life Sci 1977;21:1–22.

Sturman JA. Physiol Rev 1993;73:119–47.

Mansoor MA, Bergmark C, Svardal AM, et al. Arterioscler Thromb Vasc Biol 1995;15:232–40.

Andersson A, Isaksson A, Brattstrom L, et al. Clin Chem 1993;39:1590–7.

Mansoor MA, Ueland PM, Svardal AM. Am J Clin Nutr 1994;59:631–5.

Guttormsen AB, Schneede J, Fiskerstrand R, et al. J Nutr 1994;124:1934–41.

Selhub J, Jacques PF, Wilson PWF, et al. JAMA 1993;270:2693–8.

Dalery K, Lussier-Cacan S, Selhub J, et al. Am J Cardiol 1995;75:1107–11.

Nygard O, Vollset SE, Refsum H, et al. JAMA 1995;274:1526–33.

Rasmussen K, Moller J, Lyngbak M, et al. Clin Chem 1996;42:630–6.

Trautwein EA, Hayes KC. Am J Clin Nutr 1990;52:758–64.

Rana SK, Sanders TAB. Br J Nutr 1986;56:17–27.

Laidlaw SA, Shultz TD, Cecchino JT, et al. Am J Clin Nutr 1988;47:660–3.

Malinow MR, Axthelm MK, Meredith MJ, et al. J Lab Clin Med 1994;123:421–9.

National Research Council. Recommended dietary allowances. 10th ed. Washington, DC: National Academy Press, 1989.

US Department of Agriculture. Agricultural handbook no. 8. Agriculture Research Service, United States Department of Agriculture, 1976–1986.

Young VR, Wagner DA, Burini R, et al. Am J Clin Nutr 1991;54:377–85.

Storch KJ, Wagner DA, Burke JF, et al. Am J Physiol 1988;255:E322–31.

FAO/WHO/UNI. Energy and protein requirements. WHO technical report #724. Geneva: World Health Organization, 1985.

Laidlaw SA, Grosvenor M, Kopple JD. JPEN 1990;14:183–8.

Pasantes-Morales H, Quesada O, Alcocer L, et al. Nutr Rep Int 1989;40:793–801.

Roe DA, Weston MO. Nature 1965;203:287–8.

Kataoka H, Ohnishi N. Agric Biol Chem 1986;50:1887–8.

Huxtable RJ. Physiol Rev 1992;72:101–63.

Rassin DK, Sturman JA, Gaull GE. Early Hum Dev 1978;2:1–13.

Paauw JD, Davis AT. Am J Clin Nutr 1994;60:203–6.

Martensson J, Hermansson G. Metabolism 1984;33:425–8.

Palacin M, Chillaron J, Mora C. Biochem Soc Trans 1996;24:856–63.

Mora C, Chillaron J, Calonge MJ, et al. J Biol Chem 1996;271:10569–76.

Chillaron J, Estevez R, Mora C, et al. J Biol Chem 1996;271:17761–70.

Sakhaee K. Miner Electrolyte Metab 1994;20:414–23.

Refsum H, Helland S, Ueland PM. Clin Chem 1985;31:624–8.

Erbe RW. Inborn errors of folate metabolism. In: Blakley RL, Whitehead VM, eds. Folates and pterins, vol 3. New York: John Wiley & Sons, 1986;413–65.

Jensen H. Biochim Biophys Acta 1994;1194:44–52.

Sturman JA, Chesney RW. Pediatr Nutr 1995;42:879–97.

McKeever MP, Weir DG, Molloy A, et al. Clin Sci 1991;81:551–6.

Poole JR, Mudd SH, Conerly E, et al. J Clin Invest 1975;55:1033–48.

Stipanuk MH. Annu Rev Nutr 1986;6:179–209.

Backlund PS Jr, Smith RA. J Biol Chem 1981;256:1533–5.

Storch KJ, Wagner DA, Burke JF, et al. Am J Physiol 1990;258:E790–8.

Selhub J, Miller J. Am J Clin Nutr 1992;55:131–8.

Stipanuk MH, Benevenga NJ. J Nutr 1977;107:1455–67.

Storch KJ, Wagner DA, Young VR. Am J Clin Nutr 1991;54:386–94.

Shih VE, Fringer JM, Mandell R, et al. Am J Hum Genet 1995;57:34–9.

Mudd SH, Levy HL, Skovby F. Disorders of transsulfuration. In: Scriver CR, Beaudet AL, Sly WS, Valle D, eds. The metabolic and molecular bases of inherited disease, vol 1. New York:

McGraw-Hill, 1995;1279–327.

Kraus JP. J Inherited Metab Dis 1994;17:383–90.

Sebastio G, Sperandeo MP, Panico M, et al. Am J Hum Genet 1995;56:1324–33.

Goyette P, Frosst P, Rosenblatt DS, et al. Am J Hum Genet 1995;56:1052–9.

Rozen R. Clin Invest Med 1996;19:171–8.

Fenton WA, Rosenberg LE. Inherited disorders of cobalamin transport and metabolism. In: Scriver CR, Beaudet AL, Sly WS, Valle D, eds. The metabolic and molecular bases of inherited

disease, vol 3. New York: McGraw-Hill, 1995;3129–49.

Engbersen AMT, Franken DG, Bower GHJ, et al. Am J Hum Genet 1995;56:142–50.

Jacques PF, Bostom AG, Williams RR, et al. Circulation 1996;93:7–9.

Kang S-S, Wong PWK, Bock H-GO, et al. Am J Hum Genet 1991;48:546–51.

Rozen R, Jacques P, Bostom A, et al. Am J Hum Genet 1995;57S:A250.

Kang S-S, Wong PWK, Susmano A, et al. Am J Hum Genet 1991;48:536–46.

Van der Put NMJ, Steegers-Theunissen RPM, Frosst P, et al. Lancet 1995;346:1070–1.

Guttormsen AB, Schneede J, Ueland PM, et al. Am J Clin Nutr 1996;63:194–202.

Verhoef P, Stampfer MJ, Buring JE, et al. Am J Epidemiol 1996;143:845–59.

Ubbink JB, van der Merwe A, Delport R, et al. J Clin Invest 1996;98:177–84.

Boushey CJ, Beresford SAA, Omenn GS, et al. JAMA 1995;274:1049–57.

Ubbink JH, Becker PJ, Vermaak WJH, et al. Clin Chem 1995;41:1033–7.

Schmitz C, Lindpaintner K, Verhoef P, et al. Circulation 1996;94:1812–4.

Arnadotti M, Hultberg B, Nilsson-Ehle P, et al. Scand J Clin Invest 1996;56:41–6.

Chauveau P, Chadefaux B, Conde M, et al. Miner Electrolyte Metab 1996;22:106–9.

den Heijer M, Koster R, Blom HJ, et al. N Engl J Med 1996;334:759–62.

Mills JL, Scott JM, Kirke PN, et al. J Nutr 1996;126:756S–60S.

Selhub J, Jacques PF, Bostom AG, et al. N Engl J Med 1995;332:286–91.

Goddijn-Wessel TAW, Wouters MGAJ, Van de Molen EF. Eur J Obstet Gynecol Reprod Biol 1996;66:23–9.



82.

83.

84.

85.

86.

87.

88.

89.

90.

91.

92.

93.

94.

95.

96.

97.

98.

99.

100.

101.

102.

103.

104.

105.

106.

107.

108.

109.

110.

111.

112.

113.

114.

115.

116.

117.

118.

119.

120.

121.

122.

123.

124.

125.

126.

127.

128.

129.

130.

131.

132.

133.

134.

135.

136.

137.

138.

139.



Kluijtmans LAJ, van den Heuvel LPWJ, Boers GHJ, et al. Am J Hum Genet 1996;58:35–41.

Harpel PC, Zhang X, Borth W. J Nutr 1996;126:1285S–9S.

Blom HJ, Van der Molen EF. Fibrinolysis 1994;8(Suppl 2):86–7.

Lentz SR, Sobey CG, Piegors DJ, et al. J Clin Invest 1996;98:24–9.

Tsai JC, Wang H, Perrella MA, et al. J Clin Invest 1996;97:146–53.

Moephuli SR, Klein NW, Baldwin MT, et al. Proc Natl Acad Sci USA 1997;94:543–8.

Eskes TKAB. Eur J Obstet Gynecol Reprod Biol 1997;71:105–11.

Bostom AG, Jacques PF, Nadeau MR, et al. Atherosclerosis 1995;116:147–51.

Tsai MY, Garg U, Key NS, et al. Atherosclerosis 1996;122:69–77.

Dudman NPB, Guo X-W, Gordon RB, et al. J Nutr 1996;126:12295S–300S.

Zeisel J, Blusztajn JK. Annu Rev Nutr 1994;14:269–96.

Rosenblatt DS. Inherited disorders of folate transport and metabolism. In: Scriver CR, Beaudet AL, Sly WS, Valle D, eds. The metabolic and molecular bases of inherited disease, vol 3. 7th ed.

New York, McGraw-Hill, 1995;3111–28.

Brattstrom LE, Israelsson B, Jeppsson JO, et al. Scand J Clin Lab Invest 1988;48:215–21.

Ubbink JB, Vermaak WJH, van der Merwe A, et al. J Nutr 1994;124:1927–33.

Kang S-S. J Nutr 1996;126:1273S–5S.

Fortin LJ, Genest J Jr. Clin Biochem 1996;28:155–62.

Lieberman MW, Wiseman AL, Shi ZZ, et al. Proc Natl Acad Sci USA 1996;93:7923–6.

Lauterburg BH, Mitchell JR. J Hepatol 1987;4:206–211.

Fukagawa NK, Ajami AM, Young VR. Am J Physiol 1996:270:E209–14.

Crawhall JC. Clin Biochem 1985;18:139–42.

Bella DL, Kwon YH, Stipanuk MH. J Nutr 1996;126:2179–87.

Martensson J. Metabolism 1982;31:487–92.

Koide T, Watanabe M, Shimada M. Acta Histochem Cytochem 1994;27:384.

Davies MH, Ngong JM, Pean A, et al. J Hepatol 1995;22:551–60.

Bradley H, Gough A, Sokhi RS, et al. J Rheumatol 1994;21:1192–6.

Gaull GE, Rassin DK, Raiha NCR, et al. J Pediatr 1977;90:348–55.

Irving CS, Marks L, Klein PD, et al. Life Sci 1986;38:491–5.

Bella DL, Stipanuk MH. Am J Physiol 1995;269:E910–7.

DeLeve LD, Kaplowitz N. Pharmacol Ther 1991;52:287–305.

Meister A. Pharmacol Ther 1991;51:155–94.

Hinchman CA, Ballatori N. J Toxicol Environ Health 1994;41:387–409.

Sasse CE, Baker DH. Dev Brain Dysfunct 1994;7:230–6.

Brueton MJ, Berger HM, Brown GA, et al. Gut 1978;19:95–8.

Wasserhess P, Becker M, Staab D. Am J Clin Nutr 1993;58:349–53.

Miller RG, Jahoor F, Jaksic T. J Pediatr Surg 1995;30:953–8.

Vina J, Vento M, Garcia-Sala F, et al. Am J Clin Nutr 1995;61:1067–9.

Martensson J, Finnstrom O. Early Hum Dev 1985;11:333–9.

Zelikovic I, Chesney RW, Friedman AL, et al. J Pediatr 1990;116:301–6.

Helms RA, Christensen ML, Storm MC, et al. J Nutr Biochem 1995;6:462–6.

Chawla RK, Berry CJ, Kutner MH, et al. Am J Clin Nutr 1985;42:577–84.

Martensson J, Foberg U, Fryden A, et al. Scand J Gastroenterol 1992;27:405–11.

Stegink LD, den Besten L. Science 1972;178:514–6.

McLean AEM, Armstrong GR, Beales D. Biochem Pharmacol 1989;38:347–52.

Blackledge HM, O'Farrell JO, Minton NA, et al. Hum Exp Toxicol 1991;10:159–65.

Mathisen GA, Fonnum F, Paulsen RE. Neurochem Res 1996;21:293–8.

Santucci AC, Spincola L-J. Dev Brain Dysfunct 1994;7:230–6.

Porter RHP, Roberts PJ. Neurosci Lett 1993;154:78–80.

Zerangue N, Kavanaugh MP. J Physiol 1996;493:419–23.

Sandberg M, Orwar O, Hehmann A. J Neurochem 1991;57:S152.

Pedersen OO, Lund-Karlsen R. Invest Ophthalmol Vis Sci 1980;19:886–92.

Lund-Karlsen R, Grofova I, Malthe-Sorenssen D, et al. Brain Res 1981;208:167–80.

Imaki H, Sturman JA. Nutr Res 1990;10:1385–400.

Serougne C, Ferezou J, Rukaj A. Biochim Biophys Acta 1987;921:522–30.

Yang B-S, Wan Q, Kato N. Biosci Biotech Biochem 1994;58:1177–8.

Sugiyama K, Akai H, Muramatsu K. J Nutri Sci Vitaminol 1986;32:537–49.

Sturman JA, Messing JM. J Nutr 1992;122:82–8.

Murakami S, Yamagishi I, Asami Y, et al. Pharmacology 1996;52:303–13.

Kamata K, Sugiura M, Kojima S, et al. Eur J Pharmacol 1996;303:47–53.



SELECTED READINGS

Boushey CJ, Beresford SAA, Omenn GS, et al. A quantitative assessment of plasma homocysteine as a risk factor for vascular disease. Probable benefits of increasing folic acid intakes. JAMA

1995;274:1049–57.

DeLeve LD, Kaplowitz N. Glutathione metabolism and its role in hepatotoxicity. Pharmacol Ther 1991;52:287–305.

Kang S-S. Treatment of hyperhomocyst(e)inemia: physiological basis. J Nutr 1996;126:1273S–5S.

Rozen R. Molecular genetic aspects of hyperhomocysteinemia and its relation to folic acid. Clin Invest Med 1996;19:171–8.

Stipanuk MH. Metabolism of sulfur-containing amino acids. Annu Rev Nutr 1986;6:179–209.

Sturman JA. Taurine in development. Physiol Rev 1993;73:119–47.



Chapter 35. Glutamine and Arginine

Modern Nutrition in Health and Disease



Chapter 35. Glutamine and Arginine

STEVE F. ABCOUWER and WILEY W. SOUBA

Glutamine Metabolism

Glutamine as a Nitrogen Carrier

Glutamine as a Metabolic Intermediate

Glutamine as a Supporter of Glutathione Metabolism

Glutamine as an Energy Source

Glutamine in Catabolic States

Interorgan Glutamine Transfer

Hormonal Influences on Glutamine Synthesis

Muscle-Sparing Effect of Glutamine

Glutamine Effect on Intestinal Repair and Function

Glutamine and Cancer

Arginine Metabolism

Arginine and Nitrogen Balance

Arginine as a Metabolic Intermediate

Arginine as a Source of Nitric Oxide

Arginine in Catabolic States

Arginine Deficiency in Catabolic States

Effect of Arginine upon Intestinal Function and Repair

Effect of Arginine on Wound Healing

Effect of Arginine on Endocrine Systems

Arginine and the Immune System

Arginine in Cancer

Chapter References



The amino acids glutamine (GLN) and arginine (ARG) are of special interest because they have been recently classified as conditionally essential amino acids,

indicating that they may be required in increased amounts by patients suffering from a catabolic insult such as injury, cancer, or severe infection (see Chapter 1).

However, the utility of including glutamine or arginine in the nutritional support of patients recovering from surgery or suffering from trauma, infection, or cancer has

yet to be conclusively demonstrated in clinical settings. Most studies examining the metabolism of these two amino acids have used cultured cells and animal models

of trauma or infection. A wealth of experimental data proves the ability of glutamine-supplemented parenteral nutrition to support gut function, improve nitrogen

balance, and alleviate catabolic demands upon muscle mass. In addition, limited clinical trials have found that glutamine feeding improves patient outcome in a

number of settings (Table 35.1). Similarly, experimental data have shown that arginine can promote intestinal function and repair, promote wound healing, stimulate

endocrine hormone production, and improve immune system function. Limited clinical trials have confirmed that inclusion of arginine in enteral and parenteral nutrition

can improve nitrogen balance and immune defenses of patients (see [ 1] for a recent review). This chapter reviews a selection of these experimental and clinical

studies and examines the limitations of these two nutrients as therapeutic adjuvants.



Table 35.1 Clinical Studies Demonstrating Benefits of Supplementing TPN with Glutamine (GLN)



GLUTAMINE METABOLISM

Glutamine as a Nitrogen Carrier

The presence of two nitrogen atoms, an a-amino group and an amide group, as well as the abundance of glutamine in plasma combine to make this amino acid the

most important “nitrogen shuttle” in the body. In fact, approximately one-third of all amino acid–derived nitrogen transported by the blood is in the form of glutamine.

This amino acid serves as a vehicle for transportation of ammonia in a nontoxic form from peripheral tissues to visceral organs where it can be excreted as ammonium

by the kidneys or converted to urea by the liver. The vast majority of nitrogen supplied by muscle in the postprandial period is exported as glutamine ( 2). Ammonia

derived from glutamine in the mitochondria (by hydrolysis of glutamine to glutamate by the enzyme glutaminase or by hydrolysis of glutamate to a-ketoglutarate by the

enzyme glutamate dehydrogenase) can combine with CO 2 to form carbamoyl phosphate that then enters the urea cycle (3). The amino group of glutamate can be

used in the synthesis of aspartate from oxaloacetate. Through aspartate, the amino nitrogen of glutamine can also enter the urea cycle. In fact, glutamine is the major

source of both nitrogen used in hepatic ureagenesis ( 4) and nitrogen excreted in urine ( 5). In addition, perfusion of isolated rat livers with glutamine greatly stimulated

urea output (6, 7).

The liver has evolved an elegant mechanism for concerted ammonia detoxification and glutamine supply ( 8). In the periportal region, high incoming concentrations of

ammonia from the splanchnic bed are utilized for urea synthesis while at the same time ammonia and glutamate are being generated from incoming glutamine by the

hepatic isozyme of the enzyme glutaminase. In the perivenous portion of the liver, high levels of glutamine synthetase convert glutamate and excess ammonia into

glutamine, effectively reducing the ammonia concentrations in hepatic venous blood to non-toxic levels. With this intraorgan glutamine cycle, the liver is able to

effectively detoxify incoming blood of ammonia and control output of glutamine. In contrast, the kidney counteracts acidosis by utilizing a the “kidney-type” isozyme of

glutaminase to generate ammonia from glutamine (9). The kidney disposes of excess nitrogen by consuming glutamine and excreting the ammonia produced ( 10).

Glutamine as a Metabolic Intermediate

Glutamine also plays a key role in cellular energetics and metabolism by functioning as an important source of cellular fuel as well as a source of carbon and nitrogen

for metabolic intermediates and macromolecular synthesis ( Fig. 35.1). Glutamine acts as a fuel through its partial oxidation to form lactate or its complete oxidation to

form CO2. In fact, glutamine can serve as the primary respiratory substrate in enterocytes, lymphocytes, and cells in culture (discussed below). Glutamine plays a key

role in regulating protein synthesis in tissues and cultured cells ( 11, 12 and 13). Glutamine itself is used in peptide synthesis and is a precursor to many other amino

acids. In addition, glutamine is converted to numerous metabolic intermediates and donates an amine group in many metabolic pathways including those leading to

purine and pyrimidine nucleotide synthesis and the formation of glucosamine. Acetyl groups derived from glutamine are used in the synthesis of fatty acids and

therefore incorporated into membrane phospholipids. In the kidney and liver, the carbon skeleton of glutamine is used for gluconeogenesis during times of starvation

(14, 15). Thus, glutamine has many important metabolic and synthetic roles essential for cellular viability.



Figure 35.1. Metabolic functions of glutamine (GLN). Glutamine enters the cell by an active carrier-mediated process. Most utilized glutamine is converted to

glutamate (GLU) through hydrolysis catalyzed by the enzyme glutaminase (GA) and by a number of transaminase enzymes (TAs). Glutamate is used in protein

synthesis, used for glutathione (GSH) synthesis, and potentially converted back into glutamine by the enzyme glutamine synthetase (GS). GLU is also exported from

the cell in exchange for import of cystine (CYS2). Imported cystine is converted to cysteine, which together with glutamate and glycine (GLY) form glutathione. GLU is

converted to proline or converted to a-ketoglutarate by glutamate dehydrogenase (GDH) or transaminases. As a-ketoglutarate, the carbon backbone of glutamine

enters the TCA cycle. Once in the TCA cycle the glutamine-derived carbons can be oxidized to CO 2 or serve as a backbone for formation of numerous metabolites

and amino acids. In addition to glutamate and lactate, appreciable amounts of glutamine carbons are exported from the cell as aspartate, citrate, malate, pyruvate,

alanine, and proline ( 16).



The first step in the use of glutamine as a respiratory fuel, metabolic precursor, and nitrogen donor is its conversion to glutamate (GLU). Glutamate is the most

abundant intracellular amino acid in most tissues, with the notable exception of muscle, in which intracellular glutamine levels exceed those of glutamate ( 16, 17).

Glutamate and ammonia are formed by hydrolysis of glutamine's amide group by the enzyme glutaminase. Glutamate is also formed by transfer of glutamine's amide

group, catalyzed by a number of transaminase enzymes. Enzymatic transfer of glutamine's amide nitrogen occurs in various synthetic reactions. For example,

glutamine is the nitrogen donor in formation of cytosolic carbamoyl phosphate, which is then used with aspartic acid in the committed step in pyrimidine synthesis

(formation of N-carbamoylaspartate). Transamination from glutamine occurs in synthesis of the pyrimidine nucleotide CTP from UTP. In the purine biosynthetic

pathway, glutamine contributes two amine groups in reactions leading to the synthesis of IMP and donates a third amine group in the conversion of IMP to GMP.

Glutamine also provides an amide group for synthesis of asparagine, glucosamine, and nicotinamide adenine dinucleotide (NAD +). Glutamate formed from glutamine

can in turn be further deaminated, converted to proline or back into glutamine, used for glutathione synthesis, or exported. As glutamate, the a-amino group of

glutamine is used in several transamination reactions including synthesis of alanine from pyruvate, synthesis of aspartate from oxaloacetate, and formation of

phosphoserine, which is subsequently hydrolyzed to serine.

The five-carbon backbone of glutamine enters the tricarboxylic acid (TCA) cycle through conversion of glutamine-derived glutamate to a-ketoglutarate (2-oxoglutarate)

by glutamate dehydrogenase or one of several transaminases. Via the TCA cycle, glutamine serves as a respiratory fuel source and the precursor of many synthetic

intermediates. For example, through oxaloacetate, a TCA cycle intermediate, glutamine is used for gluconeogenesis or converted to aspartate or alanine. Aspartate

can in turn be converted to asparagine, methionine, threonine, isoleucine, and lysine, be a precursor in pyrimidine synthesis, and again be an amine donor in purine

synthesis (discussed above). Through malate and then pyruvate (the precursor of alanine), the carbon backbone of glutamine can be converted to lactate or

acetyl-CoA. The acetyl group of acetyl-CoA can reenter the TCA cycle, eventually leading to the complete respiratory oxidation of glutamine carbons to CO 2.

Acetyl-CoA also provides glutamine-derived carbon in the synthesis of fatty acids that can be used in production of phospholipids for cellular membranes.

Glutamine as a Supporter of Glutathione Metabolism

Considerable evidence suggests that glutamine may play a key role in the support of glutathione synthesis ( Fig. 35.1) (see also Chapter 34). Glutathione, a tripeptide

composed of glutamate, cysteine, and glycine, is the major store of cellular reducing equivalents and serves to protect cells from oxidative stress ( 18). As a source of

intracellular glutamate, glutamine provides one of the constituents of glutathione. Glutamine carbons are incorporated into glutathione by rat kidney glomeruli ( 19),

and the decline of hepatic glutathione levels during acetaminophen-induced liver injury was inhibited by treating rats with glutamine-supplemented parenteral nutrition

(20). In fact, radioactive glutamine carbons are incorporated into glutathione in several animal tissues ( 20). Glutamine may also be essential for maintenance of

cellular cysteine levels. Bannai and Ishii demonstrated a functional link between the use of glutamine and cystine (CYS2) by human diploid fibroblasts when they

found that both the quantity of glutamine used and the quantity of glutamate released by these cells depended upon the concentration of cystine in the culture media

(21). They concluded that approximately one-third of the glutamine used by these fibroblasts was used to produce extracellular glutamate. Tracer studies

demonstrated that exported glutamate is a major end product of glutamine used by kidney cells ( 22) as well as several tumor cell lines ( 16). In our laboratory, we have

found that approximately 40% of glutamine consumed by human breast cell lines and breast cancer cells is released as glutamate, and the rate of glutamate release

by these cells also depends upon the extracellular concentration of cystine ( 16a). A working model predicts that glutamine, by serving as a source of glutamate,

provides two of the three precursors for glutathione synthesis: glutamate itself, which is directly utilized in glutathione synthesis, and cysteine, which is derived from

cystine imported via the export of glutamate. This role in glutathione synthesis suggests that glutamine availability may have profound effects upon cellular redox

control and that glutamine utilization may be increased under conditions of oxidative stress.

Glutamine as an Energy Source

A substantial portion of glutamine can be used for cellular respiration. For example, HeLa and CHO cells each convert approximately 30% of catabolized glutamine

carbons to CO 2 and 15% to lactate, while approximately 20% of its carbons are incorporated into macromolecules ( 23, 24). The total amount of cellular energy derived

from glutamine depends upon the extent of oxidation and the rate of glutamine utilization. These factors in turn depend largely on the absolute amounts and relative

proportions of glutamine and glucose available as well as the type and proliferative state of the cell. For example, glutamine is a primary fuel source for the intestine

and for enterocytes in culture (25, 26). Likewise, thymocytes derive a large portion of their energy from glutamine oxidation, especially after mitogenic stimulation ( 27).

Under physiologic conditions, glutamine oxidation can account for as much as one-third of the cellular ATP production in cultured cells of many types ( 28, 29 and 30),

and relative glutamine oxidation increases at lower glucose levels ( 24, 31). Cells survive and even grow in glucose-free media containing sufficient glutamine and

nucleic acid precursors (32, 33). Thus, glutamine is an important source of cellular energy. However, little is known about what determines the rate of glutamine use or

oxidation for a cell or tissue type.



GLUTAMINE IN CATABOLIC STATES

Interorgan Glutamine Transfer

During catabolic states, a combination of increased glutamine use and decreased nutrient uptake creates a glutamine demand that is met primarily by increased

glutamine efflux from lung and muscle tissue (see also Chapter 98). Although plasma glutamine levels are usually maintained in septic rats and injured patients ( 34,

35 and 36), lung and muscle glutamine stores can be rapidly depleted after injury or infection ( 34, 37, 38, 39 and 40). Muscle is a major producer of glutamine during

normal and catabolic states. In the normal postabsorptive 200-g rat, glutamine is released from the hindquarter at a rate of approximately 0.4 mmol/min ( 35). Given

that skeletal muscle glutamine concentrations are approximately 6 to 8 mmol/L intracellular water ( 41), even a normal rate of release cannot be sustained for long

without de novo glutamine synthesis. During catabolic states the efflux of glutamine from rat and dog muscle increases markedly ( 34, 37, 42, 43), and increased net

production rate must eventually compensate for this heightened release. The muscle can maintain a rapid efflux of glutamine in part by increasing the rate of

proteolysis while decreasing the rate of protein synthesis ( 44). This increases the intracellular pool of amino acids for export and for production of glutamine, which

can be derived from a large number of amino acids through their conversion to glutamate, either directly or through a-ketoglutarate ( Fig. 35.1). Thus, any amino acid

that can feed into the TCA cycle can support glutamine production. Glutamate can then be converted to glutamine by the action of glutamine synthetase (GS).

Muscle glutamine levels may be an indication of severe catabolic stress. Investigators have shown a significant correlation between survival in septic patients and

skeletal muscle glutamine stores ( 45). In addition, an association between reduction of plasma and muscle glutamine levels and disease severity has been reported

for pancreatitis patients ( 46). Presumably, muscle glutamine stores and glutamine supplied by muscle proteolysis do not suffice to meet demand for glutamine in some