Vitamin saturation test

Saturation and Intradermal Tests. Ascorbic acid saturation tests, based on observations that subjects with low tissue reserves excreted less of a test dose of vitamin C in the urine than subjects with adequate stores, did not distinguish between varying degrees of deficiency at the lower levels of nutrition. Relatively high plasma levels must be attained before the effect of the renal tubular reabsorption is surpassed and significant excretion occurs (see Section 5). 

A water soluble vitamin which cannot be synthesized by man and therefore has to be obtained from the diet. It is found extensively in vegetables and fruit, especially the citrus varieties. Since the vitamin is carried mainly in the leukocytes, its measurement in these cells gives some indication of the vitamin C status of the body. The ascorbic acid saturation test can also be used to assess the vitamin status. The biochemical role of the vitamin is obscure although it does seem to be required for collagen formation. Deficiency of the vitamin causes scurvy, the symptoms of which can be related to poor collagen formation. These include poor wound-healing, osteoporosis (due to bone matrix deficiency), a tendency to bleed (due to deficiences in the vascular walls) and anaemia. 

Another problem associated with saturation analysis is that abnormally low results may be obtained unless cyanide is present when the vitamin is freed from its binder. It appears that forms other than cyanocobalamin are difficult to separate completely from the binding protein. Early studies that failed to recognize this not infrequently found that results from patients with pernicious anemia gave negative values (R9). A recent study by Brown et al. (B6) examined the effect of varying the concentration of cyanide used in the test. They found that an excess of cyanide resulted in a significant increase in apparent vitamin B12 levels in sera from patients who were deficient in the vitamin, but it had little effect on sera from normal patients. They found the mean of 12 vitamin B12-deficient sera to be 49 ng/liter when 3 mg/liter of cyanide was used in the extraction mixture, 104 ng/liter in the presence of 30 mg/liter of cyanide, and 196 ng/liter when 300 mg/liter of cyanide was used. The authors emphasized that cyanide was necessary to convert all of the several forms of vitamin B12 present in serum to cyanocobalamin but warned that the concentration should not be greater than 5 mg/liter. 

In the early stages of reduced vitamin B12 absorption, the amount of vitamin B12 bound to TCII decreases rapidly (10-21 days) without any decrease in the total serum B12 level. The total vitamin B12 concentration begins to decline only when the saturation of TCII falls below 5%. Reduced saturation of TCII is thus one of the earliest indicators of reduced intake of vitamin B12 and is detectable in advance of clinical disease. This makes holo-TCII particularly valuable for use as a screening test in susceptible populations. Elevated serum and urine concentrations of homocysteine and MMA occur somewhat later and represent early evidence of cellular dysfunction that occurs when body (liver) stores have been greatly depleted. 

Schilling s test assesses the oral absorption of vitamin B12 and is used to diagnose pernicious anaemia. The patient is injected intramuscularly with non-labelled vitamin B12, to saturate body stores. An oral dose of vitamin B12 labelled with cobalt-58 is administered, followed by a second dose labelled with cobalt-57 bound to intrinsic factor. Prior saturation of body stores ensures any absorbed radiolabelled vitamin B12 is rapidly excreted in the urine. Urinary excretion of orally administered vitamin B12 is low in patients with pernicious anaemia due to poor absorption. Absorption is increased when it is administered with intrinsic factor. The ratio of cobalt-57 to cobalt-58 is thus raised in patients with pernicious anaemia. Intrinsic factor antibody testing is now generally used to diagnose pernicious anaemia, though the Schilling s test may occasionally be used. 

Inadequate saturation of enzymes with (vitamin-derived) coenzymes. This can be tested for three vitamins, using red blood cell enzymes thiamin (Section 6.5.3), riboflavin (Section 7.5.2), and vitamin Be (Section 9.5.3). 

Metabolic loading tests and the determination of enzyme saturation with cofactor measure the ability of an individual to meet his or her idiosyncratic requirements from a given intake, and, therefore, give a nearly absolute indication of nutritional status, without the need to refer to population reference ranges. A number of factors other than vitamin intake or adequacy can affect responses to metabolic loading tests. This is a particular problem with the tryptophan load test for vitamin Be nutritional status (Section 9.5.4) a number of drugs can have metabolic effects that resemble those seen in vitamin deficiency or depletion, whether or not they cause functional deficiency. 

Estimation of the vitamm Be requirements of infants presents a problem, and there is a clear need for further research. Human mUk, which must be assumed to be adequate for infant nutrition, provides only 2.5 to 3.5 //g of vitamin Be per g of protein-lower than the requirement for adults. Although their requirement for catabolism of amino acids may be lower than in adults (because they have net new protein synthesis), they must also increase their body content of the vitamin as they grow. Coburn (1994) noted that the requirement for growth in a number of animal species was less than that to maintain saturation of transaminases or rniriimum excretion of tryptophan metabolites after a test dose and was about 15 nmol per g of body weight gain across a range of species. 

It is relatively easy to assess the state of body reserves of vitamin C by measuring the excretion after a test dose. A subject whose reserves are saturated will excrete the whole of a test dose of500 mg of ascorbate over 6 hours. A more precise method involves repeating the loading test daily until complete excretion is achieved, thus giving an indication of how depleted the body stores were. 

The purpose of the rarely used Schilling urinary excretion test is to diagnose vitamin B12 deficiency anemia caused by a B12 absorption defect resulting from a lack of intrinsic factor (pernicious anemia). The patient first receives an oral dose of radiolabeled vitamin B12. Two hours later, the patient receives a large intramuscular dose of nonlabeled vitamin B12 to saturate plasma transport proteins. Any excess vitamin B12 that is not taken up by the transport proteins or stored in the liver will be excreted in the urine. A 24-hour urine collection is then measured for radioactivity. If sufficient gastrointestinal intrinsic factor is being produced, the radiolabeled B12 will be absorbed. 

Thiamine is excreted in the urine, the amount being dependent on dietary intake and the relative saturation of the tissue stores. Determination of thiamine excretion in the urine, especially after a test dose of thiamine has been administered, is one of the methods used in evaluating nutritive status relative to this vitamin. After intramuscular injection of 1 mg. of thiamine, persons who are adequately nourished excrete at least 100 fig. in the subsequent 4 hr., whereas patients with signs of thiamine deficiency usually excrete less than 50 /xg. during this period. Estimation of the concentration of thiamine in blood has also been used in nutritional appraisal. Mean... 

The absorption of vitamin B can be determined by the Schilling test. An oral dose of Co- or Co-labelled vitamin B is given with a parenteral flushing dose of 1 mg of non-radioactive vitamin to saturate body reserves, and the urinary excretion of radioactivity is followed as an index of absorption of the oral material. Normal subjects excrete 16—45% of the radioactivity over 24 hours, whereas patients lacking intrinsic factor excrete less than 5%. 

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