Evaluation of Anemia.

Anemia is a common problem in primary care. Classification based on mean cell volume narrows the differential diagnosis and directs testing. A marked macrocytosis is characteristic of vitamin B12 and folate deficiencies, certain medications, and primary bone marrow disorders. The three most common causes of microcytic anemia are iron deficiency, thalassemia trait, and anemia of inflammation. Additional laboratory testing is required for diagnosis. Determination of the rate of development of anemia and examination of a blood smear may provide diagnostic clues to guide more specialized testing. Diagnosis of iron, vitamin B12, or folate deficiency mandates determination of the underlying cause.

Evaluation of macrocytosis.

Macrocytosis, generally defined as a mean corpuscular volume greater than 100 fL, is frequently encountered when a complete blood count is performed. The most common etiologies are alcoholism, vitamin B12 and folate deficiencies, and medications. History and physical examination, vitamin B12 level, reticulocyte count, and a peripheral smear are helpful in delineating the underlying cause of macrocytosis. When the peripheral smear indicates megaloblastic anemia (demonstrated by macro-ovalocytes and hyper-segmented neutrophils), vitamin B12 or folate deficiency is the most likely cause. When the peripheral smear is non-megaloblastic, the reticulocyte count helps differentiate between drug or alcohol toxicity and hemolysis or hemorrhage. Of other possible etiologies, hypothyroidism, liver disease, and primary bone marrow dysplasias (including myelodysplasia and myeloproliferative disorders) are some of the more common causes.

Nitrous oxide and formiminoglutamic acid: excretion in surgical patients and anaesthetists.

We have investigated the possible toxicity of nitrous oxide on vitamin B12 and its sequelae upon folic acid metabolism using the urine formiminoglutamic acid excretion test, an index of the functional state of folate metabolism. Ten control subjects not exposed to nitrous oxide and five patients receiving limb surgery under local anaesthesia excreted normal amounts of formiminoglutamic acid in urine for 6 days. Fifty patients received nitrous oxide anaesthesia for similar surgery and, of these, 22 had a dose-dependent increase in excretion on the first 2 days after operation. There were large individual variations. Exposure to 70% nitrous oxide appeared to cause abnormal metabolism of folate when exposure was greater than 90 min. Ten anaesthetists demonstrated normal excretion of formiminoglutamic acid; their exposure to nitrous oxide was typical of that in other studies of theatre environmental pollution.

Experimental maternal and neonatal folate status relationships in nonhuman primates.

The influence of maternal dietary folic acid intake on folate status was studied in Cebus albifrons monkeys by feeding 10 or 250 micrograms/100 kcal dietary folic acid during pregnancy and 4 wk postpartum. Maternal, infant, and nonpregnant hematologic indices; blood and liver folate concentrations; and urinary formiminoglutamic acid excretion all varied with dietary folate intake and pregnancy status as did milk folate concentration in lactating dams. Maternal folate status, determined by plasma, red blood cell, and milk folate concentrations, as well as urinary formiminoglutamic acid excretion, all were correlated significantly with liver folate concentrations in neonates (r = 0.740, r = 0.919, r = 0.936, and r = -0.851, respectively). Results in these primates showed that neonatal folate status was related significantly to the dietary folate intake and folate status of the mother during pregnancy and lactation.

Folate requirement and metabolism in nonpregnant women.

Folate metabolism and requirements were studied in 10 adult nonpregnant women maintained for 92 d in a metabolic unit. After a folate depletion period of 28 d, the subjects received increasing supplements of folate from food items or as pteroylmonoglutamic acid (PGA). Plasma folate levels fell 60% during the depletion period and continued to fall until 200 micrograms/d of naturally occurring food folates were provided. Supplements of 300 micrograms/d of naturally occurring folates produced a small rise in plasma folate levels although erythrocyte folate levels continued to fall. Lymphocyte deoxyuridine suppression, neutrophil hypersegmentation, and other measurements related to folate metabolism were performed. When compared with PGA, dietary folates appeared to be no more than 50% available. A daily intake of 200-250 micrograms of dietary folates appears to meet the folate requirements of nonpregnant adult women whereas an intake of 300 micrograms/d provides an allowance for storage.

Folate deficiency in rats fed amino acid diets.

Growth rate, hematological changes, serum, erythrocyte and liver folate levels, and urinary excretion of formiminoglutamic acid (FIGLU) were measured in rats fed p-aminobenzoic acid (PABA)-free, sulfonamide-supplemented purified amino acid diets with and without folic acid and fiber, individually and in combination. Fiber had no effect on growth but was necessary to prevent mortality in the absence of folic acid. Folic acid did not affect growth during the first 40 days, but after this period rats failed to gain weight in the absence of folic acid. Although blood hematocrit was lower when the diet was devoid of fiber and folic acid and leucocyte counts were lower when the diets were devoid of folic acid, the hematological values remained within normal physiological limits for healthy rats of comparable age. FIGLU excretion was increased and serum, erythrocyte and liver folate levels were depressed when rats were fed folic acid-free diets for 28 days. Neutrophil hypersegmentation was clearly evident after 28 days of feeding the folic acid-free diets. The use of an amino acid diet without PABA but containing 5% fiber and 1% succinylsulfathiazole is a useful system to study chronic folic acid deficiency in the rat.

Reversible dementia and neuropathy associated with folate deficiency 16 years after partial gastrectomy.

A 57-year-old woman developed dementia and peripheral neuropathy 16 years after a partial gastrectomy (Billroth II). Serum cobalamin was 198 pmol/l (reference interval 150-550), and the vitamin B12 absorption test (Schilling) showed decreased absorption (1.7% without and 2.2% with intrinsic factor). In spite of 20 months' therapy with vitamin B12, the neurological symptoms progressed. Folate deficiency was suggested by a very low erythrocyte folate and a slightly abnormal FIGLU test. There were no other signs of general malabsorption. A few months' treatment with folic acid significantly improved the massive neurological manifestations which were verified neurophysiologically as well as histologically. A common role of vitamin B12 and folate in the development of neuropathy is suggested.

Interrelationships between Vitamin B12 and folic acid in myelomatosis: cobalamin coenzyme and tetrahydrofolic acid function.

Cobalamin and folate metabolism was investigated in 43 patients with myelomatosis, in 8 control subjects of similar age and 22 younger controls. Plasma total cobalamin was lower in myeloma patients than in either of the control groups and methylcobalamin (Me-Cbl) was disproportionately reduced. Erythrocyte levels of total cobalamin were very similar in patients and elderly controls but were half the levels in younger controls. Erythrocyte levels of Me-Cbl were slightly higher in patients than in the dlderly controls. FIGLU excretion after L-histidine was elevated in 53% of the patients but values did not correlate with serum or erythrocyte folate or with plasma total cobalamin. FIGLU excretion decreased after DL-methionine or Me-Cbl only in patients whose FIGLU excretion was initially high. The results are discussed in the light of the 'methylfolate trap hypothesis' and suggest that some patients with myelomatosis have insufficient activity of methionine synthetase to meet the additional metabolic demand for one carbon compounds.

The effects of chronic drug administration on hepatic enzyme induction and folate metabolism.

1 Patients on prolonged treatment with anticonvulsant and phenothiazine drugs exhibited lower than normal concentrations of folate in serum and erythrocytes, and showed increased urinary FIGLU excretion after histidine loading; urinary excretion of D-glucaric acid was also increased suggesting induction of the hepatic microsomal enzymes. 2 Folate deficiency by enzyme-inducing drugs was seen to be determined more by the duration of therapy than by the nature of the drugs. Excretion of FIGLU was increased by 70% by 2-5 years of treatment with anticonvulsant, phenothiazine or tricyclic drugs, and by 200% after 6 or more years. 3 Hepatic microsomal enzyme induction, as measured by D-glucaric acid excretion, was greatest after 2-5 years treatment. 4 It is suggested that the increased requirements for folate, resulting from microsomal enzyme induction, lead to folate deficiency and this subsequently limits enzyme induction, leading to adverse drug side-affects. 5 The dietary folate of hospitalized patients would seem to be generally inadequate for patients on long term treatment with enzyme-inducing drugs.

Fromino-glutamic acid (Figlu) excretion and abortion.

PIP: The relationship between excretion of forminoglutamic acid (figlu) in the urine and spontaneous abortion in the first and second trimesters of pregnancy was investigated in 135 nonanemic women after spontaneous abortion and curettage. The 135 were subdivided into Group A, 95 cases with no known reason for spontaneous abortion, and Group B, 40 women in whom probable abortion cause was known; Group C was control. In Group A, figlu was detected in urine of 40 (42.1%). In Group B, where abortion causes ranged from diabetes, to toxoplasmosis, to cervical incompetence, figlu was positive in only 2 (5%) cases. 3/35 controls (8.6%) were positive for figlu. The percentage of positive figlu findings in Group A was significantly higher when compared with Group B (P .001) and with Group C (P .001). In Group A, the positive figlu excretion was statistically unrelated to previous abortions (P .05), previous pregnancies (P .05), or age groups (P .05). Further, when the 40 Group B cases and the 35 Group C cases were compared with the above, there was no change in the results. If increased figlu excretion can be considered an index of folic acid deficiency, then it is possible that this deficiency may be an etiological factor in cases of spontaneous abortion.^ieng

Urinary formiminoglutamate in man. Normal values related to sex and age. Effects of low calorie intake and alcohol consumption.

A modification of the enzymic method of Tabor and Wyngarden for formiminoglutamate (FIGLU) estimation in urine is described. Alkaline degradation of FIGLU in blanks enhanced the sensitivity of the method. FIGLU excretion in 94 healthy persons indicated a sex and age dependence. Short term calorie restriction or fasting led to a decrease of FIGLU excretion while acute alcohol intoxication or chronic alcohol administration were accompanied by enhanced FIGLU excretion. It is suggested that decreased FIGLU excretion was due to histidine deficiency.

Hydantoin-5-propionic aciduria in folic acid nondependent formiminoglutamic aciduria observed in two siblings.

Hydantion-5-propionic acid was detected in massive amounts in the urine of two previously described sisters with folic acid nondependent formiminoglutamic aciduria. HPA was identified rigorously, e.g. by gas chromatography-mass spectrometry, and was measured quantitatively by selected ion monitoring (mass fragmentography) using deuterated HPA as internal standard. Before histidine loading, both girls with the postulated formiminotransferase deficiency excreted an amount of HPA more than 50 times greater than the control subjects. After histidine ingestion, HPA excretion was still doubled or tripled. With the exception of the father, the values for the other members of the family also markedly exceeded the normal range and were of the same order of magnitude as in folate deficiency. Measurement of HPA excretion in urine seems to be a valuable supplement or alternative to the enzymatic FIGu test for the detection of general or functional folate deficiency or impaired folate utilization and it will be indispensable for the detection of (as yet unknown) 4-imidazolone-5-propionic acid hydrolase deficiency.

Serum vitamin B12 levels in the aged.

In an attempt to throw light on the question of age-related variations in the normal blood content of cobalamin and on the frequency of deficiencies of antimegaloblastic nutriments in the elderly, 273 geriatric patients have been investigated. Low serum vitamin B12 values were found in one third of these patients, due to latent pernicious anaemia in five and malabsorption in seven cases, and probably caused by nutritional deficiency of folate or cobalamin in 78 cases. In that part of the series with apparently normal vitamin B12 levels, the mean value (379+/-14 pg/ml) was lower than the mean (456+/-20 pg/ml) for a younger control group. However, this cannot be taken as a sign of a physiological lowering of the cobalamin values with age, as nutritional deficiencies could not be ruled out in this part of the series. It is concluded that serum vitamin B12 assays should be performed rather liberally in the aged. Patients with nutritional deficiency of cobalamin or folate should be treated, even if frank megaloblastic anaemia is not present.

Investigation into diagnosis and treatment of cobalt deficiency in lambs.

The development of cobalt deficiency was studied in 30 Scottish Blackface lambs grazing pasture on a soil containing 0-17 ppm cobalt. By the end of an eight-week period 50 per cent of lambs were subjectively appraised as showing signs of cobalt deficiency. After a further period lasting four weeks, during which three groups of lambs were studied (one group, least affected by cobalt deficiency, acted as control, the second received a single cobalt bullet and the third received oral doses of 200 mg cobalt chloride at the beginning of the period and three weeks later). Mean urinary formiminoglutamic acid (FIGLU) concentrations were inversely related to serum vitamin B12 concentrations and increased from 0-08 to 0-20 mumole per ml in group 1, and decreased to virtually zero within one week of treatment in groups 2 and 3. The use of serum vitamin B12 and urinary FIGLU concentrations in the diagnosis of cobalt deficiency in sheep are discussed.

Metabolic studies of a family with massive formiminoglutamic aciduria.

We have described two siblings who excrete massive amounts (up to 3.89 mmol/24 hr) of N-formiminoglutamic acid (FIGLU) in their urine. This unusual compound was isolated from urine, purified, and firmly identified as FIGLU by combined gas chromatography-mass spectrometry. The patients presumably have a deficiency in activity of the hepatic enzyme, glutamate formiminotransferase, which carries out the fourth sequential step in the main pathway of histidine degradation. Unlike children reported previously with this disorder, our patients had normal serum folate levels, had no hematologic abnormalities, and were not mentally retarded. Very small amounts of FIGLU were present in the plasma of one of the patients, but FIGLU was not detectable in the cerebrospinal fluid of either patient. Administration of pharmacologic doses of folic or folinic acid produced a decrease in excretion of FIGLU in urine. Histidine loading caused a small and comparable urinary excretion of FIGLU in the children's parents and in control adult subjects.

[Radioanalytical methods of determination in vitro of folates and vitamin B 12: diagnostic use in disorders of absorption and utilization]. / Le metodiche radioanalitiche di dosaggio "in vitro" dei folati e della vitamina B12: utilità diagnostica nei difetti di assorbimento ed utilizazione

The blood concentrations of vitamin B12 and folate, which are very useful in diagnosis of megaloblastic anemia and of these factors' dificiencies, are actually measured by precise, rapid, and specific competitive binding radioassays. Futher clinical advantages can be reached with the application of other in vitro radioisotope techniques, such as radioassay of IF, of antibodies anti-IF, of transcobalamins, and of FABP (folic acid binding protein). The major impact of the vitamin B12, folates and other related radioassays has been to permit more Hospitals and laboratories to do these determinations, replacing the more time-consuming, relatively imprecise, and often artifactual microbiological assays.