Folate bioavailability from foods rich in folates assessed in a short term human study using stable isotope dilution assays.

Different sources of folate may have different bioavailability and hence may impact the standard definition of folate equivalents. In order to examine this, a short term human study was undertaken to evaluate the relative native folate bioavailabilities from spinach, Camembert cheese and wheat germs compared to pteroylmonoglutamic acid as the reference dose. The study had a single-centre, randomised, four-treatment, four-period, four-sequence, cross-over design, i.e. the four (food) items to be tested (referred to as treatments) were administered in sequences according to the Latin square, so that each experimental treatment occurred only once within each sequence and once within each study period. Each of the 24 subjects received the four experimental items separated by a 14-day equilibrium phase and received a pteroylmonoglutamic acid supplement for 14 days before the first testing and between the testings for saturation of body pools. Folates in test foods, plasma and urine samples were determined by stable isotope dilution assays, and in urine and plasma, the concentrations of 5-methyltetrahydrofolate were evaluated. Standard non-compartmental methods were applied to determine the biokinetic parameters C(max), t(max) and AUC from baseline corrected 5-methyltetrahydrofolate concentrations within the interval from 0 to 12 hours. The variability of AUC and C(max) was moderate for spinach and oral solution of pteroylmonoglutamic acid but high for Camembert cheese and very high for wheat germs. The median t(max) was lowest for spinach, though t(max) showed a high variability among all treatments. When comparing the ratio estimates of AUC and C(max) for the different test foods, highest bioavailability was found for spinach followed by that for wheat germs and Camembert cheese. The results underline the dependence of folate bioavailability on the type of food ingested. Therefore, the general assumption of 50% bioavailability as the rationale behind the definition of folate equivalents has to be questioned and requires further investigation.

Folate-status response to a controlled folate intake in nonpregnant, pregnant, and lactating women.

BACKGROUND: Folate dose-response studies in women of childbearing age who consumed a folic acid (FA)-containing multivitamin in the era of FA fortification are lacking. OBJECTIVE: We sought to investigate folate-status response to a known folate dose comprising an FA-containing prenatal supplement (750 µg/d) plus natural food folate (400 µg/d) in third-trimester pregnant women, lactating women 5-15 wk postpartum, and nonpregnant women. DESIGN: Pregnant (n = 26), lactating (n = 28), and nonpregnant (n = 21) women consumed the study folate dose under controlled intake conditions for 10-12 wk. Blood, urine, and breast milk were collected at baseline, study midpoint, and study end. RESULTS: Study-end serum total folate concentrations averaged ~30 ng/mL and did not differ by physiologic group (P = 0.876). Study-end urinary folate excretion represented ~9-43% of total folate intake and ranged from 100 to 500 µg/d. Third-trimester pregnant women excreted less urinary folate than did lactating (P = 0.075) and nonpregnant (P < 0.001) women. Lactating women excreted less (P < 0.001) urinary FA than did nonpregnant women. Breast-milk total folate concentrations remained constant (P = 0.244; 61.8 ng/mL at study end), whereas breast-milk FA concentrations increased (P = 0.003) to 24.1 ng/mL at study end. CONCLUSIONS: The consumption of the study folate dose yielded a supranutritional folate status regardless of the physiologic state. Based on urinary folate excretion, folate use was greatest to least: pregnant > lactating > nonpregnant women. Breast-milk folate species were responsive to maternal folate intake, and FA made up ~40% of breast-milk total folate at study end. These findings warrant revisiting prenatal supplement FA formulation in populations exposed to FA-fortification programs.

Analysis of 5-methyltetrahydrofolate in human blood, serum and urine by on-line coupling of capillary isotachophoresis and zone electrophoresis.

The predominant circulating folate coenzyme in plasma/serum, 5-methyltetrahydrofolate (5-MTHF) was determined in human blood, serum and urine using a method based on the hyphenation of capillary ITP and zone electrophoresis. Measurements were done with a commercially available instrument for capillary isotachophoresis equipped with a column-switching system. The choice of electrolytes was limited by the instability of 5-MTHF and volatility of electrolytes for the potential coupling of the instrumentation with MS detector. To get an insight into the separability of individual sample components in an isotachophoretic analysis, we constructed zone existence diagrams for isotachophoretic electrolyte systems having a leading electrolyte composed of acetate and ammonium of pH 4.5 and 7.0, hydrocarbonate and ammonium, pH 7.8, chloride and ammonium, pH 5.6, and chloride and creatinine, pH 5.0, with hydroxide ion as the terminator. For isotachophoretic preseparation, the non-volatile leading electrolyte with good buffering capacity composed of 1 × 10(-2) M HCl and 2.5 × 10(-2) M creatinine, pH 5.0, and terminating electrolyte composed of 1 × 10(-2) M MES was selected as the most suitable. The optimum BGE for CZE analysis from the standpoint of analyte stability, separability and volatility for MS coupling was 1 × 10(-2) M acetate with 3.5 × 10(-2) M ammonium, pH 4.5. Using this combination of electrolytes, LODs reached with optical detection at 220 nm were 1.6 × 10(-7) M in human blood, 1.1 × 10(-7) M in human serum and 4.7 × 10(-6) M in human urine. Estimated content of 5-MTHF in blood and serum samples of women following oral daily administration of 0.8 mg of folic acid was 1.2 × 10(-5) and 5.8 × 10(-6) M, respectively.

Determination of 5-methyltetrahydrofolate (13C-labeled and unlabeled) in human plasma and urine by combined liquid chromatography mass spectrometry.

The association of folates with the prevention of neural tube defects and reduced risk of other chronic diseases has stimulated interest in the development of techniques for the study of their bioavailability in humans. Stable isotope protocols differentiate between oral and/or intravenous test doses of folate and natural levels of folate already present in the body. An liquid chromatography/mass spectrometry (LC/MS) procedure is described that has been validated for the determination of [13C]5-methyltetrahydropteroyl monoglutamic acid ([13C]5-CH3H4PteGlu) in plasma and urine, following oral dosing of volunteers with different labeled folates. Folate binding protein affinity columns were used for sample purification prior to LC/MS determination. Chromatographic separation was achieved using a Superspher 100RP18 (4 microm) column and mobile phase of 0.1 mol/L acetic acid (pH 3.3):acetonitrile (90:10; 250 microL/min). Selected ion monitoring was conducted on the [M-H](-) ion: m/z 458 and 459 for analyzing 5-CH3H4PteGlu; m/z 464 [M+6-H](-) to determine 5-CH3H4PteGlu derived from the label dose; m/z 444 for analysis of 2H4PteGlu internal standard, and m/z 446 and 478 to confirm that there was no direct absorption of unmetabolized compounds. Calibration was linear over the range 0-9 x 10(-9) mol/L; the limits of detection and quantification were 0.2 x 10(-9) and 0.55 x 10(-9) mol/L, respectively. The mean coefficient of variation of the ratios (m/z 463/458) was 7.4%. The method has potential applications for other key folates involved in one-carbon metabolism.

Conversion of 5-formyltetrahydrofolic acid to 5-methyltetrahydrofolic acid is unimpaired in folate-adequate persons homozygous for the C677T mutation in the methylenetetrahydrofolate reductase gene.

Methylenetetrahydrofolate reductase (MTHFR) catalyzes the synthesis of 5-methyltetrahydrofolic acid (5-CH(3)-H(4) folic acid), the methyl donor for the formation of methionine from homocysteine. A common C677T transition in the MTHFR gene results in a variant with a lower specific activity and a greater sensitivity to heat than the normal enzyme, as measured in vitro. This study was undertaken to determine the capacity of homozygotes for the MTHFR C677T transition to convert 5-formyltetrahydrofolic acid (5-HCO-H(4) folic acid) to 5-CH(3)-H(4) folic acid, a process that requires the action of MTHFR. Six subjects homozygous for the C677T transition (T/T) and 6 subjects with wild-type MTHFR (C/C) were given a 5-mg oral dose of (6R:,S:)-5-HCO-H(4) folic acid. Plasma and urine were analyzed for 5-CH(3)-H(4) folic acid concentrations using affinity/HPLC coupled with fluorescence or UV detection. The mean areas under the curves created by the rise and fall of plasma 5-CH(3)-H(4) folic acid after the oral dose did not differ between the two genotypes, 424.5 +/- 140.3 (T/T) vs. 424.1+/- 202.4 h.nmol/L (C/C). There also was no significant difference in the mean cumulative 7-h urinary excretion of 5-CH(3)-H(4) folic acid between the T/T (2.5 +/- 1.4 micromol) and C/C (1.9 +/- 1.0 micromol) genotypes. Under the conditions employed, the conversion of oral 5-HCO-H(4) folic acid to 5-CH(3)-H(4) folic acid is not impaired in persons with the T/T MTHFR genotype. Possible reasons for these findings are discussed.

Kinetic profile of overall elimination of 5-methyltetrahydropteroylglutamate in rats.

The in vivo biliary and urinary excretion kinetics of 5-methyltetrahydropteroylglutamate (5-CH3-H4PteGlu) were studied in rats. During infusion at various rates (48-965 nmol. h-1. kg-1), the total body clearance (CLtotal) of 5-CH3-H4PteGlu could be attributed almost entirely to the sum of the biliary and urinary (CLurine,p) excretion clearances. After a 4-h infusion at the highest rate, the 5-CH3-H4PteGlu in the liver was 10 times higher than the endogenous level, whereas its polyglutamate form did not increase, suggesting that most of the infused 5-CH3-H4PteGlu is not incorporated in the polyglutamate pool but is eliminated by excretion. The parallel increase in CLtotal and CLurine,p with the increase in infusion rate might result from saturation of reabsorption at the renal proximal tubules, since the urinary excretion clearance, defined with respect to the kidney concentration, also increased while the biliary excretion clearance, defined with respect to the liver concentration, remained almost constant. We conclude that the hepatobiliary excretion is a relatively low-affinity process with a constant clearance, whereas the renal tubular reabsorption is saturated at higher plasma 5-CH3-H4PteGlu concentration ( approximately 0.5 microM). Urinary excretion becomes the predominant elimination route for any excess 5-CH3-H4PteGlu in the body.

Determination of folate transport pathways in cultured rat proximal tubule cells.

Deficiency of the vitamin folic acid has recently been linked with increased incidence of neural tube defects and of cardiovascular disease, through elevated plasma homocysteine levels. The kidney has an important role in conserving folate to counteract development of deficiency. Urinary folate excretion is regulated by the degree of reabsorption of folate by the proximal tubule cell. To evaluate an in vitro model for studies of the regulation of urinary folate excretion, the present studies examined the transport of 5-methyltetrahydrofolate (5-CH3-H4PteGlu), the primary form of folate in the glomerular filtrate, by normal rat proximal tubule (RPT) cells in confluent monolayer cultures. Specific binding of 5-CH3-H4PteGlu to the apical membrane was saturable (K(D) = 27 nM), but intracellular transport was not saturated up to 100 nM concentrations. 5-CH3-H4PteGlu transport was decreased 50% by concentrations of folic acid that completely blocked 5-CH3-H4PteGlu binding by the apical folate receptor. Probenecid (10 mM), an anion exchange (reduced folate carrier) inhibitor, reduced 5CH3-H4PteGlu transport by 50% without significantly affecting binding. Aspirin (3 mM) did not alter 5-CH3-H4PteGlu transport, but significantly enhanced the inhibition due to probenecid. Similarly, indomethacin (5 microM) potentiated the inhibition of 5-CH3-H4PteGlu transport by probenecid. These data suggest that RPT cells take up 5-CH3-H4PteGlu by both the folate receptor and the reduced folate carrier, implying a role for both pathways in regulating urinary folate excretion.

Potentiated decrease of plasma folate levels caused by the coadministration of folic acid in rats treated with methotrexate.

The decrease of plasma 5-methyltetrahydrofolic acid (5-MF) levels, postulated as an indicator of folate status, was studied following the administration of both methotrexate (MTX) alone and MTX with folic acid (FA) using rats as our experimental model. Blood and urine samples were serially collected over a 9 hr period after the administration of MTX, MTX with FA and from a control group to examine the plasma kinetics and the renal clearance of 5-MF. The pharmacokinetics of MTX and the plasma protein binding of 5-MF were also examined. The concentrations of these analytes were assayed using high performance liquid chromatography (HPLC). MTX administration produced decreased plasma 5-MF levels. This observed decrease was potentiated by oral FA administration, suggesting that the folate status was more severely altered by the coadministration of FA. The renal clearance of 5-MF also increased dose-dependently with FA (0.05-5 mg/kg) coadministration. The plasma protein binding of 5-MF was not affected by the FA administration, which indicates that the fraction of 5-MF that was filtered through the glomerular apparatus appeared to be unchanged. In addition, the pharmacokinetic profiles of MTX also appeared not to be affected by the addition of FA. We conclude that the inhibition of reabsorption of 5-MF in the renal tube by concurrent administration of MTX and FA must be one of the causal factors for the demonstrated decrease in the plasma 5-MF levels in rats.

Folate status response to controlled folate intake in pregnant women.

A metabolic study (84-d) was conducted to investigate the folate status response of pregnant subjects (n = 12) during their second trimester and nonpregnant controls (n = 12) to folate intakes approximating the current (400 microg/d) and former (800 microg/d) recommended dietary allowance (RDA). The overall goal of the study was to provide metabolic data to assist in the interpretation of the current RDA for folate. Subjects were fed a controlled diet containing 120 +/- 15 microg/d (mean +/- SD) folate and either 330 or 730 microg/d synthetic folic acid. Outcome variables between and within supplementation groups were compared at steady state. Serum folate was higher (P 0.05) were detected in serum folate between pregnant and nonpregnant women within the same supplementation group. Urinary 5-methyl-tetrahydrofolate excretion was greater (P 0.05) in 5-methyl-tetrahydrofolate excretion were detected between pregnant and nonpregnant women within supplementation groups. Differences (P

Folate synthesized by bacteria in the human upper small intestine is assimilated by the host.

BACKGROUND & AIMS: Some intestinal flora are known to synthesize folate. The aim of this study was to determine whether folate synthesized by small intestinal flora is assimilated by the human host. METHODS: Subjects with atrophic gastritis and healthy volunteers were studied before and after omeprazole administration. A double-lumen perfusion tube was placed in the duodenum. 3H-labeled P-aminobenzoic acid, a precursor substrate for bacterial folate synthesis, was perfused. Downstream intestinal aspirates and a 48-hour urine collection were obtained. RESULTS: Atrophic gastritis and omeprazole administration were associated with increases in duodenal pH and in small intestinal flora. Bacterially synthesized folates were isolated from the intestinal aspirates. Tritiated 5-methyltetrahydrofolate, a major metabolite of folate, was isolated from the urine of omeprazole-treated subjects in greater quantities than from drug-free subjects (P<0.01); the quantity of tritiated 5-methyltetrahydrofolate in the urine of the subjects with atrophic gastritis was similarly elevated. CONCLUSIONS: (1) Mild bacterial overgrowth caused by atrophic gastritis and administration of omeprazole are associated with de novo folate synthesis in the lumen of the small intestine; (2) the human host absorbs and uses some of these folates; and (3) the contribution to folate nutriture from this source remains unclear.

Folate transport pathways regulate urinary excretion of 5-methyltetrahydrofolate in isolated perfused rat kidney.

The reabsorption of 5-methyltetrahydrofolic acid (5-CH3-H4PteGlu) by the renal proximal tubule has an important role in the maintenance of plasma folate concentrations. However, the mechanism by which this vitamin traverses the renal epithelium remains to be determined. Studies in cultured cells have suggested that the folate receptor in association with a probenecid-sensitive anion carrier may be involved in the transmembrane transport of the vitamin. Because 5-CH3-H4PteGlu is reabsorbed and metabolized in the isolated perfused rat kidney (IPRK) in a smaller manner to in vivo models, the IPRK was used to evaluate pathways involved in folate reabsorption. Reabsorption of 5-CH3-H4PteGlu could not be saturated in the isolated perfused rat kidney, even at concentrations up to 2 mumol/L. Folic acid (PteGlu) was used as a competitive inhibitor of FR-dependent reabsorption of 5-CH3-H4PteGlu. When 5-CH3-H4PteGlu was maintained at 1 nmol/L (a concentration at which receptor-mediated transport should be maximal), PteGlu (up to 100 nmol/L) had no effect on reabsorption. The addition of probenecid (1 mmol/L) did not affect the reabsorption of 5-CH3-H4PteGlu but inhibited the fractional excretion of the anion para-aminohippurate. Probenecid also inhibited the urinary excretion of 5-CH3-H4PteGlu metabolites, indicating that reabsorbed 5-CH3-H4PteGlu was metabolized to products that were subsequently secreted into the urine by anion exchange pathways. The physiological importance of a folate receptor-mediated reabsorption of 5-CH3-H4PteGlu appears to be minor in the isolated perfused rat kidney, whereas nonspecific pathways appear to play a major role in the renal folate reabsorption.

Clinical pharmacokinetics of the antipurine antifolate (6R)-5,10- dideaza-5,6,7,8-tetrahydrofolic acid (Lometrexol) administered with an oral folic acid supplement.

(6R)-5,10-Dideaza-5,6,7,8-tetrahydrofolic acid (lometrexol) is an antipurine antifolate which selectively inhibits glycinamide ribonucleotide formyltransferase. Lometrexol pharmacokinetics were evaluated in 17 patients (32 courses) as part of a Phase I study in which folic acid supplementation was used to improve tolerance to the drug, its clinical utility being previously limited by severe cumulative toxicity. Lometrexol was administered as an i.v. bolus every 4 weeks at a starting dose of 12 mg/m2, with subsequent interpatient dose escalation to 16, 30, and 45 mg/m2. p.o. folic acid (5 mg/day) was given for 7 days before and 7 days after lometrexol administration. The disposition of total lometrexol in plasma was best described by a biexponential model for data acquired up to 12 h after drug administration, although triexponential plasma pharmacokinetics were often found to give a more adequate description when data were available at later time intervals (24 h and greater). Mean plasma half-lives (+ SD) for model-dependent analysis were t1/2alpha 19 +/- 7 min, t1/2beta 256 +/- 96 min, and t1/2gamma (where measurable) 1170 +/- 435 min. Lometrexol area under plasma concentration versus time curve was proportional to the dose administered. Moderate plasma protein binding of lometrexol was evident (78 +/- 3%) with an inverse linear relationship between fraction of unbound lometrexol and the concentration of serum albumin. The volume of distribution of lometrexol at steady state was between 4.7 and 15.8 l/m2. Renal elimination of lometrexol, studied in 19 patients (21 courses), was considerable, accounting for 56 +/- 17% of the total dose administered within 6 h of treatment, and 85 +/- 16% within 24 h of treatment. These recoveries of unchanged lometrexol indicate that the drug does not appear to undergo appreciable systemic metabolism at the range of concentrations studied. Lometrexol pharmacokinetics were also examined in seven patients who received 45 or 60 mg/m2 lometrexol as part of a separate study of the drug given with folinic acid rescue 5-7 days after treatment. No marked differences were evident in lometrexol plasma half-lives, plasma clearance, or the extent of plasma protein binding, indicating that there is not a pronounced pharmacokinetic interaction between lometrexol and folic acid.

Highly sensitive coupled-column high-performance liquid chromatographic method for the separation and quantitation of the diastereomers of leucovorin and 5-methyltetrahydrofolate in serum and urine.

A column-switching chiral HPLC assay was developed that allows the separation and quantitation of the diastereomers of leucovorin (LV, 5-formyltetrahydrofolic acid) and its metabolite 5-methyltetrahydrofolate (METHF) in serum and urine by means of fluorescence detection. The analysis procedure consists of an on-line concentration of the folates in the HPLC system which is followed by the elution and separation of folates on an achiral 3-microns Microbore C18 column in (6R,S)-LV and (6R,S)-METHF. (6R,S)-LV and (6R,S)-METHF are subsequently transferred on-line onto a chiral 7-microns bovine serum albumin column through a Rheodyne valve system and are separated into their diastereometers. Time of analysis is 70 min. Detection limit is 5 ng/ml for each diastereometer. The within-day variation ranges between 3.2 and 15.8% in relation to the measured concentration. Between-day variation is 4.4-12.1% for a concentration of 100 ng/ml for each diastereometer. (6R,S)-LV and (6S)-LV pharmacokinetics were assessed by analyzing serum and urine samples of four-healthy volunteers.

Specific and sensitive high-performance liquid chromatographic method with fluorescence detection for measurement of lometrexol and its polyglutamates in biologic samples.

A reversed-phase high-performance liquid chromatographic (HPLC) assay is described for the quantitative determination of lometrexol in biological samples; the assay is rapid, simple, specific, and highly sensitive. The method requires the dissociation of lometrexol from folate-binding proteins present in blood and formation of a fluorescent oxidized derivative of the compound. The dissociation of lometrexol from folate-binding proteins was achieved by acidification to pH 3.5 using ammonium formate, followed by serum protein precipitation with perchloric acid. The protein-free lometrexol was subsequently oxidized by MnO2 at 90 degrees C for 10 min. Chromatographic separation of lometrexol without interference was achieved on a C18 reversed-phase column with a convex gradient, using acetonitrile-0.1% ammonium formate, pH 7.0, as the mobile phase. In human serum and urine the calibration curve was linear between 5 and 300 nM. The lower limit of quantification was 5 nM. The method has been applied successfully to measure serum and urinary levels of lometrexol in patients.

5-methyltetrahydrofolate urinary excretion: modeling by cultured human kidney cells.

Initial attempts to model urinary folate reabsorption using cultures of HPT cells on porous filter inserts produced disappointing results in that large amounts of 5M were transported across the epithelial monolayer in a nonspecific manner. Since the impermeable molecule inulin was also transported, there apparently existed a significant leakage pathway in the way that the cultured cells were used for transport studies. 5M was bound to the AP membrane and taken up into the HPT cell by specific processes, while inulin was excluded, suggesting that the HPT cells were nevertheless operating functionally. TER values from cultured HPT cells plateaued at a high level when cells became confluent, suggesting an epithelial layer with functional tight junctions. However, when growth media were removed and replaced with transport buffers, there was an immediate loss of TER that fully recovered if the transport buffers were preincubated for 60 min. Under these conditions, transport studies showed the expected results--no movement of inulin through the cell layer and much reduced transfer of folate through the paracellular pathway. These results suggest that a transient opening of tight junctions occurs when growth media are replaced with biological buffers (or with fresh growth media), but that recovery of tight junction function occurs with time.

Folic acid, 5-methyl-tetrahydrofolate and 5-formyl-tetrahydrofolate exhibit equivalent intestinal absorption, metabolism and in vivo kinetics in rats.

The intestinal absorption and in vivo kinetics of (6S)-[3H]-5-methyl-tetrahydrofolate (5-methyl-H4folate), (6S)-[3H]-5-formyl-H4folate and [3H]folic acid were investigated to determine whether inherent differences exist in the overall bioavailability of these folates in rats. Adult rats (n = 9 per group) were given an intragastric dose of the appropriate folate (50 pmol/100 g body wt) in 50 mmol/L ascorbate (pH 7). Each compound underwent nearly complete absorption within 8 h, and there was no significant difference in the excretion kinetics in relation to the form of folate administered. A biphasic pattern of excretion was observed over the following 8 d. Both urine and feces were important excretory routes. The rapid phase of total isotopic excretion (urinary and fecal) exhibited a half time (t1/2) of 0.11-0.12 d, whereas the t1/2 of the slower phase was 13.4-15.9 d. Isotopic distributions and the pattern of labeled folates in urine and tissues were similar regardless of the form administered. These results indicate that the bioavailability of orally administered folic acid, 5-methyl-H4folate and 5-formyl-H4folate is equivalent in rats under the conditions of this study.

Renal folate absorption and the kidney folate binding protein. I. Urinary clearance studies.

The kidney possesses a high concentration of a folate binding protein (FBP) that resides primarily in the brush-border membrane (BBM) of the proximal tubular cells. To assess the possible involvement of this protein in renal conservation of folate we determined the urinary clearance, in rats, of three forms of folates with sharply different affinities for FBP. After single intravenous injections of 0.1 to 1.0-nmol doses of radioactive folates the urinary clearance based on radioisotope determination was in the sequence: folic acid less than 5-methyltetrahydrofolate (5-CH3 THF) much less than methotrexate. At higher doses the urinary folate clearance was increased and the differences between the three injected forms were narrowed and were no longer noticeable at 100-nmol doses. Under conditions of continuous infusion to attain plasma folate levels of 2.3-5.7 pmol/ml, the urinary clearance based on chromatographic analyses of plasma and urine after correction for plasma folate binding was 0.20 ml/min for folic acid, 0.37 ml/min for 5-CH3 THF, and 1.76 ml/min for methotrexate. These chromatographic analyses have also shown the presence in both plasma and urine of metabolites formed from infused folates. Metabolites found after infusion of folic acid include 5-CH3 THF with a urinary clearance of 0.3 ml/min and an unknown with a urinary clearance of 0.8 ml/min. The latter metabolite appears also to occur in plasma and urine after infusion of 5-CH3 THF. Infusion of methotrexate was associated with the appearance of a metabolite with a urinary clearance of 2.5 ml/min. This sequence of urinary clearance is in inverse order to the affinities of these three forms of folate for the kidney BBM FBP.(ABSTRACT TRUNCATED AT 250 WORDS)

Folate metabolism in man: the effect of malignant disease.

The metabolism of [2-14C]+[3', 5', 7, 9-3H] folic acid and [214C]+[3', 5', 7, 9-3H] 10-formylfolate was studied in hospital inpatients. Metabolites detected in the urine after folic acid feeding included the unchanged compound, other folates and a number of breakdown products, such as p-acetamidobenzoyl-L-glutamate and p-acetamidobenzoate. This confirms the existence of a folate catabolic pathway in man. Patients with malignant disease excreted less of the dose in urine, incorporated more into the reduced folate pool, and showed decreased catabolism of folate, when compared to controls. 10-Formylfolate was excreted largely unchanged, and appears not to be reduced by man. Also 10-formylfolate interfered with the reduction of folic acid given simultaneously.

The metabolism of 5-methyltetrahydropteroyl-L-glutamic acid and its oxidation products in the rat.

Folate metabolism in the rat was investigated using radiolabelled 5-methyltetrahydropteroylglutamate (5-CH3-H4PteGlu) and its oxidation products. 5-CH3-H4PteGlu is absorbed completely from the intestine, although in some preparations it is an equimolecular mixture of C-6 epimers, only one of which is naturally present in biological systems. The methyl group is incorporated into non-folate compounds, including methionine and creatine. No evidence was observed for the oxidation of the methyl group of 5-CH3-H4PteGlu to form other folate types. The tetrahydrofolate moiety of 5-CH3-H4PteGlu is metabolized in a similar manner to folic acid, forming formyl folates and tissue polyglutamates, and is catabolized by scission. The triazine oxidation product of 5-CH3-H4PteGlu is not metabolized by the rat or its gut microflora. 5-Methyl-5,6-dihydropteroylglutamate, however, is assimilated into the folate pool, but is substantially broken down by passage through the gut. The possible implication of this in scorbutic diets is discussed.