THE JOURNAL OF BIOLOGICAL CHEMISTRY: The Biochemical Behavior of LeadL I. Influence of Calcium, Phosphorus, and Vitamin D on Lead in Blood and Bone

Background: Current clinical methods for folate give different results and cannot measure the various forms of folate. We developed an isotope-dilution tandem mass spectrometric method coupled to liquid chromatography (LC/MS/MS) as a candidate reference method for 5-methyltetrahydrofolic acid (5MeTHF), 5-formyltetrahydrofolic acid (5FoTHF), and folic acid (FA) in human serum. Methods: We quantitatively isolated folates from 275 μL of serum with a phenyl solid-phase extraction cartridge, then detected and quantified them in stabilized serum extracts by positive-ion electrospray ionization LC/MS/MS. We used an isocratic mobile phase of acetic acid in organic solvent on a C 8 analytical column. 13 C-labeled folates were used as internal standards. Results: Limits of detection in serum were 0.13 (5MeTHF), 0.05 (5FoTHF), and 0.07 (FA) nmol/L. Within- and between-run imprecision (CV) was 0.5 nmol/L, and 2.0 nmol/L. Total folate (TFOL) concentrations determined by competitive protein binding radioassay were ∼9% lower than results obtained with LC/MS/MS. The microbiologic assay gave ∼15% higher TFOL results with FA calibrator and no difference with 5MeTHF calibrator. The mean (SD) [range] TFOL in 42 sera was 35.5 (17.8) [6.5–75.6] nmol/L. Thirty-two samples with TFOL 50 nmol/L had, on average, 81.7% 5MeTHF, 15.7% FA, and 2.5% 5FoTHF. Conclusions: This stable-isotope-dilution LC/MS/MS method can quantify 5MeTHF, 5FoTHF, and FA in serum. Currently used clinical assays agree with this candidate reference method.

/pdf/determination-of-folate-vitamers-in-human-serum-by-stable-22kl2598j7.pdf

Determination of Folate Vitamers in Human Serum by Stable-Isotope-Dilution Tandem Mass Spectrometry and Comparison with Radioassay and Microbiologic Assay

Food composition tables were studied from nine European countries participating in the European Prospective Investigation into Cancer and Nutrition (EPIC): Denmark, France, Germany, Greece, Great Britain, Italy, The Netherlands, Spain and Sweden. They were compared from the point view of availability, definition, analytical methods, and mode of expression of the nutrients of interest for EPIC, and it was seen that most of the nutrients in the tables are analysed and expressed in a compatible way. For some nutrients, however, common methods and definitions (folate, dietary fibre), or modes of expression (energy, protein, carbohydrates, carotenes, vitamin A and E) have not yet been agreed upon, so values are not comparable. For vitamin C a wide range of values are found due to the high natural variation in foods. For compiled tables, an additional problem is the use of several sources which may mean that the nutritional values are not comparable within the same table; and these values cannot be converted if the source is not stated. In addition, some tables were compiled using food composition values produced over 20 years ago with outdated analytical methods. In view of the inconsistent values for some nutrients and due to the large amount of foods reported within EPIC, it was concluded that standardised food composition tables have to be developed for the nine European countries involved in EPIC in order to provide comparable nutrient intake data.

Comparison of nutrients in the food composition tables available in the nine European countries participating in EPIC. European Prospective Investigation into Cancer and Nutrition.

Publisher Summary This chapter reviews the current knowledge concerning: (1) the inherent chemical properties of folates and methods of synthesis of experimentally important forms of the vitamin, (2) analytical methods for measurement of folates in foods and other biological materials, (3) the stability and chemical behavior of folates in foods, (4) the folate requirement of humans and the nutritional adequacy of human diets, ( 5 ) methods of assessment of the bioavailability of the vitamin, and (6) the mechanism of folate absorption and factors affecting the bioavailability of dietary folate. The application of these methods in the near future will provide significant new information with respect to the identity, content, and bioavailability of folates in foods. Although the chemical behavior of folates in foods is reasonably well characterized, common methods of food preparation and processing often yield inevitable losses of folate by oxidative or other modes of degradation or by aqueous extraction.

Chemical and nutritional aspects of folate research: analytical procedures, methods of folate synthesis, stability, and bioavailability of dietary folates.

Folates and its derivatives occur as polyglutamates in nature. The multiplicity of forms and the generally low levels in foods makes quantitative analysis of folate a difficult task. The assay of folates from foods generally involves three steps: liberation of folates from the cellular matrix; deconjugation from the polyglutamate to the mono and di-glutamate forms; and the detection of the biological activity or chemical concentration of the resulting folates. The detection methods used are the microbiological assay relying on the turbidimetric bacterial growth of Lactobacillus rhamnosus which is by far the most commonly used method; the HPLC and LC/MS techniques and bio-specific procedures. This review attempts to describe the methods along with the merits and demerits of using each of these methods.

Folate: Methods of analysis

1. The response of Lactobacillus casei was measured for a number of the monoglutamyl forms of folate derivatives. 2. At the concentrations of folate commonly used in the assay of folate vitamin in foods the response of L. casei to folic acid, (pteroylglutamic acid) and 5-formyl-tetrahydrofolic acid was similar, but 5-methyl-tetrahydrofolic acid gave as little as half the response of folic acid. 3. The response was modified by altering pH but not by concentration of ascorbate. 4. These results have implications for the assays of foods for folate where mixtures of folate derivatives are present. 5. A modified procedure is suggested in which the monoglutamates give similar responses.

/pdf/studies-on-the-response-of-lactobacillus-casei-to-different-486cfrmpah.pdf

Studies on the response of Lactobacillus casei to different folate monoglutamates.

1 Folate was measured microbiologically with Lactobacillus casei in extracts from a range of different foods at two incubation pH values, 61 and 68 2 The values for folate content obtained at pH 62 were, in several instances, considerably higher than at pH 68 The 'positive drift' seen in the values for dilutions incubated at pH 68 were absent from results at pH 62 3 A comparison was made of the ability of the two main sources (hog-kidney and chicken pancreas) of deconjugase enzyme to produce measurable folate and the hog-kidney enzyme was shown to produce consistently higher values 4 The results presented here will have significance for studies of folate intake using calculations from food composition tables, in the light of the apparent discrepancy between calculated folate intake and the recommended daily allowances for the UK

/pdf/studies-on-the-response-of-lactobacillus-casei-to-folate-3jjm3vezhh.pdf

Studies on the response of Lactobacillus casei to folate vitamin in foods.

The comparative and absolute growth response of Lactobacillus casei was measured nephelometrically for time periods of 17-23 h in the microbiological assay of folic acid and 5-methyl-tetrahydrofolic acid at concentrations of 0-8 ng/10 ml basal medium at pH 6.8. At concentrations of 0-1 ng/10 ml the comparative growth response to 5-methyl-tetrahydrofolic acid was markedly depressed whereas growth was the same at 2 ng/10 ml and above. Comparative growth was unaffected by the length of assay incubation, depressed growth being due to differences in log-phase growth rates with the rate-plot for 5-methyl-tetrahydrofolic acid being sigmoidal and for folic acid being a rectangular hyperbola with linearity only in the 0-1 ng/10 ml range. The reciprocal rate-plot for folic acid was linear whereas that for 5-methyl-tetrahydrofolic acid was coincidental only in part, giving rise to the same estimate of maximum velocity and substrate concentration for half-maximum velocity, with the exhibition of a strong threshold at low concentration. A previous observation (Phillips & Wright, 1982) that the L. casei growth response to 5-methyl-tetrahydrofolic acid may be significantly less than that to folic acid is confirmed as is the long-established view that the response to both folates may be equal. In the light of current knowledge regarding folate-binding, transport and metabolism by L. casei, it is argued that the intracellular oxidation of 5-methyl-tetrahydrofolic acid to 5, 10-methylene-tetrahydrofolic acid is a rate-limiting step at low substrate concentrations, subsequently giving rise to a threshold growth response peculiar to 5-methyl-tetrahydrofolic acid.(ABSTRACT TRUNCATED AT 250 WORDS)

/pdf/the-threshold-growth-response-of-lactobacillus-casei-to-5-51ewr3dl0j.pdf

D. R. Phillips

Papers

Studies on the response of Lactobacillus casei to different folate monoglutamates.

Studies on the response of Lactobacillus casei to folate vitamin in foods.

The threshold growth response of Lactobacillus casei to 5-methyl-tetrahydrofolic acid: implications for folate assays.